期刊
ARCHIVES OF BIOCHEMISTRY AND BIOPHYSICS
卷 435, 期 1, 页码 42-49出版社
ELSEVIER SCIENCE INC
DOI: 10.1016/j.abb.2004.12.016
关键词
glutathione; S-glutathionylation; glutathione S-transferase; nitrosoalutathione; reactive oxygen species; reactive nitrogen oxide species
Oxidative and nitrosative stress lead to the S-glutathionylation of proteins and subsequent functional impairment. Glutathione S-transferase (GST) from Schistosoma japonicum was found to bind to the glutathione moiety of S-glutathionylated proteins, thus establishing a convenient method for detecting S-glutathionylated proteins by biotinylated GST. Applications of this method to proteins that were prepared from Cultured cells and blotted onto a membrane exhibited numerous positive bands, which were abolished by treatment with dithiothreitol. Treatment of a cellular extract with nitrosoglutathione led to enhanced staining of the bands in a dose-dependent manner. The method was also applicable for the histochemical detection of S-glutathionylated proteins in situ. The positive staining by biotin-GST became faint in the presence of S-glutathionylated ovalbumin, Suggesting that the reaction is specific to S-glutathionylated proteins. Collectively, these data indicate that the method established here is simple and useful for detecting S-glutathionylated proteins on blotted membrane and in Situ. (C) 2005 Elsevier Inc. All rights reserved.
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