INHIBITION OF ENZYMES 801 



are all combined with the mercurial the enzyme is completely inhibited, 

 the less readily reacting SH groups being now available. 



(2) Only a small number of SH groups are normally reactive, but follow- 

 ing mercaptide formation the enzyme unfolds and exposes other SH groups, 

 the groups reacted after total inhibition being secondarily released. 



As discussed under Case A, the titration to 100% inhibition does not 

 necessarily provide the number of SH groups involved in the catalysis. 

 Reversal experiments run in parallel to the titration may give some infor- 

 mation as to which mechanism is involved. 



Case E: Inhibition parallels the SH group reaction but levels off. 



One must assume that complete combination of the enzyme with the 

 mercurial does not produce complete inhibition, which could easily be the 

 case if the SH groups reacted are sufficiently far from the active center so 

 that the introduced side chain would modify only the catalysis. A control 

 without the enzyme is, of course, mandatory to eliminate nonenzymic com- 

 ponents to the rate. 



Case F: Inhibition develops without the reaction of SH groups. 



(1) The mercurial, which was assumed to be specific for SH groups, is 

 not, and is inhibiting by another mechanism. 



(2) Reaction of SH groups leads to structural changes by which more 

 SH groups appear; lack of reaction might be concluded from titrations in 

 which residual SH groups are determined, not by direct spectrophotometric 

 methods. 



(3) The method for the estimation of SH group disappearance may be 

 faulty, i.e., those groups reacted by the mercurial may be resistant to the 

 titrating agent. 



There are some instances in which non-SH enzymes are inhibited by 

 mercurials, but in general a result of this type should suggest a re-examina- 

 tion of the methods used. 



Let us now examine a simple system in greater detail. An enzyme having 

 2 SH groups per molecule, one of these groups involved in the catalysis and 

 one not, is treated with increasing amounts of a mercurial; how will the 

 relative reactivities of the SH groups affect the results? Representing the 

 mercurial by M, the two equilibria may be written: 



