FACTORS AFFECTING ENZYME ACTIVITY

FACTORS AFFECTING ENZYME ACTIVITY

EFFECT OF TEMPERATURERaising the temperature increases the rate of both uncatalyzed and enzyme-catalyzed reactions by increasing the kinetic energy and the collision frequency of the reacting molecules. of velocity with higher THowever, heat energy can also increase the kinetic energy of the enzyme to a point that exceeds the energy barrier for disrupting the noncovalent interactions that maintain its three dimensional structure.

Enzymes from humans generally exhibit stability at temperatures up to 35-40C. By contrast, enzymes from the thermophilic microorganisms that reside in volcanic hot springs or undersea hydrothermal vents may be stable up to or even above 100C.

The Q10, or temperature coefficient, is the factor by which rate of a biologic process increases for a 10C increase in temperature.For the temperatures over which enzymes are stable, the rates of most biologic processes typically double for a 10C rise in temperature (Q10 = 2).

EFFECT OF PHThe rate of almost all enzyme-catalyzed reactions exhibits a significant dependence on hydrogen ion concentration. Most intracellular enzymes exhibit optimal activity at pH values between 5-9.For enzymes whose mechanism involves acid-base catalysis, the residues involved must be in the appropriate state of protonation for the reaction to proceed. The binding and recognition of substrate molecules with dissociable groups also typically involves the formation of salt bridges with the enzyme.

EFFECT OF PHThe most common charged groups are carboxylate groups (negative) and protonated amines (positive). Gain or loss of critical charged groups adversely affects substrate binding and thus will retard or abolish catalysis.

The pH at which maximal enzyme activity is achieved is different for different enzymes, and often reflects the [H+] at which the enzyme functions in the body. For example, pepsin, a digestive enzyme in the stomach, is maximally active at pH 2,

EFFECT OF PH

SUBSTRATE CONCENTRATIONThe rate of an enzyme-catalyzed reaction increases with substrate concentration until a maximal velocity (Vmax) is reached. The leveling off of the reaction rate at high substrate concentrations reflects the saturation with substrate of all available binding sites on the enzyme molecules present. Most enzymes show Michaelis-Menten kinetics, in which the plot of initial reaction velocity (vo) against substrate concentration ([S]), is hyperbolic.

EFFECT OF ENZYME CONC.

EFFECT OF PRODUCT CONC.Product formed as result of enzymatic reaction may accumulate & may lower the enzymatic reaction by occupying active site of E.Under certain conditions of high conc. of P a reverse reaction may be favored from PS.

EFFECT OF INHIBITORSAny substance that can diminish the velocity of an enzyme-catalyzed reaction is called an inhibitor. In general, irreversible inhibitors bind to enzymes through covalent bonds. Reversible inhibitors typically bind to enzymes through noncovalent bonds.TYPES OF ENZYME INHIBITION COMPETITIVE INHIBITION NONCOMPETITIVE INHIBITION SUICIDE INHIBITION

COMPETITIVE INHIBITIONThis type of inhibition occurs when the inhibitor binds reversibly to the same site that the substrate would normally occupy and, therefore, competes with the substrate for that site.In such inhibition both EI & ES complexes are formed during the reaction.Actual amount of ES & EI will depend on;Affinity b/w E & S/I.Actual conc. of S & I present.Time of preincubation of E with the S or I.

Effect on Vmax: The effect of a competitive inhibitor is reversed by increasing [S]. At a sufficiently high substrate concentration, the reaction velocity reaches the Vmax observed in the absence of Inhibitor.2. Effect on Km: A competitive inhibitor increases the apparent Km fora given substrate. This means that, in the presence of a competitiveinhibitor, more substrate is needed to achieve 12Vmax.

EFFECT ON M-M GRAPH

EFFECT ON LINE WEAVER BURK PLOT

Statin drugs as examples of competitive inhibitors: This group of antihyperlipidemic agents competitively inhibits the first committed step in cholesterol synthesis. This reaction is catalyzed by hydroxymethylglutarylCoA reductase (HMG-CoA reductase) Statin drugs, such as atorvastatin (Lipitor) and pravastatin (Pravachol),are structural analogs of the natural substrate for this enzyme, and compete effectively to inhibit HMG-CoA reductase.By doing so, they inhibit de novo cholesterol synthesis, thereby lowering plasma cholesterol levels.

NATURALLY OCCURING SUBSTANCE /ANALOGDRUD NAMECHIEF ACTION & CLINICAL USEHYPOXANTHINEALLOPURINOLINHIBIT FORMATION OF URIC ACIDTREATMENT OF GOUTPABA(PARA AMINO BENZOIC ACID)SULFONAMIDESINHIBIT FORMATION OF FOLIC ACID BY BACTERIAANTIBACTERIAL DRUGVITAMIN KDICOUMAROLANTICOAGULANTFOLIC ACID METHOTREXATEINHIBIT FH2 REDUCTASEINHIBIT DNA SYNTHESIS ANTICANCER DUUG

NATURALLY OCCURING SUBSTANCE /ANALOGDRUD NAMECHIEF ACTION & CLINICAL USE

ACETYL CHOLINEPHYSIOSTIGMINEINHIBIT Ach ESTERASEMYESTHENIA GRAVISURACILFIUOROURACILINHIBIT RNA SYNTHESISEFFECTIVE AGAINST RNA VIRUSGLUTAMINEAZASERINEINHIBIT PURINE SYNTHESISANTICANCER DRUGCATECHOLAMINESPHENELZINEINHIBIT MAO ENZYMEMENTAL DEPRESSION

NONCOMPETITIVE INHIBITION This type of inhibition is recognized by its characteristic effect on Vmax. Noncompetitive inhibition occurs when the inhibitor and substrate bind at different sites on the enzyme. It may be reversible & irreversible. The noncompetitive inhibitor can bind either free enzyme or the ES complex, thereby preventing the reaction from occurring.This probably brings about changes in 3D structure of E ,inactivating it catalytically.If I can be removed from its site of binding without affecting activity of E, it is called reversible NCI.If inhibitor can be removed only at loss of E activity, it is known as irreversible NCI.

Effect on Vmax: Noncompetitive inhibition cannot be overcome byincreasing the concentration of substrate. Thus, noncompetitiveinhibitors decrease the apparent Vmax of the reaction.2. Effect on Km: Noncompetitive inhibitors do not interfere with thebinding of substrate to enzyme. Thus, the enzyme shows the sameKm in the presence or absence of the noncompetitive inhibitor.

EFFECT ON M-M GRAPH

EFFECT ON LINE WEAVER BURK PLOT

ENZYME INHIBITEDDRUG NAMECLINICAL USEENOLASEFLUORIDE IONREMOVE Mg & Mn IONINHIBIT GLYCOLYSISALDEHYDE DEHYDROGENASEDISULFIRAMALCOHOLISMCHELATE IONIC CALCIUMEDTAANTICOAGULANTFERROCHELATASELEADINHIBIT HEME SYNTHESIS

ENZYME INHIBITEDDRUG NAMECLINICAL USE

INHIBIT Ach ESTERASE DFP(DI-ISOPROPYL-FLUOROURACIL)INSECTIDEHAS MANY SH GROUPSBIND HEAVY METALS BAL(DIMERCAPROL)ANTIDOTE FOR HEAVY METAL POISONING

COMPETITIVE INHIBITIONNON COMPETITIVE INHIBITIONREVERSIBLEREVERSIBLE OR IRREVERSIBLEINHIBITOR & SUBSTRATE RESEMBLE EACH OTHERDOES NOT RESEMBLEINHIBITOR BINDS THE ACTIVE SITEINHIBITOR DOESNOT BIND THE ACTIVE SITEINHIBITOR CANNOT BIND WITH ES COMPLEXINHIBITOR CAN BIND WITH ES COMPLEX

COMPETITIVE INHIBITIONNON COMPETITIVE INHIBITION

Vmax IS SAMEVmax LOWEREDKm INCREASEDKm UNALTEREDLOWERS THE SUBSTRATE AFFINITY TO ENZYMEDOESNOT CHANGE SUBSTRATE AFFINITY FOR ENZYMECOMPLEX IS E-ICOMPLEX IS E-S-I OR E-I