CHAPTER 5 



OXIDANTS 



Many reagents have been used to oxidize protein and enzyme SH groups 

 for the purpose of either estimating these groups or determining the relation- 

 ship of the groups to the enzyme activity. Most of these oxidants at present 

 are of little importance in the study of enzymes or metabolism, mainly be- 

 cause of their lack of specificity for SH groups. General over-all oxidation 

 of an enzyme, involving several types of group and ending in partial or com- 

 plete denaturation, provides no useful information. If oxidants are to be 

 used for the specific modification of SH groups it is necessary that the choice 

 of oxidant and the experimental conditions be made very carefully. The 

 oxidizing activity of the reagent must be neither too high nor too low (i.e., 

 its oxidation-reduction potential must be in the proper range relative to the 

 SH groups under the selected conditions) and the ability of the substance 

 to react in other ways with the enzyme must be minimal. Of the factors 

 determining the rates of oxidation of SH groups and the specificity of an 

 oxidant, the pH and the temperature are the most important. Some of the 

 oxidants that have been abandoned in enzyme work might well be applicable 

 in certain studies if the optimal conditions for their use were known. 



The formation of enzyme disulfide groups during oxidation requires SH 

 groups that are close enough to link together in S — S bonds, or are so located 

 as to be able to approach each other readily. The SH groups may be on the 

 same enzyme molecule or on different molecules: 



SH S 



Intramolecular oxidation: R, ^ Xqx > '- R 



SH 



\ 



^red 



Intermolecular oxidation: 



2 R— SH -r Xox •* * R— S-S — R ^ Xred 



The hydrogen atoms may be transferred directly to the oxidant, or may 

 form H+ ions, the oxidant accepting only electrons. A lone SH group on an 

 enzyme, although fully exposed, may not be oxidized if sterically it cannot 

 associate with another SH group. The formation of enzyme aggregates, or 



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