704 6. O-IODOSOBENZOATE 



nature of the clots from soft and opaque to firm, elastic, and almost trans- 

 parent, and simultaneously the water binding increases from 14.3 to 36.5 

 g/g. It is quite possible that more linear proteins, which may be reasonably 

 flexible, can be altered quite markedly by the formation of disulfide bonds, 

 and that particular regions on the surface may be made unavailable for 

 other reactions. 



Further evidence for structural changes induced by o-iodosobenzoate is 

 the decrease in the viscosity of G-actin brought about by 2 mJf of the 

 reagent acting for 30 min at 25^ and pH 7.8-8 (Barany et al., 1962). This 

 is interpreted as an inhibition of polymerization. Simultaneously there is a 

 decrease in the ability to bind Ca++, as shown by the loss of G-actin-bound 

 Ca^^ upon tretment with o-iodosobenzoate. Although there appears to be 

 some correlation between changes in viscosity and Ca++ binding as pro- 

 duced by various SH reagents, the mechanisms involved are not yet under- 

 stood. 



INHIBITION OF ENZYMES 



Most SH enzymes are inhibited by o-iodosobenzoate (see Table 6.1) and 

 in a few instances the inhibition is very marked. Enzymes without SH 

 groups at or near their active centers, as shown by failure to be inhibited 

 by SH reagents in general, are not affected by o-iodosobenzoate, except 

 possibly in the single instance where it has been claimed to act as a com- 

 petitive inhibitor on D-amino acid oxidase, although in such cases the con- 

 centration would usually have to be a good deal higher than for the oxi- 

 dation of susceptible SH groups (Frisell et al., 1949) (see page 342). Inas- 

 much as o-iodosobenzoate is used up in the reaction, inhibition is of a titra- 

 tion type and spontaneously irreversible; thus the degree of inhibition will 

 often depend on the amount of enzyme present, or the amount of some SH 

 containing impurity that also reacts with the reagent. In most complex 

 systems, many enzymes will be inactivated to varying degrees, and probably 

 little specificity is possible. However, the remarkable sensitivity of creatine 

 kinase — definite inhibition at 0.00001 mM and complete inhibition at 

 0.00013 mM — might well make it possible to block this enzyme selectively 

 (Ennor and Rosenberg, 1954). This inhibition may explain the observation 

 of Bailey and Marsh (1952) that the fall in creatine phosphate in muscle 

 homogenates is almost completely prevented by o-iodosobenzoate. It might 

 be worthwhile to consider the use of o-iodosobenzoate in glycerinated and 

 similar muscle preparations to determine the role of creatine kinase in the 

 initiation of contraction or relaxation. Phosphoglyceraldehyde dehydro- 

 genase seems to be less sensitive to o-iodosobenzoate than to iodoacetate or 

 iodoacetamide, so that a specific block of glycolysis at this step would be 

 impossible. It may be noted that several dehydrogenases are quite well 

 inhibited by o-iodosobenzoate, notably the xanthine, malate, and aldehyde 



