A METHOD AND KIT FOR DETECTING S-NITROSOTHIOL REDUCTION OF S-NITROSYLATED PROTEINS

摘要

We disclose a method and kit for selectively labeling S-nitrosylated proteins. After blocking free thiol groups with an alkylthiolating agent (e.g, methyl methanethiosulfonate), we reduce S-nitrosylated cysteines with a reducing agent (e.g., ascorbate), thereby producin free thiol groups from the S-nitrosylated cysteines. Thereafter, the free thiol groups can be labeled and detected with maleimide- derivatized fluorescent dyes (e g., maleimide conjugated cyanine dyes). Our method finds use in detecting, quantifying, in situ visualizing, and a means for site-specific identification of S-nitrosylation events.