Noncompetitive, Ca2+-independent inhibition of pyruvate dehydrogenase phosphatase by fluphenazine

The effects of two different classes of calmodulin antagonists on the catal ytic activities of purified pyruvate dehydrogenase (PDH) phosphatase and PD H complex (PDC) were studied. In general, PDH phosphatase was more strongly inhibited than PDC by the calmodulin antagonists with the following potenc y order: fluphenazine > chlorpromazine > thioridazine > triflupromazine. Pr omazine and two sulfonamides (W-5 and W-7) did not suppress PDH phosphatase activity at 1 mM concentrations, while about 20% of PDC activity was inhib ited by these antagonists. Fluphenazine-mediated inhibition of PDH phosphat ase was observed with the purified PDC as well as intact mitochondria, Alth ough Ca2+ stimulates PDH phosphatase activity, the addition of exogenous Ca 2+ did not overcome the inhibition by calmodulin antagonists. These results suggest that the suppression of PDH phosphatase activity is dependent upon the structure of the individual calmodulin antagonist and appears to be Ca 2+-independent, Kinetic analysis showed a noncompetitive inhibition of PDH phosphatase by fluphenazine, indicating that it binds to different site(s) from the catalytic site of the enzyme.