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Brain vs. Brawn: The Ultimate Showdown. Pin your opponent down for 3 seconds. Investigation Term Papers? After a long and painful season of grueling challenges, only two contestants remain in the competition, and their mission is to battle it out for the million-dollar prize! They are taken to a Colosseum to on marfan fight in the ultimate final challenge, which features numerous obstacles and the return of term papers a few old faces. Essay? One contestant wins the support of the entire the crowd, with one exception. In the end, one contestant wins the challenge and the money, and someone gets his long-awaited karma. Lightning arrogantly bullies Cameron before the final challenge. The episode begins after the events of term papers The Enchanted Franken-Forest, and analytical essay image, Cameron is disheartened that Zoey didn't make it to the finale. However, Lightning is criminal confident, because it will be much easier to win the million dollars due to Cameron's lack of athleticism. About? Cameron tells him that even though he isn't that strong, he still has as they call it game.
Lightning shows his game by lifting a log over his head and saying how he's got it in the bag. Cameron points out though that he could not win if the last challenge is an intellectual one, but Lightning points out criminal investigation papers, that past Total Drama finales have always been physical and states that he can crush Cameron like a grape. In the ap us history confessional, Cameron is shown not to be worried, but terrified. However, he mentions that he has come a long way and how he can do anything if he can beat Lightning. After he says that, Lightning scares him saying the same grape threat. Cameron says on the other hand if Lightning wins, he will still have a bubble to investigation stay in for the next sixteen years growing his skin back while eating his mom's sandwiches through a straw. Lightning screams the word grape making Cameron even more terrified.
He says to essay his mom if she's watching this to start pumping out the oxygen. Lightning is term papers then in the confessional saying while doing curl-ups at the same time, if he was a gentleman he would let Cameron win, (referring to him as a little girl). But he states that his father didn't raise a gentleman and that the victory will be his before getting scared and falling down due to the horns playing in analysis essay the background. Investigation Term Papers? Lightning hurts himself when he hammers on his helmet. Later that day, Chris is seen being pulled in a carriage with a television; it shows the two finalist's parents, being able to analytical image video chat with them. Cameron's mom is first seen talking to him. She had told him since purified air costs so much, she won't be able to afford his bubble. Cameron seems shocked while his mother needs him to win the million. Criminal Investigation Papers? Next is Lightning's father, but only of an effective essay, seen from his backside. Criminal Investigation? He is also saying that his son is essay on marfan syndrome going to win the million while he's being interviewed. He even bet his championship rings.
Lightning seems nervous with the heavy toll he holds now, while his father says he's nothing without them. The video then shuts off, while Lightning never got to actually speak to him. Investigation Papers? The final two and Chris are seen in the carriage as they approach the Chrisiseum. Once inside, Chris then shows the eliminated contestants. Of An Effective Analysis? Cameron then asks what happened to Scott, who is now in a machine. Term Papers? Chris says that his machine helps him say yes and no, since he can't talk.
Sam then asks if he has to cheer, while Chris says it's mandatory to cheer, or else Fang will be released on them. Chris then announces the final challenge, a gladiator battle royal. The first part is to expository essay create armor out of a massive junk pile dumped next to them. Chris then says they have fifteen minutes to investigation papers assemble their armor, but to history essay question also be careful for booby traps. Lightning first starts out pushing Cameron as he goes back and forth in front of the investigation Peanut Gallery, who are cheering and assignment students, helping him build. Cameron then finds a computer and a HazMat suit. When he pulled on term, the suit, a bomb goes off in front of essay image him. Lightning then makes fun of him and continues working.
But Cameron then spots a blowtorch that is next to Lightning, which he could use. Later on, Lightning has his suit armor almost complete, while. Cameron uses a computer, powered by the Marshmallow of Toxic Loserdom, to program his armor. A quick montage is then seen between the criminal investigation papers two preparing, until the fifteen minutes were up. Lightning is on marfan seen with his armor which everyone cheers him on, while he makes fun of Cameron, who only has a suitcase. Cameron then kicks the case, releasing a which self assembles onto him.
Lightning is stunned by his suit of high-tech, power armor. Dakota fights a mutant turtle to save Sam. Criminal Investigation Term Papers? Chris then announces part two, where the rules are, whoever pins the other for three seconds first win. The battle then starts with Lightning going after Cameron. Cameron then blasts him, causing him to fall down. Chris then brings in the mutant beetle that shoots fire at Lightning, while he runs towards Cameron and punches him. He then falls down, powering his rockets on the bottom of his feet, blasting the beetle out of the ring. Cameron and Lightning fight in the challenge. Lightning then attacks Cameron multiple times with his weapon, which causes Cameron's suit to lower in armor strength. Features Of An Expository Essay? Chris gets bored of the battle and brings out a mutant alligator. Cameron blasts out of the criminal investigation term alligators mouth, while he picks up Lightning who punches him from history essay question 2011 below.
Cameron then punches the alligator in the stomach, sending them both flying. Lightning is able to land on the ground in a split formation, injuring his groin in criminal investigation term papers the process. Of An Effective Analysis Essay? Mike then tells Cameron to pin him, but Lightning gets back up, keeping the investigation term fight going on. Cameron fire his blaster arm at Lightning, but he manage to deflect one of the shots out of qualities his way with his club. One of the shots ended up in Chris's chair hitting his remote. The remote then malfunctions, causing all the mutant creatures on investigation term, the island (and Ezekiel) to appear. Zoey, Mike, and Scott are seen traumatized in the confessional after seeing all the animals again. Assignment Log For? The animals then attack the investigation term papers eliminated contestants, causing them to go into features essay, the ring with Lightning and Cameron.
Anne Maria is seen confronting a mutant squirrel, and the squirrel shoots her hair. Investigation Term? Ezekiel then comes to save her and leans in for a kiss as she screams. Lightning gets struck by actual lightning. Cameron then gets worried about his endangered friends, while Lightning attacks him from behind and almost pins him. Cameron then throws him off causing his helmet to also fall off.
Cameron then uses the helmet to save Anne Maria, knocking out Ezekiel. Assignment Log For? Anne Maria then hairsprays her hair to fix the damage done to it.Dakota is seen attacking a mutant turtle holding Sam, while Dawn says her tea leaves didn't predict B getting punched by criminal term papers the octopus in the tree. Staci is essay also seen getting eaten by Larry, talking about her ancestors as usual. Cameron then blasts Larry in the privates, as he runs away. Lightning finally comes back to criminal papers hit Cameron once more, getting rid of all of Cameron's armor strength all together.
Cameron then orders the armor to transfer all power to essay on marfan syndrome create a magnetic field, which finally shuts down Cameron's armor. Everyone is then stunned when it appears nothing has happened Lightning then tries to criminal investigation attack Cameron one last time, but everything starts to get attracted to Lightning's armor. Cameron then tells him he turned him into a magnet, causing everything magnetic in the field to be pulled towards him. A storm cloud nearby strikes Lightning, causing everything to get blasted away from him and turns his hair white. However, Cameron also couldn't move as he uses the last of his armor's energy for the last attack. Depending on the ending, one contestant fall over the other and wins the challenge. After collecting his reward, the winner leaves with the other contestants on a boat driven by Chef. Chris then starts to sign off, but before he does, he reveals that he placed a bomb at the bottom of the on marfan boat. When he pressed the investigation term papers detonator however, the history question 2011 dock explodes underneath him explodes instead.
Chris realizes that Chef has switch the bomb's position and yells at him. Chris then resumes his sign off, when suddenly a flock of helicopters surround the island, saying that the island is now under government protection. Papers? Chris is also ordered to be quarantined, while the cast is seen laughing as they drive away. All of the contestants cheer when Cameron declares he'll split his money with everyone of them. Ap Us History Essay 2011? Lightning falls first, followed by Cameron who landed on criminal term, top of Lightning. The contestants count to three as Cameron wins. Chris then announces Cameron has won the fourth season.
A few hours later, the cast is seen at the Dock of Shame on a boat while Cameron is also seen with his million dollars as he approaches the boat with everyone else. Cameron then realizes he no longer needs the money for his bubble. He then decides that because they all helped him no longer need his bubble, that he'd share the money with them. Sam is essay image seen talking about buying video games, Jo talking about starting a gym, and Brick shouts out that he gets to go to fashion school. Everyone looks at criminal investigation term him awkwardly as the boat pulls away. Essay On Marfan? Lightning with the criminal investigation million dollar case. Lightning falls on top of ap us essay question Cameron in a daze.
Chris counts to three, and criminal term papers, then declares Lightning the winner, much to everyone's chagrin. In the confessional, Lightning boasts about his win, saying not even lightning can stop the Lightning! He receives the million dollars on of an, the Dock of Shame while Zoey comforts Cameron over his loss, which Cameron says was more of a victory due to him still being alive. Term? When Mike asks what Lightning will spend the money on, Jo asks for question 2011, 10% due to her getting him to the finals in the first place. Sam offers to play any video games that Lightning needs to beat, but Lightning rejects both of them and says that he's spending it on his own stadium so that he can get in shape for football season. Criminal Investigation Papers? Chris pulls out an X-ray of Lightning's body following the challenge and advises him to avoid physical workouts for a while, but Lightning says that pain doesn't matter to him. He then takes a step forward and breaks his leg as the boat pulls away.
The screen then switches to the confessional with Jo in it. She states how she carried these morons to the finale and thinks she should have won the money, as she then says that she got ripped off.
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What comes to mind when you hear the word ‘Indonesia’? Indonesia is the one and only country in the . universe that has so many islands, languages, tribes, traditional dishes, culture or should I say cultures – because the Indonesian people them selves can not count the culture that they have in Indonesia – different dialects, the variety of skin tones and criminal so many variety that we can not count. If we ask foreigners what the first thing that pops up when they heard. Culture of ap us essay question Indonesia , Demographics of criminal investigation papers Indonesia , Indonesia 1762 Words | 5 Pages. The American Freedom The Civil Rights Movement was an era devoted to activism for equal rights and essay 2011 treatment of African . Americans in the United States. During this period, people rallied for criminal investigation term papers social, legal, political and cultural changes to prohibit discrimination and end segregation.
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Shannon Mooningham Professor Lisa Orta English 190 September 28, 2011 Proud to Be a Woman “First Bone of a Woman” by on marfan Patricia . Wellingham-Jones and criminal term “Afternoon in the Garden” are two poems that present the creation of the first woman and explore the assignment log for nature of term womanhood through the behavior, speech and descriptions of her. History 2011. The authors addressed the story of criminal term papers creation differently through the features expository use of papers alliteration, figurative language and symbol. “First Bone of a Woman,” describes in full detail the. Adam , Adam and Eve , Book of Genesis 1430 Words | 4 Pages. ? Proud to essay, be an Indian 20 26 Oct 2010 | India India, a country with a culture having more than ten thousand years has enriched the global . scientific, educational, economic and cultural scenario significantly. That is the reason why mark Twain has stated “India is the cradle of the human race, the birthplace of human speech, the mother of history, the grandmother of term legend and about life the great grandmother of tradition.” Being born in such a great country, you have millions of reasons to criminal papers, feel proud as.
Company rule in India , Dutch East India Company , East India Company 1225 Words | 6 Pages. ? Proud To Be a Human Being When I opened the television, I was surprised because I heard a news that some friendly aliens from a highly . developed civilization decided to visit our planet, they would encounter a contradictory race of being-us. I am so happy for their visiting. I hope that I can meet them and image communicate with them. I am proud of the human race because of the wonderful inventions, entertainment and unique culture of different countries. It is a good opportunity to introduce the criminal investigation earth.
Hominidae , Human , Mobile phone 1071 Words | 3 Pages. ? American Dad Casey Jones February 10, 2015 Dr. Chuck Tryon English 120 Fayetteville State University Abstract The main character . Stan Smith, a CIA agent and father who is always on the lookout for terrorist and extraterrestrial activity, will go to the absolute extreme to make sure his country is safe from log for students harm. In addition to criminal term papers, his regular normal family, his wife Francine and his two teenage children Haley and Steve, they also has two “different and special” members. Essay On Marfan Syndrome. Roger, the sarcastic.
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United States has always been recognized throughout the criminal investigation term . world as the “Land of Opportunity.” The attraction of the essay about in prison promise of freedom, wealth, prosperity, and success had drawn millions of immigrants to come to America and search for a better life. One of the most important aspects is that the U.S is an egalitarian nation which opportunities are given based upon investigation term ability or achievement, rather than social status. Cultural assimilation , Culture , Family 1815 Words | 6 Pages. American and North Vietnamese Perception of War. Austin Beeler American and North Vietnamese Perceptions of War To the American Soldiers and the NVA/VC soldiers, the Vietnam . War was both very similar and of an essay very different. Most of soldiers in investigation papers, Vietnam went into the war with a purpose, and they had a very high morale levels. Both the Americans and the NVA soldiers left with many problems. The war left most soldiers with very severe cases of mental issues. There were also many things about the war that differed to the soldiers. One was the analytical image type.
French Indochina , Military , Nationalism 918 Words | 3 Pages. African Americans in the American Revolution. ?African Americans in term, the American Revolution Leading into the American Revolution, African . Americans were placed in a dilemma of whether to rebel or remain faithful to the crown. There were two sides of the American Revolution; the Patriots (also known as the analytical essay image rebels), were the Americans that wanted to gain their independence from Great Britain. The Loyalist was the people who wanted to criminal investigation term papers, stay under the control of Great Britain. Some African Americans joined the Loyalist because they were promised. American Revolutionary War , Battles of Lexington and Concord , Black people 1563 Words | 5 Pages.
American Literature: The Meaning of American. sprout up out of nowhere. The culture of the African- Americans , the Native Americans , and the Europeans provided us with the essay syndrome . customs and culture that we today call our own. Through centuries of exposure to different cultures and ways, America has become, in criminal term papers, the words of Jimmy Carter, “not a melting pot, but a beautiful mosaic.” (Carter and Shapiro 137) In this sense, the word “ American ” takes on a unique meaning, in essay about life, that to be an American means to be part of this mosaic, and to criminal papers, have the ability. African American , American Civil War , Americas 832 Words | 3 Pages. Week 1 Conference 1. What is African- American history? African- American history are those events that started with the . first slave ships from Africa to log for, the Caribbean Islands and carry through their journey as a people both individually and investigation term papers collectively to today's societies across the log for students Americas. 2. How have your ideas about investigation, African- American history in features expository essay, particular and history in general been shaped by the contexts in which you encountered these histories? I was born and raised in.
African American , African diaspora , Afro-Latin American 893 Words | 3 Pages. American Dream or American Nightmare. Criminal Investigation Papers. Speech Analysis. American Dream or American Nightmare Be The American Dream is something that can be easily defined for some . people, and for others it can be very difficult to put into words. The American Dream is a combination of a few things. Essay On Marfan. I firmly believe that Americans should have equal access to this ideal of an papers, American Dream in order for it not to be considered just mere notion. Of An. My idea of the American Dream is also built on freedom.
Our fore fathers who built this great nation were on the right. African American , Black people , Haiti 1507 Words | 4 Pages. American History American Imperialism. ?The March of the Flag: American reasoning to interventionism Aleksander Trzebunia HIST2402A Professor: Andrew Johnston TA: Emily Cuggy . In the late 1800’s, at the turn of the 20th century, European empires controlled around 60% of the criminal investigation globe. The decaying Spanish and essay image French empires had relinquished most of their Imperialist goals to control more overseas territories and were left with the remains of what they could hold onto. Papers. The British Empire however was still going strong and syndrome as the. British Empire , Empire , Foreign policy of the criminal term United States 2182 Words | 9 Pages. Native Americans in the American Revolution.
November 16th, 2012 While most think of the American Revolution only essay on marfan as a battle between the American colonists and the . British Empire, Native Americans were a major factor in the war. Criminal Term Papers. The British and Americans clamored for war alliances from various Native American tribes and in features expository essay, most cases, the British came out victorious. This presented the rebellious Americans with the dilemma of criminal term how to essay about, confront hostile Native American tribes as American settlers moved steadily westward into Indian country. American Revolution , American Revolutionary War , Massachusetts 1280 Words | 4 Pages. “Al-Shaitan” “The American Sniper” by Chris Kyle is an account of the deadliest American sniper ever, called “the devil” by criminal investigation term the . enemies he hunted and “the legend” by his Navy SEAL brothers.
From 1999 to 2009, U.S. On Marfan Syndrome. Navy SEAL Chris Kyle recorded the most career sniper kills in United States military history. Criminal Term. The Pentagon has officially confirmed more than 150 of Kyle's kills (the previous American record was 109). Iraqi insurgents feared Kyle so much they named him al-Shaitan (“the devil”) and placed. Armed forces , Conscription , Military 1323 Words | 3 Pages. American Culture in My Eyes What is assignment students really American culture? Because of criminal investigation short American history, some people . in my country still have a question: What makes the United States develop rapidly? The answer is analytical unique American culture.
On the small aspects, it can be attributed to the living standard, which is American life culture, such as daily diet, clothes, and transportation. To the papers major aspects, American culture can be attributed to values and social-orientation. For example, the main content. Americas , Caribbean , Immigration to the United States 780 Words | 3 Pages. American Revolution: American Romanticism. American Romanticism American Romanticism was a literary and qualities analysis artistic movement of the nineteenth century that placed a premium . on fancy, imagination, emotion, nature, individuality, and exotica. It was the Romantic period in American Literature, which included writers like Washington Irving, Emerson, Thoreau, Emily Dickinson, Edgar Allan Poe, Walt Whitman, Herman Melville, and others. One characteristic of American Romanticism involves humanity's relationship to nature. Much of the criminal investigation term papers writing during. American poets , Edgar Allan Poe , James Russell Lowell 1883 Words | 5 Pages.
African American and American Society. T: Th. 3:30p.m.-4:45p.m. D’Angelo, Raymond and Herbert Douglas, eds. Taking Sides: Clashing Views in Race and Ethnicity, 7th edition (Dubuque, IA: . McGraw-Hill, 2009) “Issue #9: Is Racism a Permanent Feature of American Society?” Is Racism a Permanent feature of American Society? Derrick Bell argues in this issue that the prospects for achieving racial equality in the United States are “illusory for Blacks. Bell reminds us despite the fact of the essay about life in prison progress of blacks in United States; the. African American , Black people , Discrimination 1438 Words | 4 Pages.
Asian Americans in American Cinema. Abstract With a recent surge of Asian Americans in American cinema, we are noticing that the stereotypes that we have used in . the past are actually being used by their culture to criminal, break into American culture. Days of portraying Asian Americans as unintelligible and subservient are gone and ap us history essay question 2011 are being replaced with a whole new outlook. The introduction of Martial Arts into American film has evolved the way Asians are viewed in American cinema. Quentin Tarantino introduced a whole new outlook to term papers, Asian. Asian American , Bruce Lee , Film 1394 Words | 4 Pages. Characteristics of Americans/American Culture.
101 Characteristics of Americans / American Culture To help you compare and contrast what you observe of American . About Life. culture and criminal investigation term your own, mark the similarities and differences between your culture and what you have read about in this book. 1. America is enormous: the third largest country in the world with a population of more than 300 million people. Essay. 2. Criminal Investigation Papers. Americans come in history question, all colors, have all types of religions, and investigation papers speak many languages from all over of an effective essay, the world. 3. Investigation. Americans are extremely independent. American , Anna Faris , United States 1763 Words | 5 Pages. The Exploitation of the Native Americans. the Native Americans The Native Americans are a prime example of the essay life in prison repression, poverty, and discrimination many minority . Criminal Investigation Term. groups have had to endure throughout the features expository essay years. The Native Americans had their own land, culture and language.
They were people able to criminal term, adapt well to their particular region by hunting, fishing and farming crops. Assignment Log For. Their cultures primarily rested on wise use of all natural resources available. Many historians believe there were between 6 and 10 million Native Americans living in. Dawes Act , Federal government of the United States , Indian reservation 1596 Words | 5 Pages. American Inequality in American Psycho. Derek Naegle Dr. Cammie Sublette ENGL 2033 September 7, 2012 American Inequality in papers, American Psycho Set in the Manhattan of . 1989, Brett Easton Ellis's novel American Psycho sketches the life of Patrick Bateman, an attractive 26-year-old Harvard graduate who earns a six-figure income on Wall Street.
Bateman and assignment log for students his Ivy League educated friends enjoy all the luxury Manhattan has to criminal term, offer, including expensive restaurants, exclusive nightclubs and excessive amounts of of an expository cocaine. However, what their. American Psycho , Bret Easton Ellis , Manhattan 682 Words | 3 Pages. ?The Native American population has been seen as a minority ever since the first white settlers arrived to America in 1492. With over 500 . Criminal Investigation Term. federally recognized tribes in the United States, it is unfair to group their traditions and culture as one. In Montana, there are seven federally recognized reservations today, which include the Northern Cheyenne, Crow, Fort Peck, Fort Belknap, Blackfeet, and Flathead (Reservations). Features. In this paper, I am focusing on statistics and data gathered from investigation all Montana. Demographics of the United States , Great Plains , Indian reservation 2090 Words | 8 Pages. accents of English or new vocabulary created by essay image mixing two languages together. Investigation Term Papers. However, misunderstanding between Americans and qualities effective analysis essay Mexicans were . obviously unavoidable due to language barriers.
Besides, we also had several stereotypes related to cultural differences between people who came from different countries because language is an important part of criminal investigation term papers culture. To be more specific, while American people were perceived to have better life with appropriate salary and essay question 2011 high standard of living, Mexicans were. Americas , Anthropology , Culture 1965 Words | 7 Pages. The American Dream is criminal investigation term papers not a paradise but a nightmare. ?“The American Dream is history question 2011 not a paradise but a nightmare.” By comparing A Streetcar Named Desire and The Great Gatsby to investigation papers, what extent do you . agree with this statement? According to James Truslow Adams, who wrote ‘The Great Epic of expository America’, the American Dream is “not a dream of motor cars and high wages merely, but a dream of criminal papers social order in which each man and essay each woman shall be able to attain regardless of circumstances of birth or position”1. A Streetcar Named Desire and The Great Gatsby both. Arnold Rothstein , F. Scott Fitzgerald , James Truslow Adams 2381 Words | 7 Pages. Assimilation Into American Society-Immigrants.
June 26, 2011 Assimilation into American society: “Immigrants” written by criminal investigation papers Pat Mora In the poem, “Immigrants”, it talks about how . immigrants want their children to be welcomed into American society. They will do whatever they need to, to get their children to be as American as possible; even if they lose some of their own culture in the process. The poem, “Immigrants” by Pat Mora, has many underlying themes. The main theme of about life in prison how immigrants assimilate into American culture has a deep connection to. Flag of the investigation term United States , Immigration to the United States , The Star-Spangled Banner 876 Words | 3 Pages. North Dakota is currently in one of the biggest debates over a Native American team mascot. The Standing Rock Sioux Tribe is fighting with . the University of North Dakota regarding the Fighting Sioux mascot.
The Native American students have been increasing the pressure on the University to change its name. Assignment Log For. “We’re seeing more educators around the county, in middle Schools, high schools and at universities, concerned about the racial climate in schools dropping these symbols” (Johansen. Great Plains , Indigenous peoples of the Americas , Mascot 2165 Words | 6 Pages. Asian American /Chinese Culture Theresa Chambers Webster University Abstract A Chinese American is an . American who is term of ethnic Chinese descent. Most Chinese Americans are descended from China's majority ethnic group, the history essay Han. The rest are usually members of one of China's 56 minorities, such as the Hui.
Chinese Americans constitute one group of papers overseas Chinese and are also one group of Asian Americans . Asian American / Chinese Culture Immigration Chinese immigration to the United. Asian American , Chicago , China 925 Words | 3 Pages. The American Dream then and analysis now Imagine you are one of the investigation term early settlers in essay about life, America. You left Europe, a world full of criminal investigation term papers religious persecution, . Ap Us History 2011. political oppression and poverty behind you. You have dreams like: - Personal freedom, self-fulfillment, dignity and happiness, - Economic things, like prosperity and criminal investigation term success - Rising from log for poverty to fame and fortune (from rags to term, riches) - Social dreams of equality and a classless society - Religious dreams of religious freedom in a “promised land”. Americas , Anarchism , Human rights 760 Words | 3 Pages. My Historical Report on African American I choose to go with African American Race; I know more about African . American culture personally. I grew up with mostly African American and bi-racial friends as a child and enjoyed being around them. What have been the ap us history essay experiences of this racial group throughout U.S. history?
Throughout the U.S. History the criminal term African American race have experienced Martin Luther King Jr. (1929-1968) who pushed strides for freedom. He believed that people should have the. African American , American Civil War , Black people 817 Words | 3 Pages.
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34 things you should remove from criminal papers your resume immediately. Hiring managers rarely have the time or resources to look at each resume closely, and they typically spend about six seconds on their initial fit/no fit decision. If you want to pass that test, you need to have some solid qualifications — and the perfect resume to ap us history 2011, highlight them. Here are 34 things you should strike from your resume right now. Criminal? If you applied, it's already obvious you want the job. The exception: If you're in a unique situation, such as changing industries completely, it may be useful to include a brief summary. Yes, you might have been the king of making milkshakes at the restaurant you worked for in high school. But unless you are planning on redeeming that title, it is time to get rid of all that clutter. But as Alyssa Gelbard, career expert and founder of essay on marfan career-consulting firm Resume Strategists, points out: Past work experience that might not appear to be directly relevant to the job at criminal investigation term hand might show another dimension, depth, ability, or skill that actually is relevant or applicable. Only include this experience if it really showcases additional skills that can translate to the position you're applying for. Don't include your marital status, religious preference, or Social Security number.
This might have been the standard in essay, the past, but all this information could lead to investigation term, discrimination, which is illegal, so there's no need to include it. A full street address is the first thing Amanda Augustine, a career-advice expert for TopResume, looks for to immediately cut from a resume . Essay Syndrome? Nobody needs to have that on their resume anymore, and, to be quite honest, it's a security concern, she tells Business Insider. Augustine suggests including only one phone number on your resume, and that number should really be your cellphone, so that you can control who answers your incoming phone calls, when, and what the voice mail sounds like. Also, you don't want employers trying to contact you in five different places, because then you have to keep track of that, she says. In many cases, nobody cares. If it's not relevant to the job you're applying for, it's a waste of space and a waste of the company's time. A CareerBuilder survey asked 2,000 hiring managers for memorable resume mistakes, and blatant lies were a popular choice. One candidate claimed to be the former CEO of the company to which he was applying, another claimed to be a Nobel Prize winner, and criminal investigation one more claimed he attended a college that didn't exist. Rosemary Haefner, chief human-resources officer at qualities of an effective essay CareerBuilder, says these lies may be misguided attempts to criminal, compensate for lacking 10o% of the qualifications specified in the job posting. But Haefner says candidates should concentrate on the skills they can offer, rather than the skills they can't offer. Hiring managers are more forgiving than job seekers may think, Haefner explains.
About 42% of employers surveyed said they would consider a candidate who met only on marfan syndrome, three out of criminal term papers five key qualifications for a specific role. When you use a 0.5-inch margin and eight-point font in an effort to get everything to analytical image, fit on investigation papers one page, this is an epic fail, says J.T. O'Donnell, a career and ap us history essay question 2011 workplace expert, founder of career-advice site Careerealism.com, and author of term Careerealism: The Smart Approach to a Satisfying Career. She recommends lots of white space and no more than a 0.8 margin. Augustine agrees, warning particularly against dense blocks of text. Let's be honest: You're looking this over quickly, you're glancing through it, your eyes glaze over analytical image when you get to a big, long paragraph, she says.
In the same vein, you can also overload your resume with too many bullet points, which Augustine calls death by bullets. If absolutely everything is bulleted, it has the same effect as big dense blocks of term text — your eyes just glaze over it, she says. Qualities Of An Effective Analysis? Augustine explains that bullets are only to be used to draw attention to the most important information. If you bullet everything, everything is important, which means really nothing stands out, she says. If you took time off to travel or raise a family, Gelbard doesn't recommend including that information on your resume. In some countries, it is acceptable to include this information, especially travel, but it is not appropriate to include that in the body of a resume in the US. 11. Investigation Papers? Details that give away your age. If you don't want to be discriminated against for a position because of your age, it's time to remove your graduation date, says Catherine Jewell, author of New Resume, New Career. Another surprising way your resume could give away your age: double spaces after a period. If your employers want to speak to your references, they'll ask you.
Also, it's better if you have a chance to tell your references ahead of time that a future employer might be calling. If you write references upon request at the bottom of assignment your resume, you're merely wasting a valuable line, career coach Eli Amdur says. The format of your resume is just as important as its content, Augustine says. She says the best format is the format that will make it easiest for term papers the hiring manager to assignment log for students, scan your resume and still be able to pick out criminal investigation papers, your key qualifications and career goals. Once you pick a format, stick with it. If you write the day, month, and ap us history 2011 year for one date, then use that same format throughout the rest of the resume.
Your resume shouldn't include the words I, me, she, or my, says Tina Nicolai, executive career coach and founder of investigation Resume Writers' Ink. Don't write your resume in the third or first person. It's understood that everything on your resume is qualities of an about you and criminal investigation your experiences. Never describe past work experience using the features of an expository essay present tense. Only your current job should be written in the present tense, Gelbard says. 16. Term Papers? A less-than-professional email address. Analytical Essay Image? If you still use an old email address, like BeerLover123@gmail.com or CuteChick4life@yahoo.com, it's time to term papers, pick a new one. It only takes a minute or two, and it's free. Amdur says there is no reason to put the word phone in front of the actual number.
It's pretty silly. They know it's your phone number. The same rule applies to email. 18. Essay In Prison? Your current business-contact info. This is not only dangerous; it's stupid. Do you really want employers calling you at work? How are you going to handle that? Oh, and by the way, your current employer can monitor your emails and criminal investigation phone calls. So if you're not in log for students, the mood to criminal papers, get fired, or potentially charged with theft of services (really), then leave the business info off.
19. Headers, footers, tables, images, charts. These fancy embeddings will have hiring managers thinking, Could you not? While a well-formatted header and log for students footer may look professional, and some cool tables, images, or charts may boost your credibility, they also confuse the criminal investigation term papers applicant-tracking systems that companies use nowadays, Augustine tells Business Insider. The system will react by scrambling up your resume and spitting out a poorly formatted one that may no longer include your header or charts. Even if you were an ideal candidate for the position, now the hiring manager has no way to contact you for an interview. Image? Don't include your boss' name on your resume unless you're OK with your potential employer contacting him or her.
Even then, Gelbard says the only reason your boss' name should be on your resume is criminal if the life person is someone noteworthy, and if it would be really impressive. Companies often have their own internal names for criminal investigation term papers things like customized software, technologies, and processes that are only essay on marfan syndrome, known within that organization and investigation term not by those who work outside of it, Gelbard says. Be sure to exclude terms on your resume that are known only to on marfan, one specific organization. Criminal Term Papers? 22. Social-media URLs that are not related to the targeted position. Links to your opinionated blogs, Pinterest page, or Instagram account have no business taking up prime resume real estate. Candidates who tend to analysis essay, think their personal social media sites are valuable are putting themselves at criminal investigation term papers risk of landing in the 'no' pile, Nicolai says. But you should list relevant URLs, such as your LinkedIn page or any others that are professional and directly related to the position you are trying to acquire, she says. Assignment? 23. Term? More than 15 years of experience. When you start including jobs from before 2000, you start to lose the hiring manager's interest.
Your most relevant experience should be from the past 15 years, so hiring managers only of an, need to see that, Augustine says. On the same note, never include dates on education and certifications that are older than 15 years. Some people include past hourly rates for jobs they held in college, Nicolai says. This information is unnecessary and criminal investigation papers may send the wrong message. Amy Hoover, president of Talent Zoo, says you also shouldn't address your desired salary in essay on marfan, a resume. This document is intended to criminal term, showcase your professional experience and assignment skills. Salary comes later in investigation, the interview process. Don't use Times New Roman and serif fonts, as they're outdated and old-fashioned, Hoover says. Use a standard, sans-serif font like Arial. Expository Essay? Also, be aware of the font size, she says.
Your goal should be to criminal term papers, make it look nice and of an essay sleek — but also easy to read. Curly-tailed fonts are also a turn-off, according to O'Donnell. Criminal Investigation? People try to make their resume look classier with a fancy font, but studies show they are harder to read and the recruiter absorbs less about you. CareerBuilder asked 2,201 US hiring managers: What resume terms are the biggest turnoffs? They cited words and essay 2011 phrases such as, best of breed, go-getter, think outside the box, synergy, and people pleaser. Terms employers do like to criminal investigation term, see on resumes include: achieved, managed, resolved, and launched — but only if they're used in moderation. Essay Life In Prison? 28. Reasons you left a company or position. Investigation Papers? Candidates often think, If I explain why I left the position on ap us essay 2011 my resume, maybe my chances will improve. Wrong, Nicolai says. Listing why you left is irrelevant on your resume.
It's not the criminal term time or place to bring up transitions from one company to essay about in prison, the next. Investigation Papers? Use your interview to address this. Once you're out of school, your grades aren't so relevant. If you're a new college graduate and your GPA was a 3.8 or higher — it's OK to essay, leave it. But, if you're more than three years out of school, or if your GPA was lower than a 3.8, ditch it. This may become the norm at some point in the future, but it's just weird — and tacky and criminal term distracting — to include a photo with your resume for now. 31. An explanation of why you want the job.
That's what the cover letter and interviews are for! Your resume is not the place to assignment log for, start explaining why you'd be a great fit or why you want the job. Your skills and qualifications should be able to do that for you — and if they don't, then your resume is criminal investigation term papers either in bad shape, or this isn't the right job for you. Don't try to sell yourself by using all sorts of essay syndrome subjective words to describe yourself, O'Donnell says. I'm an excellent communicator or highly organized and motivated are opinions of yourself and not necessarily the truth. Term? Recruiters want facts only. They'll decide if you are those things after they meet you, she says. 33. Generic explanations of accomplishments.
Don't just say you accomplished X, Y, or Z — show it by quantifying the facts. For instance, instead of, Grew revenues try, X project resulted in an Y% increase in revenues. Avoid including a job on features of an expository your resume if you only held the position for papers a short period of time, Gelbard says. You should especially avoid including jobs you were let go from or didn't like. Vivian Giang and of an essay Natalie Walters contributed to earlier versions of this article. SEE ALSO: 9 phrases on your resume that make hiring managers cringe.
DON'T MISS: We took a real resume from a mid-level employee and turned it into something fantastic. Recommended For You Powered by Sailthru. 34 things you should remove from your resume immediately. Hiring managers rarely have the time or. Criminal Investigation Term? Get the best of Business Insider delivered to your inbox every day.
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Enzymes: principles and biotechnological applications. Enzymes are biological catalysts (also known as biocatalysts) that speed up biochemical reactions in living organisms, and which can be extracted from criminal term papers cells and then used to catalyse a wide range of features of an, commercially important processes. This chapter covers the basic principles of enzymology, such as classification, structure, kinetics and inhibition, and also provides an overview of industrial applications. In addition, techniques for the purification of term, enzymes are discussed. Enzymes are biological catalysts (also known as biocatalysts) that speed up biochemical reactions in history essay 2011, living organisms. They can also be extracted from cells and then used to criminal investigation papers, catalyse a wide range of commercially important processes. For example, they have important roles in the production of sweetening agents and image, the modification of antibiotics, they are used in washing powders and various cleaning products, and they play a key role in analytical devices and assays that have clinical, forensic and investigation term, environmental applications. The word ‘enzyme’ was first used by the German physiologist Wilhelm Kuhne in 1878, when he was describing the ability of yeast to produce alcohol from sugars, and it is essay, derived from the Greek words en (meaning ‘within’) and zume (meaning ‘yeast’). In the late nineteenth century and early twentieth century, significant advances were made in the extraction, characterization and commercial exploitation of criminal investigation, many enzymes, but it was not until the essay question 1920s that enzymes were crystallized, revealing that catalytic activity is associated with protein molecules.
For the next 60 years or so it was believed that all enzymes were proteins, but in the 1980s it was found that some ribonucleic acid (RNA) molecules are also able to term papers, exert catalytic effects. These RNAs, which are called ribozymes, play an important role in gene expression. In the same decade, biochemists also developed the technology to generate antibodies that possess catalytic properties. These so-called ‘abzymes’ have significant potential both as novel industrial catalysts and in therapeutics. Notwithstanding these notable exceptions, much of classical enzymology, and the remainder of this essay, is focused on the proteins that possess catalytic activity. As catalysts, enzymes are only of an expository, required in very low concentrations, and they speed up reactions without themselves being consumed during the reaction.
We usually describe enzymes as being capable of catalysing the conversion of substrate molecules into product molecules as follows: The enormous catalytic activity of enzymes can perhaps best be expressed by a constant, k cat , that is variously referred to as the turnover rate, turnover frequency or turnover number. This constant represents the number of substrate molecules that can be converted to product by a single enzyme molecule per unit time (usually per minute or per second). Examples of turnover rate values are listed in Table 1. For example, a single molecule of carbonic anhydrase can catalyse the conversion of over half a million molecules of its substrates, carbon dioxide (CO 2 ) and water (H 2 O), into the product, bicarbonate (HCO 3 ? ), every second—a truly remarkable achievement. As well as being highly potent catalysts, enzymes also possess remarkable specificity in that they generally catalyse the conversion of only one type (or at most a range of similar types) of criminal investigation, substrate molecule into product molecules. Some enzymes demonstrate group specificity. For example, alkaline phosphatase (an enzyme that is analytical essay image, commonly encountered in first-year laboratory sessions on enzyme kinetics) can remove a phosphate group from a variety of substrates.
Other enzymes demonstrate much higher specificity, which is described as absolute specificity. For example, glucose oxidase shows almost total specificity for its substrate, ?-D-glucose, and virtually no activity with any other monosaccharides. As we shall see later, this specificity is of paramount importance in many analytical assays and criminal term, devices (biosensors) that measure a specific substrate (e.g. glucose) in a complex mixture (e.g. a blood or urine sample). Enzymes typically have common names (often called ‘trivial names’) which refer to the reaction that they catalyse, with the suffix -ase (e.g. Analytical Essay? oxidase, dehydrogenase, carboxylase), although individual proteolytic enzymes generally have the investigation papers suffix - in features, (e.g. trypsin, chymotrypsin, papain). Often the trivial name also indicates the substrate on which the enzyme acts (e.g. glucose oxidase, alcohol dehydrogenase, pyruvate decarboxylase). However, some trivial names (e.g. Investigation Term? invertase, diastase, catalase) provide little information about the analytical image substrate, the product or the reaction involved.
Due to term, the growing complexity of and inconsistency in the naming of enzymes, the International Union of about in prison, Biochemistry set up the Enzyme Commission to address this issue. The first Enzyme Commission Report was published in 1961, and provided a systematic approach to the naming of term, enzymes. The sixth edition, published in 1992, contained details of nearly 3 200 different enzymes, and supplements published annually have now extended this number to over 5 000. Within this system, all enzymes are described by a four-part Enzyme Commission (EC) number. For example, the essay enzyme with the trivial name lactate dehydrogenase has the papers EC number 18.104.22.168, and qualities analysis, is more correctly called l –lactate: NAD + oxidoreductase. The first part of the EC number refers to the reaction that the enzyme catalyses (Table 2).
The remaining digits have different meanings according to the nature of the reaction identified by the first digit. For example, within the oxidoreductase category, the second digit denotes the hydrogen donor (Table 3) and investigation papers, the third digit denotes the hydrogen acceptor (Table 4). Thus lactate dehydrogenase with the EC number 22.214.171.124 is an oxidoreductase (indicated by the first digit) with the essay on marfan alcohol group of the lactate molecule as the hydrogen donor (second digit) and NAD + as the hydrogen acceptor (third digit), and is the 27th enzyme to criminal investigation term, be categorized within this group (fourth digit). Fortunately, it is now very easy to find this information for any individual enzyme using the Enzyme Nomenclature Database (available at image http://enzyme.expasy.org). Enzyme structure and substrate binding. Amino acid-based enzymes are globular proteins that range in size from less than 100 to investigation papers, more than 2 000 amino acid residues. These amino acids can be arranged as one or more polypeptide chains that are folded and bent to form a specific three-dimensional structure, incorporating a small area known as the active site (Figure 1), where the substrate actually binds. The active site may well involve only a small number (less than 10) of the constituent amino acids. It is the shape and charge properties of the active site that enable it to bind to history question, a single type of substrate molecule, so that the papers enzyme is able to demonstrate considerable specificity in its catalytic activity.
The hypothesis that enzyme specificity results from the complementary nature of the substrate and analytical essay, its active site was first proposed by criminal investigation term papers, the German chemist Emil Fischer in 1894, and became known as Fischer's ‘lock and essay, key hypothesis’, whereby only a key of the correct size and criminal papers, shape (the substrate) fits into the keyhole (the active site) of the lock (the enzyme). It is astounding that this theory was proposed at a time when it was not even established that enzymes were proteins. As more was learned about enzyme structure through techniques such as X-ray crystallography, it became clear that enzymes are not rigid structures, but are in fact quite flexible in shape. In the light of this finding, in 1958 Daniel Koshland extended Fischer's ideas and presented the ‘induced-fit model’ of substrate and enzyme binding, in which the enzyme molecule changes its shape slightly to accommodate the binding of the substrate. The analogy that is log for, commonly used is the ‘hand-in-glove model’, where the hand and glove are broadly complementary in shape, but the term glove is moulded around the hand as it is inserted in analytical essay image, order to investigation term papers, provide a perfect match. Since it is the active site alone that binds to the substrate, it is logical to ask what is the role of the essay syndrome rest of the protein molecule.
The simple answer is that it acts to stabilize the criminal term papers active site and provide an appropriate environment for interaction of the life site with the criminal substrate molecule. Life? Therefore the active site cannot be separated out term papers from the rest of the protein without loss of catalytic activity, although laboratory-based directed (or forced) evolution studies have shown that it is sometimes possible to generate smaller enzymes that do retain activity. It should be noted that although a large number of history question, enzymes consist solely of protein, many also contain a non-protein component, known as a cofactor, that is term papers, necessary for the enzyme's catalytic activity. A cofactor may be another organic molecule, in which case it is called a coenzyme, or it may be an inorganic molecule, typically a metal ion such as iron, manganese, cobalt, copper or zinc. A coenzyme that binds tightly and permanently to the protein is generally referred to as the prosthetic group of the features expository enzyme. When an enzyme requires a cofactor for investigation term papers, its activity, the syndrome inactive protein component is criminal investigation term, generally referred to as an apoenzyme, and life, the apoenzyme plus the cofactor (i.e. the active enzyme) is called a holoenzyme (Figure 2).
The need for minerals and vitamins in the human diet is partly attributable to their roles within metabolism as cofactors and coenzymes. How do enzymes work? The broad answer to this question is that they do not alter the equilibrium (i.e. the papers thermodynamics) of a reaction. On Marfan? This is because enzymes do not fundamentally change the investigation term structure and energetics of the products and reagents, but rather they simply allow the analytical reaction equilibrium to be attained more rapidly. Let us therefore begin by clarifying the concept of chemical equilibrium. In many cases the equilibrium of a reaction is far ‘to the right’—that is, virtually all of the substrate (S) is converted into criminal product (P). For this reason, reactions are often written as follows: This is a simplification, as in all cases it is ap us essay question, more correct to write this reaction as follows: This indicates the presence of an equilibrium.
To understand this concept it is investigation term papers, perhaps most helpful to look at a reaction where the equilibrium point is quite central. In this reaction, if we start with a solution of 1 mol l ?1 glucose and add the enzyme, then upon of an essay, completion we will have a mixture of term, approximately 0.5 mol l ?1 glucose and 0.5 mol l ?1 fructose. This is the equilibrium point of this particular reaction, and although it may only take a couple of seconds to reach this end point with the enzyme present, we would in fact come to assignment log for students, the same point if we put glucose into criminal papers solution and waited many months for the reaction to occur in assignment log for students, the absence of the enzyme. Interestingly, we could also have started this reaction with a 1 mol l ?1 fructose solution, and it would have proceeded in the opposite direction until the criminal investigation term same equilibrium point had been reached. The equilibrium point for this reaction is expressed by the equilibrium constant K eq as follows: Thus for a reaction with central equilibrium, K eq = 1, for an equilibrium ‘to the right’ K eq is 1, and for an equilibrium ‘to the left’ K eq is 1. Therefore if a reaction has a K eq value of 10 6 , the equilibrium is very far to the right and can be simplified by denoting it as a single arrow. We may often describe this type of reaction as ‘going to analytical, completion’. Conversely, if a reaction has a K eq value of 10 ?6 , the equilibrium is very far to the left, and for all practical purposes it would not really be considered to proceed at all. It should be noted that although the concentration of reactants has no effect on the equilibrium point, environmental factors such as pH and temperature can and do affect the position of the criminal papers equilibrium. It should also be noted that any biochemical reaction which occurs in vivo in a living system does not occur in isolation, but as part of a metabolic pathway, which makes it more difficult to conceptualize the relationship between reactants and reactions.
In vivo reactions are not allowed to proceed to their equilibrium position. If they did, the reaction would essentially stop (i.e. the forward and qualities of an effective essay, reverse reactions would balance each other), and there would be no net flux through the pathway. However, in many complex biochemical pathways some of the individual reaction steps are close to equilibrium, whereas others are far from equilibrium, the latter (catalysed by regulatory enzymes) having the greatest capacity to control the overall flux of materials through the pathway. Enzymes form complexes with their substrates. We often describe an enzyme-catalysed reaction as proceeding through three stages as follows: The ES complex represents a position where the substrate (S) is bound to the enzyme (E) such that the reaction (whatever it might be) is made more favourable. As soon as the reaction has occurred, the product molecule (P) dissociates from the enzyme, which is then free to bind to another substrate molecule.
At some point during this process the substrate is criminal term, converted into an intermediate form (often called the transition state) and then into the product. The exact mechanism whereby the enzyme acts to essay, increase the rate of the reaction differs from one system to another. However, the general principle is that by binding of the criminal papers substrate to the enzyme, the reaction involving the substrate is made more favourable by lowering the activation energy of the reaction. In terms of energetics, reactions can be either exergonic (releasing energy) or endergonic (consuming energy). However, even in an exergonic reaction a small amount of energy, termed the activation energy, is needed to students, give the reaction a ‘kick start.’ A good analogy is that of a match, the head of which contains a mixture of papers, energy-rich chemicals (phosphorus sesquisulfide and potassium chlorate). When a match burns it releases substantial amounts of analytical, light and heat energy (exergonically reacting with O 2 in term, the air). However, and perhaps fortunately, a match will not spontaneously ignite, but rather a small input of energy in the form of heat generated through friction (i.e. striking of the match) is question 2011, needed to initiate the criminal investigation term papers reaction. Of course once the match has been struck the amount of energy released is considerable, and greatly exceeds the of an expository small energy input during the criminal investigation term papers striking process. As shown in Figure 3, enzymes are considered to features expository essay, lower the activation energy of criminal investigation term, a system by making it energetically easier for the transition state to form.
In the presence of an assignment students enzyme catalyst, the formation of the transition state is energetically more favourable (i.e. it requires less energy for the ‘kick start’), thereby accelerating the rate at which the reaction will proceed, but not fundamentally changing the energy levels of either the reactant or the product. Properties and mechanisms of enzyme action. Enzyme kinetics is the study of factors that determine the speed of enzyme-catalysed reactions. It utilizes some mathematical equations that can be confusing to students when they first encounter them. However, the theory of kinetics is both logical and investigation, simple, and it is essential to develop an understanding of this subject in ap us essay question 2011, order to be able to appreciate the investigation papers role of enzymes both in on marfan syndrome, metabolism and in biotechnology. Assays (measurements) of term papers, enzyme activity can be performed in either a discontinuous or continuous fashion. Discontinuous methods involve mixing the substrate and enzyme together and measuring the product formed after a set period of time, so these methods are generally easy and quick to essay, perform. In general we would use such discontinuous assays when we know little about the system (and are making preliminary investigations), or alternatively when we know a great deal about the system and are certain that the time interval we are choosing is appropriate. In continuous enzyme assays we would generally study the investigation papers rate of an enzyme-catalysed reaction by mixing the enzyme with the substrate and continuously measuring the appearance of product over time. Of course we could equally well measure the rate of the reaction by on marfan, measuring the criminal disappearance of of an effective, substrate over time. Apart from the actual direction (one increasing and one decreasing), the two values would be identical.
In enzyme kinetics experiments, for convenience we very often use an artificial substrate called a chromogen that yields a brightly coloured product, making the reaction easy to follow using a colorimeter or a spectrophotometer. However, we could in fact use any available analytical equipment that has the capacity to term, measure the concentration of either the product or the substrate. In almost all cases we would also add a buffer solution to the mixture. Essay Image? As we shall see, enzyme activity is investigation papers, strongly influenced by features of an expository essay, pH, so it is important to set the pH at a specific value and keep it constant throughout the experiment. Our first enzyme kinetics experiment may therefore involve mixing a substrate solution (chromogen) with a buffer solution and criminal term, adding the enzyme. Assignment? This mixture would then be placed in a spectrophotometer and the appearance of the coloured product would be measured. This would enable us to follow a rapid reaction which, after a few seconds or minutes, might start to slow down, as shown in Figure 4. A common reason for this slowing down of the speed (rate) of the reaction is that the substrate within the mixture is being used up and thus becoming limiting. Alternatively, it may be that the enzyme is unstable and is denaturing over criminal investigation papers, the course of the experiment, or it could be that the pH of the assignment mixture is changing, as many reactions either consume or release protons.
For these reasons, when we are asked to specify the rate of term papers, a reaction we do so early on, as soon as the enzyme has been added, and when none of the above-mentioned limitations apply. We refer to ap us, this initial rapid rate as the criminal investigation papers initial velocity ( v 0 ). Measurement of the reaction rate at essay image this early stage is criminal investigation papers, also quite straightforward, as the rate is effectively linear, so we can simply draw a straight line and measure the gradient (by dividing the concentration change by the time interval) in order to syndrome, evaluate the reaction rate over this period. We may now perform a range of similar enzyme assays to evaluate how the initial velocity changes when the substrate or enzyme concentration is altered, or when the pH is changed. Investigation Papers? These studies will help us to essay, characterize the properties of the enzyme under study. The relationship between enzyme concentration and investigation term, the rate of the reaction is usually a simple one.
If we repeat the experiment just described, but add 10% more enzyme, the essay in prison reaction will be 10% faster, and if we double the enzyme concentration the reaction will proceed twice as fast. Thus there is a simple linear relationship between the reaction rate and the amount of criminal, enzyme available to catalyse the reaction (Figure 5). This relationship applies both to enzymes in vivo and to those used in biotechnological applications, where regulation of the amount of enzyme present may control reaction rates. When we perform a series of enzyme assays using the same enzyme concentration, but with a range of essay on marfan syndrome, different substrate concentrations, a slightly more complex relationship emerges, as shown in investigation papers, Figure 6. Effective Essay? Initially, when the investigation substrate concentration is increased, the rate of essay image, reaction increases considerably. However, as the substrate concentration is increased further the effects on the reaction rate start to decline, until a stage is reached where increasing the substrate concentration has little further effect on the reaction rate. At this point the enzyme is considered to be coming close to saturation with substrate, and demonstrating its maximal velocity ( V max ). Note that this maximal velocity is in fact a theoretical limit that will not be truly achieved in any experiment, although we might come very close to it. The relationship described here is a fairly common one, which a mathematician would immediately identify as a rectangular hyperbola. The equation that describes such a relationship is as follows: The two constants a and b thus allow us to describe this hyperbolic relationship, just as with a linear relationship ( y = mx + c ), which can be expressed by investigation, the two constants m (the slope) and of an essay, c (the intercept).
We have in fact already defined the constant a — it is V max . The constant b is a little more complex, as it is the value on the x -axis that gives half of the maximal value of y . In enzymology we refer to this as the Michaelis constant ( K m ), which is defined as the substrate concentration that gives half-maximal velocity. Our final equation, usually called the Michaelis–Menten equation, therefore becomes: In 1913, Leonor Michaelis and Maud Menten first showed that it was in fact possible to derive this equation mathematically from first principles, with some simple assumptions about the way in criminal term, which an enzyme reacts with a substrate to qualities of an effective analysis essay, form a product. Central to their derivation is the concept that the reaction takes place via the formation of an ES complex which, once formed, can either dissociate (productively) to release product, or else dissociate in the reverse direction without any formation of product. Thus the criminal investigation term papers reaction can be represented as follows, with k 1 , k ?1 and k 2 being the rate constants of the three individual reaction steps: The Michaelis–Menten derivation requires two important assumptions. Analytical Essay? The first assumption is that we are considering the initial velocity of the reaction ( v 0 ), when the product concentration will be negligibly small (i.e. [S] ? [P]), such that we can ignore the possibility of any product reverting to substrate. The second assumption is that the concentration of substrate greatly exceeds the concentration of enzyme (i.e. [S]?[E]). The derivation begins with an equation for the expression of the initial rate, the rate of formation of criminal, product, as the assignment log for students rate at which the term papers ES complex dissociates to form product.
This is based upon the rate constant k 2 and the concentration of the ES complex, as follows: 1. Since ES is an of an intermediate, its concentration is unknown, but we can express it in terms of criminal, known values. In a steady-state approximation we can assume that although the concentration of substrate and product changes, the concentration of the ES complex itself remains constant. The rate of formation of the ES complex and the rate of its breakdown must therefore balance, where: and. Hence, at essay life steady state: This equation can be rearranged to yield [ES] as follows: 2. The Michaelis constant K m can be defined as follows: Equation 2 may thus be simplified to: 3. Since the concentration of substrate greatly exceeds the concentration of enzyme (i.e. [S] ? [E]), the concentration of uncombined substrate [S] is almost equal to the total concentration of substrate. The concentration of uncombined enzyme [E] is papers, equal to the total enzyme concentration [E] T minus that combined with substrate [ES].
Introducing these terms to Equation 3 and essay in prison, solving for ES gives us the following: 4. We can then introduce this term into term Equation 1 to give: 5. The term k 2 [E] T in fact represents V max , the maximal velocity. Thus Michaelis and Menten were able to derive their final equation as: A more detailed derivation of the Michaelis–Menten equation can be found in qualities of an effective, many biochemistry textbooks (see section 4 of Recommended Reading section). There are also some very helpful web-based tutorials available on the subject. Michaelis constants have been determined for many commonly used enzymes, and are typically in the lower millimolar range (Table 5). It should be noted that enzymes which catalyse the same reaction, but which are derived from different organisms, can have widely differing K m values. Furthermore, an enzyme with multiple substrates can have quite different K m values for each substrate.
A low K m value indicates that the enzyme requires only criminal term, a small amount of substrate in order to essay, become saturated. Therefore the maximum velocity is reached at investigation term papers relatively low substrate concentrations. Qualities Effective? A high K m value indicates the term need for high substrate concentrations in order to achieve maximum reaction velocity. Thus we generally refer to K m as a measure of the log for affinity of the criminal enzyme for its substrate—in fact it is an inverse measure, where a high K m indicates a low affinity, and qualities effective essay, vice versa. The K m value tells us several important things about a particular enzyme. An enzyme with a low K m value relative to the physiological concentration of substrate will probably always be saturated with substrate, and will therefore act at a constant rate, regardless of variations in investigation term papers, the concentration of substrate within the physiological range. An enzyme with a high K m value relative to the physiological concentration of substrate will not be saturated with substrate, and its activity will therefore vary according to the concentration of substrate, so the rate of formation of product will depend on the availability of substrate. If an enzyme acts on several substrates, the substrate with the lowest K m value is frequently assumed to be that enzyme's ‘natural’ substrate, although this may not be true in all cases. If two enzymes (with similar V max ) in different metabolic pathways compete for the same substrate, then if we know the K m values for effective, the two enzymes we can predict the term papers relative activity of the two pathways.
Essentially the essay about in prison pathway that has the enzyme with the lower K m value is likely to be the ‘preferred pathway’, and more substrate will flow through that pathway under most conditions. For example, phosphofructokinase (PFK) is the enzyme that catalyses the first committed step in investigation papers, the glycolytic pathway, which generates energy in the form of ATP for the cell, whereas glucose-1-phosphate uridylyltransferase (GUT) is an enzyme early in the pathway leading to the synthesis of glycogen (an energy storage molecule). Both enzymes use hexose monophosphates as substrates, but the K m of PFK for its substrate is lower than that of GUT for its substrate. Analytical Image? Thus at lower cellular hexose phosphate concentrations, PFK will be active and GUT will be largely inactive. At higher hexose phosphate concentrations both pathways will be active. Investigation Term Papers? This means that the cells only store glycogen in times of plenty, and history question, always give preference to the pathway of ATP production, which is the more essential function. Very often it is not possible to estimate K m values from a direct plot of velocity against substrate concentration (as shown in Figure 6) because we have not used high enough substrate concentrations to come even close to estimating maximal velocity, and therefore we cannot evaluate half-maximal velocity and thus K m . Fortunately, we can plot our experimental data in a slightly different way in order to obtain these values. Papers? The most commonly used alternative is the Lineweaver–Burk plot (often called the double-reciprocal plot). This plot linearizes the hyperbolic curved relationship, and the line produced is easy to extrapolate, allowing evaluation of V max and features of an expository, K m . For example, if we obtained only the first seven data points in investigation papers, Figure 6, we would have difficulty estimating V max from a direct plot as shown in essay syndrome, Figure 7a.
However, as shown in Figure 7b, if these seven points are plotted on investigation, a graph of 1/velocity against 1/substrate concentration (i.e. a double-reciprocal plot), the data are linearized, and the line can be easily extrapolated to the left to provide intercepts on both the y -axis and the x -axis, from which V max and K m , respectively, can be evaluated. One significant practical drawback of ap us history question, using the Lineweaver–Burk plot is the excessive influence that it gives to measurements made at the lowest substrate concentrations. These concentrations might well be the most prone to error (due to difficulties in making multiple dilutions), and result in reaction rates that, because they are slow, might also be most prone to criminal investigation term papers, measurement error. Often, as shown in Figure 8, such points when transformed on the Lineweaver–Burk plot have a significant impact on the line of best fit estimated from the data, and therefore on the extrapolated values of both V max and K m . The two sets of points shown in Figure 8 are identical except for the single point at the top right, which reflects (because of the plot's double-reciprocal nature) a single point derived from a very low substrate concentration and a low reaction rate. However, this single point can have an enormous impact on the line of best fit and the accompanying estimates of kinetic constants. In fact there are other kinetic plots that can be used, including the Eadie–Hofstee plot, the Hanes plot and the Eisenthal–Cornish-Bowden plot, which are less prone to such problems. However, the expository Lineweaver–Burk plot is still the papers most commonly described kinetic plot in the majority of enzymology textbooks, and about life, thus retains its influence in undergraduate education. Enzymes are affected by pH and temperature. Various environmental factors are able to affect the rate of enzyme-catalysed reactions through reversible or irreversible changes in the protein structure. Term Papers? The effects of pH and temperature are generally well understood.
Most enzymes have a characteristic optimum pH at which the essay about velocity of the catalysed reaction is maximal, and above and criminal investigation, below which the velocity declines (Figure 9). The pH profile is dependent on a number of factors. As the pH changes, the ionization of groups both at history essay question 2011 the enzyme's active site and on the substrate can alter, influencing the rate of binding of the substrate to criminal papers, the active site. These effects are often reversible. For example, if we take an enzyme with an optimal pH (pH opt ) of 7.0 and place it in an environment at pH 6.0 or 8.0, the charge properties of the enzyme and the substrate may be suboptimal, such that binding and essay image, hence the reaction rate are lowered. If we then readjust the pH to 7.0, the optimal charge properties and hence the maximal activity of the enzyme are often restored. However, if we place the enzyme in a more extreme acidic or alkaline environment (e.g. at pH 1 or 14), although these conditions may not actually lead to changes in the very stable covalent structure of the protein (i.e. its configuration), they may well produce changes in term papers, the conformation (shape) of the protein such that, when it is returned to pH 7.0, the original conformation and hence the enzyme's full catalytic activity are not restored. It should be noted that the of an essay optimum pH of an enzyme may not be identical to that of its normal intracellular surroundings. Criminal? This indicates that the local pH can exert a controlling influence on log for students, enzyme activity. The effects of temperature on enzyme activity are quite complex, and can be regarded as two forces acting simultaneously but in opposite directions. As the papers temperature is raised, the history 2011 rate of molecular movement and hence the rate of investigation, reaction increases, but at the same time there is a progressive inactivation caused by denaturation of the enzyme protein.
This becomes more pronounced as the temperature increases, so that an assignment apparent temperature optimum (T opt ) is investigation papers, observed (Figure 10). Thermal denaturation is time dependent, and for analytical essay, an enzyme the term term ‘optimum temperature’ has little real meaning unless the duration of essay syndrome, exposure to that temperature is criminal term papers, recorded. The thermal stability of an enzyme can be determined by first exposing the protein to a range of temperatures for a fixed period of assignment students, time, and subsequently measuring its activity at one favourable temperature (e.g. 25°C). The temperature at which denaturation becomes important varies from one enzyme to another. Normally it is negligible below 30°C, and starts to criminal investigation term, become appreciable above 40°C. Essay About Life? Typically, enzymes derived from microbial sources show much higher thermal stability than do those from mammalian sources, and enzymes derived from extremely thermophilic microorganisms, such as thermolysin (a protease from Bacillus thermoproteolyticus ) and Taq polymerase (a DNA polymerase from criminal investigation papers Thermus aquaticus ), might be completely thermostable at 70°C and still retain substantial levels of activity even at 100°C. Enzymes are sensitive to ap us history, inhibitors. Substances that reduce the activity of an term enzyme-catalysed reaction are known as inhibitors.
They act by either directly or indirectly influencing the catalytic properties of the active site. Inhibitors can be foreign to the cell or natural components of it. Those in the latter category can represent an important element of the regulation of cell metabolism. Many toxins and assignment log for students, also many pharmacologically active agents (both illegal drugs and prescription and over-the-counter medicines) act by inhibiting specific enzyme-catalysed processes. Inhibitors are classified as reversible inhibitors when they bind reversibly to an enzyme. A molecule that is structurally similar to the normal substrate may be able to bind reversibly to the enzyme's active site and therefore act as a competitive inhibitor.
For example, malonate is a competitive inhibitor of the enzyme succinate dehydrogenase, as it is capable of binding to the enzyme's active site due to its close structural similarity to the enzyme's natural substrate, succinate (see below). When malonate occupies the active site of criminal investigation papers, succinate dehydrogenase it prevents the natural substrate, succinate, from binding, thereby slowing down the rate of oxidation of succinate to fumarate (i.e. inhibiting the reaction). One of the characteristics of competitive inhibitors is that they can be displaced from the about life in prison active site if high concentrations of investigation term papers, substrate are used, thereby restoring enzyme activity. Thus competitive inhibitors increase the K m of a reaction because they increase the concentration of substrate required to saturate the enzyme. However, they do not change V max itself.
In the case of certain enzymes, high concentrations of either the substrate or the product can be inhibitory. For example, invertase activity is analytical, considerably reduced in the presence of high concentrations of sucrose (its substrate), whereas the ?-galactosidase of Aspergillus niger is strongly inhibited by galactose (its product). Products of an enzyme reaction are some of the most commonly encountered competitive inhibitors. Other types of reversible inhibitor also exist. Non-competitive inhibitors react with the enzyme at a site distinct from the active site. Term Papers? Therefore the binding of the inhibitor does not physically block the substrate–binding site, but it does prevent subsequent reaction. Most non-competitive inhibitors are chemically unrelated to the substrate, and their inhibition cannot be overcome by increasing the substrate concentration. Such inhibitors in effect reduce the concentration of the essay syndrome active enzyme in solution, thereby reducing the V max of the reaction. Criminal Investigation Papers? However, they do not change the value of K m . Uncompetitive inhibition is rather rare, occurring when the inhibitor is only able to about, bind to the enzyme once a substrate molecule has itself bound. As such, inhibition is most significant at high substrate concentrations, and results in a reduction in the V max of the reaction. Uncompetitive inhibition also causes a reduction in criminal investigation term papers, K m , which seems somewhat counterintuitive as this means that the affinity of the enzyme for its substrate is essay syndrome, actually increased when the investigation term inhibitor is present.
This effect occurs because the binding of the inhibitor to the ES complex effectively removes ES complex and thereby affects the overall equilibrium of the reaction favouring ES complex formation. It is noteworthy however that since both V max and K m are reduced the observed reaction rates with inhibitor present are always lower than those in the absence of the uncompetitive inhibitor. Irreversible inhibitors and poisons. If an image inhibitor binds permanently to an enzyme it is investigation, known as an in prison irreversible inhibitor. Criminal Investigation? Many irreversible inhibitors are therefore potent toxins. Organophosphorus compounds such as diisopropyl fluorophosphate (DFP) inhibit acetylcholinesterase activity by reacting covalently with an important serine residue found within the active site of the enzyme. Essay? The physiological effect of this inactivation is interference with neurotransmitter inactivation at the synapses of nerves, resulting in the constant propagation of nerve impulses, which can lead to criminal investigation papers, death.
DFP was originally evaluated by ap us history 2011, the British as a chemical warfare agent during World War Two, and criminal, modified versions of this compound are now widely used as organophosphate pesticides (e.g. parathione, malathione). Allosteric regulators and the control of enzyme activity. Having spent time learning about enzyme kinetics and the Michaelis–Menten relationship, it is often quite disconcerting to essay life, find that some of the criminal investigation term papers most important enzymes do not in fact display such properties. Allosteric enzymes are key regulatory enzymes that control the activities of metabolic pathways by responding to inhibitors and activators. These enzymes in fact show a sigmoidal (S-shaped) relationship between reaction rate and substrate concentration (Figure 11), rather than the usual hyperbolic relationship. Essay? Thus for criminal, allosteric enzymes there is an on marfan area where activity is lower than that of an equivalent ‘normal’ enzyme, and also an criminal investigation papers area where activity is higher than that of an equivalent ‘normal’ enzyme, with a rapid transition between these two phases. Of An Analysis Essay? This is rather like a switch that can quickly be changed from ‘off’ (low activity) to criminal investigation papers, ‘on’ (full activity). Most allosteric enzymes are polymeric—that is, they are composed of at least two (and often many more) individual polypeptide chains. They also have multiple active sites where the substrate can bind. Much of our understanding of the analytical essay function of allosteric enzymes comes from studies of haemoglobin which, although it is investigation, not an enzyme, binds oxygen in history, a similarly co-operative way and term, thus also demonstrates this sigmoidal relationship. Ap Us 2011? Allosteric enzymes have an initially low affinity for papers, the substrate, but when a single substrate molecule binds, this may break some bonds within the enzyme and thereby change the shape of the protein such that the remaining active sites are able to bind with a higher affinity.
Therefore allosteric enzymes are often described as moving from a tensed state or T-state (low affinity) in which no substrate is bound, to a relaxed state or R-state (high affinity) as substrate binds. Other molecules can also bind to allosteric enzymes, at essay about life in prison additional regulatory sites (i.e. not at the active site). Molecules that stabilize the protein in its T-state therefore act as allosteric inhibitors, whereas molecules that move the investigation term papers protein to its R-state will act as allosteric activators or promoters. A good example of an allosteric enzyme is aspartate transcarbamoylase (ATCase), a key regulatory enzyme that catalyses the first committed step in the sequence of reactions that produce the pyrimidine nucleotides which are essential components of DNA and RNA. Ap Us History Essay? The reaction is as follows: The end product in the pathway, the pyrimidine nucleotide cytidine triphosphate (CTP), is an active allosteric inhibitor of the enzyme ATCase. Therefore when there is criminal term, a high concentration of CTP in the cell, this feeds back and inhibits the ATCase enzyme, reducing its activity and thus lowering the rate of log for, production of further pyrimidine nucleotides.
As the concentration of CTP in the cell decreases then so does the inhibition of criminal investigation term papers, ATCase, and the resulting increase in enzyme activity leads to the production of more pyrimidine nucleotides. This negative feedback inhibition is an important element of biochemical homeostasis within the of an expository essay cell. However, in investigation term, order to synthesize DNA and RNA, the cell requires not only qualities effective analysis, pyrimidine nucleotides but also purine nucleotides, and these are needed in roughly equal proportions. Purine synthesis occurs through a different pathway, but interestingly the final product, the purine nucleotide adenosine triphosphate (ATP), is a potent activator of the enzyme ATCase. This is logical, since when the criminal term cell contains high concentrations of purine nucleotides it will require equally high concentrations of pyrimidine nucleotides in order for these two types of log for students, nucleotide to combine to form the polymers DNA and RNA. Thus ATCase is able to criminal, regulate the production of pyrimidine nucleotides within the cell according to cellular demand, and also to ensure that pyrimidine nucleotide synthesis is synchronized with purine nucleotide synthesis—an elegant biochemical mechanism for the regulation of an extremely important metabolic process.
There are some rare, although important, cases of monomeric enzymes that have only features expository, one substrate-binding site but are capable of demonstrating the sigmoidal reaction kinetics characteristic of term, allosteric enzymes. Particularly noteworthy in this context is the monomeric enzyme glucokinase (also called hexokinase IV), which catalyses the phosphorylation of essay syndrome, glucose to criminal investigation term, glucose-6-phosphate (which may then either be metabolized by essay, the glycolytic pathway or be used in glycogen synthesis). It has been postulated that this kinetic behaviour is a result of individual glucokinase molecules existing in one of two forms—a low-affinity form and a high-affinity form. The low-affinity form of the enzyme reacts with its substrate (glucose), is investigation, then turned into the high-affinity form, and remains in that state for a short time before slowly returning to its original low-affinity form (demonstrating a so-called slow transition). Essay? Therefore at criminal investigation papers high substrate concentrations the essay about in prison enzyme is likely to criminal investigation, react with a second substrate molecule soon after the first one (i.e. while still in its high-affinity form), whereas at lower substrate concentrations the enzyme may transition back to essay on marfan syndrome, its low-affinity form before it reacts with subsequent substrate molecules. This results in its characteristic sigmoidal reaction kinetics. Origin, purification and uses of enzymes. Enzymes are essential components of animals, plants and term, microorganisms, due to the fact that they catalyse and features of an essay, co-ordinate the complex reactions of cellular metabolism.
Up until the 1970s, most of the commercial application of enzymes involved animal and criminal investigation, plant sources. At that time, bulk enzymes were generally only used within the life in prison food-processing industry, and enzymes from animals and plants were preferred, as they were considered to be free from the problems of term papers, toxicity and contamination that were associated with enzymes of microbial origin. However, as demand grew and as fermentation technology developed, the competitive cost of microbial enzymes was recognized and they became more widely used. Compared with enzymes from plant and animal sources, microbial enzymes have economic, technical and ethical advantages, which will now be outlined. The sheer quantity of enzyme that can be produced within a short time, and in a small production facility, greatly favours the use of microorganisms. For example, during the production of rennin (a milk-coagulating enzyme used in assignment, cheese manufacture) the traditional approach is to use the enzyme extracted from the stomach of a calf (a young cow still feeding on its mother's milk). The average quantity of rennet extracted from a calf's stomach is 10 kg, and it takes several months of intensive farming to produce a calf. Investigation? In comparison, a 1 000-litre fermenter of recombinant Bacillus subtilis can produce 20 kg of enzyme within 12 h. Thus the microbial product is expository essay, clearly preferable economically, and is free from the ethical issues that surround the use of criminal term, animals. Indeed, most of the cheese now sold in supermarkets is made from of an essay milk coagulated with microbial enzymes (so is suitable for vegetarians).
A further advantage of investigation term papers, using microbial enzymes is their ease of extraction. Many of the microbial enzymes used in biotechnological processes are secreted extracellularly, which greatly simplifies their extraction and purification. Microbial intracellular enzymes are also often easier to essay 2011, obtain than the equivalent animal or plant enzymes, as they generally require fewer extraction and purification steps. Animal and plant sources usually need to be transported to the extraction facility, whereas when microorganisms are used the investigation same facility can generally be employed for production and extraction. In addition, commercially important animal and log for, plant enzymes are often located within only one organ or tissue, so the term papers remaining material is essentially a waste product, disposal of which is required.
Finally, enzymes from plant and animal sources show wide variation in yield, and may only be available at certain times of year, whereas none of about life in prison, these problems are associated with microbial enzymes. Microbial enzymes often have properties that make them more suitable for commercial exploitation. Criminal? In comparison with enzymes from of an essay animal and plant sources, the stability of microbial enzymes is usually high. For example, the high temperature stability of criminal term, enzymes from thermophilic microorganisms is often useful when the process must operate at high temperatures (e.g. during starch processing). Microorganisms are also very amenable to genetic modification to produce novel or altered enzymes, using relatively simple methods such as plasmid insertion.
The genetic manipulation of animals and plants is technically much more difficult, is essay, more expensive and papers, is still the subject of in prison, significant ethical concern, especially in the U.K. Enzymes may be intracellular or extracellular. Although many enzymes are retained within the cell, and may be located in specific subcellular compartments, others are released into the surrounding environment. The majority of enzymes in industrial use are extracellular proteins from either fungal sources (e.g. Aspergillus species) or bacterial sources (e.g.
Bacillus species). Examples of these include ?-amylase, cellulase, dextranase, proteases and amyloglucosidase. Many other enzymes for investigation term, non-industrial use are intracellular and are produced in much smaller amounts by the cell. Examples of these include asparaginase, catalase, cholesterol oxidase, glucose oxidase and glucose-6-phosphate dehydrogenase. Within the cell, enzymes are generally found along with other proteins, nucleic acids, polysaccharides and on marfan, lipids. The activity of the enzyme in relation to the total protein present (i.e. the specific activity) can be determined and used as a measure of enzyme purity. A variety of methods can be used to remove contaminating material in order to purify the papers enzyme and increase its specific activity. Enzymes that are used as diagnostic reagents and in clinical therapeutics are normally prepared to a high degree of purity, because great emphasis is placed on on marfan syndrome, the specificity of the reaction that is being catalysed. Clearly the higher the level of purification, the greater the cost of enzyme production. Criminal? In the case of many bulk industrial enzymes the degree of purification is less important, and such enzymes may often be sold as very crude preparations of culture broth containing the growth medium, organisms (whole or fragmented) and enzymes of analytical image, interest.
However, even when the cheapest bulk enzymes are utilized (e.g. Criminal Term Papers? proteases for essay, use in criminal investigation papers, washing powders), the enzyme cost can contribute around 5–10% of the final product value. At the features of an expository end of a fermentation in which a microorganism rich in the required enzyme has been cultured, the broth may be cooled rapidly to 5°C to prevent further microbial growth and stabilize the criminal term enzyme product. The pH may also be adjusted to optimize enzyme stability. If the enzyme-producing organism is a fungus, this may be removed by centrifugation at low speed. Log For Students? If the enzyme source is bacterial, the bacteria are often flocculated with aluminum sulfate or calcium chloride, which negate the charge on the bacterial membranes, causing them to criminal investigation, clump and thus come out of suspension. Extracellular enzymes are found in the liquid component of the pretreatment process. However, intracellular enzymes require more extensive treatment. The biomass may be concentrated by centrifugation and washed to remove medium components.
The cellular component must then be ruptured to release the enzyme content. This can be done using one or more of the following processes: • ball milling (using glass beads) • enzymic removal of the analytical essay cell wall. • liquid shearing through a small orifice at high pressure (e.g. within a French press) Separation of enzymes from the resulting solution may then involve a variety of criminal investigation term, separation processes, which are often employed in a sequential fashion. The first step in essay, an enzyme purification procedure commonly involves separation of the proteins from the non-protein components by a process of investigation, salting out. Qualities Essay? Proteins remain in aqueous solution because of interactions between the hydrophilic (water-loving) amino acids and the surrounding water molecules (the solvent). If the ionic strength of the solvent is term, increased by adding an assignment students agent such as ammonium sulfate, some of the water molecules will interact with the salt ions, thereby decreasing the number of water molecules available to criminal investigation, interact with the protein.
Under such conditions, when protein molecules cannot interact with the solvent, they interact with each other, coagulating and coming out of essay on marfan, solution in the form of a precipitate. This precipitate (containing the criminal investigation term enzyme of interest and other proteins) can then be filtered or centrifuged, and separated from the supernatant. Since different proteins vary in the extent to which they interact with water, it is possible to perform this process using a series of additions of ammonium sulfate, increasing the ionic strength in a stepwise fashion and removing the essay about life in prison precipitate at each stage. Thus such fractional precipitation is not only capable of separating protein from non-protein components, but can also enable separation of the criminal investigation term enzyme of interest from some of the other protein components. Subsequently a wide variety of techniques may be used for further purification, and steps involving chromatography are standard practice. Ion-exchange chromatography is often effective during the early stages of the purification process. The protein solution is added to a column containing an log for students insoluble polymer (e.g. cellulose) that has been modified so that its ionic characteristics will determine the type of mobile ion (i.e. cation or anion) it attracts.
Proteins whose net charge is opposite to that of the criminal investigation term papers ion-exchange material will bind to it, whereas all other proteins will pass through the column. A subsequent change in about life in prison, pH or the introduction of a salt solution will alter the electrostatic forces, allowing the retained protein to be released into solution again. Gel filtration can be utilized in the later stages of a purification protocol to separate molecules on investigation term, the basis of molecular size. Ap Us History Essay Question 2011? Columns containing a bed of cross-linked gel particles such as Sephadex are used. These gel particles exclude large protein molecules while allowing the criminal term entry of essay on marfan, smaller molecules. Separation occurs because the larger protein molecules follow a path down the column between the investigation term Sephadex particles (occupying a smaller fraction of the column volume). Larger molecules therefore have a shorter elution time and are recovered first from the gel filtration column. Affinity chromatography procedures can often enable purification protocols to be substantially simplified.
Typically, with respect to enzyme purification, a column would be packed with a particulate stationary phase to which a ligand molecule such as a substrate analogue, inhibitor or cofactor of the enzyme of interest would be firmly bound. As the sample mixture is about, passed through the criminal investigation term column, the enzyme interacts with, and binds, to the immobilised ligand, being retained within the column as all of the other components of the mixture pass through the column unrewarded. Subsequently a solution of the ligand is introduced to the column to release (elute) and thereby recover the bound enzyme from the column in a highly purified form. Nowadays numerous alternative affinity chromatography procedures exist that are able to separate enzymes by binding to areas of the molecule away form their active site. Advances in molecular biology enable us to purify recombinant proteins, including enzymes, through affinity tagging. In a typical approach the gene for essay on marfan syndrome, the enzyme of interest would be modified to code for a further short amino acid sequence at either the investigation N- or C- terminal. For example, a range of features, polyhistidine tagging procedures are available to criminal papers, yield protein products with six or more consecutive histidine residues at their N- or C- terminal end. When a mixture containing the tagged protein of interest is subsequently passed through a column containing a nickel-nitrilotriacetic acid (Ni-NTA) agarose resin, the histidine residues on ap us essay question, the recombinant protein bind to criminal term, the nickel ions attached to the support resin, retaining the protein, whilst other protein and non-protein components pass through the column. Assignment Log For Students? Elution of the bound protein can then be accomplished by adding imidazole to the column, or by reducing the pH to 5-6 to displace the criminal His-tagged protein from the nickel ions. Such techniques are therefore capable of rapidly and analytical essay, highly effectively isolating an criminal papers enzyme from a complex mixture in only one step, and typically provide protein purities of features, up to 95%. If more highly purified enzyme products are required, other supplemental options are also available, including various forms of preparative electrophoresis e.g. disc-gel electrophoresis and criminal investigation, isoelectric focusing.
Enzymes are antigenic, and since problems occurred in the late 1960s when manufacturing workers exhibited severe allergic responses after breathing enzyme dusts, procedures have now been implemented to ap us history question 2011, reduce dust formation. These involve supplying enzymes as liquids wherever possible, or increasing the particle size of term papers, dry powders from 10 ?m to 200–500 ?m by either prilling (mixing the enzyme with polyethylene glycol and preparing small spheres by atomization) or marumerizing (mixing the essay on marfan syndrome enzyme with a binder and criminal investigation term, water, extruding long filaments, converting them into spheres in a marumerizer, drying them and covering them with a waxy coat). Although many industrial processes, such as cheese manufacturing, have traditionally used impure enzyme sources, often from animals or plants, the development of much of modern industrial enzymology has gone hand in hand with the commercial exploitation of features expository, microbial enzymes. These were introduced to the West in around 1890 when the Japanese scientist Jokichi Takamine settled in the U.S.A. and set up an investigation enzyme factory based on Japanese technology. The principal product was Takadiastase, a mixture of amylolytic and proteolytic enzymes prepared by cultivating the fungus Aspergillus oryzae on rice or wheat bran. Takadiastase was marketed successfully in the U.S.A. as a digestive aid for the treatment of dyspepsia, which was then believed to of an effective, result from the incomplete digestion of starch. Bacterial enzymes were developed in criminal investigation term papers, France by August Boidin and Jean Effront, who in 1913 found that Bacillus subtilis produced a heat-stable ?-amylase when grown in a liquid medium made by assignment log for students, extraction of term, malt or grain. The enzyme was primarily used within the textile industry for the removal of the ap us 2011 starch that protects the warp in the manufacture of cotton.
In around 1930 it was found that fungal pectinases could be used in the preparation of fruit products. In subsequent years, several other hydrolases were developed and sold commercially (e.g. Criminal Investigation Term? pectosanase, cellulase, lipase), but the technology was still fairly rudimentary. After World War Two the fermentation industry underwent rapid development as methods for the production of antibiotics were developed. These methods were soon adapted for the production of enzymes. In the 1960s, glucoamylase was introduced as a means of analytical essay, hydrolysing starch, replacing acid hydrolysis. Subsequently, in the 1960s and 1970s, proteases were incorporated into criminal papers detergents and then glucose isomerase was introduced to produce sweetening agents in the form of high-fructose syrups.
Since the 1990s, lipases have been incorporated into washing powders, and a variety of immobilized enzyme processes have been developed (see section on enzyme immobilization), many of which utilize intracellular enzymes. Currently, enzymes are used in essay life, four distinct fields of commerce and technology (Table 6): • as industrial catalysts. • as therapeutic agents. • as analytic reagents. • as manipulative tools (e.g. in genetics). Of the thousands of different types of enzymes, about 95% are available from suppliers in quantities ranging from ?g to kg, provided essentially for research purposes. Around 40–50 enzymes are produced on an industrial scale (i.e. ranging from criminal investigation multiple kilograms to tonnes per students annum).
The global enzyme market is currently dominated by the hydrolases, especially the criminal term papers proteases, together with amylases, cellulases and lipases supplied either as liquid concentrates or as powders or granules that release the soluble enzyme on dissolution. Global production is dominated by essay, two companies, which between them supply more than two-thirds of the global enzyme market, namely the Danish company Novozymes, with a market share of 47%, and the U.S. company DuPont (which recently acquired Genencor), with 21%. The value of the world enzyme market has increased steadily from criminal investigation term ?110 million in essay on marfan syndrome, 1960 to ?200 million in 1970, ?270 million in 1980, ?1 000 million in 1990 and over ?2 000 million in 2010. Food and beverage enzymes represented the largest sector of the industrial enzymes market in 2010, with a value of ?750 million, and the market for enzymes for technical applications (including diagnostic applications, research and criminal, biotechnology) accounted for a further ?700 million. Estimates of qualities of an effective, future demand are in the range of ?4 000–5 000 million between 2015 and papers, 2016, growing at a rate of 6–7% annually. The developing economies of the Asia-Pacific Region, the Middle East and Africa are now seen to be emerging as the fastest growing markets for industrial enzymes. Microbial enzymes are typically produced in essay life, batches by criminal investigation term, culturing the producing organism within a batch fermenter. Fermentation typically lasts between 30 and 150 h, with the optimum enzyme yield for the process falling somewhere between the assignment students optimum biomass yield and the point of maximal enzyme activity within the cells. Relatively small fermenters with a volume of 10–100 m 3 are generally employed, allowing flexibility where a number of criminal investigation term, different products are being produced.
Many production systems are optimized by means of a fed-batch process, in which substrates are gradually fed into the reactor over the course of the fermentation, rather than being provided all at once at the start of the process. True continuous culture techniques have been used in laboratory-scale studies, but have not been widely implemented on a commercial scale, although Novozymes does have a continuous process for the production of glucose isomerase, since this is a larger-volume market and essay, the company has a very strong market share. During the production of criminal investigation papers, commercially important products via enzymatic catalysis, soluble enzymes have traditionally been used in ap us history essay question 2011, batch processes that employ some form of stirred-tank reactor (STR). Papers? In these processes, at the end of the batch run the product must be separated from any unused substrate, and also from the enzyme catalyst. Removal of the enzyme at this stage can be achieved by thermal denaturation (only if the product is thermostable) or by history essay question 2011, ammonium sulfate precipitation or ultrafiltration. These processes represent a costly downstream processing stage and generally render the enzyme inactive, so when a new batch run is to be started a fresh batch of enzyme is required. Immobilized enzyme systems, in contrast, ‘fix’ the enzyme so that it can be reused many times, which has a significant impact on production costs. As a very simple example, if an criminal investigation enzyme is assignment log for students, mixed with a solution of warm (but not too hot) agar and this is allowed to investigation, set, the enzyme will be entrapped (for the purposes of this example let us ignore the fact that the enzyme will gradually leak out of log for, this gel). The agar can then be cut up into cubes and these can be placed in a STR, together with substrate, as shown in Figure 12. Again the reaction would be allowed to proceed (and it might actually be slower due to diffusional constraints and other effects described later).
At the end of the batch run the criminal investigation catalyst can now be easily separated from the product by passing the reactor contents through a coarse mesh. Immediately an ap us history 2011 important downstream processing step has been carried out and, just as importantly, the active enzyme has been recovered so that it can be reused for the next batch run. This ease of criminal investigation papers, separation of essay syndrome, enzyme from product is a major advantage of all immobilized systems over their counterparts that use free (i.e. soluble) enzyme. This physical advantage of ease of criminal, reuse of about, immobilized biocatalysts is one of the main reasons why such systems are favoured commercially. However, immobilization may also produce biochemical changes that lead to enhanced biocatalyst stability, which may be manifested as: • an criminal investigation papers increased rate of catalysis.
• prolonged duration of catalysis. • greater operational stability to image, extremes of pH, temperature, etc. The particular advantage(s) conferred by criminal investigation term papers, immobilization will therefore differ from one system to another. It should be noted that often there may be no biochemical advantage at all, and the simple physical advantage of ease of separation of the biocatalyst from the product may be sufficient to favour the commercial development of an immobilized process. At this point one problem that will immediately spring to mind for most students is that they have always been taught to fully mix all of the features expository reagents of a reaction, yet the basic principle of immobilization is to partition the biocatalyst into a distinct phase, rather than mix it homogeneously with the substrate. Criminal Term? Will this not cause reaction rates to be low? The answer to this question is yes, and the relationship between the activity of an immobilized system and a non-immobilized system can be expressed as the history essay effectiveness factor (?), where: Thus an immobilized system with an effectiveness factor of 0.1 would show only 10% of the activity of term papers, a non-immobilized system with the same amount of enzyme and operating under the same conditions. At first sight this might appear to essay syndrome, be a major problem. However, if it is possible to reuse the biocatalyst many times this is still economically viable, even with systems that have a low effectiveness factor.
In principle, therefore, for economic viability: Thus if an immobilized system has an effectiveness factor of 0.1 (i.e. 10%) and criminal, we can reuse the biocatalyst 10 times, we essentially achieve the same overall catalytic activity with both the non-immobilized system and the immobilized one. However, if we are able to reuse the biocatalyst 100 times we in fact obtain 10 times more total activity from the immobilized system than from the 2011 equivalent non-immobilized system, so the term papers immobilized system may be economically preferable. Once a biocatalyst has been immobilized it can also be put in a range of continuous-flow reactors, enabling a continuous supply of substrate to be turned into product as it passes through the reactor. The control of such continuous-flow reactors can be highly automated, leading to considerable savings in production costs. For example, a STR can be easily modified to produce a continuous-flow stirred-tank reactor (CSTR) (Figure 13a), in expository essay, which the enzyme is held within the reactor by a coarse mesh, and criminal papers, the product continuously flows out on marfan syndrome of the reactor as substrate is pumped in. It is also possible to produce a packed-bed reactor (PBR) (Figure 13b), in which the agar cubes are packed into a column and the substrate is pumped through the bed without any need for stirring. CSTRs and PBRs enable the enzyme to be reused many times before it needs to criminal investigation term papers, be replaced. Ap Us? For example, in investigation term papers, the production of about life, high-fructose syrups, the immobilized glucose isomerase enzyme would typically be used continuously for between 2 and 4 months, and only after this time (when its activity would have dropped to 25% of the original level) would it need to criminal investigation papers, be replaced. The overall operating costs of continuous-flow reactors are often significantly lower than those of equivalent batch processes.
Batch reactors need to be emptied and image, refilled frequently at regular intervals. Not only is this procedure expensive, but it also means that there are considerable periods of time when such reactors are not productive (so-called ‘downtime’). Criminal Term? In addition, batch processes make uneven demands on both labour and services. They may also result in qualities effective analysis, pronounced batch-to-batch variations, as the reaction conditions change with time, and they may be difficult to scale up, due to investigation term, the changing power requirements for ap us 2011, efficient mixing. Due to their higher overall process efficiency, continuous processes using immobilized enzymes may be undertaken in production facilities that are around 10 to 100 times smaller than those required for equivalent batch processes using soluble enzymes. Term Papers? Therefore the capital costs involved in setting up the facility are also considerably lower. It should be noted that although the agar entrapment method described here has provided a useful example, it is not a particularly effective form of immobilization. The high temperature required to prevent the agar from setting may lead to thermal inactivation of the enzyme, and the agar gel itself is very porous and will allow the enzyme to leak out into the surrounding solution. There are in fact thousands of different techniques of immobilization, all of which are much more effective than our example.
In general these techniques can be classified as belonging to about in prison, one of three categories (Figure 14): The physical adsorption of an enzyme to a supporting matrix is the oldest method of immobilization. As early as 1916, J.M. Nelson and Edward G. Griffin described the adsorption of yeast invertase on investigation papers, to activated charcoal, and the subsequent use of this preparation for sucrose hydrolysis. Assignment Log For? Over the criminal term papers years a variety of adsorbents have been used, including cellulose, Sephadex, polystyrene, kaolinite, collagen, alumina, silica gel and glass.
Such immobilization procedures are extremely easy to perform, as the adsorbent and enzyme are simply stirred together for a time (typically minutes to hours). The binding forces that immobilize the catalyst on the support may involve hydrogen bonds, van der Waals forces, ionic interactions or hydrophobic interactions. Essay Syndrome? Such forces are generally weak in comparison with covalent bonds—for example, a hydrogen bond has an energy content of about 20 kJ mol ?1 , compared with 200–500 kJ mol ?1 for investigation, a covalent bond. Thus, when using such methods, yields (i.e. the amount of enzyme bound per unit of adsorbent) are generally low. In addition, adsorption is generally easily reversed, and image, can lead to desorption of the enzyme at a critical time.
However, despite these limitations, such a method was used in the first commercial immobilized enzyme application, namely DEAE–Sephadex-immobilized l -amino acid acylase, in 1969. Papers? DEAE–Sephadex is an ion-exchange resin that consists of an inert dextran particle activated by analytical essay image, the addition of numerous diethylaminoethyl groups. Particles of this material remain positively charged at pH 6–8 (see Figure 15a) and thus bind strongly to proteins, which are generally negatively charged in this pH range. If the pH is kept constant, the enzyme and criminal term papers, support will remain ionically linked. About In Prison? However, when over time the investigation term enzyme loses its activity through denaturation, the pH can be adjusted to students, a more acidic value, the old enzyme will be desorbed, and the pH can then be readjusted back to pH 6–8 and a fresh batch of criminal term, enzyme bound. Thus the support matrix may be used many times, giving the process significant economic benefits. Clearly DEAE–Sephadex immobilization is only of about life in prison, value for enzymes that have a neutral-to-alkaline pH optimum. For enzymes that function best under acidic conditions, CM–Sephadex is more suitable.
This contains carboxymethyl groups that remain negatively charged at pH 3.5–4.5 (Figure 15b). Proteins at investigation term papers this pH are generally positively charged and will thus ionically bind to the support. Desorption of the enzyme will occur when the pH is adjusted to a more alkaline value. Due to the simplicity and controllability of this immobilization procedure, combined with the economic benefits of reuse of the of an essay support, ion-exchange materials are now widely used as the method of choice in many industrial settings. Immobilization of enzymes by covalent bonding to activated polymers is a widely used approach since, although it is often a tedious procedure, it is capable of producing an immobilized enzyme that is firmly bound to its support. The range of polymers and chemical coupling procedures that are used is enormous. The history of covalent bonding for enzyme immobilization dates back to 1949, when F. Michael and J. Ewers used the criminal investigation term azide derivative of carboxymethylcellulose to immobilize a variety of proteins. Activated cellulose supports continue to be popular due to features of an expository, their inherent advantages of high hydrophilicity, ready availability, potential for derivatization, and the ease with which cellulose-based polymers can be produced either as particulate powders or as membranous films.
It is often more effective not to build the criminal reactive group into the cellulose itself, but instead to use a chemical ‘bridge’ between the cellulose and the enzyme molecule. The requirements for such a bridging or linking molecule are that it must be small, and that once it has reacted with the support it must have a further reactive group capable of reacting with the enzyme. An example of such a bridging molecule is glutaraldehyde, which contains two aldehyde groups, one at either end of features expository essay, its (CH 2 ) 3 moiety. At neutral pH values the aldehyde groups will react with free amino groups. Thus one end of the criminal papers glutaraldehyde molecule may be attached to essay life, the support, and the other to the enzyme. Covalently immobilized enzymes are strongly bound to their support, so when the proteins denature they are difficult to remove (in contrast to adsorption, as described earlier). Therefore it is usual for criminal investigation papers, both the enzyme and the support to be replaced. This may result in higher operational costs compared with adsorption techniques in which the support may be reused.
The entrapment of an enzyme can be achieved in a number of ways: • inclusion within the matrix of a highly cross-linked polymer. • separation from the bulk phase by analysis, a semi-permeable ‘microcapsule’ • dissolution in a distinct non-aqueous phase. An important feature of entrapment techniques is investigation term papers, that the enzyme is not in of an effective essay, fact attached to criminal term papers, anything. Consequently there are none of the steric problems associated with covalent or adsorption methods (i.e. the possibility of the enzyme binding in such a way that its active site is obstructed by part of the supporting polymer matrix). The example of an enzyme retained in agar, described earlier, is a useful illustration of entrapment. A preferable alternative involves mixing the catalyst with sodium alginate gel and extruding this into a solution of calcium chloride to produce solid calcium alginate particles. This technique has the advantage of not requiring the use of high temperatures. Assignment Log For Students? However, although it is a popular activity in teaching laboratories, outside that setting it is investigation papers, generally unsuitable for the immobilization of purified enzymes, as these are often able to leak out of the gel. Entrapment techniques for features, purified enzymes are more likely to involve retaining the term papers enzyme behind some form of ultrafiltration membrane. However, gel entrapment procedures may be useful when dealing with larger catalysts, such as whole cells.
For example, gel-immobilized living yeast cells have been used successfully in the manufacture of champagne by Moet Chandon. Immobilization: changes in enzyme properties. Earlier in this essay it was suggested that immobilization might change the properties of an enzyme to enhance its stability. Initially it was believed that such enhanced stability resulted from the formation of bonds between the enzyme and the supporting matrix that physically stabilize the structure of the protein. Indeed there are some published reports that describe this phenomenon.
With regard to the stabilization of essay life, proteolytic enzymes, which often exhibit more prolonged activity in the immobilized state, this is most probably explained by the fact that such proteases in free solution are prone to autodigestion (i.e. enzyme molecules cleave the peptide bonds of adjacent enzyme molecules), a process that is largely prevented when they are fixed to a supporting matrix. However, the effects of immobilization are more often due to the supporting matrix changing the microenvironment around the enzyme and/or introducing diffusional constraints that modify the activity of the catalyst. Consider, for example, immobilization of the enzyme by adsorption on to a polyanionic (negatively charged) support such as cellulose. If the substrate is a cation (i.e. positively charged), it will be attracted to the support and thus to the enzyme. Criminal Investigation Term Papers? In this case the enzyme might well display higher activity, as the substrate concentration in analytical image, its microenvironment would be higher than that in the surrounding bulk phase. Other cations would also be attracted, and importantly these would include H + ions. Thus the microenvironment would also be enriched in H + ions, so the pH surrounding the enzyme would be lower than the pH of the term bulk phase. Consequently the enzyme would also exhibit an altered pH profile compared with that of features of an, its soluble counterpart. In addition, the immobilization matrix might act as a barrier to the diffusion of substrates, products and criminal investigation term, other molecules.
For example, if a high enzyme loading was put into of an effective analysis essay a gel particle and criminal term papers, this was then immersed in substrate solution, the substrate would diffuse into the gel and rapidly be converted into product. Enzyme molecules entrapped deeper within the gel particle might therefore be inactive simply because they had not received any substrate to work on (i.e. all of the substrate was converted to product in effective, the outer layers of the particle). Criminal Papers? Although this is of an expository essay, obviously somewhat inefficient, it does have one useful effect. When over time the enzyme within the system denatures, the loss of activity of the criminal investigation enzyme in the outer part of the in prison particle means that substrate will now diffuse deeper into the particle to reach the previously unused core enzyme molecules. In effect this inner reserve of enzyme will offset the loss of enzyme activity through denaturation, so the system will show little or no overall loss of activity.
This explains the observation that immobilized systems often have a longer operational lifetime than their soluble equivalents. In addition, it is of interest that enzymes bound to natural cell membranes (phospholipid bilayers) within living cells will also probably demonstrate these effects, and immobilized systems thus provide useful models for the study of such membrane-bound proteins in living cells. The major industrial processes that utilize immobilized enzymes are listed in Table 7. Sales of immobilized enzymes peaked in investigation papers, 1990, when they accounted for about 20% of all industrial enzyme sales, almost entirely due to the use of glucose isomerase for the production of sweetening agents. Other commercial applications utilize penicillin acylase, fumarase, ?–galactosidase and amino acid acylase. Since 2000, although there has been consistent growth in enzyme markets, few new processes employing immobilized enzymes have been introduced. The following three examples highlight many of the biochemical, technological and economic considerations relating to of an essay, the use of immobilized enzymes on a commercial and term, industrial scale. Undoubtedly the most significant large-scale application of immobilized enzymes involves the production of high-fructose corn syrup (HFCS). Although most of the general public believe that sucrose is responsible for the ‘sweetness’ of food and features essay, drinks, there have been significant efforts to replace sucrose with alternative, and often cheaper, soluble caloric sweetening agents. HFCS is a soluble sweetener that has been used in term papers, many carbonated soft drinks since the 1980s, including brand-name colas such as Coca-Cola and essay in prison, Pepsi-Cola.
HFCS is produced by the enzymatic digestion of starch derived from corn (maize). Developments in HFCS production have been most prominent in countries such as the criminal term U.S.A., which have a high capacity to produce starch in the form of corn, but which do not cultivate significant amounts of sugar cane or sugar beet, and must therefore import either the raw products (for processing) or the refined sugar (sucrose) itself. Simple corn syrups can be manufactured by breaking down starch derived from corn using the enzyme glucoamylase alone or in combination with ?-amylase. These enzymes are cheap and can be used in a soluble form. Features Of An Essay? Since starch has to be extracted from corn at high temperatures (because starch has poor solubility at low temperatures and forms very viscous solutions), the process utilizes enzymes from thermophilic organisms, which have very high temperature optima. Simple corn syrup is therefore composed predominantly of glucose, which unfortunately has only 75% of the sweetness of sucrose. However, in order to make the investigation term papers syrup sweeter the enzyme glucose isomerase, which catalyses the of an effective analysis following reaction, can be employed: This enzyme (described previously in the section on properties and mechanisms of enzyme action) will produce a roughly 50:50 mixture of glucose and criminal term, fructose at equilibrium, and since fructose has 150% of the sweetness of sucrose, this glucose:fructose mixture will have a similar level of sweetness to sucrose.
However, glucose isomerase is an intracellular bacterial enzyme, and would be prohibitively expensive to use in a soluble form. This makes it an ideal candidate for analytical image, use in an immobilized process. The first glucose isomerase enzyme to be isolated was obtained from species of Pseudomona s in 1957, and more useful enzymes were isolated throughout the 1960s from species of term papers, Bacillus and Streptomyces. Ap Us Essay? In 1967, the Clinton Corn Processing Company of term papers, Iowa, U.S.A. (later renamed CPC International) introduced a batch process that utilized an immobilized glucose isomerase enzyme, and by 1972 the of an effective analysis essay company had developed a continuous process for the manufacture of HFCS containing 42% fructose using a glucose isomerase enzyme immobilized on a DEAE ion-exchange support. During the late 1970s, advances in enzymology, process engineering and fractionation technology led to the production of syrups with a higher fructose content, and today HFCS containing 55% fructose is generally produced, and is commonly used in soft drinks, although 42% fructose syrups are still also produced for use in some processes, including the production of bakery foodstuffs. In 2010, the U.S. production of HFCS was approximately 8 million metric tons, accounting for term, 37% of the U.S. caloric sweetener market, and it is estimated that today about 5% of the entire corn crop in the U.S.A. is used to produce HFCS. Within the dairy industry the life in prison production of 1 kg of criminal investigation term papers, cheese requires about 10 litres of milk, and produces about life in prison 9 litres of whey as a waste product. Whey is a yellowish liquid containing 6% dry matter, of which nearly 80% is lactose. The enzyme lactase (?-galactosidase) may be used to investigation term papers, break down lactose to of an analysis essay, its constituent monosaccharides, namely glucose and galactose, which are more soluble than lactose, and have potential uses as carbon sources in microbial fermentation, and can also be used as caloric sweeteners. Valio Ltd of Finland has developed arguably the most successful commercial process for the treatment of whey.
Using a lactase enzyme obtained from Aspergillus , immobilized by adsorption and cross-linked on to a support resin, whey syrups are produced that have been utilized as an ingredient in drinks, ice cream and confectionery products. The Aspergillus enzyme has an acid pH optimum of 3–5, and by papers, operating at low pH the process avoids excessive microbial contamination. Treatment plants that utilize 600-litre columns have been built in Finland, and these are used to treat 80 000 litres of whey per day. This technology has also been used to produce whey syrups in England (by Dairy Crest) and in Norway. Similar technology can also be used to remove lactose from qualities essay milk. Lactose-free milk is produced for consumption by those who have lactose intolerance (a genetic condition), and also for consumption by pets such as cats, which are often unable to digest lactose easily. The first industrial processing facility to use immobilized lactase to investigation term, treat milk was opened in 1975, when Centrale del Latte of Milan, Italy, utilized a batch process in which yeast ( Saccharomyces ) lactase, with a neutral pH optimum of 6–8, was immobilized within hollow permeable fibres.
This process was capable of treating 10 000 litres of milk per day, and was operated at low temperature to prevent microbial contamination. Production of semi-synthetic penicillins. High yields of natural penicillins are obtained from features expository essay species of the term papers fungus Penicillium through fermentation processes. However, over the years many microbial pathogens have become resistant to natural penicillins, and are now only treatable with semi-synthetic derivatives. These are produced through cleavage of natural penicillin, such that the G or V side chain is removed from the 6-aminopenicillanic acid (6-APA) nucleus of the molecule: Thereafter, by students, attachment of a chemically different side chain, a semi-synthetic penicillin product (e.g. ampicillin, amoxicillin) can be formed. In addition, the 6-APA can undergo chemical ring expansion to yield 7-aminodesacetoxycephalosporanic acid (7-ADCA), which can then be used to generate a number of important cephalosporin antibiotics (e.g. cephalexin, cephradine, cefadroxil). The development of immobilized penicillin G acylase dates back to research conducted in 1969 by University College London and Beecham Pharmaceuticals in the U.K. Penicillin G acylases are intracellular enzymes found in E. coli and investigation term papers, a variety of other bacteria, and the Beecham process immobilized the E. coli enzyme on a DEAE ion-exchange support.
Later systems used more permanent covalent bonding to attach the enzyme to the support. In the 1980s and essay, 1990s, world production of penicillins was dominated by European manufacturers, which accounted for production of around 30 000 tonnes of penicillin per annum, 75% of criminal term papers, which was used for the manufacture of semi-synthetic penicillins and history essay question 2011, cephalosporins. Term Papers? However, over the past 10 years, due to essay life, increasing costs of labour, energy and raw materials, more bulk manufacturing has moved to criminal term papers, the Far East, where China, Korea and India have become major producers. Essay About? The market currently suffers from significant overcapacity, which has driven down the unit cost of penicillin and cephalosporin products. Criminal? However, penicillins and essay, cephalosporins still represent one of the world's major biotechnology markets, with annual sales of papers, about ?10 000 million, accounting for qualities, 65% of the entire global antibiotics market. Enzymes have a wide variety of uses in criminal, analytical procedures. Their specificity and potency allow both detection and amplification of a target analyte. ‘Wet chemistry’ enzyme-based assays for the detection and quantification of a variety of substances, including drugs, are widespread. Enzymes also play a key role in immunodiagnostics, often being used as the agent to amplify the signal—for example, in enzyme-linked immunosorbent assays (ELISAs). Within DNA-fingerprinting technology, the enzyme DNA polymerase plays a key role in the amplification of DNA molecules in the polymerase chain reaction. However, ‘wet chemistry’ analytical methods are increasingly being replaced by the use of biosensors—that is, self-contained integrated devices which incorporate a biological recognition component (usually an immobilized enzyme) and an electrochemical detector (known as a transducer). Much of the of an technological development of biosensors has been motivated by the need to measure blood glucose levels.
In 2000, the criminal investigation term World Health Organization estimated that over 170 million people had diabetes, and predicted that this figure will rise to over 360 million by 2030. In view of assignment log for, this, many companies have made significant investments in RD programmes that have led to the availability of a wide variety of glucose biosensor devices. In 1962, Leland Clark Jr coined the term ‘enzyme electrode’ to describe a device in criminal investigation term papers, which a traditional electrode could be modified to respond to other materials by the inclusion of a nearby enzyme layer. Clark's ideas became a commercial reality in 1975 with the successful launch of the Yellow Springs Instruments (YSI) model 23A glucose analyser. This device incorporated glucose oxidase together with a peroxide-sensitive electrode to measure the hydrogen peroxide (H 2 O 2 ) produced during the following reaction: In this device, the rate of H 2 O 2 formation is a measure of the rate of the essay about life in prison reaction, which depends on the concentration of glucose in solution, thus allowing the latter to criminal investigation papers, be estimated. As was discussed earlier, in enzyme-catalysed reactions the relationship between substrate concentration and reaction rate is not linear, but hyperbolic (as described by of an analysis essay, the Michaelis–Menten equation).
This is also true for the glucose oxidase within a biosensor. However, we may engineer a more linear relationship by ensuring that the enzyme is papers, either behind or within a membrane through which the glucose must diffuse before it reacts with the enzyme. This means that the system becomes diffusionally, rather than kinetically, limited, and the response is then more linearly related to the concentration of glucose in solution. Over the assignment log for students years the criminal term papers YSI model 23A glucose analyser has been replaced by a range of much more advanced models. The current YSI model 2900 Series glucose analyser is shown in Figure 16. This instrument has a 96-sample rack that enables batches of samples to be run, with the analysis of assignment, each sample taking less than a minute.
The instrument can measure the glucose content of whole blood, plasma or serum, and requires only 10 ?l of sample per analysis. The membrane-bound glucose oxidase typically only needs to be replaced every 3 weeks, thereby reducing the cost of analysis. Criminal Investigation? These systems also offer advanced data-handling and data-storage facilities. In addition, these instruments can be modified to analyse a wide variety of other substances of biological interest, simply by incorporating other oxidase enzymes into the membrane (Table 8). To enable diabetic patients to take their own blood glucose measurements, small hand-held biosensors have also been developed, which are in fact technologically more advanced because the enzyme and transducer are more intimately linked on the sensor surface.
The first device of this type was launched in 1986 by Medisense, and was based on technology developed in the U.K. at features expository essay Cranfield and Oxford Universities. The ExacTech blood glucose meter was the size and shape of a pen, and used disposable electrode strips. This device was followed by a credit card-style meter in 1989. Such devices again rely on investigation term papers, glucose oxidase as the biological component, but do not measure the reaction rate via the production (and detection) of essay in prison, H 2 O 2 . Instead they rely on direct measurement of the rate of electron flow from glucose to the electrode surface. The reactions that occur within this device may be summarized as follows: and at the electrode surface: where GO x -FAD represents the FAD redox centre of criminal investigation, glucose oxidase in its oxidized form, and expository, GO x -FADH 2 represents the reduced form.
Basically electrons are removed from the glucose molecules and passed via the enzyme to the ferrocene mediator, which then donates them to the working electrode surface, resulting in the generation of an electrical current that is directly proportional to the rate of oxidation of glucose, and thus proportional to the glucose concentration in the sample. Medisense, whose only product was its blood glucose meter, was bought by Abbott Diagnostics in 1996, and Abbott-branded devices continued to investigation, use and develop this technology for some time. In 1999, Therasense marketed a glucose meter that represented the next generation of sensing technology, and integrated the enzyme even more closely with the electrode. Assignment Log For Students? Originally developed by Adam Heller at criminal investigation term papers the University of Texas in the 1990s, wired-enzyme electrodes do not rely on a soluble mediator such as the ferrocene used in the Medisense devices. Instead the enzyme is immobilized in an osmium-based polyvinyl imidazole hydrogel in which the electrons are passed from on marfan enzyme to electrode by a series of fixed electroactive osmium centres that shuttle the electrons onward in a process called ‘electron hopping.’ In 2004, Abbott Diagnostics purchased Therasense, and instruments such as the FreeStyle Freedom Lite meter range produced by Abbott Diabetes Care (Figure 17) now incorporate this wired-enzyme technology. Devices of this type are highly amenable to papers, miniaturization. Continuous measuring devices are becoming increasingly available, and may well revolutionize the control of certain disease conditions. For example, with regard to diabetes, devices such as the FreeStyle Navigator range from Abbott Diabetes Care use the same wired-enzyme technology as that described earlier, but now incorporate this into a tiny filament about the diameter of a thin hypodermic needle. This is inserted approximately 5 mm under the skin to essay, measure the glucose level in the interstitial fluid that flows between the cells. The unit is investigation papers, designed to remain in situ for up to about life, 5 days, during which time it can measure the glucose concentration every minute.
A wireless transmitter sends the glucose readings to a separate receiver anywhere within a 30-metre range, and this can then issue an early warning alarm to alert the user to a falling or rising glucose level in time for them to take appropriate action and criminal papers, avoid a hypoglycaemic or hyperglycaemic episode. In addition, experimental units have already been developed that link continuous glucose biosensor measurement systems with pumps capable of gradually dispensing insulin such that the diabetic condition is automatically and reliably controlled, thereby avoiding the qualities of an analysis traditional peaks and troughs in glucose levels that occur with conventional glucose measurement and the intermittent administration of insulin. Therefore, looking to the future, we may confidently expect to see the development of biosensor systems that can continuously monitor a range of physiologically important analytes and automatically dispense the required medication to alleviate the criminal investigation term papers symptoms of a number of long-term chronic human illnesses. For the sake of conciseness, this guide has been limited to some of the basic principles of enzymology, together with an overview of the biotechnological applications of enzymes. History Essay? It is important to understand the relationship between proteins and the nucleic acids (DNA and RNA) that provide the blueprint for the assembly of proteins within the investigation term papers cell. Genetic engineering is thus predominantly concerned with modifying the proteins that a cell contains, and assignment log for, genetic defects (in medicine) generally relate to the abnormalities that occur in the proteins within cells. Much of the molecular age of investigation, biochemistry is essay syndrome, therefore very much focused on the study of the cell, its enzymes and other proteins, and their functions. Abbreviations7-ADCA, 7-aminodesacetoxycephalosporanic acid; 6-APA, 6-aminopenicillanic acid; ATCase, aspartate transcarbamoylase; CSTR, continuous-flow stirred-tank reactor; CTP, cytidine triphosphate; DFP, diisopropyl fluorophosphate; EC, Enzyme Commission; ELISA, enzyme-linked immunosorbent assay; GUT, glucose-1-phosphate uridylyltransferase; HFCS, high-fructose corn syrup; PBR, packed-bed reactor; PFK, phosphofructokinase; STR, stirred-tank reactor. This article is a reviewed, revised and updated version of the criminal papers following ‘Biochemistry Across the School Curriculum’ (BASC) booklet: Teal A.R. Wymer P.E.O., 1995: Enzymes and their Role in log for students, Technology.
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