Gas-phase cleavage of PTC-derivatized electrosprayed tryptic peptides in an FT-ICR trapped-ion cell: Mass-based protein identification without liquidchromatographic separation

Condensed phase protein sequencing typically relies on N-terminal labeling with phenylisothiocyanate ("Edman" reagent), followed by cleavage of the N- terminal amino acid. Similar Edman degradation has been observed in the gas phase by collision-activated dissociation of the N-terminal phenyl thiocar bamoyl protonated peptide [1] to yield complementary b(1) and y(n-1) fragme nts, identifying the N-terminal amino acid. By use of infrared multiphoton (rather than collisional) activation, and Fourier transform ion cyclotron r esonance (rather than quadrupole) mass analysis, we extend the method to di rect analysis of a mixture of tryptic peptides. We validate the approach wi th bradykinin as a test peptide, and gp on to analyze a mixture of 25 pepti des produced by tryptic digestion of apomyoglobin. A b(1)(+) ion is observe d for three of the Edman-derivatized peptides, thereby identifying their N- terminal amino-acids. Search of the SWISS-PROT database gave a single hit ( myoglobin, from the correct biological species), based on accurate-mass FT- ICR MS for as few as one Edman-derivatized tryptic peptide. The method is r obust-it succeeds even with partial tryptic digestion, partial Edman deriva tization, and partial MS/MS IRMPD cleavage. Improved efficiency and automat ion should be straightforward. (C) 2001 American Society for Mass Spectrome try.