inhibitors of enzymes, mechanism of action of enzymes and inhibitors

2. EFFECT OF TEMPERATURE
Raising 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 T
However, 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.

3. Enzymes from humans generally exhibit stability at temperatures
up to 35-40°C.
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 100°C.

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

5. EFFECT OF PH
The 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.

6. EFFECT OF PH
The 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,

7. EFFECT OF PH

8. SUBSTRATE CONCENTRATION
The 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.

10. EFFECT OF ENZYME CONC.
Velocity of enzymatic reactions is
α enzyme conc.
Providing more E molecules
enables conversion of large no. of
substrate molecules into product.

11. 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 P→S.

12. EFFECT OF INHIBITORS
Any 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

13. COMPETITIVE INHIBITION
This 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.

15. 1. 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 for
a given substrate. This means that, in the presence of a competitive
inhibitor, more substrate is needed to achieve 1⁄2Vmax.

16. EFFECT ON M-M GRAPH

17. EFFECT ON LINE WEAVER BURK PLOT

18. 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 hydroxymethylglutaryl–CoA
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.

20. NATURALLY OCCURING
SUBSTANCE /ANALOG
DRUD NAME CHIEF ACTION &
CLINICAL USE
HYPOXANTHINE ALLOPURINOL INHIBIT FORMATION
OF URIC ACID
TREATMENT OF GOUT
PABA(PARA AMINO
BENZOIC ACID)
SULFONAMIDES INHIBIT FORMATION
OF FOLIC ACID BY
BACTERIA
ANTIBACTERIAL DRUG
VITAMIN K DICOUMAROL ANTICOAGULANT
FOLIC ACID METHOTREXATE INHIBIT FH2
REDUCTASE
INHIBIT DNA
SYNTHESIS
ANTICANCER DUUG

21. NATURALLY
OCCURING SUBSTANCE
/ANALOG
DRUD NAME CHIEF ACTION &
CLINICAL USE
ACETYL CHOLINE PHYSIOSTIGMINE INHIBIT Ach ESTERASE
MYESTHENIA GRAVIS
URACIL FIUOROURACIL INHIBIT RNA
SYNTHESIS
EFFECTIVE AGAINST
RNA VIRUS
GLUTAMINE AZASERINE INHIBIT PURINE
SYNTHESIS
ANTICANCER DRUG
CATECHOLAMINES PHENELZINE INHIBIT MAO ENZYME
MENTAL DEPRESSION

22. 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.

25. 1. Effect on Vmax:
2. Noncompetitive inhibition cannot be overcome by
increasing the concentration of substrate. Thus, noncompetitive
inhibitors decrease the apparent Vmax of the reaction.
2. Effect on Km:
Noncompetitive inhibitors do not interfere with the
binding of substrate to enzyme. Thus, the enzyme shows the same
Km in the presence or absence of the noncompetitive inhibitor.

26. EFFECT ON M-M GRAPH

27. EFFECT ON LINE WEAVER BURK PLOT

28. ENZYME INHIBITED DRUG NAME CLINICAL USE
ENOLASE FLUORIDE ION REMOVE Mg & Mn ION
INHIBIT GLYCOLYSIS
ALDEHYDE
DEHYDROGENASE
DISULFIRAM ALCOHOLISM
CHELATE IONIC
CALCIUM
EDTA ANTICOAGULANT
FERROCHELATASE LEAD INHIBIT HEME
SYNTHESIS

29. ENZYME INHIBITED DRUG NAME CLINICAL USE
INHIBIT Ach ESTERASE DFP(DI-ISOPROPYL-
FLUOROURACIL)
INSECTIDE
HAS MANY –SH GROUPS
BIND HEAVY METALS
BAL(DIMERCAPROL) ANTIDOTE FOR HEAVY
METAL POISONING

32. COMPETITIVE INHIBITION NON COMPETITIVE INHIBITION
REVERSIBLE REVERSIBLE OR IRREVERSIBLE
INHIBITOR & SUBSTRATE RESEMBLE
EACH OTHER
DOES NOT RESEMBLE
INHIBITOR BINDS THE ACTIVE SITE INHIBITOR DOESNOT BIND THE ACTIVE
SITE
INHIBITOR CANNOT BIND WITH ES
COMPLEX
INHIBITOR CAN BIND WITH ES
COMPLEX

33. COMPETITIVE INHIBITION NON COMPETITIVE INHIBITION
Vmax IS SAME Vmax LOWERED
Km INCREASED Km UNALTERED
LOWERS THE SUBSTRATE AFFINITY TO
ENZYME
DOESNOT CHANGE SUBSTRATE
AFFINITY FOR ENZYME
COMPLEX IS E-I COMPLEX IS E-S-I OR E-I