Endothelium-derived relaxing factor (EDRF) is an important modulator of vascular function, causing both vasodilation and platelet inhibition by stimulation of quanylyl cyclase. The chemical nature of EDRF has been studied using a variety of pharmacologic and analytic techniques that are largely indirect; most investigators in the field currently believe that the preponderance of evidence supports the view that this bioactive molecule is chemically equivalent to nitric oxide (No). Nitric oxide is a highly reactive, relatively short-lived free radical species that is synthesized by the ubiquitous family of nitric oxide synthase enzymes from L-arginine. Inasmuch as NO must be transferred from its enzymatic source of synthesis to its principal effector site, guanylyl cyclase, by way of a cellular environment rich in potential coreactants or enzymes that either facilitate its stabilization, promote it biologic activity, or hasten its metabolic fate, many investigators are of the opinion that NO is stabilized through the formation of an adduct with another molecular species in the cellular milieu. Among the most reactive and available species in the cellular environment that can produce adducts with NO having biologic effects similar to EDRF and NO are biomolecules containing thiol groups. Thiols react spontaneously (under physiologic conditions) with NO (directly or by way of N2O3 or NO+) to form S-nitroso-thiols (thionitrites, or RSNOs), as we and others have shown. In this proposal, we plan to study in detail t he effects of RSNOs formed from both low-molecular-weight thiols and, most interestingly, protein thiols on vascular smooth muscle and platelet function; establish with newly developed chemical techniques the identify of endogenous low-molecular-weight RSNOs that form, and, in particular, the specific intracellular and extracellular S-nitroso-proteins that form by a novel mechanism of posttranslational modification, S- nitros(yl)ation; and dissect the biochemical and cellular mechanism(s) by which RSNO adducts evolve, are transported, manifest biologic activity, and are metabolized. The overall purpose of these experiments is, therefore, to elucidate the role of this class of biologically active compounds in the vasculature.
| Scharfstein, J S; Keaney Jr, J F; Slivka, A et al. (1994) In vivo transfer of nitric oxide between a plasma protein-bound reservoir and low molecular weight thiols. J Clin Invest 94:1432-9 |
