DISULFIDES 663 



normally contains disuL&de groups and can be inactivated by cysteine, GSH 

 and other thiols; reactivation occurs with cystine or GSSG (Cooperstein, 

 1963). This may also be the case with lens aminopeptidase, which is inhib- 

 ited rather strongly by cysteine and GSH but not at all by GSSG (Spec- 

 tor, 1963). Another reason for the lack of response to a disulfide is that 

 the environment of the enzyme SH group may be unfavorable for the ap- 

 proach of the disulfide or may affect the redox potential in such a manner 

 as to deter the interaction. If seralbumins are incubated with excess cystine, 

 the SH content as determined by p-mercuribenzoate titration drops to zero; 

 half the SH groups are lost from bovine hemoglobin (Isles and Jocelyn, 

 1963). GSSG has no effect on either type of protein. Incidentally, the reac- 

 tion of cystine with bovine seralbumin leads to the disappearance of 1 mol- 

 ecule of cystine for each pair of SH groups lost, so it is likely that the 

 cysteine formed is reoxidized and mixed disulfides are formed according to: 



2 Cy— S— S— Cy - 2 Prot— SH ±-. 2 Cy— SH - 2 Trot— S— S— Cy 

 2 Cy— SH ^ Cy— S— S— Cy i- 2 H+ + 2 e- 



Cy— S— S— Cy f 2 Prot— SH ^-. 2 Prot— S— S— Cy + 2 H^ -f 2 e- 



Some evidence for the formation of mixed disulfides during the inhibition 

 of enzymes has been reported. Beef liver catalase contains 8 SH groups ti- 

 tratable with p-chloromercuribenzoate. Reaction with cystine-S^^ results in 

 the oxidation of 4 of these groups to disulfides and the formation of 4 mixed 

 disulfides {YM et al, 1961): 



SH^ SH SH SH X— S— S S— S S — S— X 



\ I I / _^ \ I I / 



catalase^ -^ C— S — S— C > .: catalase 



/ I I \ / I I \ 



SH SH SH SH X— S— S S— S S— S— X 



This inhibition is spontaneously reversible; cystine at 0.037 mM, pH 7, and 

 370 inhibits maximally 17% around 5 min and then the inhibition decreases. 

 Cystamine monosulfoxide likewise appears to form mixed disulfides with 

 phosphoglyceraldehyde dehydrogenase, and this is reversible by thiols (Pihl 

 and Lange, 1962). Cystamine itself inhibits glucose utilization in erythro- 

 cytes and this block is believed to be at hexokinase (Eldjarn and Bremer, 

 1962). The inhibition is reversible by thiols, and mixed disulfide formation 

 was postulated; however, there was no direct evidence for this as opposed 

 to simple oxidation. Dithioglycolate and cystine were reported to form 

 mixed disulfides with myosin ATPase, but inhibition of the enzyme activity 

 does not occur until the last 2 or 3 SH groups are altered (Barany, 1959). 

 The optimal temperatures and pH's for reaction of enzyme SH groups 

 with disulfides are difficult to determine in our present state of knowledge. 

 Apparently the temperature coefficient is quite high; succinate dehydrogen- 



