INHIBITION OF ENZYMES 773 



mercurial are bound to the same SH group. In most cases, increase in the 

 substrate concentration does not reduce the inhibition once established, but 

 Robert et al. (1952) claim that acetylcholine is able to displace Hg++ from 

 horse serum cholinesterase, the affinity of the enzyme for the acetylcholine 

 actually being greater than for the Hg++; it is not known if mercaptide 

 formation is involved. One suspects that the inhibition may sometimes ap- 

 pear to be competitive where actually the mercurial is reacting with the 

 substrate, either exclusively or in addition to the enzyme (compare curves 

 in Figs. 1-5-1 and 1-5-14), and such might be the case in the inhibition of 

 /^-amylase by p-MB and PM (Ghosh, 1958), although the extent of reaction 

 of mercurials with starch is not known. 



Noncompetitive inhibition may be observed when the mercurial reacts 

 with groups, SH or other, adjacent to the active center, and thus suppresses 

 the rate of breakdown of the ES complex, and when the affinity for the 

 enzyme is not so high that mutual depletion kinetics hold. The interference 

 with ES breakdown may be steric through the side chains introduced or 

 secondarily by alteration of the protein structure. It must be remembered 

 that mutual depletion systems usually indicate formally noncompetitive 

 behavior if the common plotting procedures are used (compare Figs. 1-5-3 

 and 1-5-24), despite the fact that the inhibition may be fundamentally 

 competitive. No pure instances of uncompetitive or coupling inhibition have 

 been reported, but it is not unlikely that preferred reaction with the ES 

 complex occurs. The inhibition of alkaline phosphatase is actually mixed 

 (noncompetitive and uncompetitive), but p-MB reacts more readily with 

 the ES complex {K,' = 0.163 mM) than with E {K, = 2.5 milf) (Lazdunski 

 and Ouellet, 1962). There are some examples in which such reaction with 

 the ES complex is possible, e.g., the inhibition of urease by Hg++ (Evert, 

 1952), of acid phosphatase by p-MB (Newmark and Wenger, 1960), and of 

 succinate dehydrogenase by p-MB (Warringa and Giuditta, 1958). The in- 

 hibition of lactate dehydrogenase from Propionibacterium pentosaceum by 

 p-MB is greater in the presence of lactate than when no substrate is present 

 during incubation, and this was postulated to be due to the greater number 

 of free SH groups, presumably arising through reduction by lactate (Moli- 

 nari and Lara, 1960), and a similar situation may occur with glutathione 

 reductase and NADPH, the presence of the reduced coenzyme increasing 

 the inhibition markedly (Mapson and Isherwood, 1963). 



Another approach to the classification of mercurial inhibitions, and per- 

 haps the primary one, is the determination of the component — enzyme, 

 substrate, coenzyme, or cofactor — with which the mercurial reacts. Reac- 

 tion with the apoenzyme has generally been assumed above and with respect 

 to the molecular mechanism might be divided into three types: (1) binding 

 to an SH group at the active center, preventing complexing of the apo- 

 enzyme with any of the other components, (2) binding with an SH group 



