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Isolation, Purification, Characterization and Direct Conjugation of the Lipid A‐Free Lipopolysaccharide of Vibrio cholerae O139
Author(s) -
Xu Peng,
Korcová Jana,
Baráth Peter,
Čížová Alžbeta,
Valáriková Jana,
Qadri Firdausi,
Kelly Meagan,
O'Connor Robert D.,
Ryan Edward T.,
Bystrický Slavomír,
Kováč Pavol
Publication year - 2019
Publication title -
chemistry – a european journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.687
H-Index - 242
eISSN - 1521-3765
pISSN - 0947-6539
DOI - 10.1002/chem.201902263
Subject(s) - vibrio cholerae , chemistry , strain (injury) , lipopolysaccharide , chromatography , high performance liquid chromatography , lipid a , hydrolysis , vibrionaceae , conjugate , microbiology and biotechnology , biochemistry , bacteria , biology , mathematical analysis , genetics , mathematics , gene , anatomy , endocrinology
The lipopolysaccharide (LPS) of Vibrio cholerae O139, strain CIRS245, was isolated conventionally, and the lipid A was removed by mild acid hydrolysis (0.1 m NaOAc buffer containing 1 % SDS, pH 4.2, 95 °C, 8 h). The crude product was a complex mixture consisting mainly of constituent fragments of the O‐specific polysaccharide‐core (OSPc). The OSPc was only a minor component in the mixture. Two‐stage purification of the crude OSPc by HPLC gave pure OSPc fragment of the LPS, as shown by NMR spectroscopy, analytical HPLC and ESI‐MS. This material is the purest OSPc fragment of the LPS from Vibrio cholerae O139 reported to date. The purified OSPc was readily converted to the corresponding methyl squarate derivative and the latter was conjugated to BSA. The conjugate, when examined by ELISA, showed immunoreactivity with sera from patients in Bangladesh recovering from cholera caused by V. cholerae O139, but not O1.