kinetics are obtained. Mixed inhibition is characterised by a linear Lineweaver–Burk
plot that does not fit any of the patterns shown in Fig. 15.5a. The plots for the
uninhibited and inhibited reactions may intersect either above or below the 1/[S]
axis. The associatedKican be obtained from a secondary plot of the slope either of the
primary plot or of 1/Vmaxfor the primary plots against inhibitor concentration. In
both cases the intercept on the inhibitor concentration axis isKi. Non-competitive
inhibition may be regarded as a special case of mixed inhibition.Substrate inhibition
A number of enzymes at high substrate concentration displaysubstrate inhibition
characterised by a decrease in initial rate with increased substrate concentration. The
graphical diagnosis of this situation is shown in Fig. 15.5b. It is explicable in terms of
the substrate acting as an uncompetitive inhibitor and forming a dead-end complex.End-product inhibition
The first enzymes in an unbranched metabolic pathway are commonly regulated by
end-product inhibition. Here the final product of the pathway acts as an inhibitor of
the first enzyme in the pathway thus switching off the whole pathway when the final
product begins to accumulate. The inhibition of aspartate carbamyltransferase by
cytosine triphosphate (CTP) in the CTP biosynthetic pathway is an example of this
form of regulation. In branched pathways, product inhibition usually operates on the
first enzyme after the branch point.Irreversible inhibition
Irreversible inhibitors, such as the organophosphorus and organomercury com-
pounds, cyanide, carbon monoxide and hydrogen sulphide, combine with the enzyme
to form a covalent bond. The extent of their inhibition of the enzyme is dependent
upon the reaction rate constant (and hence time) for covalent bond formation and
upon the amount of inhibitor present. The effect of irreversible inhibitors, which
cannot be removed by simple physical techniques such as dialysis, is to reduce the
amount of enzyme available for reaction. The inhibition involves reactions with a
functional group, such as hydroxyl or sulphydryl, or with a metal atom in the active
site or a distinct allosteric site. Thus the organophosphorus compound, diisopropyl-
phosphofluoridate, reacts with a serine group in the active site of esterases such as
acetylcholinesterase, whilst the organomercury compoundp-hydroxymercuribenzoate
reacts with a cysteine group, in both cases resulting in covalent bond formation and
enzyme inhibition. Such inhibitors are valuable in the study of enzyme active sites
(Section 15.4.1).Applications of enzyme inhibition
The study of the classification and mechanism of enzyme inhibition is of importance
in a number of respects:- it gives an insight into the mechanisms by which enzymes promote their catalytic
activity (Section 15.4.1);
595 15.2 Enzyme steady-state kinetics