The Biotin Switch Method for the Detection of S-Nitrosylated Proteins

Science's STKE  12 Jun 2001:
Vol. 2001, Issue 86, pp. pl1
DOI: 10.1126/stke.2001.86.pl1

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Many of the effects of nitric oxide are mediated by the direct modification of cysteine residues resulting in an adduct called a nitrosothiol. Here, we describe a novel method for detecting proteins that contain nitrosothiols. In this three-step procedure, nitrosylated cysteines are converted to biotinylated cysteines. Biotinylated proteins can then be detected by immunoblotting or can be purified by avidin-affinity chromatography. We include examples of the detection of S-nitrosylated proteins in brain lysates after in vitro S-nitrosylation, as well as the detection of endogenous S-nitrosothiols in selected neuronal proteins.

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