ABSTRACT
Background: As a drug target and an antigenic agent, HIV-1 protease [HIV-1 PR] is at the center of attention for designing anti-AIDS inhibitors and diagnostic tests. In previous studies, the production of the recombinant protease has been faced with several difficulties; therefore, the aims of this study were the easy production, purification of the soluble form of protease in E. coli and investigation of its immunoreactivity
Methods: Protease coding region was isolated from the serum of an infected individual, amplified by RT-PCR and cloned into PTZ57R using TA-cloning. Protease coding frame was isolated by PCR and cloned in pET102/D. TOPO expression vector and cloned protease was expressed in Escherichia coli [E. coli] BL21. Produced recombinant protein was purified by affinity Ni-NTA column and protein concentration was checked by BCA protein assay kit. Subsequently, immunoreactivity of recombinant protease [rPR] was assayed by Western blotting and ELISA
Results: Cloning of the HIV protease by TOPO cloning system in pET102/D.TOPO was confirmed with PCR and sequencing. The concentration range of purified recombinant protein was 85 to 100 micro g/ml. Immunogenicity of rPR was confirmed by Western blotting and ELISA
Conclusion: Soluble production of recombinant HIV-1 protease [HIV-1 rPR] was performed successfully. This recombinant protein disclosed 86% specificity and 90% sensitivity in immunoassay tests