Mechanism of enzyme action

Presentation given by Dr. Karthikeyan at Department of Biochemistry, Maulana Azad Medical College on 04.04.2013.

1. Mechanism of Enzymatic Catalysis Moderator: Dr. S. K. Gupta Presenter: Dr. Karthikeyan

2. Enzymes increases the rate of reaction

3. Higher reaction rates, 10 6 -10 12 Milder reaction conditions (temp, pH, …) Greater reaction specificity (no side products) Capacity for regulation Chemical catalyst Vs. Biological catalyst

4. Chemical catalyst Vs. Biological catalyst Temperature of 400 ° C Pressure of 200 atm Iron catalyst

5. Proximity Straining Orientation Change Change of environment Transition state stabilization How does enzymes increase the rate ?

6. Acid Base Catalysis Covalent Catalysis Metal Ion Catalysis What are all the modes of catalysis ?

7. Active Site 3 dimensional cleft Unique microenvironment Nature of binding Specificity

8. Changes during and after E-S complex E-S Complex Multiple, Reversible, Weak Interactions Binding Energy Helps in attainment of transition state Products

9. Transition state Fleeting, Momentary, Unstable chemical species Evidence for existence Meaning of Transition State stabilization

10. Enzymes lower Δ G ‡

11. Stabilization of transition state is the mechanism of enzyme action.

12. Transition State Vs. Intermediate

13. Active site is more complementary to transition state than substrate. So, Transition state analogues are better competitive inhibitors than substrate analogues

14. higher their concentration , the more frequently they will encounter one another Concept of effective molarity Proximity Effect

15. Proximity Effect

16. Straining

17. Chemical Reactions Nucleophilic Substitutions Cleavage Reactions Oxidation–Reduction Reactions

18. Nucleophilic Substitutions

19. denotes movement of electrons

20. Acid-Base catalysis

21. Why fossils of RNA world NOT found ?

22. Structure of RNA

23. RNAse A

24. RNAse A

25. h -oh

26. -oh

27. -oh

28. <- -o h

29. <- -o h

30. <- -o h

31. Why RNAse can’t act on DNA ?

32. Covalent catalysis

33. Serine proteases

34. Chymotrypsin Hydrophobic pocket

35. Trypsin Deep groove with negatively charged aspartate inside

36. Elastase Shallow groove

37. Catalytic Triad

38. Ser-195 becomes a nucleophile

39. Tetrahedral intermediate Ser-195 attacks the carbonyl group

40. Acyl enzyme intermediate

41. Binding of Water

42. Tetrahedral intermediate

43. Regeneration of enzyme

44. Other Serine Proteases Subtilisin Wheat germ Serine carboxy peptidase II E.coli Clp protease ( caseinolytic peptidase)

45. Serine proteases are example for both convergent and divergent evolution !

46. Aspartic protease HIV PROTEASE Pepsin Rennin Lysosomal cathepsins .

47. Aspartic protease Catalysis involves two conserved aspartyl residues which act as acid-base catalysts 2 different active site aspartates act simultaneously as a general base or as a general acid . This is possible because their immediate environment favours ionization of one but not the other.

48. Metal Ion catalysis

49. Charge Shielding by metals

50. Binding to zinc lowers the p K a of water from 15.7 to 7

51. Ogston’s 3 Point Attachment Theory

52. Enzymes react stereo-specifically with chiral compounds

53. Worked under Nobel laureate  H. Gobind Khorana A prominent scientific journal rejected his work as "a technological development of no general interest“. Michael Smith

54. Vitalistic Theory

55. thank you!