REGULATION OF RAT-LIVER S-ADENOSYLMETHIONINE SYNTHETASE DURING SEPTICSHOCK - ROLE OF NITRIC-OXIDE

We investigated the modulation of rat liver S-adenosylmethionine (SAM) synthetase in a model of acute sepsis, Our results show that animals treated with bacterial lipopolysaccharide experience a marked decrease in liver SAM synthetase activity. No changes were detected in the hep atic levels of SAM synthetase protein, suggesting that inactivation of the existing enzyme was the cause of the observed activity loss. Lipo polysaccharide treatment resulted in the expression of calcium-indepen dent/cytokine-inducible nitric oxide (NO) synthase in liver and the ac cumulation in plasma of the NO-derived species nitrite and nitrate. NO implication in the in vivo regulation of SAM synthetase was evaluated in animals treated with the NO donor molecule 3-morpholinosydnonimine . The analysis of liver enzymatic activity, along with protein and mes senger RNA levels yielded results similar to those obtained with lipop olysaccharide treatment. To assess directly the sensitivity of SAM syn thetase to NO, the rat liver-purified high- and low-molecular weight f orms of the enzyme were exposed to various doses of 3-morpholinosydnon imine and other NO donors such as S-nitroso-N-acetylpenicillamine, res ulting in a dose-dependent inhibition of enzymatic activity. This effe ct was reversed by addition of the reducing agents beta-mercaptoethano l and glutathione. Finally, cysteine 121 was identified as the site of molecular interaction between NO and rat liver SAM synthetase that is responsible for the inhibition of the enzyme. To reach this conclusio n, the 10 cysteine residues of the enzyme were changed to serine by si te-directed mutagenesis, and the effect of NO on the various recombina nt enzymes was measured.