Title:Engineering of Chimeric Protein Based on E Protein Domain III of Tick- Borne Encephalitis Virus and OmpF Porin of Yersinia pseudotuberculosis
Volume: 24
Issue: 10
Author(s): Anna M. Stenkova*, Natalia S. Chopenko, Ludmila A. Davydova, Andrey N. Mazeika, Evgeniya P. Bystritskaya, Olga Y. Portnyagina, Stanislav D. Anastyuk, Dmitrii S. Kulbatskii, Ekaterina N. Lyukmanova, Dmitriy A. Dolgikh, Eduard Y. Kostetsky and Nina M. Sanina
Affiliation:
- Far Eastern Federal University, Vladivostok,Russian Federation
Keywords:
Domain III, envelope protein, flavivirus vaccine, OmpF porin, TBEV, tick-borne encephalitis.
Abstract: Background: Tick-borne encephalitis poses a serious public health threat in the endemic
regions. The disease treatment is restricted to symptomatic therapy, so great expectations are in the
development of the prophylactic and therapeutic vaccines. The domain III of E protein of the tickborne
encephalitis virus is the main antigenic domain which includes virus-specific epitopes recognized
by neutralizing antibodies.
Objectives: The main objective of this study was to design, express, isolate and characterize the
chimeric protein based on the fusion of domain III of E protein of the tick-borne encephalitis virus
and bacterial porin OmpF from Yersinia pseudotuberculosis.
Methods: The chimeric gene was obtained by the PCR based fusion method from two fragments
containing overlapping linker sequences. Resulting plasmids were transformed into BL21(DE3)
pLysS electrocompetent cells for subsequent heterologous protein expression. All recombinant
proteins were purified using immobilized metal affinity chromatography under denaturing conditions.
The identity of the chimeric protein was confirmed by MALDI-TOF mass spectrometry and
immunoblot analysis. The content of antibodies against the EIII protein was estimated in mice
blood serum by ELISA.
Results: The bacterial partner protein was used for decreasing toxicity and increasing immunogenicity
of antigen. The chimeric protein was successfully expressed by the Escherichia coli cells.
The purified protein was recognized with immunoblots by anti-E protein of tick-borne encephalitis
virus monoclonal antibodies. Furthermore, the protein was able to elicit antibody response against
domain III of E protein in immunized mice.
Conclusion: The newly obtained chimeric antigen could be valuable for the development of the
preventing tick-borne encephalitis subunit vaccines.
