Generation and detection of S-nitrosothiols.

摘要: Nitric oxide (NO) plays a pivotal role in cellular signaling many different organisms as the result of modification protein activities/functions by S-nitrosylation. This NO-dependent posttranslational is based on attachment NO to sulfur moiety cysteine residues. However, instability S-nitrosothiols makes it difficult analyze this type vitro well vivo. Jeffrey and colleagues developed method--named biotin switch method--that allows detection purification S-nitrosylated proteins. The principle behind technology substitution group linker three-step procedure. First, all free thiol groups are blocked with thiol-reactive agent, followed selective reduction residues using ascorbate. In final step, reduced labeled linker, so that previously finally biotinylated. Afterwards, biotinylated proteins can be detected anti-biotin antibodies or purified affinity chromatography neutravidin agarose. chapter, we give detailed description method, which used for proteomics approach identify candidates S-nitrosylation analyse selected