Abstract

Soluble guanylyl cyclase (sGC) is the major physiological target of sydnonimine-based vasodilators such as molsidomine. Decomposition of sydnonimines results in the stoichiometric formation of nitric oxide (NO) and superoxide (O₂⨪), which rapidly react to form peroxynitrite. Inasmuch as sGC is activated by NO but not by peroxynitrite, we investigated the mechanisms underlying sGC activation by 3-morpholinosydnonimine (SIN-1). Stimulation of purified bovine lung sGC by SIN-1 was found to be strongly dependent on glutathione (GSH). By contrast, GSH did not affect sGC activation by NO released from 2,2-diethyl-1-nitroso-oxyhydrazine, indicating that NO/O₂⨪ released from SIN-1 converted GSH to an activator of sGC. High performance liquid chromatography identified this product as the thionitrite S-nitrosoglutathione. Further, the reaction product decomposed to release NO upon addition of Cu(NO₃)₂ in the presence of GSH. Activation of sGC was antagonized by the Cu(I)-specific chelator neocuproine, whereas the Cu(II)-selective drug cuprizone was less potent. Carbon dioxide (delivered as NaHCO₃) antagonized S-nitrosation by peroxynitrite but not by SIN-1. Thus, NO/O₂⨪ released from SIN-1 mediates a CO₂-insensitive conversion of GSH to S-nitrosoglutathione, a thionitrite that activates sGC via trace metal-catalyzed release of NO. These results may provide novel insights into the molecular mechanism underlying the nitrovasodilator action of SIN-1.