Expression, purification and renaturation of Pol P51 antigen of HIV-1 strain CN54 and its application in antibody detection.
- Author:
Jue HOU
1
;
Jing SUN
;
Zhiyong XU
;
Wenling FAN
;
Yixuan ZHANG
;
Yong LIU
;
Yanling HAO
Author Information
1. College of Life Science and Biopharmaceutics, Shenyang Pharmaceutical University, Shenyang, China.
- Publication Type:Journal Article
- MeSH:
Escherichia coli;
genetics;
metabolism;
HIV Antibodies;
blood;
immunology;
HIV Infections;
immunology;
virology;
HIV Reverse Transcriptase;
biosynthesis;
genetics;
immunology;
HIV-1;
classification;
immunology;
Humans;
Protein Renaturation;
Recombinant Proteins;
biosynthesis;
genetics;
immunology;
Sensitivity and Specificity
- From:
Chinese Journal of Biotechnology
2010;26(2):201-206
- CountryChina
- Language:Chinese
-
Abstract:
To obtain the pure and soluble P51 antigen of HIV-1 strain CN54, we transformed the Escherichia. coli strain BL21 codonplus-RIL with recombinant plasmid pTHioHisA51 which carries a gene encoding the Polymerase (Pol) P51 antigen of HIV-1 CN54 formerly, and induced protein expression by IPTG. We purified the recombinant protein with Chelating Sepharose FF-Ni and DEAE-Sepharose FF column chromatography, then renatured the recombinant protein by dialyzation. Purified protein was identified by Western blotting. We labeled and coated antigen P51 in a dual-antigen sandwich system, and tested it with serum samples from HIV-infected individuals. The results showed that P51 was expressed as inclusion body, and represented about 50% of total cellular protein. After purification and renaturation, the purity of P51 was up to 95%. Western blotting and sandwich ELISA demonstrated that recombinant P51 had good anti-HIV antibody specificity and sensitivity. The results suggested that recombinant HIV-1 P51 can be prepared as diagnostic reagent, and provides valuable support for HIV-1 detection and vaccine research.
