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Use of N-glycanase to release asparagine-linked oligosaccharides for structural analysis

✍ Scribed by Shirish Hirani; Richard J. Bernasconi; James R. Rasmussen

Publisher: Elsevier Science
Year: 1987
Tongue: English
Weight: 800 KB
Volume: 162
Category: Article
ISSN: 0003-2697
DOI: 10.1016/0003-2697(87)90424-6

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✦ Synopsis

An enzymatic procedure for releasing asparagine-linked oligosaccharides from glycoproteins by treatment with IV-glycanase (peptide-N4-(N-acetyl-&3lucosaminyl)asparagine amidase) has been investigated. Ribonuclease B, transferrin, fetuin, and q-acid glycoprotein were treated with N-glycanase and the released oligosaccharides were radiolabeled with NaB3&. Lectin staining of the iV-glycanase-treated proteins indicated that the deglycosylation reactions had proceeded to completion. The labeled carbohydrate chains were analyzed by HPLC on Micro-Pak AX-5 and AX-10 columns. The proportion of high-mannose and bi-, tri-, and tetraantennary complex chains obtained from each glycoprotein was in agreement with literature values. These results demonstrate that N-glycanase provides a simple method to release all common classes of asparagine-linked oligosaccharides from a glycoprotein in a form that can be radiolabeled directly for structu~ analysis. Q 1987 Academic press Inc.

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