1. Enzyme Mechanisms

2. Oxidoreductases catalyze oxidation-reduction reactions.
Transferases catalyze transfer of functional groups from one molecule to another.
Hydrolases catalyze hydrolytic cleavage.
Lyases catalyze removal of a group from or addition of a group to a double bond, or other cleavages involving electron rearrangement.
Isomerases catalyze intramolecular rearrangement.
Ligases catalyze reactions in which two molecules are joined.
Types of Enzymes

3. Two Models for Enzyme-Substrate Interaction

4. Induced Conformational Change in Hexokinase

5. Coenzymes

6. Example of a Coenzyme Involved in Oxidation/Reduction Reactions
Hydride (H:-) transfer
(nicotinamide adenine dinucleotide)

7. Stereospecificity of Yeast Alcohol Dehydrogenase
ethanol
acetaldehyde
pro-S
pro-R
pro-R
pro-S
Vennesland and Westheimer, 1950

8. Stereospecificity Conferred by an Enzyme

9. Catalytic Mechanisms Acid-base catalysis Covalent catalysis
Metal ion catalysis
Electrostatic catalysis
Proximity and orientation effects
Preferential binding to transition state (transition state stabilization)

10. Acid-Base Catalysis

11. Keto-Enol Tautomerism: Uncatalyzed vs. Acid- or Base-Catalyzed

12. Example of Acid-Base Catalysis: Bovine Pancreatic RNase A

13. Covalent Catalysis: Nucleophiles and Electrophiles
Protonated

14. Example of Covalent Catalysis: Decarboxylation of Acetoacetate
Lysine side chain e-amino group on enzyme is nucleophile in attack on substrate.
Electrophilic “electron sink”

15. Example of Metal Ion Catalysis: Carboxypeptidase A

16. Example of Metal Ion Catalysis: Carbonic Anhydrase
Carbonic anhydrase catalyzes the reaction:
CO2 + H2O HCO3− + H+  

17. Enolase Mechanism

18. Entropic and Enthalpic Factors in Catalysis
Proximity and orientation effects
Transition state stabilization through preferential binding of transition state

19. Proximity and Orientation Effects

20. Enzymes Are Complementary to Transition State

21. Serine Protease Mechanism: Multiple Catalytic Mechanisms at Work

22. Structure of the Serine Protease Chymotrypsin

23. Serine Protease Substrate Specificity and Active-Site Pockets
Trypsin cleaves amide bond immediately C- terminal to basic amino acid residues.
Substrate specificity in serine proteases through active-site binding of side chain of amino acid residue adjacent to amide bond that will be cleaved.
Chymotrypsin cleaves amide bond immediately C-terminal to hydrophobic amino acid residues.

24. Serine Nucleophile in Serine Proteases

25. The Pre-Steady State in Chymotrypsin-Catalyzed Hydrolysis of p-Nitrophenyl Acetate
H2O k1
E + S ES  EP2  E + P2  
k-1
k k3
vo = kcat[E]t[S]/(KM + [S])
Steady-state velocity, where
kcat = k2k3/(k2 + k3)
KM = KSk3/(k2 + k3)
KS = k-1/k1
For chymotrypsin with ester substrates: k2 >> k3
Release of P1 faster than EP2 breaks down to E + P2

26. Serine Protease Mechanism
Catalytic triad:
Acid-base catalysis
Covalent catalysis
Proximity/orientation effects
Also (not depicted here) - electrostatic catalysis and transition state stabilization

32. carboxylic acid,

34. The Oxyanion Hole In Serine Proteases
Role of oxyanion hole in serine protease mechanism:
Electrostatic catalysis
Preferential binding of transition state

35. Trypsin/Bovine Pancreatic Trypsin Inhibitor (BPTI) Complex
Trypsin-BPTI complex resembles tetrahedral transition state.

36. Transition State in Proline Racemase Reaction and Transition State Analogs
Proline racemase preferentially binds transition state, stabilizing it, and is potently inhibited by transition state analogs.

37. RNA-Based Catalysts (Ribozymes)

38. Cleavage of a Typical Pre-tRNA by Ribonuclease P
Ribonuclease P is a ribonucleoprotein (RNA- and protein-containing complex), and the catalytic component is RNA.
An even more complex example of an RNA- and protein-containing enzyme system is the ribosome. The central catalytic activity of the ribosome (peptide bond formation) is catalyzed by an RNA component.
tRNA substrate of ribonuclease P

39. Catalysis by the Intervening Sequence in Tetrahymena Preribosomal RNA
RNA by itself without any protein can be catalytic.

40. Enzyme Regulation

42. Effect of Cooperative Substrate Binding on Enzyme Kinetics
Cooperative enzymes do not obey simple Michaelis-Menten kinetics.

43. Effect of Extreme Homoallostery
Homotropic allosteric regulation by substrate: S at one active site affects catalysis of S P at other sites on enzyme complex.  
Extreme positive cooperativity depicted here
[S]c = critical substrate concentration

44. Heteroallosteric Control of an Enzyme
(positive allosteric effector)
Heterotropic allosteric regulation: non-S effectors modulate catalysis of S P.  
(negative allosteric effector)

45. A Model for Enzyme Regulation: Aspartate Transcarbamoylase (Aspartate Carbamoyltransferase) in Pyrimidine Synthesis

46. Aspartate Transcarbamoylase (ATCase)-Catalyzed Reaction

47. Feedback Inhibition of ATCase by CTP

48. Regulation of ATCase by ATP and CTP
ATP is a positive heterotropic allosteric effector of ATCase, while CTP is a negative heterotropic allosteric effector.

49. Detailed Structure of One Catalytic Subunit and Adjacent Regulatory Subunit of ATCase

50. Quaternary Structure of ATCase in T State and R State
CTP and ATP bind at regulatory site, but CTP preferentially binds in T state, while ATP preferentially binds R state.

51. X-Ray Structure of Aspartate Transcarbamoylase
T State
T State
“top” view
“side” view
R State
“side” view