INHIBITION OF ENZYMES 35 



Adenosinetriphosphatase and Related Enzymes 



The behavior of ATPase with respect to iodoacetate becomes important 

 in considering the effects of iodoacetate on muscle. ATPase is classed as an 

 SH enzyme due to its inhibition primarily by mercurials. However, it ap- 

 pears to be one of the enzymes whose SH groups are relatively unreactive 

 to iodoacetate. Needham (1942) reported that myosin ATPase is not inhib- 

 ited by 3 mM iodoacetate even after 2 hr incubation, although all of the 

 SH groups giving the nitroprusside test are reacted, and Singer and Barron 

 (1944) found no inhibition on reaction of myosin ATPase with 3 mM iodo- 

 acetamide. Engelhardt and Lyubimova (1942) observed a 30% inhibition 

 with 10 mM iodoacetate, and Engelhardt (1946) remarked on the strange 

 unreactivity of myosin SH groups with iodoacetate. The results of Polls 

 and Meyerhof (1947) were very erratic, but they found in general some 

 stimulation at low concentrations (below 1 mM) of iodoacetate and iodo- 

 acetamide, and small inhibitions at higher concentrations, the response ap- 

 parently depending on the state of the myosin. Bailey and Perry (1947) 

 observed a very slow reaction of myosin with iodoacetamide, some 15 hr 

 being required for completion at 16° and concentrations between 1.7 and 

 42 mM. A similar sluggishness was noted by Barany and Barany (1959 a), 

 iodoacetate at 100 mM and pH 8 reducing the SH groups by one third in 

 2 hr, and iodoacetamide reacting about twice as rapidly, the slower reac- 

 tion with iodoacetate being attributed to the negative charge on the en- 

 zyme. In contrast to the early work of Needham, they found that myosin 

 loses most of its ATPase activity before half of its SH groups are reacted. 

 There are at least two different types of SH group in myosin. Of the 15 SH 

 groups (per molecular weight unit of 200,000), only 14 can exchange with 

 the disulfide reagent, bis-(/?-carboxyethyl)disulfide; if the myosin treated 

 with this substance is then exposed to iodoacetamide, the activity is not 

 restored by /5-mercaptoethanol, as it is in the absence of iodoacetamide 

 (Stracher, 1963). Thus there seems to be one SH group at the active site, 

 and this was confirmed by incubating myosin with labeled iodoacetamide 

 and isolating 5-carboxymethylcysteine. It is difficult to generalize on the 

 behavior of myosin ATPase in the presence of these inhibitors, but it seems 

 safe to say that there will be little if any inhibition at the concentrations 

 used to produce the characteristic changes in muscle contractility, unless 

 the susceptibility of myosin ATPase is greater in vivo. 



The results with the ATPases from other sources are equally variable, 

 due perhaps to the different types of ATPase activity measured. In general, 

 rather small inhibitions have been reported: 29% by 2 mM iodoacetate for 

 the guinea pig brain enzyme (Gore, 1951), 40% by 10 mM iodoacetate for 

 insect muscle enzyme (Gilmour, 1955), 30% by 10 mM iodoacetate for 

 human erythrocyte enzyme (Herbert, 1956), and 17% by 100 mM iodo- 

 acetate for the potato apyrase (Barany and Barany, 1959 b). Mitochondrial 



