S-NITROSATION OF SERUM-ALBUMIN BY DINITROSYL-IRON COMPLEX

The objective of this study was to identify a potential mechanism for S-nitrosation of proteins. Therefore, we assessed S-nitrosation of bov ine serum albumin by dinitrosyl-iron-di-L-cysteine complex [(NO)(2)Fe( L-cysteine)(2)], a compound similar to naturally occurring iron-nitros yls. Within 5-10 min, (NO)(2)Fe(L-cysteine)(2) generated paramagnetic albumin-bound dinitrosyl-iron complex and S-nitrosoalbumin in a ratio of 4:1. Although S-nitroso-L-cysteine was concomitantly formed in low amounts, its concentration was not sufficient to account for formation of S-nitrosoalbumin via a trans-S-nitrosation reaction. Low oxygen te nsion did not affect S-nitrosation by the dinitrosyl-iron complex thus excluding the involvement of oxygenated NOx-species in the nitrosatio n reaction. Blockade of albumin histidine residues by pyrocarbonate, w hich prevented formation of dinitrosyl-iron-albumin complex, did not i nhibit S-nitrosation of albumin. Thus, S-nitrosation of albumin by (NO )(2)Fe(L-cysteine), can proceed by direct attack of a nitrosyl moiety on the protein thiolate, without previous binding of the iron. We conc lude that protein-bound dinitrosyl-iron complexes detected in high con centrations in certain tissues provide a reservoir of S-nitrosating sp ecies, e.g. low molecular dinitrosyl iron complexes.