Denitration of peroxynitrite-treated proteins by 'protein nitratases' from rat brain and heart

Abstract

Putative 'protein nitratases,' which catalyze denitration of peroxynitrite (PN)-treated proteins, were detected in the homogenate/crude extract of rat brains and hearts. Nitratase activity was monitored by the decreased intensity of nitrotyrosine immunoreactive-bands in Western blot and increased nitrate level in dialysate of incubation mixture, which contained homogenate/crude extract, protease inhibitors and a PN-treated substrate, such as treated histone (III-S), BSA or invertase. Enhanced activity of nitratases was noted by preincubating crude extract with Ca2+. In addition, at least two types of nitratases may occur: type I, reductant-dependent, and type II, reductant- independent. Furthermore, upon denitration, the activity of PN-treated invertase increased to the same activity level of the untreated invertase. The overall reaction catalyzed by nitratases for denitration of nitrotyrosine residues in protein could be as follows: Protein-Tyr-NO2 + H2O --> Protein-Tyr-H + H+ + NO3-. The nitration/denitration of protein-tyrosine may be crucial in regulating signal transduction.