Nitric oxide induced S-glutathionylation and inactivation of glyceraldehyde-3-phosphate dehydrogenase

S-Nitrosylation of protein thiol groups by nitric oxide (NO) is a widely re cognized protein modification. In this study we show that nitrosonium tetra fluoroborate (BF4NO), a NO+ donor, modified the thiol groups of glyceraldeh yde-3-phosphate dehydrogenase (GAPDH) by S-nitrosylation and caused enzyme inhibition. The resultant protein-S-nitrosothiol was found to be unstable a nd to decompose spontaneously, thereby restoring enzyme activity. In contra st, the NO-releasing compound S-nitrosoglutathione (GSNO) promoted S-glutat hionylation of a thiol group of GAPDH both in vitro and under cellular cond itions. The GSH-mixed protein disulfide formed led to a permanent enzyme in hibition, but upon dithiothreitol addition a functional active GAPDH was re covered. This S-glutathionylation is specific for GSNO because GSH itself w as unable to produce protein-mixed disulfides. During cellular nitrosative stress, the production of intracellular GSNO might channel signaling respon ses to form protein-mixed disulfide that can regulate intracellular functio n.