Abstract
BackgroundA major discovery in human etiology recognized that cervical cancer is a consequence of an infection caused by some mucosatropic types of human papillomavirus (HPV). Since L1 protein of HPV is able to induce the formation of neutralizing antibodies, it becomes a protein target to develop HPV vaccines. Therefore, this study aims to obtain and analyze the expression of HPV subunit recombinant protein, namely L1 HPV 52 in E. coli BL21 DE3. The raw material used was L1 HPV 52 protein, while the synthetic gene, which is measured at 1473 bp in pD451-MR plasmid, was codon-optimized (ATUM) and successfully integrated into 5643 base pairs (bps) of pETSUMO. Bioinformatic studies were also conducted to analyze B cell epitope, T cell epitope, and immunogenicity prediction for L1HPV52 protein. ResultsThe pETSUMO-L1HPV52 construct was successfully obtained in a correct ligation size when it was cut with EcoRI. Digestion by EcoRI revealed a size of 5953 and 1160 bps for both TA cloning petSUMO vector and gene of interest, respectively. Furthermore, the right direction of construct pETSUMO-L1HPV52 was proven by PCR techniques using specific primer pairs then followed by sequencing, which shows 147 base pairs. Characterization of L1 HPV 52 by SDS-PAGE analysis confirms the presence of a protein band at a size of ~55 kDa with 6.12 mg/L of total protein concentration. Observation under by transmission electron microscope demonstrates the formation of VLP-L1 at a size between 30 and 40 nm in assembly buffer under the condition of pH 5.4. Based on bioinformatics studies, we found that there are three B cell epitopes (GFPDTSFYNPET, DYLQMASEPY, KEKFSADLDQFP) and four T cell epitopes (YLQMASEPY, PYGDSLFFF, DSLFFFLRR, MFVRHFFNR). Moreover, an immunogenicity study shows that among all the T cell epitopes, the one that has the highest affinity value is DSLFFFLRR for Indonesian HLAs. ConclusionRegarding the achievement on successful formation of L1 HPV52-VLPs, followed by some possibilities found from bioinformatics studies, this study suggests promising results for future development of L1 HPV type 52 vaccine in Indonesia.
Highlights
A major discovery in human etiology recognized that cervical cancer is a consequence of an infection caused by some mucosatropic types of human papillomavirus (HPV)
This study focused on the production of Virus-like particles (VLP) L1 protein that is highly recommended as a future vaccine to overcome for HPV type 52 infection in Indonesia
Construction of truncated L1 HPV type 52 in E. coli BL21 DE3 The complete genome used as template for HPV 52 L1 protein sequence was obtained by the genome analysis from the database of the National Centre for Biotechnology Information (NCBI)
Summary
A major discovery in human etiology recognized that cervical cancer is a consequence of an infection caused by some mucosatropic types of human papillomavirus (HPV). Around 40 types of those HPV’s variants infect mucosal epithelium, and epidemiology studies found its strong association with cervical cancer. The HPV 52 is known to be the most prevalent type in Indonesia, while another study identified it to be the frequent cause of cervical cancer in East Asian Countries (Japan, South Korea, Taiwan, and China) than other parts of the world [2]. Based on these reports, HPV 52 prophylactic vaccine is suggested to be introduced in Indonesia, in order to curtail the prevailing human papillomavirus
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