Glutathiolation of proteins by glutathione disulfide S-oxide derived from S-nitrosoglutathione - Modifications of rat brain neurogranin/RC3 and neuromodulin/GAP-43

S-Nitrosoglutathione (GSNO) undergoes spontaneous degradation that generate s several nitrogen-containing compounds and oxidized glutathione derivative s. We identified glutathione sulfonic acid, glutathione disulfide S-oxide ( GS(O)SG), glutathione disulfide S-dioxide, and GSSG as the major decomposit ion products of GSNO. Each of these compounds and GSNO were tested for thei r efficacies to modify rat brain neurogranin/RC3 (Ng) and neuromodulin/GAP- 43 (Nm), Among them, GS(O)SG was found to be the most potent in causing glu tathiolation of both proteins; four glutathiones were incorporated into the four Cys residues of Ng, and two were incorporated into the two Cys residu es of Nm. Ng and Nm are two in vivo substrates of protein kinase C; their p hosphorylations by protein kinase C attenuate the binding affinities of bot h proteins for calmodulin. When compared with their respective unmodified f orms, the glutathiolated Ng was a poorer substrate and glutathiolated Nm a better substrate for protein kinase C. Glutathiolation of these two protein s caused no change in their binding affinities for calmodulin. Treatment of [S-35]cysteine-labeled rat brain slices with xanthine/xanthine oxidase or a combination of xanthine/xanthine oxidase with sodium nitroprusside result ed in an increase in cellular level of GS(O)SG. These treatments, as well a s those by other oxidants, all resulted in an increase in thiolation of pro teins; among them, thiolation of Ng was positively identified by immunoprec ipitation. These results show that GS(O)SG is one of the most potent glutat hiolating agents generated upon oxidative stress.