Optimization the expression of human papilloma virus E6 and E7 polytopic construct in E. coli expression system

• Main Authors: Arian Rahimi, Arash Arashkia, Amir Mirzaie, Hassan Noorbazargan, Seyed Ataollah Sadat Shandiz, Roghayeh Rahimi, Mehdi Mahdavi
• Format: Article
• Language: fas
• Published: Tehran University of Medical Sciences 2015-12-01
• Series: Tehran University Medical Journal
• Subjects: human papilloma virus; E6 and E7 proteins; polytopic vaccine; E.coli
• Online Access: http://tumj.tums.ac.ir/browse.php?a_code=A-10-25-5390&slc_lang=en&sid=1

Background: Human papilloma virus is a DNA virus from the papillomavirus family that is most prevalent in human cervical cancers and many studies showed the E6 and E7 proteins are present in the majority of cervical cancer cases. Development of universal HPV peptide-based vaccine with more serotypes coverage has considerable value. The aim of the study was to design a multi-epitope universal vaccine for major HPV based on E6 and E7 proteins and optimization the expression of polytopic construct contains E6 and E7 genes from different genotypes of human papilloma virus as a candid vaccine. Methods: In this experimental study that was carried out in Pasteur Institute of Iran, Virology Department from October 2013 to November 2014. In order to design the polytypic construct, we predicted the most probable immunogenic epitopes of E6 and E7 from common high risk HPV16, 18, 31, 45 along with high prevalent type 6 and 11 using bioinformatics methods. The synthetic pET28a expression vector harboring E6 and E7 protein was transformed into Escherichia coli hosts and its expression was analyzed by SDS-PAGE and western blotting. Finally, in order to expression optimization of recombinant protein, cell density, induction time, growth temperature, IPTG (Isopropyl β-D-1-thiogalactopyranoside) concentration and cultures media were studied. Results: In the present study the recombinant fusion protein was expressed successfully and the highest expression of target protein was achieved in super broth medium containing 0.1% glucose and 0.2% L-arabinose. In Super broth medium, the optimum condition for recombinant protein expression was occurred at OD600 of 0.8, 0.1mM IPTG, one hour’s incubation time at 37 °C and BL21 (A1) host. Conclusion: The results of this study show that the optimum expression of E6 and E7 proteins from different genotypes of human papilloma virus can be performed. Moreover, by purification of recombinant protein and evaluation of its immunogenicity in mice, it can be used as a vaccine candidate against the human papilloma virus.