SIMPLIFICATION OF HIGH-ENERGY COLLISION SPECTRA OF PEPTIDES BY AMINO-TERMINAL DERIVATIZATION

Four-sector tandem mass spectrometry proves extremely useful for provi ding sequence information for peptides. The complexity of ion fragment ations, however, makes data interpretation difficult and time consumin g. Attachment of a fixed positive charge to the peptide amino terminus forces production of only N-terminal fragment ions to yield simplifie d, predictable fragmentation. Reaction of a peptide at pH 6 with iodoa cetic anhydride selectively modifies the N-terminus by exploiting the pK(a) differences between the alpha-amino group and any lysine side-ch ain epsilon-amino groups. The iodoacetyl peptide can react with many r eagents to form a fixed positive charge. We find reaction with dimethy loctylamine forms a quaternary ammonium derivative with good surface a ctivity properties and concomitant increased sensitivity. The high-ene rgy CAD fragment ion spectra of the N-terminally derivatized peptides show predominantly a(n) and d(n) ions. The abundant d(n) ions permit r eady distinction of leucine and isoleucine. Fewer fragment ions make d ata interpretation simpler and lead to more intense peaks since the io n intensity is spread among fewer peaks. The method is particularly us eful for peptides which do not otherwise yield sufficient fragmentatio n to provide either the complete sequence or the locations of modified amino acids.