IDENTIFICATION OF PHOSPHOCYSTEINE BY ELECTROSPRAY MASS-SPECTROMETRY COMBINED WITH EDMAN DEGRADATION

Phosphocysteine is an intermediate in the phasphoenolpyruvate-dependen t phosphotransferase system (PTS) and in the dephosphorylation of phos photyrosine residues by protein tyrosine phosphatases. A method is des cribed for the direct and non-radioactive localization of phosphocyste ine. The phosphorylated Glu-C peptide of the EII(Mtl) protein of the P TS of Staphylococcus carnosus was identified by LC-electrospray MS and isolated for further analysis. Following chemical modification with a lkaline ethanethiol, S-ethylcysteine was identified during Edman degra dation, demonstrating that phosphocysteine reacts like phosphoserine. As a control, the unphosphorylated cysteine could not be modified by t he alkaline ethanethiol.