Effects of methyl mercury and cadmium on the kinetics of substrate activation of (K+)-paranitrophenyl phosphatase.

Previous studies from this laboratory have indicated that methyl mercuric chloride (CH3HgCl) and cadmium chloride (CdCl2) are potent inhibitors of K+-p-nitrophenyl phosphatase (K+-PNPPase). The present studies were undertaken to study the effects of CH3HgCl and CdCl2 on the substrate activation kinetics of K+-PNPPase to understand the mechanism of inhibition of Na+ pump by these heavy metals. Uncompetitive inhibition with regard to activation by PNPP was indicated by altered Vmax and Km values by both the heavy metals. Substrate activation kinetics of heavy metal inhibited K+-PNPPase in the presence of 25 microM dithiothreitol and glutathione indicated mixed type of activation by altering apparent Vmax and Km. Absence of competition between PNPP site and heavy metals appear to indicate absence of reactive-SH groups in the active site. Failure of added iodacetate, in concentrations ranging from 5 X 10(-8) to 5 X 10(-5) M, to inhibit K+-PNPPase further substantiate this conclusion. The results suggest that CH3HgCl and CdCl2 inhibit Na+ pump by inducing conformational changes in the enzyme and thereby decrease catalytic velocity of dephosphorylation of the enzyme-phosphoryl complex. Hydrolysis of PNPP was linear with time with or without either heavy metal and the inhibition exerted by CH3HgCl or CdCl2 on free or heavy metal loaded enzyme indicated absence of heavy metal interaction. The results suggest that CH3HgCl and CdCl2 inhibit K+-PNPPase possibly by binding at two different sites.