New method of peptide sequencing by modification with Fluorescent reagent using MALDI-TOF-MS PSD method

抄録

There are many peptides and proteins whose amino acid sequence cannot be analyzed, because full fragmentation ions are not detected by PSD method in MALDI- TOF- MS.In our proteome researches, antigen peptides produced by digestion of nucleic protein of influenza virus were in the case. Sequences of the amino acid residues in peptides modified in thiol group of N-terminal cysteine by fluorescene were completely analyzed with sufficient fragment ions by PSD method. Similar analyses of antigen peptides with cysteine residue at C-terminal or non-terminal positions were performed. Sufficient numbers of fragment ions for the peptide sequencings were detected in MS spectra and the sequence analyses were perfectly made.Furthermore, modifications were also made for N-terminal position of other general peptides without cysteine residue. Similar modification reaction gave satisfactory results of MS analyses of the peptide sequencings.Since there is the possibility that amino group at the side chain of amino acid residue is also modified by the fluorescene along with the N-terminal position, we examined reactivity of the fluorescent reagent against an amino group at side chain of arginine, asparagine, glutamine, or lysine residue in peptides to conform the useful common application of the method to all peptides. The obtained MS spectra clearly showed that all modified peptides gave completely full fragment ions for the sequence analyses. The modification by fluorescent reagent to general peptides is a complete and reliable technique for the sequence analysis of peptides by using MALDI-TOF-MS PSD method in proteome research.