Aim and Background: HIV protease plays an important role in the maturation of the virus and host immune response stimulation; therefore it is important as a therapeutic target and can be used in diagnostic tests. In previous studies, producing rPR has been faced with problems such as toxicity, hydrophobic and purification. The purpose of this study is the use of expression system pET102/D.TOPO to overcome the mentioned problems.  

Methods: Protease gene was isolated from the serum of an infected individual. Cloning process was performed by TOPO Cloning system. Protease gene transformed into E.coli BL21and induced with IPTG. Produced recombinant protein was purified by affinity chromatography (Ni-NTA column). Protein concentration was checked by BCA protein assay kit. Recombinant protein was identified by SDS-PAGE and western-blotting.

Results: HIV-1 protease gene Cloning using TOPO Cloning vector expression system pET102 / D was confirmed by PCR and sequencing. rPR concentration after purification was 60 to 85 micrograms per milliliter. rPR Expression and Immunogenicity by SDS-PAGE and Western blotting were confirmed.

Conclusion: Expression and production of rPR in soluble form was a success. In used Cloning system, because of the expression of rPR in fused form with thioredoxin and histidine tags, problems of toxicity, hydrophobicity and purification were noticeably solved. According to the generated rPR, it can be used to evaluate the diagnostic tests and for designing protease inhibitors in subsequent studies.