Wednesday, 25 September 2019

What is Competitive Inhibition?

Competitive Inhibition

Competitive Inhibition


Before I tell you, what is competitive inhibition lets to introduce what is Inhibition? Inhibitors are substances which tend to decrease the rate of an enzyme-catalyzed reaction. Although some act on a substrate or cofactor, we will restrict our discussion here to those which combine directly with an enzyme.  Reversible inhibitors bind to an enzyme in a reversible fashion and can be removed by dialysis (or simply dilution) to restore full enzymic activity, whereas irreversible inhibitors cannot be removed from an enzyme by dialysis. Sometimes it may be possible to remove an irreversible inhibitor from an enzyme by introducing another component to the reaction mixture, but this would not affect the classification of the original interaction.

Competitive Inhibition occurs through Reversible inhibitors. Competitive inhibitors often resemble the substrates whose reactions they inhibit, and because of this structural similarity, they may compete for the same binding site on the enzyme. The enzyme-bound inhibitor then either lacks the appropriate reactive group or it is held in an unsuitable position with respect to the catalytic site of the enzyme or to other potential substrates for a reaction to occur. 

In either case, a  dead-end complex is formed, and the inhibitor must dissociate from the enzyme and be replaced by a molecule of the substrate before a reaction can take place at that particular enzyme molecule.
For example,  malonate ( -O2C.CH2.CO2- )  is a competitive inhibitor  of  the reaction catalyzed by  succinate dehydrogenase:
Competitive Inhibition
Malonate has two carboxyl groups, like the substrate, succinate, and can fill the succinate-binding site on the enzyme. However, the subsequent reaction involves the formation of a double bond, and since malonate, unlike succinate, has only one carbon atom between the carboxyl groups, it cannot react.
The effect of a competitive inhibitor depends on the inhibitor concentration, the substrate concentration and the relative affinities of the substrate and the inhibitor for the enzyme. In general, at a particular inhibitor and enzyme concentration, if the substrate concentration is low, the inhibitor will compete favorably with the substrate for the binding sites on the enzyme and the degree of inhibition will be great. However, if, at this same inhibitor and enzyme concentration, the substrate concentration is high, then the inhibitor will be much less successful in competing with the substrate for the available binding sites and the degree of inhibition will be less marked. At very high substrate concentrations, molecules of the substrate will greatly outnumber molecules of inhibitor and the effect of the inhibitor will be negligible. Hence Vmax for the reaction is unchanged.

However, the apparent  Km,  the substrate concentration when V0  =  1/2  Vmax,  is clearly increased as a result of the inhibition and is given the symbol K'm·

Competitive Inhibition

Let us investigate the steady-state kinetics of a simple single-substrate single-binding-site single intermediate enzyme-catalyzed reaction in the presence of a competitive inhibitor,  I.


Competitive Inhibition
Competitive Inhibition

This is an equation of the same form as the Michaelis-Menten equation, the only difference being that Km has been increased by a factor  (1+([Io]/ Ki)).  (Note that the inhibitor concentration will usually be of the same order of magnitude as the substrate concentration and thus much greater than the enzyme concentration, so  [I]≈[I0] just as  [S][S0].)  Therefore, for simple competitive inhibition, Vmax is unchanged but Km is altered so that  K'm = Km[I]+[I0]/Ki)),  where K'm is the apparent Km in the presence of an initial concentration [I0of a competitive inhibitor. It can be seen that  K'm is equal to the concentration of competitive inhibitor which apparently doubles the value of Km. The Lineweaver-Burk equation in the presence  of  a competitive inhibitor will be:
Competitive Inhibition
It must be pointed out that this identical expression would be obtained if the inhibitor-binding site was separate from the substrate-binding site, provided the binding of the substrate to the enzyme resulted in the blockage of the inhibitor-binding site by a conformational change or other mechanisms.  In this situation, the inhibitor could bind to E but not to ES, exactly as discussed above. For this reason, it has become common to classify an inhibitor as competitive if, in its presence, a Lineweaver-Burk plot is obtained with changed  Km  but unchanged  V max,  irrespective of  the actual mechanism involved
Competitive Inhibition

Once competitive inhibition has been identified, it is desirable to determine the inhibitor constant  Ki.  It is obtained from the expression  K'm  =  Km(l  +  ([I0]/Ki)),  but a graphical method is preferred to a direct substitution of numbers, to allow errors in individual determinations to be averaged out. From the above expression:
Competitive Inhibition
Competitive Inhibition

The simplest forms of competitive inhibition considered above are sometimes termed linear competitive inhibition because both primary and secondary plots are linear. In more complicated systems, the primary plots may be linear but the secondary plots non-linear. For example, if not one but two molecules of an inhibitor can bind to the substrate-binding site, then parabolic competitive inhibition is said to occur, because of the shape of the secondary and Dixon plots. Similarly, if the inhibitor binds to a different site from the substrate and reduces the affinity of the enzyme for the substrate without altering the reaction characteristics of that substrate which does bind, then hyperbolic competitive inhibition results.  In each case the primary plots are linear and the inhibition patterns are indistinguishable from those for linear competitive inhibition.
Competitive inhibitors, like other types of inhibitors, may be used to help elucidate metabolic pathways by causing accumulation of intermediates. In this way, Hans Krebs and colleagues used inhibition by malonate to investigate the tricarboxylic acid cycle, of which succinate dehydrogenase is a component.
Provided they are not dangerous to humans, competitive inhibitors may be used in medicine or agriculture as chemotherapeutic drugs, insecticides or herbicides, to destroy or prevent the growth of unwanted organisms. For example, sulphonamides such as sulphanilamide, once widely used in medicine, are competitive inhibitors  of bacterial enzymes involved in the biosynthesis  of  the coenzyme tetrahydrofolate from p-aminobenzoic acid, the structure  of  which is similar to that  of sulphanilamide:
Competitive Inhibition

The metabolic pathway for the formation of tetrahydrofolate from p-aminobenzoic acid is not found in humans, so sulphonamides can be used to limit the growth of bacteria with relatively little risk to the patient. A detailed investigation of the binding characteristics of various competitive inhibitors that bind at the same site as a natural substrate can give useful information about factors governing the binding of the substrate. In two-substrate enzyme-catalyzed reactions, competitive inhibition studies can help elucidate the reaction mechanism.  In this context and in general, it should be noted that the product of a reaction often resembles the substrate and therefore may act as a competitive inhibitor. There are a few instances where competitive inhibition by a product may play an important role in metabolic regulation within the living cell. For example,  2,3-bisphosphoglycerate  inhibits its own formation from  3-phospho-glycerol phosphate,  a reaction catalyzed by bisphosphoglycerate mutase.

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