Prokaryotic Expression And Immunogenicity Analysis Of HPV16L2Peptide Coupled To AP205VLP

Human Papillomaviruses(HPV) infecting mainly human cutaneous and mucosa in a strict species-and tissue-specific manner and causing proliferative lesions, are non-enveloped viruses with a double-stranded DNA genome of7.8to8kilobases.More than100HPV types have been defined, and many of these HPV types are ubiquitous and globally distributed.HPV are divided into two groups: low-risk types(such as types of6and11) found mainly in benign condyloma and15high-risk types (types of16,18,31,33,35,39,45,51,52,56,58,59,66,68, and73) found in cervical cancer.The presence of HPV DNA has been reported in99.7%of cervical carcinomas worldwide, indicating that HPV infection is a necessary cause of this cancer and HPV16being the most prevalent types detected in cervical carcinoma. An icosahedral HPV capsid is composed of major capsid protein L1and minor capsid protein L2.The minor capsid proteinL2is expressed in all types of HPV and it has strong sequence’s conventionality, so it can cross-immunize different HPV genotypes infections.AP205has been recently identified as a new RNA bacteriophage infecting the Gram-negative bacteria Acinetobacter sp.The AP205protein is the major capsid protein from the RNA bacteriophage AP205, made from180subunits assembling into an icosahedral capsid in E. Coli.Since the AP205Virus-Like Particles(VLPs) are not contain RNA genome,it can’t replication.Epitopes conjugated chemically to these VLPs are displayed in a highly repetitive and organized manner, inducing strong immune responses in all species tested.In this study,the AP205coat protein gene and HPV16L2gene were connected by PCR and cloned into vector pET-30a (+) after being synthesized artificially.The constructed plasmid pET30-AP205-HPV16△L2was transformed to E.coli BL21(DH3), the recombinant protein was expressed under induction of IPTG and purified by Gel chromatography, then identified through SDS-PAGE and Western blot. The immunogenicity of purified product was measured by indirect ELISA after ICR mouse was immunized. The results show the protein was expressed solubly in E.coli BL21(DH3). The typical VLPs can be observed through Transmission electron microscopy which Showed good immunogenicity in animal experiments.