Affinity purification of recombinant cholera toxin B subunit oligomer expressed in Bacillus brevis for potential human use as a mucosal adjuvant.
For use as a mucosal adjuvant for human vaccines, a simple method has been developed for the affinity purification of recombinant cholera toxin B subunit which had been expressed in a safe host, Bacillus brevis. Recombinant cholera toxin B subunit, adsorbed quantitatively to a D-galactose-agarose column, was eluted with an 0.1-0.4 M D-galactose gradient with a yield of > 90%. The cholera toxin B subunit preparation was similar to the native cholera toxin B subunit with respect to GM1 binding ability, remarkable stability of the pentamer, and the dissociation-reassociation property by shifting pHs. Cross-linking experiments with glutaraldehyde demonstrated that the pentameric form was predominant; tetrameric, trimeric, dimeric and monomeric forms were detected to a lesser extent, and additionally 10- and 15-mers were observed depending on the concentration of the cholera toxin B subunit.
Yasuda
Y
Y
Department of Microbiology, Nagoya City University Medical School, Nagoya, Japan. yyasuda@med.nagoya-cu.ac.jp
Matano
K
K
Asai
T
T
Tochikubo
K
K
D000276:Adjuvants, Immunologic / Q000737:chemistry / Q000302:isolation & purification*; D001407:Bacillus / Q000378:metabolism*; D002772:Cholera Toxin / Q000737:chemistry / Q000302:isolation & purification*; D002846:Chromatography, Affinity / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification; D016622:Silver Staining
D000276:Adjuvants, Immunologic; D011994:Recombinant Proteins; D002772:Cholera Toxin