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Mapping the glycation sites in the neoglycoconjugate from hexasaccharide antigen of Vibrio cholerae , serotype Ogawa and the recombinant tetanus toxin C‐fragment carrier
Author(s) -
Jahouh Farid,
Xu Peng,
Vann Willie F.,
Kováč Pavol,
Banoub Joseph H.
Publication year - 2013
Publication title -
journal of mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.475
H-Index - 121
eISSN - 1096-9888
pISSN - 1076-5174
DOI - 10.1002/jms.3258
Subject(s) - chemistry , vibrio cholerae , glycation , cholera toxin , recombinant dna , toxin , biochemistry , stereochemistry , chromatography , microbiology and biotechnology , bacteria , genetics , receptor , gene , biology
We report herein the glycation sites in a vaccine candidate for cholera formed by conjugation of the synthetic hexasaccharide fragment of the O‐specific polysaccharide of Vibrio cholerae , serotype Ogawa, to the recombinant tetanus toxin C‐fragment (rTT–Hc) carrier. Matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry analysis of the vaccine revealed that it is composed of a mixture of neoglycoconjugates with carbohydrate : protein ratios of 1.9 : 1, 3.0 : 1, 4.0 : 1, 4.9 : 1, 5.9 : 1, 6.9 : 1, 7.9 : 1 and 9.1 : 1. Liquid chromatography tandem mass spectrometry (LC‐MS/MS) analysis of the tryptic and GluC V8 digests allowed identification of 12 glycation sites in the carbohydrate–protein neoglycoconjugate vaccine. The glycation sites are located exclusively on lysine (Lys) residues and are listed as follows: Lys 22, Lys 61, Lys 145, Lys 239, Lys 278, Lys 318, Lys 331, Lys 353, Lys 378, Lys 389, Lys 396 and Lys 437. Based on the 3‐D representation of the rTT–Hc protein, all the glycation sites correspond to lysines located at the outer surface of the protein. Copyright © 2013 John Wiley & Sons, Ltd.