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147 Cards in this Set
- Front
- Back
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What occurs in the dehydrogenation of fatty acyl degradation? What oxidizing agent is used?
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a. FAD based system
b. gives trans-alpha-beta-unsaturated CoA ester c. produces FADH_2 |
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What is ubiquinone? What step of degradation is it found in?
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a. Co-enzyme Q that has the protons and electrons of FADH_2 transferred to it.
b. occurs in the dehydrogenation step. |
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What are isozymes?
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enzyme isomers that are distinct in substrate specificity but execute the same gross reaction.
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What enzyme is responsible for the ubiquinone transfer? what type of protein it is? where is it located?
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a. acyl CoA dehydrogenase, three types each with different chain length preferences.
b. integral membrane protein c. located in the inner mitochondrial matrix. |
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What occurs in the hydration step of fatty acyl degradation? What is the stereochemistry? What enzyme is responsible?
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a. double bond formed previously is hydrated regio- and stereospecifically
b. new chiral center at C-3 is 'L' c. enoyl CoA hydrase |
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What occurs in the oxidation step of fatty acyl degradation? What is the process dependent on? What enzyme is responsible? what are the two hydrogens and two electrons collected in this step used for?
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a. 3-hydroxyacyl-S-CoA is oxidized to 3-ketoacyl CoA.
b. NAD+ dependent c. 3-hydroxyacyl CoA dehydrogenase d. Used to reduce molecular oxygen in a combustion reaction |
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What occurs in the retr-condensation step of fatty acyl degradation? What enzyme is responsible?
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a. An acetyl-S-CoA is split from the beta0ketoacyl-S-CoA
b. thiolase |
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What enzymes form the multi-enzyme complex for degradation?
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Three enzymes: the 2-enoyl-CoA hydratase, the L-3-hydrozyacyl-CoA dehydrogenase and the 3-ketoacyl-CoA thiolase
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What is another name for fatty acid degradation? Why?
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a. B-oxidation
b. because the B-Carbon is progressively converted to a secondary alcohol and a ketone in the degradative cycle. |
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What is the yield of of fatty acid chain of carbon content 2n from degradation?
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a. n units of acetyl-CoA
b. 2(n-1) hydrogen/electron pairs in the form of NADH/H+ and FADH_2 |
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What happens to the acetate and the hydrogen pairs made from degradation?
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they are futher processed in cell respiration.
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Where do odd chained carbons come from in the diet?
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cheese, yogurt, pickles, etc.
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How are odd chained fatty acids degraded? Where is the product later processed?
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a. Normally up until propinoyl-S-CoA. Then a cobalt containing cofactor vitamin B12 converts it into methylmalony-S-CoA and then succinyl-S-CoA.
b. The Krebs cycle |
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What is the naturally occuring cis/trans conformation in unsaturated fatty acids? What does Beta-oxidation work on? How is this problem overcome
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a. cis
b. trans b. enoyl CoA isomerase flips it |
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What is another problem with metabolizing naturally occurring fatty acids?
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beta-oxidation only works on even-to-odd carbon number double bonds. If a fatty acid has a double bond spanning odd-to-even double bond a mitochondrial flip via enoyl CoA isomerase occurs.
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What are six difference between fatty acid biosynthesis and degradation?
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a. Synthesis occurs in cytoplasm and degradation occurs in mitochondria.
2. ACP is used in synthesis but CoA esters are used in degradation. 3. multienzyme complexes are figured into biosynthesis but isolated enzymes are used in degradation 4. acetate/malonate figure into biosynthesis and only acetate in degradation 5. NADPH/H+ is exclusively used in synthesis and FAD and NAD+ also figure in to deg ration. 6. Palmitic acid is the end-point of synthesis and degradation can handle a wide range of input chain lengths. |
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What are the three types of ketone bodies? Which is least prevalent?
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a. beta-hydroxybuyric acid, acetoacetic acid and acetone
b. acetone |
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What are ketone bodies? Where are they formed?
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a. water soluable fat used as alternative fuel source.
b. liver |
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What uses ketone bodies the most? What prefers ketone bodies to glucose?
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a. heart muscle and other muscles
b. The heart likes acetoacetate and the brain and kidneys prefer glucose |
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When are ketone bodies synthesized?
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When glucose levels become suboptimal during fasting, starvation or uncontrolled diabetes.
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What us duabetes mellitus? What are the two types and their characteristics?
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a. caused by the lack of insulin to take glucose out of the body
b. Type I: Juvenile/ insulin-dependent form. Insulin production in pancreas is lost/reduced resulting in hyperglycemia, sugary urine, fat burned as high levels, high levels of acetyl-CoA in liver thus ketone body formation and constant unger,thirst and urination. c. Type II: Late onset. Insulin receptors lowered. Results in hyperglycemia, fat combustion and ketone body formation. |
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How is diabetes controlled?
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a. Type I: bu insulin provisions
b. Type II: diet and exercise |
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What is hyperglycemia?
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blood glucose levels that are perpetually high
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What might you smell on the breath of diabetics?
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Acetone
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What are homopolymers? heteropolymers?
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a. single monomer involved
b. two or more monomers involved |
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What bonds hold together a protein? What are these bonds generically? What do proteins make up?
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a. peptide bond
b. amide c. polypeptide chain |
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What are the six roles of proteins in cells? What are examples of each?
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1. transport/storage: hemogoblin, albumin
2. Hormones: insulin, glucagon 3. enzymes: biological catalysts 4. defensive entities: immunoglobulins 5. components of cellular organization or structure: collagen, keratin 6. contractile elements: muscle, cilia, flagella |
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What is the specific configuration of the amino acids involved in protein structure?
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L-alpha-amino acid
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How many steroisomers are possible in a molecule with n centers of chirality? How many enantiomeric pairs?
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a. 2^n called diasteroeisomerism
b. n |
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What are enantiomers? diastereoisomers? epimers?
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a. Either of two stereoisomers that are mirror images of one another but cannot be superimposed on one another and that rotate the plane of polarized light in opposite directions.
b. non-enantiomeric stereoisomers c. two diasteroisomeric structures that differ in configuration at a single chiral center |
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What is a zwitterion? What are they dependent on?
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a. internal salts formed from amino acids containing both acidic and basic functional groups.
b. dependent on the pH in which the solution is found |
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What is the isoelectric point? What is it associated with?
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a. The pH at which salt formation is complete, the amino acids have no net charge and the protein does not move in an electric field.
b. zwitterions |
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What is a mitochondria flip? when is it needed? What enzyme does it?
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a. When a double bond is in an odd-to-even position, it muct be switched before beta-oxidation can occur to prevent beta-gamma- unsaturated acyl-S-CoA
b. The oxidation of an unsaturated fatty acid c. Enoyl-S-CoA isomerase |
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What is the function of insulin?
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It stimulates the synthesis of glycogen, triacylglycerols and proteins and also inhibits their breakdown.
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Which amino acid is not chiral? Which has two chiral centers?
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a. glycine
b. isoleucine and threonine |
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What is the Henderson-Hasselbalch equation?
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pH= pKa +log[A-]/[HA]
where A- is a conjugate acid and HA is a weak acid |
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What is the general mathematical form that the HH equation follows? What is the equation characteristic of? What are the coordinates of the middle of the step?
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a. y=a constant(pKa value) + log[x/(1-x)]
b. a step shape c. the co-ordinated of the middle of the step are x=0.5 and y=pKa value |
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What does the y-value of the middle of the step using the HH equation mean?
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It is the pKa value of the side chain of the amino acid where at that pH half of the atoms are protonated and half are not.
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What does monobasic/monoprotic mean? dibasic/diprotic? tribasic/triprotic?
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Means there are __________ ionizable forms and thus _________ number of steps
a. 1 b. 2 c. 3 |
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What is a buffer? Where is the maximum buffering capacity found? What is the region of buffering?
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a. any substance or mixture of compounds that, added to a solution, is capable of neutralizing both acids and bases without appreciably changing the original acidity or alkalinity of the solution.
b. When pKa=pH c. pKa +/- 0.25 pH units |
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How many base pairs, pKa values and steps does an free amino acid have? What are two exceptions
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atleast...
a. 2 weak acid/conjugate base pairs b. two pKa values c. two steps because any free amino acid has both an amino and a carboxylic acid group. d. Glutamic acid and lysine have three ionizable groups and steps. |
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How many amino acids are in proteins polypeptides? oligopeptides? What are the exceptions?
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a. 50+
b. 30-50 c. 10-30 *chain lengths are not strict for these definitions d. Glucagon is called a proteins and only contains 29 amino acids |
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What is a multimeric protein?
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Proteins with two or more polypeptide chains
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What are the two general types of proteins? WHat are examples of each?
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a. Fibrous: Collagen and keratin
b. Globular: Hemoglobin |
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What are the characteristics of fibrous proteins? How are they structurally defined?
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a. Play structural roles in cell or tissues, are water insoluble but hydrophilic and are long linear molecules build on the cable principle.
b. Only primary structure |
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What are the characteristics of globular proteins? How are they structurally defined?
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a. proteins that play a role in enzymes, transport, immunological or hormonal control. they are either water-soluable, suspended in water or are intergral membrane proteins. They have many bends and loops in their chains.
b. defined by primary,secondary, tertiary or quaternary (but only ones that can be defined as 2' and 3') |
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What is conformation? configuration? native conformation?
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a. a unique shape/folding/coiling rotating arounf a single bond
b. the unique arrangments of a molecule c. refers to the working shape of real proteins |
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Does rotation about the carbonyl-amino groups occur? why/why not?
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No because of resonancen showing that the carbonyl-C and amino-N bond has about 40% double bond character.
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Are there cis/trans isomers in polypeptide bonds? Which is more stable? What is the exception?
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a. yes. in cis the two alpha carbons are on the same side of the peptide bond and in trans they are on opposite.
b. trans c. proline |
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What is important for the hydrogen bond formation between two non-side chain elements of the polypeptide chain?
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The higher partial positive on the nitrogen and the higher partial negative on the oxygen then is normally found
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What are the two angles used to measure rotation in a polypeptide chain? What is strange about the values? What do they follow?
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a. Phi: N-side of the alpha carbon
b. psi: C-side of the alpha carbon c. only a limited number of values are encountered in the native conformation d. Ramachandran plots |
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How is the three dimensional structure of proteins studied (2 ways)? What are their benefits?
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x-ray crystallography: time consuming but better in the long run. Gives insight to primary, secondary, tertiary and quaternary structure!
Nuclear Magnetic Resonance spectroscopy (NMR): more restricted in application but faster |
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Why are the real life values of phi and psi different from those predicted by the Ram plots?
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Because the peptide bonds are not exactly planar and can bend sometimes,the angles have a little wiggle room in value.
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Why are polypeptides called vector polymers? How do polypeptides grow?
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a. because they have a unique direction with N-->C ordering of monomers which is different than C-->N ordering.
b. Polypeptides always grow on the carboxylic acid end via dehydration reactions |
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What is the Afinsen Experiment? What does this imply?
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a. it showed that if a protein was denatured carefully an all the disulphide bonds were broken. once the denaturant was removed the native conformation would reform without mistake.
b. That tertiary structure is defined by the primary structure. |
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What are the prime drivers in bringing about a proteins native conformation? what else is involved?
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a. Hydrophobic forces
b. hydrogen bonds, salt bridges and chaperone proteins. |
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What are chaperon proteins?
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they assist the conforming of larger proteins
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What are examples of reversible denaturants? irreversible?
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a. Urea, guanidinium and sodium dodecyl sulfate
b. heat and strong acids or strong bases |
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What are salt bridges?
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charge/charge interactions between charged amino acid side chain groups (i.e. between COO- and NH_4+
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What terms are used to describe the shape of globular proteins? fibrous proteins?
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a. secondary, tertiary and quanrernary
b. primary and cabling |
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What is secondary structure tied to?
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hydrogen bonding between carbonyl and amino groups of peptide bonds in the polypeptide chain
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What are the two types of secondary structures?
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a. alpha-helix
b. beta-pleated sheets |
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What is an alpha-helix? why is it called alpha? what is the common type in peptide bonds? What is another name and what does it represent? How are the hydrogen bonds aligned? Side groups aligned? direction of the helix?
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a. Hydrogen bonds bridge the carbonyl of a peptide bond with the amino group of the peptide bond four monomer units away to form a spiral/coiled chain.
b. first possible structure predicted by Pauling and Corey c. right-handed d. 3.66_13 helix: 3.66 is number of residuals and 13 is the number of atoms involved in one turn. e. roughly parallel to the helical axis and all point in same direction f. stick out perpendicular to helical axis g. N-->C direction |
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What role do the side chains play in alpha-helix structure? What amino acids destabilize them? why?
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a. They are perpendicular to axis to reduce steric hindrance but the play no role in the formation of the alpha-helices.
b. proline because of its ring structure and glycine because only a hydrogen atom |
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What are domains?
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Super-secondary structures made up of alpha helices.
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What are beta-pleated-sheets? Why is it called beta? What are the two types? Which are more stable? How are the side chains aligned?
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a. two or more stretches of polypeptide chains that align and are held in that alignment by hydrogen bonds between peptidyl carbonyl and amino groups
b. second possible structure predicted by Pauling and Corey c. parallel (all N-->C) and antiparallel (alt. N--> and C-->N) d. antiparallel e. stick up and down perpendicular to the plane of the sheet |
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What agents can break secondary protein structure and how?
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a. urea and guanidinium chloride
b. by competing for hydrogen bonding sites of the protein. |
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What chemical bond types are responsible for primary, secondary and tertiary protein structure?
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a. primary: peptide bonds
b. Secondary: Hydrogen bonding c. tertiary: Hydrophobic ineractions, disulphide bonds between cystine residues, salt bridges between acidic and basic side chains or terminal groups and hydrogen bonding between R groups. |
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What part of the amino acid determines if it is hydrophobic or hydrophilic?
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The R group
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How is the tertiary structure of a protein affected by Urea? Detergent?
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a. urea breaks down the secondary structure by breaking/competeting for the hydrogen bonds and thus tertiary structure can't exist.
b. Detergents split up the hydrophobic interactions and thus destroy the tertiary structure. |
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What is the function of buttressing bonds? What are the three types?
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a. to stabilize the three-dimensional structure of a monomeric or multimeric protein.
b. Disulphide bridges, salt bridges, hydrogen bonds. |
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Explain how each of the three buttressing bonds work to stabilize the 3-D structure.
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1. Disulfide bonds: between two cysteine residues not necessarily close to each other
2. Salt Bridges: between acidic and basic chain groups or the N and C terminal amino acids 3. Hydrogen Bonds: Between side chain groups. (Between peptide bonds doesn't signify buttress bond). |
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Are hydrogen bonds between peptide chains buttressing bonds? hydrophobic bonds? why/why not?
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a. No, because they are secondary structure bonds
b. No, because they are the driving force behind tertiary structure and buttressing bonds help to further stabilize because the hydrophobic interactions bring distant groups closer together. |
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What forces hold the subunits of multimeric proteins together? In multimeric globular proteins do the peptide chains intertwine?
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a. A range of non-covalent forces-- hydrophobic, hydrogen bonds, and salt bridges.
b. No, because they have a native tertiary structure. Only in fibrous proteins do they intertwine. |
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What is a multimeric protein?
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It is the quaternary structure of proteins where the fully-folded subunits or monomers associate to a final product. They can be made from different or identical monomers.
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What type of biomolecule is keratin? Where is it found? What is its structure?
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a. Structural fibrous protein closely related to tropomysin that with actin is the contractile unit in muscle fibers.
b. Found in wool, hair, hooves and fingernails. c. Almost entirely constructed of helices (not alpha) with non-polar residues on one surface of the helical cylinder. Made up of two right-handed helices twisted into a left-handed cable (producing considerable strength). |
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What type of bonds control the flexibility of keratin?
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Trans-helix disulfide bonds such that more disulfide bonds means a more rigid structure.
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What type of biomolecule is collagen? Where is it found? What is its structure?
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a. Structural fibrous protein
b. Found in bone, cartilage, tendons and loose connective tissue of skin. c. triple helix of polypeptide chains with repeating Gly-X-Y where X=proline normally and Y=4-hydroxyproline normally (helps strengthen collagen). |
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What unique amino acid is found in collagen and what is its function?
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4-hydroxyproline used to strengthen collagen. Hydroxylation is accomplished after peptide chain has been formed with the aid of vitamin C and molecular oxygen to be converted from proline.
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What is avitaminosis?
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any disease caused by chronic or long-term vitamin deficiency or caused by a defect in metabolic conversion, such as tryptophan to niacin.
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What is scurvy? Why does it occur?
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a. weakness in the collagen structures due to no hydroxylation occuring in polypeptide chains leads to tooth loss, painful joints and easily damaged skin.
b. occurs due to lack of vitamin C which the body cannot make on its own but it crucial for hydroxylation. |
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What is ascorbic acid?
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Vitamin C
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What are some examples of avitarminosis?
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Scurvy (Vitamin C), Rickets (Vitamin D), Night-blindness (Vitamin A) and Pellagra (niacin)
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What is the physiological function of myoglobin? hemoglobin? Are these monomeric or multimeric proteins?
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a. Involved in the storage of oxygen in the muscle cells (monomeric)
b. Delivers oxygen throughout the body through red blood cells (Multimeric) |
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What is a prosthetic group? What type is encountered in myoglobin? hemoglobin?
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a.a non-protein (non-amino acid) component of a conjugated protein that is important in the protein's biological activity. The prosthetic group may be organic (such as a vitamin, sugar, or lipid) or inorganic (such as a metal ion). Prosthetic groups are bound tightly to proteins and may even be attached through a covalent bond. They often play an important role in the function of enzymes. A protein without its prosthetic group is called an apoprotein, while a protein combined with its prosthetic group is called a holoprotein.
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What prosthetic group is encountered in myoglobin? hemoglobin? What is its purpose?
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a. Both use an iron containing co-factor called a heme group in which the iron is in the oxidized form of Fe2+ and embedded deep in the hydrophobic interior of the protein.
b. The focus of the oxygen binding action of these proteins |
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What is a rich source of myoglobin? Why is is in high abundance there?
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a. diving animals like whales, seals, etc...
b. Meat" acts as a storage unit for oxygen and allows them to hold their breath for an extended period of time. |
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Describe how you would perform an ‘Anfinsen experiment’ on myoglobin; when during this experiment would myoglobin become non-functional; does this experiment simply confirm Anfinsen’s original work with ribonuclease?
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1. treat pure myoglobin with diethyl ether to pull the heme molecule out the the hydrophobic pocket
2. Treat with an agent that will break hydrogen bonds of the secondary structure (Urea or guanidiniun chloride) or a detergent that breaks up tertiary structure (sodium dodecyl sulfate) 3. Once unravelled, removed denaturant, add heme in excess and carry out in ether solution. b. It becomes non-functional as soon as heme is removed c. Confirms original work but further shows that tertiary structure and protein function is dictated by primary structure. |
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What is the structure of myoglobin?
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Mb: small globular protein made of primarily alpha-helices(8). It contains the prosthetic group heme for it to function in oxygen-binding/storage
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What does the oxygen-binding behavior of myoglobin require? How does it occur?
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a. Heme cofactor and two histidine residues (hist-64 and hist-93).
b. Lever action: involving the hist-93 and alpha-helix that contains it, a small movement of the iron molecule is magnified and causes the whole molecule to change shape slightly. |
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What is a conformational change?
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Shape changes in proteins that is caused by slight shifts in molecules due to binding of other molecules.
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What is the general equation of a rectangular hyperbola?
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y=[(C_1)*(X)]/(C_2+X)
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What is the value of the constants in the hyperbolic equation for myoglobin?
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c_1=1 and C_2=K
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What is the physiological significants of P_50? In what equation is this term found?
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a. It is the partial pressure of oxygen in the body at which Mb or Hb is 50% saturated which is equal to K.
b. It appears in the fractional saturation equation. |
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What is the fractional saturation equation?
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Y=pO_2/(P_50 + pO_2)
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What does the oxygen dissociation curve look like for myoglobin? hemoglobin?
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a. hyperbolic with a single equilibrium constant and a low affinity for oxygen thus a low P_50 value.
b. sigmodial due to co-operativity occuring and a higher affinity for oxygen and thus a high P_50 value. |
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What is co-operativity?
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Occuring in hemoglobin, it is the phenomenon that addition or removal of one oxygen makes subsequent additions or removals easier
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What type of secondary structures are found in myoglobin and hemoglobin?
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Only alpha-helices. There are no beta-pleated sheets
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What is the structure of hemoglobin? What does it make up?
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a. It has four subunits, two identical alpha and two identical beta. Each of the subunits contains a heme groups thus a single molecule can transport four molecules of oxygen
b. Major constituent of red blood cells |
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What are the two special properties of multimeric proteins that hemoglobin manifests?
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1. Co-operativity
2. Allostery |
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What additional molecule does hemoglobin carry? What is its function? How does it occur?
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a. D-2,3-bis-phosphoglyceric acid
b. located at the center of the hemoglobin tetramer, it keeps the P_50 value at 26 torr instead of 12 torr which means it makes hemoglobin have an increased affinity for oxygen. c. It binds to the deoxygenated form of hemoglobin thus stabilizing it and decreasing affinity for oxygen |
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What is allostery?
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Property of a small molecule to influence the behavior (shape first, the activity) of a large protein with respect to a structurally much different molecule.
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What are the two allosteric effectos of hemoglobin?
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2,3-BPG and carbon dioxide
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What is homotropic allostery? heterotropic allostery?
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a. shape changing molecule is also the protein that the molecule normally deals with (i.e. molecular oxygen)
b. shape changing molecule is different from the molecule the protein normally deals with (i.e. 2,3-BPG) |
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What are immunoglobulins?
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Antibodies which are central to the humoral immune system of vertebrates.
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What are the five types of antibodies? How are they constructed? What type of secondary structures do they have?
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a. IgA, IgD, IgG, IgEm IgM with most common being IgG's.
b. constructed from two short (light ) chains and two longer (heavy) chains c. characteristic Y shape and beta-sheets |
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Are enzymes globular or fibrous? Where does the work of an enzyme get carried out?ion?
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a. globular
b. active site often in a cleft or groove of globular structure and/or located between two of the enzymes subunits. |
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What is an enzymes function? How does it function?
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a. increases the rate of a biological reaction or allows reaction to be run at a lower pH, temperature, or pressue).
b. Decrease the activation energy needed but does not change a reactions thermodynamics! |
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Enzymes are specific for what four things?
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1. Substrate Used
2. Regiospecific: where in a given substrate the reaction will take place 3. Reaction specific: what products wll be formed from a substrate 4. Stereospecific: only some are specific for a single substrate others will accept a small range of structurally related substrates. |
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What is the function of allosteric enzymes?
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regulate the through-put of mass in a metabolic pathway crucial for anabolism and catabolism.
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What are co-factors? Why are the necessary? What can they be?
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a. additional functional groups recruited to the active site
b. Sometimes the side chain groups of amino acids isn't sufficient to provide the chemical versatility to carry out the reaction. c. inorganic or organic (co-enzymes) or metal ions, usually cations (metalloenzymes). |
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What does a progress curve show?
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The amount of product formed as a function of time
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How does the size of the enzyme compare to the size of the active site?
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a. enzyme is very large (7-50 amino acids)
b. active site is very small (1-5 amino acids) |
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What are metalloenzymes?
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An enzyme protein that uses metal ions, often cations, loosely or tightly bound to it to carry out its function.
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What is an initial reaction velocity? how is it measures? why is it initial?
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a. the slope of the curve in the linear section
b. a parameter used extensively in chemical and biochemical kinetics. c. Initial because it is the rate at which product is formed up to the point that it is recycled to substrates |
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How is the reaction rate calculated for the reaction:
A + B --> C + D What is the order of the reaction? What is k and its units? |
Forward reaction:
v=d[C or D]/dt=k [A]^n[B]^m Reverse Reaction: v=d[A or B]/dt=k [C]^o[D]^p Order: (m+n) or (o+p) k: rate constant 1/S (1st order) or L/(mol*sec) (2nd order) |
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What is meant by pseudo-first order?
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If the second substrate is in very large concentration such that small changes are undetectable (solvent) then the rate equation is independent of [B] and B becomes a constant altering k invariantly.
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What can be understood by the term "steady state aproximation"?
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The input=the output or when the ES complex is formed at the same rate that it is decomposed which means [ES] is constant.
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In Michaelis/Menton kinetics, what is Vmax? why is it a max? how does its parameter relate to an hyperbolic curve?
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a. reaction rate with a fixed amount of enzyme increases with increasing amounts of substrate until a point of saturation is reached giving Vmax.
b. Its a max because the reaction can't go any faster because all of the enzyme is involved in the ES complex c. Plots of reaction rate vs. substrate concentration show vmax as the asymptote or a in the equation y=ax/(b+x). |
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What is competitive inhibition? How is it overcome?
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a. When substrate analogs are added with proper substrate and compete for active site of enzyme thus slowing down the reaction rate.
b. Overcome by adding a lot of the proper substrate |
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What are the six systematic classes of enzymes? Their class number? The basic process they catalyze?
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1. Oxidoreductase ( Class 1): Catalyze redox reactions and have co-substrates FAD, NAD+, NADP+
2. Transferase (2): Move functional groups from one substrate to another like the transfer of phospho groups from ATP, GTP, etc to substrate commonly called kinases. 3. Hydrolases (3): Cleave esters, amides, anhydrides, etc. with water 4. Lyases (4): create or remove double bonds non-reductively with no redox co-factors. 5. Isomerases (5): change stereochemical configuration 6. Ligases (6): catalyze (A)TP-dependent substrate couplings. Also called synthetases. |
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What is the difference between lyase and ligase? oxidoreductase?
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a. lyases create or remove double bonds and ligase creates at the expense of XTP
b. Oxidoreductase gets rid of double bonds using redox cofactors. |
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For 90-95% of enzymes benchmarked by __________, how does the initial rate vary with substrate concentration? enzyme concentration?
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Myoglobin
a. initial velocity is the linear portion of a hyperbolic curve b. initial velocity is linear the entire time |
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What is the Michaelis-Menton constant? What is the biochemical significance? Why can't it ever be very, very small in size?
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a. K_m=(k_-1 + k_cat)/k_1
b. k_cat is much slower because it is the rate determining step since it requires the breaking of covalent bonds. c. The smaller the k_m the more tightly the enzyme binds the substrate and thus really small k_m would mean the enzyme doesn't release from the substrate in which case it is no longer an enzyme. |
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To what parameter does vmax give access?
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at 1/2 vmax the [S] is at k_m where the enzyme is half saturated because k_m= aprox. k_-1/k_1 which is the dissociation constant K_d.
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What is fractional velocity of an enzyme? how is it computed? what is its shape? how does the parameter relate to fractional occupation by substrate of the enzyme's active site? by oxygen of the heme site of myoglobin?
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a. v/V_max=[S]/(K + [S]) is an estimate of the fraction of enzyme active sites occupied by the substrate at any given time.
b. Its shape is hyperbolic c. V_max/2 is when 1/2 of the Enzymes sites are occupied d. Equation is in the same form of Mb's Y=Mb/(Mb + O_2) show that a small molecules react with enzymes the same as with non-enzymatic proteins. |
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What is the turn-over number? What does it measure? What is the reciprocal?
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a. k_cat
b. The catalytic power of the enzyme, the number of catalytic events that a molecule of enzyme can accomplish per unit time. c. 1/k_cat is a measure of the amount of time a single molecular transformation at an active site takes. |
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What is a double reciprocal plot? Why is it useful? What does the x and y intercept represent?
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a. plot of 1/v and 1/[S]
b. linearized form to access Km and Vmax 1/v=K_m/V_max *1/[S] + 1/V_max fits y=mx+b where 1/V_max=b and Km=m/b or 1/-[S] c. x (1/[S]): -1/K_m y (1/v): 1/V_max |
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What are the two classes of enzyme inhibition? What are they used for?
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1. reversible: critical for regulation of metabolism
2. irreversible: technologically valuable for pharmacology and chemical warfare. |
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What is meant by rational drug design? How were drugs formally obtained?
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a. Modern alternative using x-ray analytical data of enzyme structure and looking at inhibitors of the active site that will dock into it reversibly or irreversibly.
b. "suck it and see it" approach |
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What is gross irreversible inhibition? What are its uses?
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a. When irreversible protein denaturing agents work on an enzyme (eg. heat, high/low pH or agents that oxidize disulfide bonds into two sulfonic acids)
b. detroying hazardous enzymes, viruses and cells resulting from biotechnology industry by autoclaving |
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What is an autoclave? What is it used for? To what extent it is useful?
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is a pressurized device designed to heat aqueous solutions above their boiling point to achieve sterilization. The term autoclave is also used to describe an industrial machine in which elevated temperature and pressure are used in processing materials.
b. destroys potentially hazardous enzymes, viruses and cells. c. can't destroy prions (infective protein agents involved in mad cow disease for instance) because heat is not suffienciently high enough |
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What is specific irreversible inhibition? Whats an example?
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a. sets of chemical agents that specifically and with varying degrees of selectivity knock out key functional groups in an enzymes active site.
b. di-isopropylfluorophosphate |
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What is di-isopropylfluorophosphate? How does it work? What are its relatives?
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a. a specific irreversible inhibitor that is a mixed anhydride
b. It esterfied alcohol functional groups like serine which is located in the active site of many key proteolytic enzymes. It forms di-isopropyl phosphate with the alcohol in the active site, permanently inhibiting it but leaving its 3-D structure intact. c. Very effective nerve gas like Sarin and Vx gas. |
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What amino acid groups are attacked by di-isopropylfluorophosphate? What is its chemical warefare capability? why?
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a. alchol, thiol and amino groups
b. Nerve gas c. nerve synapses have serine residues in their active sites which has alcohol side chain and thus can be attacked by the inhibitors. |
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What is meant by affinity labels?
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They are inhibitors whose molecular shape only binds specifically to one enzyme or a small group of related ones and then inactivate the active site. They are important for the drug industry.
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What is site-directed mutagenesis?
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Allows one to modify the active site of amino acid(s) at the time of their formation rather then irreversibly inhibiting the residue already existing in an enzymes active site.
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What 8 steps does site-directed mutagenesis involve?
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1. isolating genes coding for the primary sequence of the enzyme under study
2. determine nucleotide sequence 3. Id the triplet that codes the amino acid you want to change 4. chemically change that codon 5. add signal sequence to allow secretion into the extracellular environment 6. insert into a plasmid to transform bacterium 7. grow bacterium and isolate modified enzyme in bulk 8. study the irreversibly modified enzyme. |
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What is reversible competitive inhibition? Where does the inhibitor bind? What does it affect? how is it overcome? How is it graphically represented?
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a. analogues of the normal substrate
b. binds to the active site preventing the normal substrate to bind. The inhibitor only binds to the native enzyme and not to the enzyme-substrate complex c. only affects K_m. V_max remains the same but is reached at a much high substrate concentration. d. add large amounts of true substrate e. rate plots w/ varying [S] gives lines radiating from the point on the y-axis corresponding to 1/V_max |
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What is reversible, uncompetitive inhibition? How common is it? How is it graphically represented?
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a. inhibitor binds only to the ES complex and never to the native enzyme
b. much less common than competitive inhibitors c. set of straight lines, one for each inhibitor concentration parallel to the plot of the uninhibited data |
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What is reversible, noncompetitive inhibition? How is it graphically represented?
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a. inhibitor can bind either the ES complex or the native enzyme and the affinity for each need not be the same
b. lines radiating from a point on x-axis if inhibitor binds both with equal affinities. if different then lines may radiate from above or below the axis. |
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What are allosteric enzymes?
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A group of enzymes representing 5-10% of total like hemaglobin demonstrate sigmodial kinetics and co-operativity. At a fixed substrate concentration, small molecules can move the sigmodial curve
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How is an allosteric inhibitor seen graphically? allosteric promotor?
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a. moves signmodial curve to the right (negative allosteric effector)
b. moves sigmodial curve to the left (positive allosteric effector) |
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What are the 7 traits of allosteric inhibitors/promotors?
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1. don't resemble normal substrate of the enzyme
2. excess of normal enzyme doesn't overcome inhibition 3. operate by changing the native conformation 4. can cause a change in K_m, V_max or both 5. catalyze the first committed step in a pathway 6. multimeric 7. do not covalently bind to their target enzyme |
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What is covalent modification?
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Regulation of enzymes by phosphorylation on serine or tyrosine residues in their primary structure often controlled by hormones
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What are protein kinases? protein phosphatases? Why is phosphorylation on tyrosine important?
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a. enzymes that phosphorylate other proteins
b. enzymes that remove phosphate from a phosphorylated protein c. it may be important in the virus-induction of cancer states in cells. |
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What are the three ways that a substrate interacts with an enzyme in a multi-substrate enzyme? Which is the norm in reactions catalyzed by hydrolases?
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1. ordered: A then B
2. Random: A/B doesn't matter 3. ping-pong: A binds, partially processed then B binds. Norm for hydrolases where the first substrate associates with the enzyme prior to being transferred to the second substrate. |
