[Expression in Escherichia coli of hepatitis B virus genes from minority nationality patients in Yunnan province with chronic hepatitis B and their antigenicity].
OBJECTIVE
To investigate the expression in Escherichia coli (E. coli) of hepatitis B virus (HBV) genes from minority nationality patients in Yunnan province with chronic hepatitis B (CHB) and their antigenicity.


METHODS
Peripheral blood samples were collected from 25 minority nationality patients with CHB in Yunnan province. Twenty-five CHB patients of Han nationality in Yunnan were used as controls. The full length HBV preS2/S and C genes were amplified by PCR, cloned, sequenced, and inserted into the prokaryotic expression vector p lambda PR. The recombinant plasmids p lambda PR-S2S and p lambda PR-C were transfected into E. coli of the line TOP10. The expression of the non-fusion proteins encoded by the HBV preS2S and C genes was determined by sodium dodecyl sulphate polyacrlamide gel electrophoresis (SDS-PAGE) and Western blotting. The antigenicity of the non-fusion proteins was examined by ELISA. Fifty samples of serum of patients with hepatitis A, 50 samples of serum of patients with hepatitis C, and 50 samples of serum of healthy blood donors were used as controls in the study of the antigenicity of non-fusion proteins.


RESULTS
SDS-PAGE showed that the recombinant plasmids p lambda PR-S2S and p lambda PR-C were both stably and highly expressed in the E. coli for all 50 CHB patients. The molecular weights of the expressed non-fusion proteins, with a concentration of 16% and a purity of 50%, were between 31,000 and 21,000. Western blotting and ELISA showed that the purified recombinant non-fusion proteins reacted strongly with the antibodies HBpreS2S/SAb and HBcAb and the serum of CHB patients, but there was no cross-activity between the non-fusion proteins and all the serum samples of controls with HA and HC, and normal controls.


CONCLUSIONS
The HBV preS2S and C genes from the minority nationality patients with CHB can be stably and highly expression in E. coli. The non-fusion proteins encoded by the HBV preS2S and C genes have high antigenicity. These findings have potential values in development of HB vaccines.
Bian
Zhong-qi
ZQ
Centre for Infectious Diseases, Kunming General Hospital of the People's Liberation Army, Kunming 650032, China. bzq@ynu.edu.cn
Hua
Zhan-lou
ZL
Yan
Wei-yao
WY
Liu
Ming-qiu
MQ
Zheng
Zhao-xin
ZX
D000293:Adolescent; D000328:Adult; D000368:Aged; D044466:Asian People / Q000208:ethnology; D015153:Blotting, Western; D002681:China / Q000453:epidemiology; D004279:DNA, Viral / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D005260:Female; D015870:Gene Expression; D006514:Hepatitis B Surface Antigens / Q000235:genetics / Q000276:immunology*; D006515:Hepatitis B virus / Q000235:genetics / Q000276:immunology*; D019694:Hepatitis B, Chronic / Q000208:ethnology / Q000235:genetics / Q000276:immunology*; D006801:Humans; D008297:Male; D008875:Middle Aged; D016133:Polymerase Chain Reaction; D011204:Poverty Areas; D011498:Protein Precursors / Q000235:genetics / Q000276:immunology*; D014758:Viral Core Proteins / Q000235:genetics
D004279:DNA, Viral; D006514:Hepatitis B Surface Antigens; D011498:Protein Precursors; D014758:Viral Core Proteins; C050807:presurface protein 2, hepatitis B surface antigen