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  1. 1. Chapter 6, page 96-103 Enzyme
  2. 2. Topics Function & Composition Catalytic Reaction Enzyme Specificity Factor affecting enzymatic rate of reaction Enzyme Regulation
  3. 3. Enzyme Function Enzyme: biological catalyst that speed up rates of reaction that would otherwise be too slow to support life Catalyst:a chemical agent that changes the rate of a reaction without being consumed by the reaction Enzymes are unaffected by the reaction and are reusable
  4. 4. Enzyme Function Enzymes act on substrates at their active site Substrate: reactant that enzyme acts on Active site: a pocket where the substrate binds; catalytic center where substrate is converted to product
  5. 5. Active Site R-groups on amino acids of the enzyme interact with the substrate at the active site
  6. 6. Enzyme Composition Almost all enzymes are composed of proteins Exception: ribozyme RNA that catalyze reactions on other RNA Hydrolysis at phosphodiester bond http://upload.wikimedia.org/wikipedia/commons/a/a9/Ribozyme.jpg
  7. 7. Enzyme Names Most enzymes have an –ase ending The root name suggests what molecule it acts upon Example: ATPase
  8. 8. Catalytic Reactions Chemical reactions between molecules involve both bond breaking and forming. Example: sucrose hydrolysis bond between glucose and fructose is broken new bonds formed with H+ and OH- Copyright © 2002 Pearson Education, Inc., publishing as Benjamin Cummings Fig. 6.11
  9. 9. Exergonic reaction A reaction that releases energy But reaction still needs an initial investment of energy to break bonds in the reactant Energy usually supplied in the form of heat (thermal energy) Fig. 6.12Copyright © 2002 Pearson Education, Inc., publishing as Benjamin Cummings
  10. 10. Activation Energy (EA) Amount of energy needed to push the reactants over an energy barrier. Reactants absorb energy becoming unstable Thermal agitation increase speed of molecules and number & strength of collisions Peak of instability = transition state Eventually bond breaks Fig. 6.12
  11. 11. Change in Free Energy (ΔG) New bonds release more energy than the initial investment to break bonds. Difference in free energy between products and reactants is the ΔG. ΔG is negative in an exergonic reaction. Fig. 6.12
  12. 12. Enzymes lower EA Allows transition state to occur at a lower temperature which speeds up the reaction. ΔG is unchanged Copyright © 2002 Pearson Education, Inc., publishing as Benjamin Cummings Fig. 6.13
  13. 13. Mechanism for Lowering EA 1. Proximity & Orientation 2. Bond strain 3. Microenvironment 4. Covalent Catalysis
  14. 14. 1. Proximity & Orientation active site brings reactants closer together and in the correct orientation
  15. 15. 2. Bond strain active site bends bonds in substrate making it easier to break
  16. 16. 3. Microenvironment Bibliotheca Alexandrina or Maktabat al-Iskandariyah is a major library and cultural center located on the shore of the Mediterranean Sea in the Egyptian city of Alexandria http://twistedsifter.com/2011/10/beautiful-libraries-around-the-world/
  17. 17. 3. Microenvironment Yale University’s Beinecke Rare Book and Manuscript Library. New Haven, Connecticut.http://twistedsifter.com/2010/08/libraries-around-the-world/
  18. 18. 3. Microenvironment  R-groups at the active site provides a favourable environment for the reaction  Eg: acidic amino acids  low pH  proton donors Thomas Fisher Rare Book Library. University of Toronto.http://twistedsifter.com/2010/08/libraries-around-the-world/
  19. 19. 4. Covalent Catalysis Enzymes may bind covalently to substrates in an intermediate step before returning to normal Increases reaction rate by: Properly orienting the substrate Changing the chemistry at the active site
  20. 20. Enzyme Specificity: Substrate Substrate specific: recognize one specific substrate can even distinguish between particular configurations (e.g. enantiomers) exception: racemase (an isomerase) which recognize both enantiomers and will interconvert between the two forms but the reverse is not true: a given substrate may be acted on by a number of different enzymes substrate specificity due to the fit between the active site and the substrate
  21. 21. Induced-Fit Model As the substrate binds, the enzyme changes shape leading to a tighter fit, bringing chemical groups in position to catalyze the reaction. Binding of a substrate induces a favourable change in the shape of the active site Copyright © 2002 Pearson Education, Inc., publishing as Benjamin Cummings Fig. 6.14
  22. 22. Video: Enzyme Specificity Tutorial Animation http://www.wiley.com//legacy/college/boyer /0470003790/animations/enzyme_binding/enzym htm
  23. 23. Fig. 6.15 Sequence of events E + S  ES  ES*  EP  E + P 1. Substrate binds to available active site pocket forming the enzyme substrate (ES) complex 2. Enzyme changes shape to envelope substrate(s): ES* (transition state) 3. Reaction occurs producing products: Enzyme product (EP) complex 4. Products lose affinity for the active site: E + P 5. Enzyme is set for another substrate: E + S
  24. 24. Fig. 6.15
  25. 25. Enzyme Specificity: Reaction Reaction specific: Enzyme catalysis is specific for one chemical reaction Example: Sucrase is an enzyme that only catalyzes the hydrolysis of sucrose Most metabolic enzymes can catalyze a reaction in both the forward and reverse direction
  26. 26. Factors Affecting Reaction Rate A single enzyme molecule can catalyze thousands or more reactions a second Limitations to enzyme activity and thus reaction rate: Substrate concentration Temperature pH Availability of cofactors
  27. 27. Saturation Curve http://upload.wikimedia.org/wikipedia/commons/thumb/5/5b/Michaelis-Menten_saturation_curve_of_an_enzyme_reaction_pl.svg/800px-Michaelis-Menten_saturation_curve_of_an_enzyme_reaction_pl.svg.png
  28. 28. Substrate concentration on Rate of Reaction Low substrate concentrations: Direct correlation between [S] and rate  [S],  speed of binding to active sites,  reaction rate High substrate concentrations: Enzyme saturation: active sites on all enzymes are engaged The only way to increase productivity at this point is to add more enzyme molecules. **Think: How will the saturation curve change?**
  29. 29. Saturation Curve http://upload.wikimedia.org/wikipedia/commons/thumb/5/5b/Michaelis-Menten_saturation_curve_of_an_enzyme_reaction_pl.svg/800px-Michaelis-Menten_saturation_curve_of_an_enzyme_reaction_pl.svg.png
  30. 30. Temperature Effects on enzyme activity  temperature,  speed of molecules,  collisions between substrate & active site Each enzyme has an optimal temperature If temperature is too high, bonds are disrupted and the protein denatures Copyright © 2002 Pearson Education, Inc., publishing as Benjamin Cummings Fig. 6.16a
  31. 31. pH Effects on enzyme activity Each enzyme has an optimal pH Most between pH 6-8 Exception: digestive enzymes those in the stomach work best at pH 2 those in the intestine are optimal at pH 8 both match their working environments. Copyright © 2002 Pearson Education, Inc., publishing as Benjamin Cummings Fig. 6.16b
  32. 32. Enzyme Type Simple enzyme: composed only of protein component Complex enzyme: an enzyme that requires a cofactor to function in addition to its protein component Apoenzyme: Inactive form of the enzyme because it is missing the cofactor Holoenzyme: Active form of the enzyme; with cofactor
  33. 33. Types of Cofactors Inorganic: metal ions Most common: Zn, Fe, Cu Organic: usually from vitamins or their derivatives Prosthetic group: covalently/permanently bonded to apoenzyme (e.g. heme) Coenzyme: non-covalently/reversibly bound to apoenzyme (e.g. NADH)
  34. 34. Types of Cofactors Cofactor Inorganic e.g. metal ions Organic (vitamin derivative) Prosthetic group (covalently bound) e.g. heme Coenzyme (non-covalently bound) e.g. NADH
  35. 35. Enzyme Regulation Inhibition Competitive Noncompetitive Allosteric Regulation Activation Inhibition Cooperativity
  36. 36. Inhibitors A molecule that binds to an enzyme preventing it from catalyzing reactions. If binding involves covalent bonds, then inhibition is often irreversible. If binding is weak, inhibition may be reversible. Reversible inhibition of enzymes is a natural part of the regulation of metabolism. Copyright © 2002 Pearson Education, Inc., publishing as Benjamin Cummings
  37. 37. Competitive Inhibition Inhibitor binds to the same site as the substrate Think: How do you overcome a competitive inhibitor? Copyright © 2002 Pearson Education, Inc., publishing as Benjamin Cummings Fig. 6.17 a,b
  38. 38. How does a competitive inhibitor change the saturation curve? http://upload.wikimedia.org/wikipedia/commons/thumb/5/5b/Michaelis-Menten_saturation_curve_of_an_enzyme_reaction_pl.svg/800px-Michaelis-Menten_saturation_curve_of_an_enzyme_reaction_pl.svg.png
  39. 39. Effect of competitive inhibitor on saturation curve http://www.examstutor.com/biology/resources/studyroom/biochemistry_and_cells/enzymes/pictures/with_without_graph.gif
  40. 40. Application of competitive inhibition: overcoming alcoholism
  41. 41. Noncompetitive Inhibition Inhibitor binds somewhere other than the active site Causes enzyme to become insensitive to substrate concentrations Brainstorm ways in which an inhibitor could decrease substrate binding without binding to the active site. Think: Can you overcome noncompetitive inhibition by adding more substrate? Copyright © 2002 Pearson Education, Inc., publishing as Benjamin Cummings Fig. 6.17a,c
  42. 42. Example of Mechanisms of Noncompetitive Inhibition http://static.newworldencyclopedia.org/7/7f/Comp_inhib3.pnghttp://upload.wikimedia.org/wikipedia/commons/thumb/5/55/Non-competitive_inhibition.svg/536px-Non-competitive_inhibition.svg.png 1. Interference with the active site 2. alters enzyme conformation rendering the active site unreceptive or less effective Inhibitor binding site Substrate binding site
  43. 43. How does a noncompetitive inhibitor change the saturation curve? http://upload.wikimedia.org/wikipedia/commons/thumb/5/5b/Michaelis-Menten_saturation_curve_of_an_enzyme_reaction_pl.svg/800px-Michaelis-Menten_saturation_curve_of_an_enzyme_reaction_pl.svg.png
  44. 44. Effect of noncompetitive inhibitor on saturation curve
  45. 45. Video: Enzyme Inhibition Tutorial Animation http://www.wiley.com//legacy/college/boyer/ 0470003790/animations/enzyme_inhibition/e nzyme_inhibition.htm
  46. 46. Allosteric Regulation Regulating an enzyme’s activity with a molecule that binds to the enzyme at a site removed from the active site The players: Allosteric enzyme: has an allosteric site Allosteric site: a specific binding site on the enzyme that is not the active site Effector: a regulator that can activate or inhibit by the enzyme by binding to an allosteric site
  47. 47. Allosteric Enzyme Most are constructed of two or more polypeptide chains (subunits) Each subunit has its own active site Allosteric sites are often located where subunits join Protein oscillates between an active and an inactive conformation Copyright © 2002 Pearson Education, Inc., publishing as Benjamin Cummings Fig. 6.18a
  48. 48. Effector: Allosteric Regulators Effectors change enzymatic activity by binding weakly to an allosteric site Allosteric activator: stabilizes the conformation that has a functional active site Allosteric inhibitor: stabilizes the conformation that lacks an active site (inactive form) Often works through cooperativity Copyright © 2002 Pearson Education, Inc., publishing as Benjamin Cummings Fig. 6.18b
  49. 49. Cooperativity Binding of a molecule (substrate or effector) stabilizes a conformational change on all the other subunits Can result in an increase or decrease in substrate affinity Can only occur in enzymes with multiple subunits Copyright © 2002 Pearson Education, Inc., publishing as Benjamin Cummings Fig. 6.20
  50. 50. Cooperativity Example of positive cooperativity: substrate binding to one active site stabilizes conformational changes at all the other subunits Example of negative cooperativity: some forms of allosteric inhibition Copyright © 2002 Pearson Education, Inc., publishing as Benjamin Cummings Fig. 6.20
  51. 51. HW Question Compare noncompetitive inhibition and allosteric inhibition. Compare allosteric activation and cooperativity. Remember: “Compare” means you’re looking at both similarities and differences. And the similarities should indicate the relationship between the 2 items and the reasons why they are being compared in the first place.