[Clone, expression and immunoreaction of napA gene of Helicobacter pylori].
OBJECTIVE
To construct a prokaryotic expression system of Helicobacter pylori(Hp) (neutrophil-activating protein) napA gene, analyze nucleic acid sequence and study its immunity and inflammation.


METHODS
napA fragment was amplified from Hp NCTC11639 chromosomal DNA by PCR. Its T-A was cloned, sequenced and compared with other Hp strains on the GenBank. Then the gene was cloned into pGEX-4T-1 fusion expression vector, expressed in E.coli and purified by GST-affinity chromatography. The purified product was used to screen 29 stains of mouse anti Hp monoclonal antibodies(mAb) and its immunity and inflammation analyzed with sera of Hp-infected patients by Western blot.


RESULTS
napA fragment was composed of 435 base pairs (GenBank No.DQ341279) and the nucleotide homology with other Hp strains on the GenBank was 94%-98%. The molecular weight of the recombinant napA-pGEX-4T-1 expressed in E.coli was 44 kDa. 3 of 29 anti-Hp mAbs were against NAP. Western blot analysis proved that the recombinant NAP was specifically recognized by the sera of Hp-infected patients.


CONCLUSIONS
The recombinant NAP has original immunoreaction. It is of great value to clinical sero-diagnosis and vaccine study of Hp.
Luo
Jun
J
Faculty of Microbiology, School of Public Health and Tropic Medicine, Southern Medical University, Guangzhou 510515, China.
Long
Bei-guo
BG
Li
Yan
Y
Lai
Jian-ping
JP
Hu
Ping
P
Zhang
Wen-bing
WB
Long
Min
M
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D015153:Blotting, Western; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005822:Genetic Vectors / Q000235:genetics; D016481:Helicobacter Infections / Q000276:immunology; D016480:Helicobacter pylori / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism
D000911:Antibodies, Monoclonal; D001426:Bacterial Proteins; D011994:Recombinant Proteins