A systematic study of glycopeptide esterification for the semi-quantitative determination of sialylation in antibodies

Rapid Communications in Mass Spectrometry : RCM
Andrey Giovanni Gomes de OliveiraHélène Perreault

Abstract

In the expression of recombinant proteins, an important parameter to control or influence is their level of sialylation. Matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometric (MS) methods tend to either underestimate (positive mode) or overestimate (negative mode) the content of sialylated vs. neutral glycans in glycoproteins. Esterification methods have been developed for free sialylated glycans and sialylated Asn-glycans, allowing these acidic groups to ionize with the same efficiency as neutral sugars. Here we describe a method which modifies glycopeptides by esterification. This simple procedure is applied to glycopeptides isolated from tryptic digests of monoclonal antibodies (mAbs), some highly sialylated. To better understand the effect of esterification on the peptide backbone, synthetic EEQYNSTYR was esterified and studied by tandem mass spectrometry (MS/MS). Acetamidation of EEQYNSTYR was also studied as some mAb samples had been overalkylated prior to tryptic digestion. As a general trend, ethyl-esterification or lactonization is observed for each sialic acid on glycoforms of EEQYNSTYR (the N-glycosylated tryptic peptide of IgG Fc), depending on the branching position of the sialic...Continue Reading

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Citations

Nov 21, 2015·Glycoconjugate Journal·Paul G LopezHélène Perreault
Jan 17, 2016·Methods : a Companion to Methods in Enzymology·Emy KomatsuHélène Perreault
Sep 13, 2015·Analytical and Bioanalytical Chemistry·Edward BodnarHélène Perreault
Mar 25, 2016·Molecular & Cellular Proteomics : MCP·Rosina PlompManfred Wuhrer
Dec 12, 2017·Mass Spectrometry Reviews·Qiwei ZhangJianjun Li
Sep 12, 2018·Electrophoresis·Aiying YuYehia Mechref
Apr 25, 2020·Mass Spectrometry Reviews·David J Harvey
Feb 5, 2021·Rapid Communications in Mass Spectrometry : RCM·Taylor BattellinoHélène Perreault
Apr 14, 2018·Journal of the American Society for Mass Spectrometry·Shuang YangJohn Cipollo
Jun 24, 2021·Mass Spectrometry Reviews·Wenjing PengYehia Mechref
Oct 7, 2021·Electrophoresis·Cristian D Gutierrez-ReyesYehia Mechref

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