1. BCOR 011 Lecture 12 9/28/2005ENZYMES

2. Last time… -  G reaction “can” go spontaneous But when will it go? And at what rate?

3. Thermodynamics Whether a reaction will occur Kinetics WHEN a reaction will occur

4. What governs WHEN a reaction will occur? The tower of blocks falling is favorable but when will it happen? Oxidation of carbohydrate polymers (starch) to carbon dioxide and water is favorable but when will it happen? Gasoline burning to carbon dioxide and water is favorable but when will it happen ?

5. For a Reaction to occur need to Destabilize Existing State to INPUT ENERGY Now In Transition Potential net usable energy Destabilization energy input “Activation Energy” Potential net usable energy

6. Need to INPUT ENERGY to Destabilize Existing State In TransitionAfter Potential net usable energy net usable energy released Regain Activation Energy Invested

7. What does activation energy represent? For a Reaction to Occur… - reactants must find each other, - meet in proper orientation - and hit with sufficient force

8. Productive Collision Many Non-productive Collisions

9. Needs of Typical chemical reactions - need large number of molecular collisions - need collide violently enough to break pre-existing bonds (not bounce) - need high concentration to find each other at significant rate HEAT !

10. The energy profile for an exergonic reaction Free energy Progress of the reaction ∆G < O EAEA Figure 8.14 A B C D Reactants A C D B Transition state A B CD Products

11. Molecules with sufficient Energy (<5%) Molecules with sufficient Energy (~40%) Temp 1 Temp 2 EAEA

12. ENZYMES make reactions easier to occur at reasonable temperature by LOWERING the ACTIVATION ENERGY E A of the reaction

13. Activation Energy Energy necessary to overcome the status quo GG Thermodynamic “favorablility” EAEA “ease” of initiating reaction GG EAEA

14. CATALYSTS: promote a specific reaction But are NOT consumed in the process Key concepts: Promotes - does not alter what would normally occur thermodynamically Specificity - promotes only one reaction, only between specific reactants to give specific products Reusable - regenerated in the process

15. ENZYMES are biological CATALYSTS - usually PROTEINS - sometimes RNA or RNA/protein complexes

16. Hard path Easy path Enzymes work as catalysts by providing an easy path to the same point HOW?

17. How do Enzymes do it? 1. Enzymes have BINDING AFFINITY for their reactants = Substrates Brings substrates in close proximity: conc

18. Enzymes act as a Specific Platform Have a very Specific 3-D Shape With a Specific Arrangement of Functional Groups Flexible OH HO + Polar Nonpolar Charged Stabilized Interactions

19. - HO OH HO OH + HO OH SPECIFICITY is the Key to Enzyme Action ENZYMES: Bind ONLY specific things Bind them ONLY in a Specific 3-D Orientation

20. 2. Enzymes ORIENT Substrates always in productive orientation

21. Productive Collision Many Non-productive Collisions ONLY Productive Collisions

22. - HO OH HO OH + HO OH With just a little nudge, can’t help but react

23. 3a. Physical Strain 3b.Chemical Strain BOND STRAIN 3. Enzymes cause BOND STRAIN - destabilize existing bonds - destabilize existing bonds “nutcracker effect” “nutcracker effect”

24. The active site –Is the region on the enzyme where the substrate binds Figure 8.16 Substate Active site Enzyme (a)

25. Induced fit of a substrate Figure 8.16 (b) Enzyme- substrate complex

26. Enzyme-substrate interactions Fischer: Lock & key Koshland: Induced fit 3a. Physical bond strain Draw an quarter - an anvil

27. The catalytic cycle of an enzyme Substrates Products Enzyme Enzyme-substrate complex 1 Substrates enter active site; enzyme changes shape so its active site embraces the substrates (induced fit). 2 Substrates held in active site by weak interactions, such as hydrogen bonds and ionic bonds. 3 Active site (and R groups of its amino acids) can lower E A and speed up a reaction by acting as a template for substrate orientation, stressing the substrates and stabilizing the transition state, providing a favorable microenvironment, participating directly in the catalytic reaction. 4 Substrates are Converted into Products. 5 Products are Released. 6 Active site Is available for two new substrate Mole. Figure 8.17

28. 3b. Chemical Bond Strain tease the bond to fall apart

29. Chemical Bond Strain Stabilize a Fictitiousstate

30. Cofactors CofactorsCofactors –Are nonprotein enzyme helpers, eg Zn ++ CoenzymesCoenzymes –Are organic cofactors Non-polypeptide things at the active site that help enzymes do their job

31. 4. Enzymes “partake” in reactions but are not consumed in them Converts MANY “A’s” into “B’s”

32. H+H+ OH - H+H+ Partakes: but start and end with the same enzyme config

33. Lysozyme

34. Lysozyme: kills bacteria Works at pH 4-5 Why?

35. Enzymes : 1. Bring reactants (substrates) in close proximity 2. Align substrates in proper orientation 3. Can act as a Lever: a press or an anvil small shape change translates to large force 4. Release products when reaction done rebind more substrates Many small steps 5. Many small steps, each easily achieved rather than one huge leap SUMMARY

36. You expect me to JUMP this? No Problem Dude Enzymes carry out reactions in a series of small steps rather than one energetic event

37. Reaction rates: Example: H 2 O 2 -> H 2 O +O 2 uncatalyzed –months Fe +++ 30,000x faster Catalase 100,000,000 x faster Enzyme kinetics- kinetikos – moving

38. Rate orvelocity # made per min Substrate Conc maximum velocity V max 1/2 V max KmKmKmKm “substrate affinity” An enzyme catalyzed rxn Can be “saturated”

39. lower the K m The lower the K m the better the enzyme recognizes substrate “finds it at low conc” higher the V max The higher the V max the more substrate an enzyme can process per min (if substrate around) “top speed” “mpg”

40. Things that affect protein structure often affect enzyme activity often affect enzyme activity temperature pH 0 1 2 3 4 5 6 7 8 9 10 0 20 40 60 80 100 º C

41. Enzyme regulation: Activity controlled Continually adjusted

42. Principal Ways of Regulating Enzymes Principal Ways of Regulating Enzymes Competitive Inhibition Allosteric Inhibition Covalent Modification (phosphorylation)

43. - HO OH HO OH + HO OH HO OH Competitive Inhibitors: bind to active site “unproductively” and block true substrates’ access I S1S1 S2S2 S & I bind to same site

44. Competitive inhibition

45. Allosteric Inhibitors “other” “site” Distorts the conformation of the enzyme Negative allosteric regulator

46. Allosteric inhibition

47. Positive Positive allosteric regulators Helps enzyme work better promotes/stabilizes an “active” conformation

48. Allosteric activation

49. Allosteric regulators change the shape conformation of the enzyme Stabilized inactive form Allosteric activater stabilizes active from Allosteric enyzme with four subunits Active site (one of four) Regulatory site (one of four) Active form Activator Stabilized active form Allosteric activater stabilizes active form Inhibitor Inactive form Non- functional active site (a) Allosteric activators and inhibitors. In the cell, activators and inhibitors dissociate when at low concentrations. The enzyme can then oscillate again. Oscillation Figure 8.20

50. A frequent regulatory modification of enzymes Phosphorylation

51. inactive + P active Phosphorylase kinase

52. Summary 1.enzymes are catalysts 2.Lower activation energy E A 3.Mechanism of action … 4.Enzyme kinetics- V max, K m 5.Regulation of enzyme activity - competitive, allosteric phosphorylation