A study of recombinant protective H.pylori antigens.
OBJECTIVE
To construct a recombinant vector which can express M (r)26000 outer membrane protein (OMP) from Helicobacter pylori (Hp), and to obtain the vaccine protecting against Hp infection and a diagnostic reagent kit quickly detecting Hp infection.


METHODS
The gene encoding the structural M(r)26000 outer membrane protein of Hp was amplified from Hp chromosomal DNA by PCR, and inserted in the prokaryotic expression vector pET32a (+), which was transformed into the Top10 E. coli strain. Recombinant vector was selected, identified and transformed into BL-21(DE3) E. coli strain. The recombinant fusion proteins were expressed. The antigenicity of recombinant protein was studied by ELISA or immunoblotting and immunized Balb/c mice.


RESULTS
The gene of M(r)26000 OMP was amplified to be 594 base pairs, 1.1% of the cloned genes was mutated and 1.51% of amino acid residues was changed, but there was homogeneity between them. The recombinant fusion protein encoded objective polypeptides of 198 amino acid residues, corresponding to calculated molecular masses of M (r)26000. The level of soluble expression products was about 38.96% of the total cell protein. After purification by Ni-NTA agarose resin columniation, the purity of objective protein became about 90%. The ELISA results showed that recombinant fusion protein could be recognized by patient serum infected with Hp and rabbit serum immunized with the recombinant protein. Furthermore,Balb/c mice immunized with the recombinant protein were protected against H.pylori infection.


CONCLUSIONS
M (r)26000 OMP may be a candidate vaccine preventing Hp infection.
Jiang
Zheng
Z
Department of Gastroenterology, the First Affiliated Hospital, Chongqing University of Medical Sciences, Chongqing 400016, China. jzh053@mail.china.com
Tao
Xiao-Hong
XH
Huang
Ai-Long
AL
Wang
Pi-Long
PL
D000818:Animals; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D001428:Bacterial Vaccines; D005798:Genes, Bacterial; D016481:Helicobacter Infections / Q000175:diagnosis / Q000276:immunology; D016480:Helicobacter pylori / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*
D001425:Bacterial Outer Membrane Proteins; D001428:Bacterial Vaccines; D011993:Recombinant Fusion Proteins