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Titolo:
Proteomics of glycoproteins based on affinity selection of glycopeptides from tryptic digests
Autore:
Geng, M; Zhang, X; Bina, M; Regnier, F;
Indirizzi:
Purdue Univ, Dept Chem, W Lafayette, IN 47907 USA Purdue Univ W LafayetteIN USA 47907 Dept Chem, W Lafayette, IN 47907 USA
Titolo Testata:
JOURNAL OF CHROMATOGRAPHY B
fascicolo: 2, volume: 752, anno: 2001,
pagine: 293 - 306
SICI:
1387-2273(20010310)752:2<293:POGBOA>2.0.ZU;2-M
Fonte:
ISI
Lingua:
ENG
Soggetto:
CAPILLARY-ELECTROPHORESIS; MASS-SPECTROMETRY; FUSION PROTEIN; CONCANAVALIN-A; GLYCOSYLATION; MICROHETEROGENEITY; CHROMATOGRAPHY; MIXTURES; LECTIN;
Keywords:
proteomics; glycoproteins; glycopeptides;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
38
Recensione:
Indirizzi per estratti:
Indirizzo: Regnier, F Purdue Univ, Dept Chem, 3164A Brown Bldg, W Lafayette, IN 47907USA Purdue Univ 3164A Brown Bldg W Lafayette IN USA 47907 47907 USA
Citazione:
M. Geng et al., "Proteomics of glycoproteins based on affinity selection of glycopeptides from tryptic digests", J CHROMAT B, 752(2), 2001, pp. 293-306

Abstract

Identification of glycoproteins in complex mixtures derived from either human blood serum or a cancer cell line was achieved in a process involving the steps of (1) reduction and alkylation, (2) proteolysis of all proteins in the mixture with trypsin, (3) affinity chromatographic selection of the glycopeptides with an immobilized lectin, (4) direct transfer of the glycopeptide fraction to a reversed-phase liquid chromatography (RPLC) column and further fractionation by gradient elution, (5) matrix-assisted laser desorption ionization mass spectrometry of individual fractions collected from the RPLC column, and (6) peptide identification based on a database search. The types of glycoproteins analyzed were; (1) N-type glycoproteins of known primary structure, (2) N-type glycoproteins of unknown structure, and (3) O-type glycoproteins glycosylated with a single N-acetylglucosamine. Identification of peptides from complex mixtures was greatly facilitated by eitherC-terminal sequencing with a carboxypeptidase mixture or by comparing chromatographic behavior and mass to standards, as in the case of a known protein. In addition, deglycosylation of peptides with N glycosidase F was necessary to identify N-type glycoproteins of unknown structure. The strength ofthis approach is that it is fast and targets specific molecular species orclasses of glycoproteins for identification. The weakness is that it does not discriminate between glycoforms. (C) 2001 Elsevier Science B.V. All rights reserved.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 03/06/20 alle ore 09:49:24