INHIBITION OF ENZYMES 869 



The interesting treatment of the kinetics of myosin ATPase inhibition 

 with p-MB recently reported by Walter (1963) should be consulted for the 

 theoretical development of the equations and appropriate plotting proce- 

 dures, but in connection with our present subject it is worth noting that 

 the kinetics are complicated by two factors which should be kept in mind 

 in such work on all enzymes. First, the reaction of the first SH groups 

 does not lead to inactivation immediately. Second, this initial reaction re- 

 duces the mercurial concentration so that the more slowly reacting groups 

 are exposed to a much lower concentration than was originally added. The 

 calculated bimolecular rate constant for these catalytically important SH 

 groups is thus quite different than that obtained from the initial rate. 



The relationship between the effects of mercurials and 2,4-dinitrophenol 

 is very interesting and the results indicate that SH groups are involved in 

 the stimulation of ATPase activity by the latter substance. Lardy and 

 Wellman (1953) noted that 0.04 mM p-MB almost completely abolishes 

 the activation of mitochondrial ATP splitting by DNP, and others not 

 only have confirmed this in general but have shown that the DNP-activated 

 enzyme is only inhibited by mercurials (Greville and Needham, 1955; Gil- 

 mour and Griffiths, 1957; Pullman et al., 1960). This is shown for myosin 

 ATPase in Fig. 7-30. However, many factors can influence these interac- 

 tions. Myers and Slater (1957 b) found that p-MB inhibits the DNP-acti- 

 vated mitochondrial ATPase from pH 6 to 8, but stimulates further at pH 9, 

 and Cooper (1958 b) has emphasized the importance of Mg++, addition of 

 this ion decreasing the inhibition by p-MB of DNP-activated enzyme, as 

 originally shown by Lardy and Wellman. Not all ATPases may behave in 

 this fashion, and the activity of a particulate preparation from Rhodospi- 

 rillum rubrum can still be stimulated by DNP in the presence of p-MB and 

 Mg++ (Cooper, 1958 a). Some investigators have postulated that DNP and 

 the mercurials activate ATPase by similar mechanisms, but it seems doubt- 

 ful if the evidence is sufficient to draw this conclusion. It has recently been 

 found that COg stimulates mitochondrial ATPase markedly and this occurs 

 in the presence of 0.005 mM Hg ++,which itself has broght about activation; 

 indeed, Hg++ activates about the same in the absence or presence of COg 

 (Fanestil et al., 1963). These stimulations thus appear to be approximately 

 additive. The K+-Na+-activated membrane ATPase contains SH groups 

 which seem to be specially involved in this activation and react readily 

 with mercurials (Skou, 1963). 



The interesting effects of the mercurials on the P,-ATP and ADP-ATP 

 exchange reactions occurring in mitochondria and the relations to ATPase 

 will be discussed later under oxidative phosphorylation (page 872). 



