We report here a method for the identification of free N-terminal peptides of in gel digested isolated proteins. It is based on the difference between the isotopic ion distribution of the N-terminal peptide and internal peptides. After guanidination of lysine residues, the primary amino groups of the gel-entrapped protein are blocked with an equimolar mixture of normal and deuterated acetic anhydride. Upon MS analysis, internal peptides display a normal isotopic ion distribution while the N-terminal peptide shows a complex isotopic ion distribution.
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