2. CH13. Enzymes http://www.youtube.com/watch?v=A cXXkcZ2jWM&feature=related

3. Essential Questions What are enzymes? What do they do?

4. Outline What characteristic features define enzymes? Can the rate of an enzyme-catalyzed reaction be defined in a mathematical way? What equations define the kinetics of enzyme-catalyzed reactions? What can be learned from the inhibition of enzyme activity? What is the kinetic behavior of enzymes catalyzing bimolecular reactions? How can enzymes be so specific? Are all enzymes proteins? Is it possible to design an enzyme to catalyze any desired reaction?

5. Virtually All Reactions in Cells Are Mediated by Enzymes Enzymes catalyze thermodynamically favorable reactions, causing them to proceed at extraordinarily rapid rates (see Figure 13.1) Enzymes provide cells with the ability to exert kinetic control over thermodynamic potentiality Living systems use enzymes to accelerate and control the rates of vitally important biochemical reactions Enzymes are the agents of metabolic function

6. Virtually All Reactions in Cells Are Mediated by Enzymes Figure 13.1 Reaction profile showing the large free energy of activation for glucose oxidation. Enzymes lower ΔG ‡, thereby accelerating rate.

7. 13.1 What Characteristic Features Define Enzymes? Enzymes can accelerate reactions as much as 10 16 over uncatalyzed rates Urease is a good example: – Catalyzed rate: 3x10 4 /sec – Uncatalyzed rate: 3x10 -10 /sec – Ratio is 1x10 14

8. What is urease?? 5PT

9. Urea is the single most abundant form of dissolved organic nitrogen present in aquatic ecosystems

10. Specificity Enzymes selectively recognize proper substrates over other molecules Enzymes produce products in very high yields - often much greater than 95% Specificity is controlled by structure - the unique fit of substrate with enzyme controls the selectivity for substrate and the product yield

11. Enzymes are the Agents of Metabolic Function Figure 13.2 The breakdown of glucose by glycolysis provides a prime example of a metabolic pathway.

12. Enzyme Nomenclature Provides a Systematic Way of Naming Metabolic Reactions

13. Coenzymes and Cofactors Are Nonprotein Components Essential to Enzyme Activity

14. 13.2 Can the Rate of an Enzyme-Catalyzed Reaction Be Defined in a Mathematical Way? Kinetics is the branch of science concerned with the rates of reactions Enzyme kinetics seeks to determine the maximum reaction velocity that enzymes can attain and binding affinities for substrates and inhibitors Analysis of enzyme rates yields insights into enzyme mechanisms and metabolic pathways This information can be exploited to control and manipulate the course of metabolic events

15. Several kinetics terms to understand rate or velocity rate constant rate law order of a reaction molecularity of a reaction

16. Chemical Kinetics Provides a Foundation for Exploring Enzyme Kinetics Consider a reaction of overall stoichiometry as shown: The rate is proportional to the concentration of A

17. Catalysts Lower the Free Energy of Activation for a Reaction A typical enzyme-catalyzed reaction must pass through a transition state The transition state sits at the apex of the energy profile in the energy diagram The reaction rate is proportional to the concentration of reactant molecules with the transition-state energy This energy barrier is known as the free energy of activation Decreasing ΔG ‡ increases the reaction rate The activation energy is related to the rate constant by:

18. Catalysts Lower the Free Energy of Activation for a Reaction Figure 13.5 Energy diagram for a chemical reaction (A→P) and the effects of (a) raising the temperature from T 1 to T 2, or (b) adding a catalyst.

19. The Transition State Understand the difference between  G and  G ‡ The overall free energy change for a reaction is related to the equilibrium constant The free energy of activation for a reaction is related to the rate constant It is extremely important to appreciate this distinction

20. The Michaelis-Menten Equation is the Fundamental Equation of Enzyme Kinetics Louis Michaelis and Maud Menten's theory It assumes the formation of an enzyme-substrate complex It assumes that the ES complex is in rapid equilibrium with free enzyme Breakdown of ES to form products is assumed to be slower than 1) formation of ES and 2) breakdown of ES to re-form E and S

21. [ES] Remains Constant Through Much of the Enzyme Reaction Time Course in Michaelis-Menten Kinetics Figure 13.8 Time course for a typical enzyme-catalyzed reaction obeying the Michaelis-Menten, Briggs- Haldane models for enzyme kinetics. The early state of the time course is shown in greater magnification in the bottom graph.

22. [E T ]=[E]+[ES] Product formation rate=k 1 ([ET]-[ES])[S] [ES] dissociation=k 2 [ES]+k -1 [ES] d[ES]=0, steady state assumption k 1 ([E T ]-[ES])[S] = k 2 [ES]+k -1 [ES] (k 2 +k -1 )/k 1 = ([E T ]-[ES])[S]/[ES] v = d[P]/dt v = k 2 [ES] v = k 2 [E T ][S]/Km+[S]

23. Understanding K m The "kinetic activator constant" K m is a constant K m is a constant derived from rate constants K m is, under true Michaelis-Menten conditions, an estimate of the dissociation constant of E from S Small K m means tight binding; high K m means weak binding Km = (k-1+k2)/k1 Km = [S]([Et]-[ES])/[ES]

24. Understanding V max The theoretical maximal velocity V max is a constant V max is the theoretical maximal rate of the reaction - but it is NEVER achieved in reality To reach V max would require that ALL enzyme molecules are tightly bound with substrate V max is asymptotically approached as substrate is increased

25. The Turnover Number Defines the Activity of One Enzyme Molecule A measure of catalytic activity k cat, the turnover number, is the number of substrate molecules converted to product per enzyme molecule per unit of time, when E is saturated with substrate. If the M-M model fits, k 2 = k cat = V max /E t Values of k cat range from less than 1/sec to many millions per sec

26. The Turnover Number Defines the Activity of One Enzyme Molecule

27. The Ratio k cat /K m Defines the Catalytic Efficiency of an Enzyme The catalytic efficiency: k cat /K m An estimate of "how perfect" the enzyme is k cat /K m is an apparent second-order rate constant It measures how the enzyme performs when S is low The upper limit for k cat /K m is the diffusion limit - the rate at which E and S diffuse together