Synthesis in Vibrio cholerae and secretion of hepatitis B virus antigens fused to Escherichia coli heat-labile enterotoxin subunit B.
A simple and effective electroporation method for the transformation of Vibrio cholerae with nonmobilizable plasmids is described. Expression plasmids directing the synthesis of fusion proteins with the subunit B of Escherichia coli heat-labile enterotoxin B (LT-B) were transformed into nontoxinogenic V. cholerae vaccine strains. A protein consisting of two overlapping immunodominant antibody-binding sites of the hepatitis B virus (HBV) middle surface antigen fused to the C terminus of full-length LT-B was secreted into the supernatant of V. cholerae cultures, whereas two other LT-B/HBV fusion proteins were mostly retained within the cells or rapidly degraded in the culture supernatant. While the secretion of fusion proteins with cholera toxin subunit B (CT-B) from V. cholerae has been described, this is to our knowledge the first report describing extracellular secretion of defined foreign epitopes fused to LT-B in V. cholerae. The fusion of guest epitopes to LT-B or CT-B and secretion in V. cholerae could be an interesting system to rapidly produce pure fusion proteins for immunisation, functional studies or diagnostic procedures. An LT-B/pre-S2 fusion protein purified from the supernatant of recombinant V. cholerae induced serum IgG antibodies against LT-B and against the HBV middle surface antigen in mice after parenteral and oral immunisation.
Schödel
F
F
Max-Planck-Institut für Biochemie, Martinsried, F.R.G.
Will
H
H
Johansson
S
S
Sanchez
J
J
Holmgren
J
J
D000818:Animals; D001427:Bacterial Toxins / Q000276:immunology / Q000378:metabolism*; D015153:Blotting, Western; D004768:Enterotoxins / Q000276:immunology / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins; D006511:Hepatitis B Antigens / Q000276:immunology / Q000378:metabolism*; D007074:Immunoglobulin G / Q000096:biosynthesis; D051379:Mice; D008810:Mice, Inbred C57BL; D010957:Plasmids; D014170:Transformation, Genetic; D014734:Vibrio cholerae / Q000378:metabolism*; D014760:Viral Fusion Proteins / Q000096:biosynthesis* / Q000276:immunology
D001427:Bacterial Toxins; D004768:Enterotoxins; D029968:Escherichia coli Proteins; D006511:Hepatitis B Antigens; D007074:Immunoglobulin G; D014760:Viral Fusion Proteins; C027538:heat-labile enterotoxin, E coli