36 1. lODOACETATE AND lODOACETAMIDE 



ATPase is either not affected (Pullman et al, 1960) or stimulated (Chari- 

 Bitron and Avi-Dor, 1959; Fanestil et al., 1963). Such effects may not be 

 directly on the ATPase, since any agent that prevents the mitochondrial 

 swelling produced by iodoacetate counteracts the stimulation. Again we 

 conclude that at concentrations blocking glycolysis little effect on the ATP- 

 ases should be observed, and that modifications in tissue function brought 

 about by iodoacetate probably do not relate to inhibition of this enzyme. 

 Certain ATPases believed to be involved in cellular activities have been 

 examined recently and none is very sensitive: the ATPase in Mytilus sper- 

 matozoa tails is inhibited 13% by 0.1 mM iodoacetate (Nelson, 1959), that 

 in the eel electric organ is depressed only 3% by 1 mM iodoacetamide 

 (Glynn, 1963), that in the cortex of sea urchin eggs, which is K+-Na+-acti- 

 vated and believed to participate in active transport, is inhibited 23% by 

 1 mM iodoacetate (Ohnishi, 1963), and the ATPase in rabbit kidney which 

 may play a role in Na+ resorption is not affected by 1 mM iodoacetate or 

 10 mM iodoacetamide (Taylor, 1963). These enzymes will be affected if 

 high concentrations of the inhibitors are used, but all the evidence indicates 

 their inhibition not to be significant in the range used to depress glycolysis. 

 The exchange reactions ADP-ATP or P,-ATP catalyzed by mitochondrial 

 or chloroplast systems are inhibited by iodoacetate only at quite high con- 

 centrations (around 10 mM) (Plant, 1957; Kahn and Jagendorf, 1961; Chiga 

 and Plant, 1959). The AMP- ATP transphosphorylase (myokinase) of rabbit 

 muscle is not inhibited by 1 mM iodoacetate without preincubation (Noda, 

 1958), although 2 SH groups are free for reaction with certain other SH 

 reagents. A thorough kinetic study of this inhibition has been made by 

 Mahowald et al. (1962), who found the rate of reaction and degree of inhi- 

 bition to be low unless fairly high ratios of iodoacetate to enzyme were 

 used. The reactions were run at pH 8 and 30^ and yet at a ratio of (I)/(E) = 

 0.48 (iodoacetate 0.27 mM) it requires 3 hr for maximal reaction, at which 

 time the inhibition is only 25%, and even at a ratio of (I)/(E) = 2.88 

 (iodoacetate 1.61 mM) it requires several hours for the reaction to be com- 

 pleted, although here the inhibition is complete. The inactivated enzyme 

 was found to contain 1.83 *S-carboxymethylcysteine residues with 0.2 half- 

 cystine residue remaining. Thus 2 SH groups per enzyme molecule react with 

 iodoacetate but the rate is so slow that the enzyme is unlikely to be signi- 

 ficantly inhibited in vivo. 



Creatine Kinase (ATP:Creatine Phosphotransferase) 



Although the phosphate-splitting and -transferring enzymes in general 

 are not readily inhibited by iodoacetate, one enzyme quite important in 

 determining the maintenance of tissue function, namely, creatine kinase, 

 is very sensitive to iodoacetate, at least in the isolated state (Table 1-7). 

 We shall discuss later the possible inhibition of this enzyme in muscles 



