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Download Page 20-1 CHAPTER 20: Enzymes 20.2
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Transcript
20.1 GENERAL CHARACTERISTICS CHAPTER 20: Enzymes Globular Globular proteins proteins that that act act as as catalysts catalysts in in biochemical biochemical processes processes • Describe the general characteristics and functions of enzymes H OH • Identify the general function of cofactors HC • Illustrate the mechanism of enzyme function −OOC • Identify factors for the regulation of enzyme activity, including zymogen precursors and allosteric regulation C CITRATE NO REACTION CH2COO− COO− aconitase SUBSTRATE CATALYST CATALYST Increases Increases reaction reaction rate rate without without itself itself undergoing undergoing any net change any net change • Compare competitive and noncompetitive inhibition HC CH2COO− C −OOC COO− cis-ACONITATE PRODUCT ENZMATIC PROCESSES UNCATALYZED • EFFICACY Reaction rates greatly (×109-1020) increased Ea ∆E Ea CATALYZED • SPECIFICITY Rates increased for one specific reaction or reaction class REACTANTS • REGULATION Reaction rates can be controlled PRODUCTS Extent of Reaction 20.2 CLASSIFICATION • TRANSFERASES Transfer of a functional group between molecules Identifying Identifying suffix suffix -ase -ase COO− • OXIDOREDUCTASES Oxidation and reduction COO− CH3 C H OH CH NH3+ CH2 COO− COO−− −2H CH3 C COO− + C O CH2 CH2 COO− O COO− C O CH2 COO− COO− + CH NH3+ CH2 CH2 COO− Page 20-1 N+(CH3)3 N+(CH3)3 • HYDROLASES Hydrolysis or esterification CH2 CH2 H2O CH2 • ISOMERASES Isomerization CH2 O OH CH3C O P O O COO− C COO− CH COO− P COO− O OH CH2OH ~H OH OH HO OH O + O C H CH2 C CH2 CH2 H C O • LIGASES (SYNTHETASES) Joining or condensation of 2 molecules COO− H2O OH HO O− O− • LYASES Addition to a double bond, or elimination to make a double bond CH2 = CH3C OH CH2O P CH2O P S CoA COO− COO− COO− HO C HO CH O CH2 C S CoA CH2 COO− COO− 20.3 ENZYME STRUCTURE: 20.4 MECHANISM of ENZYME ACTION COFACTORS AND COENZYMES • PROTEIN Globular polypeptide of 100-200 residues called an APOENZYME if it also contains • MODE OF ACTION E • COFACTOR(S) – METAL IONS (Mg2+, Fe2+, Zn2+, Mn2+, Ni2+, Se0…) and/or – COENZYMES (organic molecules ← water-soluble vitamins = B vitamins, etc) enzyme + S substrate ES COMPLEX E enzyme + P product(s) Formed Formed at at active active site site of of enzyme enzyme APOENZYME APOENZYME ++ COFACTOR(S) COFACTOR(S) == HOLOENZYME HOLOENZYME CH2 O P • Active site is a small pocket or cleft in enzyme surface ADP ATP CH2 O P CHOH CHOH C C O O P O O− • Substrate is held in active site by noncovalent interactions with amino acid side chains ACTIVE SITE • Active site shape is complementary to substrate shape • Active site shape determines specificity ACTIVE ACTIVE SITE SITE Specific Specific location location on on enzyme enzyme molecule molecule where where the the chemical chemical reaction reaction is is catalyzed catalyzed PHOSPHOGLYCERATE KINASE Page 20-2 ENZYME APOENZYME COFACTOR (COENZYME) ENZYME-SUBSTRATE COMPLEX HOLOENZYME PRODUCTS ENZYME-SUBSTRATE COMPLEX ENZYME-SUBSTRATE COMPLEX ENZYME Effect of Substrate Stereoisomerism: Lactate Dehydrogenase CH3 −OOC −OOC H OH H CH3 H OH • INDUCED-FIT Reaction of closely-related substrates (functional group, bond type, stereochemistry) are catalyzed CH3 ACTIVE SITE H OOC − OH • LOCK-AND-KEY Reaction of only one substrate is catalyzed: absolute specificity Complex formed D-LACTATE −OOC Further Reaction CH3 L-LACTATE SPECIFICITY OF ENZYME-SUBSTRATE COMPLEX OH Mismatch — no complex salt bridge Some drugs work better than the natural substance: Morphine H-bonds hydrophobic interactions Dopamine Page 20-3 20.6 FACTORS AFFECTING ACTIVITY 20.5 ENZYME ACTIVITY The ACTIVITY of an enzyme, its ability to increase the rate of a reaction, is reflected in its TURNOVER NUMBER (mol substrate consumed/min): ENZYME Carbonic anhydrase • ENZYME CONCENTRATION • SUBSTRATE CONCENTRATION TURNOVER Nº REACTION 36,000,000 CO2 + H20 ↔ H2CO3 • TEMPERATURE 60,000 Pyruvate ↔ Lactate • pH 900 Growth of DNA chains Lactate dehydrogenase DNA polymerase ENZYME CONCENTRATION Where [S] >> [E], • Increased [E] leads to increased [ES] Initial Velocity • Reaction rate increases [E] SUBSTRATE CONCENTRATION Vmax • Rate initially responsive to [S] • Rate eventually reaches a maximum (Vmax) Initial Velocity • Vmax reached when the active site of every enzyme molecule is occupied by a substrate molecule [S] Page 20-4 pH • Many structural features pH dependent (ex: COO−, NH3+) Rate • Large deviations from pH optimum irreversibly change 3-D structure • Enzymes most efficient at pH of normal environment (ex: cellular cytoplasm pH ~7) 5 7 9 pH TEMPERATURE • Enzymes most efficient at temperature of normal environment (~37°C) • Below optimum temperature, insufficient energy for normal reaction rate • Above optimum temperature, irreversible changes in 3-D structure (3° structure destroyed) Rate 0 20 40 60 T, °C ?!@x! 20.7 ENZYME INHIBITION • IRREVERSIBLE – Tight, often covalent bonding at active site – Not enzyme specific EX: Hg2+, CN−, INHIBITOR nerve gases • REVERSIBLE – Non-covalent bonding – Competitve: to enzyme active site ACTIVE EX: sulfa drugs – Non-competitve: to cofactors or on enzyme away from active site Page 20-5 INACTIVE • POSITIVE ALLOSTERISM 20.8 REGULATION OF ENZYME ACTIVITY ALLOSTERIC ENZYME ACTIVATOR (MODULATOR) Control of enzymatic processes necessary: • Conserves a cell’s chemical resources/ energy • Stops production of excess or unwanted reaction products ACTIVATED ENZYME REGULATORY SITE YES! • NEGATIVE ALLOSTERISM • END PRODUCT INHIBITION Accumulation Accumulation of of product product promotes promotes conversion conversion back back to to substrate substrate ACTIVATOR (MODULATOR) REGULATORY SITE DEACTIVATED ENZYME LeChatelier’s Principle: Chap 8 WHA? Most Most enzymatic enzymatic reactions reactions can can proceed proceed in in both both the the forward forward and and reverse reverse directions directions • FEEDBACK INHIBITION A E1 B E2 • ZYMOGEN (PROENZYME) CONVERSION C E3 D PROTEOLYTIC ENZYME “Downstream” “Downstream” product product (D) (D) inhibits inhibits own own synthesis synthesis by by acting acting as as negative negative allosteric allosteric inhibitor inhibitor of of “upstream” “upstream” enzyme enzyme (E (E11)) ZYMOGEN INACTIVE ENZYME ACTIVE • Allows transport of enzymes in inactive form Page 20-6 20.9 ENZYME APPLICATIONS • FOOD INDUSTRY: Fermentation, polysaccharide hydrolysis (“lite beer”, Beano™), meat tenderizers • MEDICINE: Post-surgery replacement of enzymes, protein breakdown as surgical procedure, clinical diagnosis of pathology • ANALYSIS: bioprocess monitoring, assaying serum contents, assaying pathogen or allergen proteins Page 20-7