ABSTRACT
<p><b>OBJECTIVE</b>To express a fusion protein of Neisseria gonorrhoeae with a mucosal adjuvant.</p><p><b>METHODS</b>The gene coding Loop VI-VIII(PL678) of porin, an out-membrane protein of Neisseria gonorrhoeae, was obtained by PCR. It was inserted into a plasmid fused with subunit B of heat labile enterotoxin. The recombinant was transformed in E. coli. The expression of fusion protein was analysed by ELISA, SDS-PAGE and Western-blot.</p><p><b>RESULT</b>Fusion protein with LTB was successfully expressed, and displayed both the ability of binding GM1 and the reactogenicity with polyclonal antibodies against Neisseria gonorrhoeae.</p><p><b>CONCLUSION</b>The expression of fusion protein laid a foundation for the study of the intramolecular vaccine against Neisseria gonorrhoeae.</p>
Subject(s)
Bacterial Outer Membrane Proteins , Bacterial Toxins , Bacterial Vaccines , Allergy and Immunology , Enterotoxins , Escherichia coli , Genetics , Escherichia coli Proteins , Neisseria gonorrhoeae , Chemistry , Allergy and Immunology , Polymerase Chain Reaction , Recombinant Fusion Proteins , Vaccines, Synthetic , Allergy and ImmunologyABSTRACT
Objective To construct a fusion expression vector with subunit B of Escherichia coli heat labile enterotoxin. Methods Gene encoding LTB without stop codon obtained by PCR was introduced to a linker and was recombined on vector Pinpoint Xa I through Eco RⅠ and Eco RⅤ sites and then fused with ureB. The recombinant was used to transform E. coli JM109. The expression of LTB was analyzed by SDS PAGE and Western blotting. Results The fusion expression plasmid was successfully constructed. The subunit B of urease of Helicobacter pylori (Hp) fusogenic protein with LTB with the ability to bind GM 1 and the reactogenicity with polyclonal antibodies against Hp was harvested. Conclusion The successfully constructed vector provides experimental base for the studies of intramolecular adjuvant vaccine.
ABSTRACT
Objective To recombine and express a component of urease B subunit transmembrane protein of helicobacter pylori. Methods A 732 bp gene fragment of urease B subunit of helicobacter pylori was cloned into pET11C and transformed into BL21（DE3）E.coli. The positive clone was induced with IPTG. The expression of target protein was analysed by SDS-PAGE and Western blot. Results It is successful to construct the recombinant plasmid pET-UreB0.7 containing urease B subunit 0.7 kb gene fragment. A protein （MW≈28 000 u） with immunoreactivity， was expressed by 19.8％ in BL21（DE3）E.coli induced with IPTG. Conclusions The recombinant component of urease B subunit transmembrane protein may play a role in the research of its biological function and might be used as the vaccine against helicobacter pylori.
ABSTRACT
Objective To study the mucosa immune responses of gastric and intestinal mucosa in Balb／c mice administered orally with Hp sonicate and mucosal adjuvant（LT）．Methods The changes of antigen specific AFC in gastric and intestinal mucosa were detected by ELISPOT assay． Results The numbers of sIgA and IgG AFC rise significantly in PP and gastric mucosa， especially the numbers of sIgA-AFC， significant differences were observed between two immunized groups and the control． Conclusions Locally synthesized specific sIgA antibodies contribute to immunity against gastric helicobacter infection．