Generation of oligomers of subunit vaccine candidate glycoprotein D of Herpes Simplex Virus-2 expressed in fusion with I
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ORIGINAL ARTICLE
Generation of oligomers of subunit vaccine candidate glycoprotein D of Herpes Simplex Virus‑2 expressed in fusion with IgM Fc domain(s) in Escherichia coli: A strategy to enhance the immunogenicity of the antigen Vikas Kumar Singh1 · Sandeep Kumar1 · Rajeev Kumar Dhaked2 · Abdul S. Ansari2 · Nirmal K. Lohiya3 · Suman Tapryal1 Received: 19 June 2020 / Accepted: 23 September 2020 © King Abdulaziz City for Science and Technology 2020
Abstract Glycoprotein D (gD) of Herpes Simplex Virus-2 is used as an antigen in various anti-herpes subunit vaccines owing to its involvement in binding the host cell receptors for host infectivity. However, most of these monomeric protein based candidates have shown low immunogenicity in animal models. To enhance the immunogenicity of gD, a fresh approach of fusing its ectodomain with the Fc domain(s) of IgM has been adopted to oligomerize the viral antigen and to exploite the immune-modulating potential of IgM Fc. Six vaccine constructs, generated by fusing three gD-ectodomain-length-variants with the Ig µ-chain domain 4 (µCH4) and µCH3-CH4 fragment, were cloned in Escherichia coli using pET28b( +) vector. The vaccine proteins were expressed in the form of inclusion bodies (IBs) and were in vitro refolded into protein oligomers of high stoichiometries of ~ 15–24, with 70–80% refolding yields. The conformations of gD and Fc components of the refolded oligomers were analyzed by ELISA and CD spectroscopy and were found to be native-like. The sizes and profiles of the size-distribution of oligomers were determined by dynamic light scattering (DLS). The candidate C2 (gD-μCH3-CH4), showing the most compact oligomer size and uniform distribution of its particles was chosen as the suitable candidate for mice immunization studies to assess the immunogenicity of the antigen gD. The C2 oligomer stimulated a strong anti-gD humoral response with an antibody titer of 102,400 and a strong, biased Th1 immune response in C57BL/6 mice, indicating its potential as a strong immunogen which may serve as an effective vaccine candidate. Keywords Glycoprotein D · Herpes simplex virus-2 · Fusion protein · Vaccine · In vitro refolding
Introduction
Electronic supplementary material The online version of this article (https://doi.org/10.1007/s13205-020-02452-6) contains supplementary material, which is available to authorized users. * Suman Tapryal [email protected] 1
Department of Biotechnology, School of Life Sciences, Central University of Rajasthan, NH‑8, Bandar Sindri, Ajmer, Rajasthan, India 305817
2
Department of Zoology, Center for Advanced Studies, University of Rajasthan, Jaipur, Rajasthan, India 302004
3
Indian Society for the Study of Reproduction and Fertility, Department of Zoology, Center for Advanced Studies, University of Rajasthan, Jaipur, Rajasthan, India 302004
Herpes viruses are a leading cause of human viral diseases, second to influenza and cold viruses (Zhu and Zhu 2014). In the family Herpesviridae, at least eight virus types are
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