Maryam-Sadat Yadavar-Nikravesh, Alireza Milani , Rouhollah Vahabpour , Mehdi Khoobi , Haleh Bakhshandeh* and Azam Bolhassani* Pages 138 - 146 ( 9 )
Background: HIV-1 TAT protein is essential for the regulation of viral genome transcription. The first exon of TAT protein has a fundamental role in the stimulation of the extrinsic and intrinsic apoptosis pathways, but its anti-HIV activity is not clear yet.
Methods: In the current study, we firstly cloned the first exon of the TAT coding sequence in the pET-24a expression vector and then protein expression was done in the Rosetta expression host. Next, the expressed TAT protein was purified by Ni-NTA column under native conditions. After that, the protein yield was determined by Bradford kit and NanoDrop spectrophotometry. Finally, the cytotoxicity effect and anti-Scr-HIV-1 activity of the recombinant TAT protein alone and along with Tenofovir drug were assessed by MTT and ELISA, respectively.
Results: The recombinant TAT protein was successfully generated in E. coli, as confirmed by 13.5% SDS-PAGE and western blotting. The protein yield was ~150-200 μg/ml. In addition, the recombinant TAT protein at a certain dose with low toxicity could suppress Scr-HIV replication in the infected HeLa cells (~30%) that was comparable with a low toxic dose of Tenofovir drug (~40%). It was interesting that the recombinant TAT protein could enhance anti-HIV potency of Tenofovir drug up to 66%.
Conclusion: Generally, a combination of TAT protein and Tenofovir drug could significantly inhibit HIV-1 replication. It will be required to determine their mechanism of action in the next studies.
HIV-1, TAT protein, prokaryotic expression system, highly active antiretroviral therapy, tenofovir drug, anti-HIV activity.
Department of Hepatitis, AIDS and Blood Borne Diseases, Pasteur Institute of Iran, Tehran, Department of Hepatitis, AIDS and Blood Borne Diseases, Pasteur Institute of Iran, Tehran, Department of Medical Lab Technology, School of Allied Medical Sciences, Shahid Beheshti University of Medical Sciences; Tehran, Department of Pharmaceutical Biomaterials and Medical Biomaterials Research Center (MBRC), Faculty of Pharmacy, Tehran University of Medical Sciences, Tehran, Nanobiotechnology Department, New Technologies Research Group, Pasteur Institute of Iran, Tehran, Department of Hepatitis, AIDS and Blood Borne Diseases, Pasteur Institute of Iran, Tehran