| Term | Definition |
|---|
| prosthetic group | A tightly bound nonprotein cofactor needed for enzyme activity. Examples include heme, flavins, metal (Ca+2, Mg+2, Zn+2, Fe+2, Fe+3). |
| apoenzyme | The protein component of an enzyme seperable from the prosthetic group. |
| holoenzyme | apoenzyme + prosthetic group |
| coenzyme | An organic cofactor. Vitamins are usually precursors to these. |
| Enzymes are classified numerically according to the chemical reactions they catalyze. | State the basis for the Enzyme Commission classification of enzymes. |
| delta G0' = -RTln[products]/[reactants] | Write the equation that relates Gibbs free energy to substrate and product concentrations. |
| delta G = delta H - T*delta S | Write the equation that relates free energy to enthalpy and entropy. |
| Transition states are the intermediate in structure between the reactant(s) and the product(s). The energy of activation is the energy difference between reactants and transition state. Catalysts help decrease the energy of activation by providing an alternate reaction pathway (stabilizes transition state). | Define the transition state and describe the effect of enzymes on the transition state. |
| The energy of activation is lowered but the standard free energy remains the same. Enzyme catalyzed reactions are must faster than uncatalyzed reactions. Catalyzed reactions may take an alternative pathway than uncatalyzed. | How does an enzyme catalyzed chemical reaction differ from the corresponding uncatalyzed reaction. |
| The standard free energy of the reaction (delta G0') is the difference between the free energy of reactants and products at equilibrium (ph = 7 and and 25 degrees C). The free energy (delta Grxn) under a set of conditions reflects the extent of displacement of the reaction from equilibrium. The energy of activation is the energy difference between the reactant(s) and the transition state. | Distinguish between the free energy of a reaction, the standard free energy and the energy of activation. |
| Enzyme has a higher affinity for transition state analogs than for substrate. | Compare binding affinity of substrates and transition state analogs for enzymes. |
| The lock and key model introduced by E. Fisher suggested that enzymes are rigid. The induced fit model proposed by Koshland suggests that enzymes are flexible and change shape to bind substrate. | Compare the lock and key model of enzyme active sites with the induced fit model. |
| Proximity, Acid-Base Catalysis, Nucleophilic (Covalent) Catalysis, and Strain or Distortion of a bond. | What general principles are used to explain enzyme catalysis (4 common mechanisms)? |
| Ribonuclease requires a 2' OH that is only found in RNA, not DNA. | Explain why ribonuclease does not digest DNA. |
| His 12 and His 119 in ribonuclease act as general acid/base catalysts by alternately taking up and releasing H+. Lysine stabilizes the negative charge on the phosphate. | Explain the role of histidyl residues and lysine residue in catalysis by ribonuclease. |
| Hydrophobic residues of the substrate bind in the hydrophobic pocket of the enzyme. His 57 takes a proton from Serine 195. Ser 195 forms a covalent ester bond with the carbonyl C of substrate. The negative charge on oxygen (oxyanion) is stabilized by H-bonding to two amide backbone hydrogens in the "oxyanion hole." C terminal peptide fragment is released and protonated by His 57. Water hydrolyzes the E-S bond. | Explain mechanism of action of chymotrypsin, emphasizing the roles of the catalytic seryl and histidyl residues in chymotrypsin. |
| Aspartyl residue take a proton from H2O. OH attacks the C=O center of peptide (substrate). Another aspartyl residue protonates the carbonyl oxygen and then another aspartyl residue takes this proton back off. The electrons on the oxygen drop back down to form a C=O group and kicks off the remainder of the peptide. Nitrogen on leaving group is protonated by another aspartyl residue. (Pepsin and Renin are also aspartyl proteases) | Explain how the active site aspartyl residues promote catalysis in aspartyl proteases (e.g. HIV protease) |
| Phosphorylation, adenylation, and methylation are all examples of covalent modifications that may regulate the enzyme. | What types of covalent modifications play a role in regulating enzyme activity? |
| 1) The availability of substrate can limit flux through a pathway. 2) Allosteric effectors may activate or inhibit the enzyme. 3) Covalent modification via phosphorylation, adenylation, methylation may activate or inhibit the enzyme. 4) Changes in enzyme concentration due to changes in rate of enzyme synthesis (mRNA transcription, translation, stability, etc). | What general mechanisms regulate enzyme activity or concentration? |
| Isozymes are two or more different enzymes catalyzing the same reaction. They can have different kinetic properties (different Km or KI values). They result from different gene products (different amino acid sequences) and may have specific tissue locations. | What are the characteristics of isoenzymes? |
| coenzymes or prosthetic groups | List some nonprotein components that can be required for an enzyme's catalytic activity. |
| v = Vmax * [S] / (Km + [S]). In this equation, v is the velocity, Vmax is the maximum velocity, [S] is the conc. Of substrate and Km is k-1/k1. | Write the Michaelis-Menten equation and define each term in the equation. |
| Hyperbolic curve (same as fractional saturation) | Describe the shape of the velocity vs. substrate curve for Michaelis-Menten kinetics. |
| 1/v = Km/(Vmax*[S]) + (1/Vmax) where y=1/v and x=1/[S] | What is the double reciprical (Lineweaver-Burke) plot? |
| Vmax is the magnitude of velocity where the graph levels off (ie where [S] is very high). Km is the [S] where v = 0.5Vmax. | How can you identify Km and Vmax on a plot of velocity vs. substrate? |
| A competitive inhibitor usually resembles the substrate. Inhibitor and substrate binding is mutually exclusive. The effects of a competitive inhibitor are reversed by raising [substrate]. Vmax remains constant but apparent Km increases. | What are features of a competitive inhibitor (include effects on Km and Vmax)? |
| A noncompetitive inhibitor usually does not resemble the substrate, but it may resemble a cosubstrate in a multireactant system. It does not prevent binding of the substrate and thus cannot be overcome by raising the [substrate]. Km remains constant but Vmax decreases. | What are the features of a noncompetitive inhibitor (include effects on Vmax and Km)? |
| ACE (angiotensin converting enzyme) normally converts angiotensin I to angiotensin II, which causes symptoms of hypertension (vasoconstriction, Na+ retention, etc). The drug lisinopril is an ACE inhibitor and thus can be used to lower blood pressure. | Give an example of how an inhibitor of an enzyme can be exploited as a drug. |
| On a double reciprical plot: Increasing competitive Inhibitor will shift graph to the right but Vmax is constant. Increasing noncompetitive inhibitor will shift graph down (Vmax changes). On a plot of v vs. [S]: Adding a competitive inhibitor will shift x intercept to the right but y intercept is unchanged. Adding noncompetitive inhibitor will shift y intercept up but x intercept is unchanged. | Distinguish between competitive and noncompetitive inhibitors on a double reciprical plot (1/v vs. 1/[S]) and on a plot of v vs. [S]. |
| Enzyme is treated with reactive reagent that forms a covalent bond with enzyme, and the modified enzyme is inactive due to modification of an amino acid that is involved in catalysis or substrate binding. Example: cyanide is a irreversible inhibitor which modifies heme enzymes by forming a stable complex with heme. | What are irreversible inhibitors? Give an example. |
| Suicide inhibitors are specialized substrates which are converted to irreversible inhibitors by the catalytic action of the enzyme. The reactive species formed by enzyme action on the suicide inhibitor inactivates the enzyme. Example: allopurinol inactivates xanthine oxidase, used to treat gout. | What are suicide inhibitors? Give an examples. |
| allosteric site | A site that binds effector molecules other than the protein's active site. |
| The first committed enzyme step in a pathway is likely to be inhibited by an end product through binding to an allosteric site. | What enzymatic step is likely to be regulated in a multi-step pathway and what types of molecules might serve as regulators? |
| Levels of troponin are elevated following an acute myocardial infarction (1-2 days afterward). CPK isoenzymes are also elevated following acute MI (1-2 days afterward). | Explain how the analysis of isoenzymes can be used in clinical diagnosis. |
| Oxidoreductase (EC 1) | This classification of enzymes includes those which catalyze oxidation/reduction reactions; transfer of H and O atoms or electrons from one substance to another. |
| Transferase (EC 2) | This classification of enzymes includes those which catalyze the transfer of a functional group from one substance to another. The group may be methyl, acyl, amino, or phosphate. |
| Hydrolase (EC 3) | This classification of enzymes includes those which catalyze the formation of two products from a substrate by hydrolysis. |
| Lyase (EC 4) | This classification of enzymes includes those which catalyze the non-hydrolytic addition or removal of groups from substrates. C-C, C-N, C-O, or C-S bonds may be cleaved. |
| Isomerase (EC 5) | This classification of enzymes include those which catalyze intramolecular rearrangement. |
| Ligase (EC 6) | This classification of enzymes include those which join together two molecules by synthesis of new C-O, C-S, C-N, or C-C bonds with simultaneous breakdown of ATP. |
| Oxidoreductase (EC 1) | What classification of enzymes would dehydrogenase fall into? |
| Transferase (EC 2) | What classification of enzyme would synthase or kinase fall into? |
| Hydrolase (EC 3) | What classification of enzyme would proteases and nucleases fall into? |
| Lyase (EC 4) | What classification of enzymes would decarboxylase fall into? |
| Isomerase (EC 5) | What classification of enzymes would mutase or alanine recimase fall into? |
| Ligase (EC 6) | What classification of enzymes would synthetase fall into? |
| Proximity | In this mechanism of catalysis, reactants are brought together on the enzyme surface, increasing their effective concentration and orienting reactive groups. Transition state analogs are good inhibitors since the enzyme binds the transition state tighter than the substrate. |
| Strain and Distortion | In this mechanism of catalysis, the transition state may have a slightly different structure induced by the enzyme that increases the reactivity of the substance. |
| Acid-Base Catalysis | In this mechanism of catalysis, imidazole, carboxyl, amino, phenolic, and sulfhydryl residues may have increased reactivity due to altered pKa values in a nonpolar environment. |
| Nucleophilic (Covalent) Catalysis | In this mechanism of catalysis, part of the substrate is covalently attached to the enzyme and part is released. The E-S complex is highly reactive. Reactive atoms on the enzyme include: S on Cys, N on Lys or His, COO- of Glu or Asp, and O of Ser-OH. |
| Schiff Base | The condensation of an amine with a carbonyl forms this. It acts as an electron sink that stabilizes negative charge on the adjacent carbon. |
| The substrate binds in the active site with the exclusion of water (and protons), which considerably lowers the dielectric constant of the medium surrounding the substrate. | What can account for changes in the apparent pKas of ionizable groups and stronger electrostatic interactions within the active site of the enzyme? |
| chymotrypsin | This serine protease catalyzes the hydrolysis of peptide bonds and cleaves at the C-terminal side of bulky, hydrophobic residues. |
| trypsin | This serine protease catalyzes the hydrolysis of peptide bonds and cleaves at arginyl- and lysyl residues. |
| elastase | This serine protease catalyzes the hydrolysis of peptide bonds and cleaves at small residues (ex. glycine). |
| Enzymes are highly specific, catalyze only biological reactions, are affected by temperature and pH, and can be regulated and inactivated. | List some important attributes of enzymes that distinguish them from other catalysts |
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