Construction, expression and immunogenicity analysis of a Tat N-terminus-deleted mutant fusion protein of human immunodeficiency virus type 1.
- Author:
Hua-Qun ZHANG
1
;
Wen-Ting LIAO
;
Qiu-Li CHEN
;
Yi-Bing GE
;
Jie YANG
;
Ping-Ping ZHANG
;
Pei-Pei QI
;
Chao LIU
;
Ting HE
;
Jin-Hong WANG
;
Wei PAN
;
Jie CAO
Author Information
1. Department of Microbiology, Shanghai Key Laboratory of Medical Biodefense, Second Military Medical University, Shanghai 200433, China.
- Publication Type:Journal Article
- MeSH:
Animals;
Female;
Gene Products, tat;
biosynthesis;
genetics;
immunology;
HIV-1;
genetics;
immunology;
Humans;
Mice;
Mice, Inbred BALB C;
Mutant Proteins;
genetics;
immunology;
Recombinant Fusion Proteins;
biosynthesis;
genetics;
immunology
- From:
Chinese Journal of Virology
2011;27(6):580-586
- CountryChina
- Language:Chinese
-
Abstract:
In the study, a gene encoding Tat protein N terminal 1- 21 amino acid residues-deleted mutant (Tat22-101) was amplified by PCR from a full length Tat gene of human immunodeficiency virus type 1, and the prokaryotic expression plasmid pET32a-Tat22-101 was constructed. After identification by digestion with endonucleases and sequencing, the recombinant plasmid pET32a-Tat22-101 was transformed into E. coli BL21(DE3) and expressed with IPTG induction. The mutant fusion protein with deleted Tat N terminal was purified by an affinity chromatography column Ni(2+)-NTA and subsequently identified by SDS-PAGE and Western blotting. The results showed that the molecular weight of the mutant protein was approximately 26.9kD. Furthermore, BALB/c mice were immunized with the mutant protein and the anti-sera were collected. ELISA results showed that the mutant protein preserved its immunogenicity, particularly it could improve the production of antibodies to other epitopes in addition to the N terminal epitope of Tat protein, which might provide some valuable information for the study of Tat functions as well as for development of potential novel HIV Tat vaccine.
