770 7. MEKCURIALS 



S — E. However, the situation is more complex, the pH being an important 

 factor in determining the type of complex occurring, and it is likely that 

 the crystalline mercuri-papain is the least soluble form of several possible 

 derivatives (Smith et al., 1954 b). Sedimentation studies at pH 4 indicate 

 a monomer or 1 : 1 complex, probably to be designated by HS — E — S — Hg+, 

 while at pH 8 there is a heavy component corresponding to a hexamer, 

 possibly cyclic with alternating — S — S — and — S — Hg — S — bonds. It is 

 interesting that there are two electrophoretic peaks at pH 4, one of unit + 

 charge greater than the other; since dissociation of the dimer must result 

 in equal proportions of HS — E — SH and HS — E — S — Hg+, this would tend 

 to confirm the dimeric structure. Oxidized papain is a mixture of 



e(^\ and E— S— S— E 



and does not react with Hg++; thus it is very important in studying the 

 combining ratios to be certain that the papain is fully reduced. Mercuri- 

 papain is actually purer than papain, as indicated by electrophoretic stud- 

 ies, has fewer N-terminal residues detected by the fluorodinitrobenzene 

 technique (Thompson, 1954), and has some 10% greater activity following 

 removal of the Hg++ with cysteine and EDTA, and it is also more stable. 

 The proteolytic enzyme, pinguinain, also forms stable complexes with Hg++ 

 which are stable for much longer times than the pure enzyme (Messing, 

 1961). Other enzymes to be crystallized as the mercury complexes are a 

 lysozyme from papaya latex (Smith et al., 1955) and 3-phosphoglyceralde- 

 hyde dehydrogenase from yeast (Velick, 1953), the latter after reaction with 

 p-MB. There is some evidence that a mercuric dimer of ficin occurs (Liener, 

 1961) while carboxy peptidase forms very stable Hg++ complexes which still 

 possess esteratic activity, although they no longer function as peptidases 

 (Vallee et al., 1961; Coleman and Vallee, 1961). There is thus sufficient 

 evidence that many enzymes form well-characterized mercurial complexes 

 and are quite stable in this state; we shall note other examples in the dis- 

 cussion of SH titrations of enzymes. 



These complexes of enzymes with Hg++ offer strong support to the con- 

 cept that completely selective reaction with SH groups can occur. How- 

 ever, if Hg++ is added in excess of that required for mercaptide formation, 

 it is quite possible that other enzyme groups may be attacked. It is likely 

 that other enzymes under the appropriate conditions can form dimers, or 

 other polymers, with Hg++, in which case the active centers may be made 

 inaccessible even though the SH group is not within the confines of the cen- 

 ter. The appearance of polymers will presumably depend strongly on the pH 

 since, at pH's progressively removed from the isoelectric point, one might 

 expect polymerization to be more and more reduced, due to the increasing 

 charge on the enzymes. 



