PMID- 26331933
OWN - NLM
STAT- MEDLINE
DCOM- 20160711
LR  - 20150903
IS  - 1097-0231 (Electronic)
IS  - 0951-4198 (Linking)
VI  - 29
IP  - 19
DP  - 2015 Oct 15
TI  - A systematic study of glycopeptide esterification for the semi-quantitative 
      determination of sialylation in antibodies.
PG  - 1817-26
LID - 10.1002/rcm.7287 [doi]
AB  - RATIONALE: 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. METHODS: 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. RESULTS: 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 acid (alpha2,3 or alpha2,6). Esterification also 
      affects the carboxyl groups in the peptide, including the C-terminal COOH. 
      CONCLUSIONS: For antibody analysis, MALDI-MS ion abundances give a better 
      semi-quantitative estimate of sialylation levels for esterified than for 
      unreacted glycopeptides. The method is simple to use and helps to differentiate 
      the branching patterns of sialic acids in antibodies.
CI  - Copyright (c) 2015 John Wiley & Sons, Ltd.
FAU - Gomes de Oliveira, Andrey Giovanni
AU  - Gomes de Oliveira AG
AD  - Chemistry Department, University of Manitoba, Winnipeg, MB, Canada, R3T2N2.
FAU - Roy, Rini
AU  - Roy R
AD  - Chemistry Department, University of Manitoba, Winnipeg, MB, Canada, R3T2N2.
FAU - Raymond, Celine
AU  - Raymond C
AD  - Human Health Therapeutics Portfolio, National Research Council Canada, Montreal, 
      QC, Canada, H4P2R2.
AD  - Departement de biochimie et medecine moleculaire, Universite de Montreal, 
      Montreal, QC, Canada, H3C3J7.
FAU - Bodnar, Edward D
AU  - Bodnar ED
AD  - Chemistry Department, University of Manitoba, Winnipeg, MB, Canada, R3T2N2.
FAU - Tayi, Venkata S
AU  - Tayi VS
AD  - Microbiology Department, University of Manitoba, Winnipeg, MB, Canada, R3T2N2.
FAU - Butler, Michael
AU  - Butler M
AD  - Microbiology Department, University of Manitoba, Winnipeg, MB, Canada, R3T2N2.
FAU - Durocher, Yves
AU  - Durocher Y
AD  - Human Health Therapeutics Portfolio, National Research Council Canada, Montreal, 
      QC, Canada, H4P2R2.
AD  - Departement de biochimie et medecine moleculaire, Universite de Montreal, 
      Montreal, QC, Canada, H3C3J7.
FAU - Perreault, Helene
AU  - Perreault H
AD  - Chemistry Department, University of Manitoba, Winnipeg, MB, Canada, R3T2N2.
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PL  - England
TA  - Rapid Commun Mass Spectrom
JT  - Rapid communications in mass spectrometry : RCM
JID - 8802365
RN  - 0 (Antibodies)
RN  - 0 (Glycopeptides)
RN  - GZP2782OP0 (N-Acetylneuraminic Acid)
SB  - IM
MH  - Antibodies/*chemistry
MH  - Esterification
MH  - Glycopeptides/*analysis/*chemistry
MH  - Humans
MH  - N-Acetylneuraminic Acid/*analysis
MH  - Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
MH  - Tandem Mass Spectrometry
EDAT- 2015/09/04 06:00
MHDA- 2016/07/12 06:00
CRDT- 2015/09/03 06:00
PHST- 2015/03/20 00:00 [received]
PHST- 2015/07/15 00:00 [revised]
PHST- 2015/07/23 00:00 [accepted]
PHST- 2015/09/03 06:00 [entrez]
PHST- 2015/09/04 06:00 [pubmed]
PHST- 2016/07/12 06:00 [medline]
AID - 10.1002/rcm.7287 [doi]
PST - ppublish
SO  - Rapid Commun Mass Spectrom. 2015 Oct 15;29(19):1817-26. doi: 10.1002/rcm.7287.