sequencedeterminationofpeptides.docVIP

Sequence determination of peptides. Katalin F. Medzihradszky Department of Pharmaceutical Chemistry, School of Pharmacy, University of California San Francisco, San Francisco, CA 94143-0446, USA and Mass Spectrometry Facility, Biological Research Center, H-6701, Szeged, POB 521, Hungary. Introduction Within the last decade mass spectrometry has become the method of choice for protein identification. Whole cell lysates, components of multi-unit protein complexes, proteins isolated by immuno-precipitation or affinity chromatography are now being routinely studied using this technique [Reviews: 1-8]. The key element in this success derives from the ease of peptide sequence determination based on interpretation of peptide fragmentation spectra. Similarly, these techniques are also used for recombinant protein characterization as well as for structure elucidation of biologically active small peptides, such as toxins, hormones, antibiotics [9-14]. Peptide sequence and structural data can be obtained by collisional activation of selected singly or multiply charged precursor ions. The principles of high energy collisional activation have been established in combination with FAB, LSIMS ionization on 4-sector instruments [15], also with MALDI on sector-orthogonal-acceleration-TOF hybrid tandem instruments [16], and with MALDI only on a TOF-TOF tandem mass spectrometer [17]. Low energy collision-induced dissociation (CID) spectra are acquired with all the other hybrid tandem instruments, for example, quadrupole-orthogonal-acceleration-time-of-flight (QqTOF) instruments, by triple quadrupole mass spectrometers and ion traps regardless of the ionization method applied [18-21]. The unimolecular decomposition of peptide ions generated by MALDI may be detected and recorded by post source decay (PSD) analysis in MALDI-TOF instruments equipped with reflectron [22]. FTMS instruments typically utilize sustained off-resonance irradiation (SORI) to generate MS/MS spectra [23], and