Abstract:

Objective To obtain antigenic epitopes of human papillomavirus（HPV） type 16 L1 protein expression and purification in prokaryotic system,and to evaluate its serological reaction. Methods The nucleotide sequence of HPV type 16 L1 protein antigenic epitopes were synthesized and cloned into prokaryotic expression vector pET-DsbC. The constructed recombinant plasmid pET-DsbC-HPV16 L1 was transformed to Escherichia coli BL21 Rosetta（DE3） for expression under the induction of isopropyl-beta-D-thiogalactoside（IPTG）. The L1 protein was expressed and purified by metal chelate affinity chromatography. The antigenicity and immunological activity of fusion protein were evaluated with clinical serum samples by western blotting and enzyme-linked immunosorbent assay（ELISA）. Results The expression vector pET-DsbC-HPV16 L1 was successfully constructed and stably expressed in Escherichia coli Rosetta （DE3）. The recombinant HPV16 L1 protein was purified by affinity chromatography,which had a relative molecular mass of 45 000 and a purity of 95%. The purified rate of recombinant protein was 22%。The results of western blotting showed that the recombinant DsbC-HPV16 L1 protein could produce specific antigen-antibody reaction with HPV16 positive serum,but there was no cross reaction with recombinant DsbC protein or healthy human serum,which had good specificity. ELISA results also indicated that recombinant protein had a higher level of reactivity to serum from patients than serum from healthy or phosphate-buffered saline（PBS）,the A₄₅₀ values were 0.56,0.24 and 0.21,respectively. Conclusions HPV type 16 L1 protein antigentic epitopes are overexpressed solubly in prokaryotic system and have a high immunogenicity and immunological activity.

Key words: Human papillomavirus type 16 L1 protein, Prokaryotic expression, Immunogenicity, Serological reaction