DIRECT AND REVERSIBLE INHIBITION OF ENDOTHELIAL NITRIC-OXIDE SYNTHASEBY NITRIC-OXIDE

The objective of this study was to investigate the regulation of endot helial nitric oxide (NO) synthase by NO. Partially purified endothelia l NO synthase was exposed to authentic NO (10-200 mu M) and to the nit rovasodilators sodium nitroprusside (SNP; 10-1,000 mu M) and S-nitroso -N-acetylpenicillamine (SNAP; 100-1,000 mu M), and enzyme activity was assayed by measuring the conversion of L-[H-3]arginine to L-[H-3]citr ulline in the presence of added cofactors. NO, SNP, and SNAP inhibited NO synthase activity in a dose-dependent manner, NO being the most po tent inhibitor. The Michaelis constant for L-arginine was not altered (4.87 mu M) by NO (50 mu M), whereas the maximal velocity of the enzym e decreased from 784 to 633 pmol . mg(-1). min(-1). Oxyhemoglobin (10 mu M) partially prevented the inhibition of NO synthase by NO (50 mu M ). The data also suggest that NO inhibits endothelial NO synthase acti vity by directly interacting with the NO synthase and not by an indire ct mechanism such as limitation of cofactor or oxygen availability. Di alysis of NO synthase treated with NO (50 mu M) partially restored the enzyme activity. This study demonstrates a direct and reversible inhi bition of NO synthase by NO, suggesting a feedback mechanism in vivo.