Effects of the position of internal histidine residues on the collision-induced fragmentation of triply protonated tryptic peptides.
The collision-induced dissociation spectra of a series of synthetic, tryptic peptides that differed by the position of an internal histidine residue were studied. Electrospray ionization of these peptides produced both doubly and triply protonated molecular ions. Collision-induced fragmentation of the triply protonated peptide ions had better efficiency than that of the doubly protonated ions, producing a higher abundance of product ions at lower collision energies. The product ion spectra of these triply protonated ions were dominated by a series of doubly charged y-ions and the amount of sequence information was dependent on the position of the histidine residue. In the peptides where the histidine was located towards the C-terminus of the peptide, a more extensive series of sequence specific product ions was observed. As the position of the histidine residue was moved towards the N-terminus of the peptide, systematically less sequence information was observed. The peptides were subsequently modified with diethylpyrocarbonate to manipulate the product ion spectra. Addition of the ethoxyformyl group to the N-terminus and histidine residue shifted the predominant charge state of the modified peptide to the doubly protonated form. These peptide ions fragmented efficiently, producing product ion spectra that contained more sequence information than could be obtained from the corresponding unmodified peptide.
Willard
B B
BB
Lerner Research Institute Mass Spectrometry Laboratory for Protein Sequencing, Department of Cell Biology, Cleveland Clinic Foundation, OH 44195, USA.
Kinter
M
M
D000595:Amino Acid Sequence; D008401:Gas Chromatography-Mass Spectrometry; D006639:Histidine / Q000737:chemistry*; D006868:Hydrolysis; D007202:Indicators and Reagents; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry*; D021241:Spectrometry, Mass, Electrospray Ionization; D014357:Trypsin / Q000737:chemistry*
D007202:Indicators and Reagents; D010455:Peptides; D006639:Histidine; D014357:Trypsin