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Comprehensive molecular characterization of a cisplatin-specific monoclonal antibody

  • Despite their immense and rapidly increasing importance as analytical tools or therapeutic drugs, the detailed structural features of particular monoclonal antibodies are widely unknown. Here, an antibody already in use for diagnostic purposes and for molecular dosimetry studies in cancer therapy with very high affinity and specificity for cisplatin-induced DNA modifications was studied extensively. The molecular structure and modifications as well as the antigen specificity were investigated mainly by mass spectrometry. Using nano electrospray ionization mass spectrometry, it was possible to characterize the antibody in its native state. Tandem-MS experiments not only revealed specific fragments but also gave information on the molecular structure. The detailed primary structure was further elucidated by proteolytic treatment with a selection of enzymes and high resolution tandem-MS. The data were validated by comparison with known antibody sequences. Then, the complexDespite their immense and rapidly increasing importance as analytical tools or therapeutic drugs, the detailed structural features of particular monoclonal antibodies are widely unknown. Here, an antibody already in use for diagnostic purposes and for molecular dosimetry studies in cancer therapy with very high affinity and specificity for cisplatin-induced DNA modifications was studied extensively. The molecular structure and modifications as well as the antigen specificity were investigated mainly by mass spectrometry. Using nano electrospray ionization mass spectrometry, it was possible to characterize the antibody in its native state. Tandem-MS experiments not only revealed specific fragments but also gave information on the molecular structure. The detailed primary structure was further elucidated by proteolytic treatment with a selection of enzymes and high resolution tandem-MS. The data were validated by comparison with known antibody sequences. Then, the complex glycan structures bound to the antibody were characterized in all detail. The Fc-bound oligosaccharides were released enzymatically and studied by matrix-assisted laser desorption/ionization mass spectrometry. Overall 16 different major glycan structures were identified. The binding specificity of the antibody was investigated by applying synthetic single and double stranded DNA oligomers harboring distinct Pt adducts. The antibody− antigen complexes were analyzed by mass spectrometry under native conditions. The stability of the complex with double stranded DNA was also investigated.zeige mehrzeige weniger

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Metadaten
Autor*innen:L. Ruhe, Stefanie Ickert, U. Hochkirch, J. Hofmann, S. Beck, J. Thomale, M. W. Linscheid
Dokumenttyp:Zeitschriftenartikel
Veröffentlichungsform:Verlagsliteratur
Sprache:Englisch
Titel des übergeordneten Werkes (Englisch):Molecular Pharmaceutics
Jahr der Erstveröffentlichung:2017
Jahrgang/Band:14
Ausgabe/Heft:12
Erste Seite:4454
Letzte Seite:4461
DDC-Klassifikation:Naturwissenschaften und Mathematik / Chemie / Analytische Chemie
Freie Schlagwörter:Antibody; Cisplatin; Native MS; PTM
DOI:10.1021/acs.molpharmaceut.7b00575
ISSN:1543-8384
ISSN:1543-8392
Verfügbarkeit des Dokuments:Datei im Netzwerk der BAM verfügbar ("Closed Access")
Datum der Freischaltung:01.02.2018
Referierte Publikation:Ja
Datum der Eintragung als referierte Publikation:01.02.2018
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