Disulfide bond mapping by cyanylation-induced cleavage and mass spectrometry.

Oxidation of sulfhydryl groups to form a disulfi de bond is one of the most common post-translational modifi cations in proteins. Disulfi de bonds play important roles in stabilizing three-dimensional structure and modulating bioactivity of the cystinyl proteins. The determination of disulfi de bond linkage is therefore an integral part of structural characterization of proteins. A mass spectrometry-based strategy utilizing chemical cleavage at cysteine residues following cyanylation reaction is described for the identifi cation of both sulfhydryl and disulfi de bond linkage in proteins. The method has been particularly powerful for the assignment of disulfi de bonds in proteins containing adjacent or closely spaced cysteines.