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37 Cards in this Set
- Front
- Back
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Enzymes |
Catalysts that increase reaction rates without being used up Most are globular proteins (some RNA also catalyze) Studies of enzyme is oldest field |
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Holoenzyme v apoenzyme |
Enzyme require molecules called cofactors or coenzymes Forms complex Vs no cofactors
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Why biocatalyst I’ve inorganic catalyst |
Greater reaction specificity and avoids side products Milder reaction conditions, conducive to cell conditions Higher reaction rates Capacity for regulation (biological pathways)
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Metabolites have many potential pathways of decomposition |
Enzymes make the desired one most favorable |
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Classes of enzymes |
TOHILL Transferases: group transfer reactions Oxidoreductases: transfer of e- Hydrolases: hydrolysis reactions Isomerases:transfer of groups in molecules to yield isomeric forms Lyases: cleavage of C-C, C-O, C-N bonds, leaving double bonds or rings, or addition of groups Ligases: formation of C-C, C-S,C-O, C-N bonds by condensation, coupled to cleave ATP or similar cofactor |
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Enzyme do not affect |
Equilibrium or free energy of reaction Help overcome activation barriers Increase reaction rates by decreasing free energy |
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How to lower free energy |
Uncaralyzed bimolecular reactions: 2 free reactants —> single restricted transition state (entropically unfavorable) Uncatalyzed unimolecular reactions: flexible reaction —> rigid transition state conversion (entropically unfavorable) Catalyze reaction: enzyme uses binding energy of substrates to organize reactants to fairly rigid complex, entropy cost is paid in binding, rigid reaction is entropically neutral |
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Free E |
See pic |
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Diagram |
Pic |
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Enzymes bind transition states best |
Enzymes active sites are complimentary to the transition state of the reaction Bind transition states better than substrates Stronger transition states lower the activation E barrier |
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Catalytic mechanisms |
Acid-base: give and take protons Covalent: change reaction paths Metal ion: use redox cofactors, pKa shifters |
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Enzyme Kinetics |
Study of the rate at which compounds react Enzymatic Rate is affected by: enzyme, substrate, effectors, temp |
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Why study enzyme kinetics? |
Quantitative description Determine order of binding substrates Elucidate acid-base catalyst Understand catalytic mechanism Understand reg of activity |
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Kinetic equations |
Start w model mechanism Identify constraints and assumptions Algebra |
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Simplest model mechanism |
One reactant, one product, no inhibitors |
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Id constraints and assumptions |
Total enzyme concentration is constant |
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Algebra: the Michaelis menten equation |
Kcat: how many substrate molecules one enzyme molecule can convert per second Km: (Michaelis constant) approx. measure of a substrates affinity for an enzyme |
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Equations for exam |
A |
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How to do kinetic measurements |
Mix enzyme and substrate Record rate of substrate disappearance and/or product formation of time Plot initial velocity vs substrate concentration Change substrate concentration and repeat |
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Determination of kinetic parameters |
A nonlinear plot used to calculate Km and Vmax A linear double/reciprocal plot is good for analysis of 2 substrate data or inhibition (line weaver plot) |
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Step 2 |
Ser and his generate ion that attacks peptide carbonyl group Forming short lived negative intermediate |
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Sequential |
Cannot easily distinguish random from ordered Will give intersection at y axis |
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Ping pong |
Lines are parallel |
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Step 5 |
Collapse of intermediate and second product formed: a carboxylate anion and displaces ser |
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Competitive inhibition |
Competes with substrate for binding Binds to active site Does not affect catalysis No change in Vmax, but increase in Km Lone intersect on lineweaver-burk |
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Uncompetitive inhibition |
Only binds to ES complex Doesn’t affect substrate binding Inhibits catalytic function Decrease in Vmax and Km Lines are parallel |
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Mixed inhibition |
Binds enzyme with or without substrate Binds to regulatory site Inhibits both substrate binding and catalysis Decrease in Vmax, change in Km Lines intersect |
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Chymotrypsin |
During digestion, proteins broken down into small peptides by protease This is one of them Able to cleave peptide bond adjacent to aromatic AAs |
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Step 1 |
Substrate binding Side chain in hydrophobic pocket |
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Step 2 |
Ser and his generate ion that attacks peptide carbonyl group Forming short lived negative intermediate |
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Step 3 |
Collapse of intermediate Reformation of double bind with C breaking peptide bond Amino leaving group is protonated |
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Step 4 |
Water molecule is deprotinated forming OH ion OH ion breaks water link forming second intermediate O is again negative in the oxyanion hole |
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Step 5 |
Collapse of intermediate and second product formed: a carboxylate anion and displaces ser |
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Step 6 |
Break off enzyme |
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Step 7 |
Dissociation of second product regenerates free enzyme |
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Sequential |
Cannot easily distinguish random from ordered Will give intersection at y axis |
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Proteases cleaving |
Back (Definition) |
