title: [Construction, expression and immunogenicity analysis of a Tat N-terminus-deleted mutant fusion protein of human immunodeficiency virus type 1].
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.
authors.affiliation: Department of Microbiology, Shanghai Key Laboratory of Medical Biodefense, Second Military Medical University, Shanghai 200433, China.
authors.forename: Hua-Qun
authors.initials: HQ
authors.lastname: Zhang
authors.forename: Wen-Ting
authors.initials: WT
authors.lastname: Liao
authors.forename: Qiu-Li
authors.initials: QL
authors.lastname: Chen
authors.forename: Yi-Bing
authors.initials: YB
authors.lastname: Ge
authors.forename: Jie
authors.initials: J
authors.lastname: Yang
authors.forename: Ping-Ping
authors.initials: PP
authors.lastname: Zhang
authors.forename: Pei-Pei
authors.initials: PP
authors.lastname: Qi
authors.forename: Chao
authors.initials: C
authors.lastname: Liu
authors.forename: Ting
authors.initials: T
authors.lastname: He
authors.forename: Jin-Hong
authors.initials: JH
authors.lastname: Wang
authors.forename: Wei
authors.initials: W
authors.lastname: Pan
authors.forename: Jie
authors.initials: J
authors.lastname: Cao
mesh_terms: D000818:Animals; D005260:Female; D015696:Gene Products, tat / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology; D015497:HIV-1 / Q000235:genetics* / Q000276:immunology*; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D050505:Mutant Proteins / Q000235:genetics* / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology
chemical_list: D015696:Gene Products, tat; D050505:Mutant Proteins; D011993:Recombinant Fusion Proteins