Inhibition of phosphatidylinositol synthase and other membrane-associated enzymes by stereoisomers of hexachlorocyclohexane.

Abstract

Hexachlorocyclohexanes (HCCH) are chlorinated analogs of inositol; the alpha, beta, gamma, and delta isomers of HCCH have the stereochemical configurations of (+/-)-, scyllo-, muco-, and myo-inositol, respectively. To assess their potential as specific tools for the study of agonist-stimulated phosphoinositide metabolism, we examined the effects of these four HCCH isomers on phosphatidylinositol (PI) synthase (CDP-1,2-diacyl-sn-glycerol:myo-inositol 3-phosphatidyltransferase), PI:inositol exchange enzyme, and several membrane-associated enzymes unrelated to inositol metabolism. In pancreas microsomes, in the presence of saturating myo-inositol, the alpha, beta, gamma, and delta isomers (4 mM) inhibited PI synthase activity by 9, 4, 22, and 69%, respectively. Half-maximal inhibition by delta-HCCH occurred at 0.25 mM. A similar pattern of HCCH inhibition was obtained using n-octylglucopyranoside-solubilized and partially purified PI synthase preparations. The inhibition by delta-HCCH was noncompetitive versus myo-inositol. The PI:inositol exchange enzyme in mouse pancreas microsomes was inhibited 90% by 1 mM delta-HCCH in the presence of 0.25% Triton X-100, but not in its absence; half-maximal inhibition occurred with 0.5 mM delta-HCCH. delta-HCCH (4 mM) also inhibited to varying extents the following enzymes: pancreas CDP-choline:1,2-diacyl-sn-glycerol cholinephosphotransferase (75%), brain and erythrocyte (Na+,K+)-ATPase (87 and 70%), brain and erythrocyte Mg2+-ATPase (38 and -5%), brain 1,2-diacyl-sn-glycerol kinase (22%), and liver glucose 6-phosphatase (16%). gamma-HCCH (4 mM) inhibited these enzymes to a lesser extent, or not at all. The order of inhibition by HCCH stereoisomers was the same as the order of their saturation level in phospholipid vesicles (delta greater than gamma greater than alpha greater than beta). This suggests that the inhibitory action is due to insertion of the compounds either into hydrophobic domains of the enzymes or into annular lipid. The results indicate that the HCCHs are not selective inhibitors of inositol metabolism.