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Characterisation of combinatorial libraries of mucin‐2 antigen peptides by high‐resolution mass spectrometry
Author(s) -
Windberg Emöke,
Hudecz Ferenc,
Marquardt Andreas,
Sebestyén Ferenc,
Kiss Andrea,
Bösze Szilvia,
MedzihradszkySchweiger Hedvig,
Przybylski Michael
Publication year - 2002
Publication title -
rapid communications in mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.528
H-Index - 136
eISSN - 1097-0231
pISSN - 0951-4198
DOI - 10.1002/rcm.649
Subject(s) - chemistry , fourier transform ion cyclotron resonance , peptide , mass spectrometry , peptide library , epitope , matrix assisted laser desorption/ionization , protein mass spectrometry , chromatography , polyclonal antibodies , peptide sequence , electrospray ionization , antigen , biochemistry , desorption , organic chemistry , genetics , adsorption , biology , gene
An epitope motif, TX 1 TX 2 T, of mucin‐2 glycoprotein was identified by means of a mucin‐2‐specific monoclonal antibody, mAb 994, raised against a synthetic mucin‐derived 15‐mer peptide conjugate. For determination of the epitope sequence recognised with highest affinity by mAb 994, a combinatorial approach was applied using the portioning‐mixing technique excluding Cys. Antibody binding of libraries was most profound when Gln was at the X 1 position. Analytical characterisation of the TQTX 2 T library was conducted by amino acid analysis and matrix‐assisted laser desorption/ionisation time‐of‐flight (MALDI‐TOF) and electrospray ionisation Fourier transform ion cyclotron resonance (ESI‐FTICR) mass spectrometric methods. Control libraries were prepared by mixing 19 individual peptides corresponding to the TQTX 2 T sequence. Thus, mixtures of 6, 10 and 19 pentapeptides were analysed and compared with the combinatorial mixture. MALDI‐TOFMS was able to detect only partially the components in the 6‐ and 10‐member mixtures, but failed to characterise a more complex 19‐member mixture. In contrast, ESI‐FTICRMS resolved all mixtures of higher complexity and provided direct identification at monoisotopic resolution, such as for a peptide library containing ‘isobaric’ lysine and glutamine (Δm = 0.0364 Da). The results of this study suggest that ESI‐FTICRMS is a powerful tool for characterisation of combinatorial peptide libraries of higher complexity. Copyright © 2002 John Wiley & Sons, Ltd.