Work

N-linked Glycan Release Efficiency: A Quantitative Comparison between NaOCl and PNGase F Release Protocols

Year: 2019

Type: article

Abstract: There are several methods, both chemical and enzymatic, to release N-linked glycans for structural characterization.One of the most common enzymatic release methods is the use of peptide:N-glycosidase F (PNGase F).A less expensive and quicker alternative has been reported for the release of N-linked glycans chemically using sodium hypochlorite (NaOCl), which hydrolyzes the peptide-glycan bond, yielding the intact glycan with a free reducing terminus.Here, we quantitatively analyzed the efficiency of the NaOCl release protocol compared with the PNGaseF release protocol for small-scale analysis (300 mg) using liquid chromatography-single reaction monitoring-mass spectrometry.We determined that the relative glycan composition of released N-linked glycans from the NaOCl protocol is similar to a typical PNGase F protocol, but the absolute recovery of N-linked glycans is significantly lower with the chemical procedure.

Source: Journal of Biomolecular Techniques JBT

DOI: 10.7171/jbt.19-3004-001

Authors: David A. Fischler, Ron Orlando

Institutions: Centers for Disease Control and Prevention, University of Georgia, National Center for Environmental Health

Language: English

Topic: Glycosylation and Glycoproteins Research

Subfield: Molecular Biology

Field: Biochemistry, Genetics and Molecular Biology

Domain: Life Sciences

Open Access status: diamond

Funder: National Institute of General Medical Sciences

Awards: Development of Isotopically Labeled Glycoproteins for use as Internal Standards, Development of Isotopically Labeled Glycoproteins for use as Internal Standards