Chapter Six The Behavior of Proteins: Enzymes

1. Chapter SixThe Behavior of Proteins: Enzymes

2. Enzyme Catalysis • Enzyme: a _____________________ • with the exception of some __________ that catalyze their own splicing (Section 10.4), all enzymes are proteins (???) • enzymes can increase the rate of a rxn by a factor of up to 1020 over an uncatalyzed rxn • some enzymes are so specific that they catalyze the rsn of only one stereoisomer; others catalyze a family of similar rxns • The rate of a reaction depends on its activation energy, DG°‡ • an enzyme provides an alternative pathway with a ______________________________

3. Enzyme Catalysis (Cont’d) • For a reaction taking place at constant temperature and pressure, e.g., in the body • the change in __________________________ is • Difference in energies between initial state and final state • The change in free energy is related to the equilibrium constant, Keq, for the reaction by

4. Enzyme Catalysis (Cont’d) • Consider the reaction H2O2→ H2O + O2

5. Temperature dependence of catalysis • Temperature can also “catalyze reaction” (increase rate) • This is dangerous, why? • Increasing temperature will lead to _______________ ______________________

6. Enzyme Kinetics • For the reaction • The rate of reaction is given by rate equation • Where k is a proportionality constant called the ___________________________________________ • ______________________________: the sum of the exponents in the rate equation: f+g

7. Enzyme Kinetics (Cont’d) • Consider the reaction Whose rate equation is given by the expression • Determined experimentally, not from _______________ • The reaction is said to be first order in A, first order in B, and second order overall • Consider this reaction of glycogen with phosphate

8. How Enzymes bind to Substrate • In an enzyme-catalyzed reaction • ____________________________, S: a reactant • ______________________: the small portion of the enzyme surface where the substrate(s) becomes bound by noncovalent forces, e.g., hydrogen bonding, electrostatic attractions, van der Waals attractions

9. Binding Models • Two models have been developed to describe formation of the_____________________ complex • __________________ model: substrate binds to that portion of the enzyme with a complementary shape • _________________ model: binding of the substrate induces a change in the conformation of the enzyme that results in a complementary fit

10. Two Modes of E-S Complex Formation

11. Formation of Product

12. An Example of Enzyme Catalysis ____________________ catalyzes • The selective hydrolysis of ___________________ where the ________ is contributed by _____ and ____ • It also catalyzes hydrolysis of the ____________ bonds

13. An Example of Enzyme Catalysis (Cont’d)

14. Non-Allosteric Enzyme Behavior Point at which the rate of reaction does not change, enzyme is __________________, maximum rate of reaction is reached

15. ATCase: An Example of Allosteric Behavior • ____________ shape - characteristic of __________ • Again max velocity reached, but different mechanism

16. Michaelis-Menten Kinetics Initial rate of an enzyme-catalyzed rxn vs [S]

17. Michaelis-Menten Model • For an enzyme-catalyzed reaction • The rates of formation and breakdown of ES are given by these equations • At steady state

18. Michaelis-Menten Model (Cont’d) • When ______________is reached, the concentration of free enzyme is the total minus that bound in ES • Substituting for the concentration of free enzyme and collecting all rate constants in one term gives • KM is called the ____________________________

19. Michaelis-Menten Model (Cont’d) • It is now possible to solve for the concentration of the enzyme-substrate complex, [ES] • Or alternately

20. Michaelis-Menten Model (Cont’d) • In the initial stages, formation of product depends only on the _______________________________________________ • If substrate concentration is ________________________ is _______________________ [ES] = [E]T • Substituting k2[E]T = Vmax into the top equation gives

21. Michaelis-Menten Model (Cont’d) When _______________ the equation reduces to

22. Linearizing The Michaelis-Menten Equation • Vmax is difficult to ___________________________________ • The equation for a hyperbola • Can be transformed into the equation for a ________ by taking __________________________________

23. Lineweaver-Burk Plot • The _______________________ plot has the form y = mx + b, and is the formula for a straight line • a plot of 1/V versus 1/[S] will give a straight line with slope of _______________ and y intercept of _______________ • known as a __________________________________________

24. Lineweaver-Burk Plot (Cont’d) • KM is the ________________________________________ • the greater the value of KM, the ________ tightly S is bound to E • Vmax is the ___________________________________

25. Turnover Numbers • Vmax is related to the ___________________________ of enzyme: also called kcat Number of moles of substrate that react to form product _____________________________________________

26. Enzyme Inhibition • ____________ inhibitor: a substance that binds to an enzyme to inhibit it, but can be released • ____________________________ inhibitor: binds to the active (catalytic) site and blocks access to it by substrate • _____________________ inhibitor: binds to a site other than the active site; inhibits the enzyme by changing its conformation • ________________________inhibitor: a substance that causes inhibition that cannot be reversed • usually involves formation or breaking of covalent bonds to or on the enzyme

27. Competitive Inhibition • Substrate competes with inhibitor for the active site; more substrate is required to reach a given reaction velocity • We can write a dissociation constant, KI for EI

28. Competitive Inhibition

29. Competitive Inhibition In a Lineweaver-Burk plot of 1/V vs 1/[S], the __________________ (and the x intercept) changes but the ______________________ does not change

30. A Lineweaver-Burke Plot, Competitive Inhibition

31. Noncompetitive Inhibition (Cont’d) • Several equilibria are involved • The maximum velocity Vmax has the form

32. Noncompetitive Inhibition (Cont’d)

33. Lineweaver-Burke Plot, Noncompetitive Inhibition • Because the inhibitor does not interfere with ______________ to the active site, KM is ______________________ • Increasing substrate concentration ____________________ noncompetitive inhibition

34. Lineweaver-Burke Plot, Noncompetitive Inhibition

35. Other Types of Inhibition • _____________________ -inhibitor can bind to the ES complex but not to free enzyme; Vmax decreases and KM decreases. • __________________ - Similar to noncompetitive, but binding of I affects binding of S and vice versa.