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Abstract

Inhibition of a major hepatic form of human cationic glutathione S-transferase by bilirubin, biliverdin, indocyanine green and chenodeoxycholic acid was investigated as a function of pH (range = 6.5 to 9.1). Changes in pH had little effect on the extent of inhibition by indocyanine green. However, inhibition by bilirubin, biliverdin and chenodeoxycholic acid was found to be pH-dependent, with markedly less inhibition at the high values of pH. The reduced inhibition at the high values of pH could not be ascribed to a failure of the enzyme to bind the nonsubstrate ligand. Instead, the complete inhibition observed at pH 6.5 became partial (hyperbolic) inhibition at pH 9.1. This behavior can be ascribed to the binding of the nonsubstrate ligands at a site other than the active site, i.e., at high values of pH there is formation of an enzyme-substrate-inhibitor complex which still retains considerable catalytic activity. At physiologic values of pH (7.0), the human transferase was completely inhibited by saturating concentrations of the tested nonsubstrate ligands. This is in contrast to our previous studies performed with the rat transferases where, although inhibition also was affected by buffer pH, some forms of the enzyme retained significant catalytic activity at pH 7.0 despite high concentrations of nonsubstrate ligands. We conclude that the ability of the human cationic glutathione S-transferases to serve as enzymes of detoxification in the presence of high intracellular concentrations of nonsubstrate ligands may be significantly reduced, and this may render the cholestatic liver unusually susceptible to injury by toxic electrophiles.