EVALUATION OF GLYCOSYLATION SITE HETEROGENEITY AND SELECTIVE IDENTIFICATION OF GLYCOPEPTIDES IN PROTEOLYTIC DIGESTS OF BOVINE ALPHA(1)-ACIDGLYCOPROTEIN BY MASS-SPECTROMETRY

Glycosylation sites in bovine alpha(1)-acid glycoprotein (AGP) have be en identified, and the inherent heterogeneity evaluated, by capillary electrophoretic and reversed-phase liquid chromatography/electrospray- mass spectrometric analyses of proteolytic digests of this glycoprotei n, The success of these methods in locating glycopeptides relied on si gnificant heterogeneity within each glycosylation site. In order to ra pidly locate sites in glycoproteins of any degree of heterogeneity, a novel mass spectrometric method was applied to selectively identify th e glycopeptides in a proteolytic digest of bovine alpha(1)-AGP. The gl ycopeptides were selectively located by the generation and detection o f characteristic oxonium ions from the carbohydrate moieties by collis ion-induced dissociation (CID) during liquid chromatography/electrospr ay-tandem mass spectrometry, and liquid chromatography/CID mass spectr ometry, in which fragmentation was induced in the supersonic expansion region of the electrospray source.