Site-specific characterization of the N-linked glycans of murine prion protein by high-performance liquid chromatography/electrospray mass spectrometry and exoglycosidase digestions.
The murine prion protein PrP gene encodes a protein of 254 amino acids with two consensus sites for Asn-linked glycosylation at codons 180 and 196. A partial site-specific study of the N-linked glycans from hamster PrP has previously been carried out by mass spectrometry [Stahl, N., Baldwin, M. A., Teplow, D. B., Hood, L., Gibson, B. W., Burlingame, A. L., and Prusiner, S. B. (1993) Biochemistry 32, 1991-2002] and revealed that the glycosylation at Ash-181 (equivalent to mouse 180) is heterogeneous, comprising over 30 glycoforms. The identification of the glycosylated peptide spanning Asn-197 was not reported. Recent technical advances in electrospray mass spectrometry now provide the sensitivity to detect low femtomole quantities of glycopeptides with >5000 mass resolution and 30 ppm mass measurement [Medzihradszky, K. F., Besman, M. J., and Burlingame, A. L. (1998) Rapid Commun. Mass Spectrom. 12, 472-478]. This performance coupled with stepwise exoglycosidase digestion has been employed tO establish the differential nature of the structural complexity (glycoforms) of the glycans at Asn-180 and Asn-196 from a single strain infected with the ME7 strain. Some sixty structures have been found characterized by neutral and sialylated bi-, tri- , and tetraantennary complex-type bearing outer-arm α(1-3)-fucosylation (the Lewis(x) and sialyl-Lewis(x) epitopes), core α(1,6) fucosylation, and the presence of terminal HexNAc residues. The Lewis(x) trisaccharide is the major nonreducing structure at Asn-180, and significant amounts of both Lewis(x) and sialyl Lewis(x) epitopes are observed at Asn-196. The abundance of the Lewis(x) and sialyl Lewis(x) epitopes on murine PrP(Sc) may indicate a role for these structures in the normal function of PrP(C) or the pathophysiology of PrP(Sc).
|Title:||Site-specific characterization of the N-linked glycans of murine prion protein by high-performance liquid chromatography/electrospray mass spectrometry and exoglycosidase digestions|
|UCL classification:||UCL > School of Life and Medical Sciences
UCL > School of Life and Medical Sciences > Faculty of Life Sciences
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