DETERMINATION OF PHOSPHORYLATION SITES IN PEPTIDES AND PROTEINS EMPLOYING A VOLATILE EDMAN REAGENT

A manual Edman degradation protocol has been developed that allows the identification of phosphorylation sites in P-32-labeled peptides at t he subpicomole level. By using both a volatile reagent, trifluoroethyl isothiocyanate, and volatile buffers, extraction steps are rendered u nnecessary and cycle times can be reduced to 45 min. The protocol was employed to identify the site of phosphorylation in phosphoserine- and phosphotyrosine-containing peptides.