Biotechnology and Bioprocess Engineering 2024; 29(3): 589-600  
Engineering potyvirus-like particles to display multiple copies of tuberculosis antigens
R. Princess1 · M. L. Stephen Raj1
1 Department of Biotechnology, Mepco Schlenk Engineering College (Autonomous), Mepco Engineering College P.O., Mepco Nagar, Sivakasi, Tamilnadu 626005, India
Correspondence to: M. L. Stephen Raj
stephenraj@mepcoeng.ac.in
Received: December 14, 2023; Revised: February 17, 2024; Accepted: February 21, 2024; Published online: March 11, 2024.
© The Korean Society for Biotechnology and Bioengineering. All rights reserved.

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
Abstract
Elicitation of antibody and cell-mediated immune responses are crucial for successful vaccine development against tuberculosis (TB). Mycobacterium tuberculosis (Mtb) antigens CFP10 and ESAT6, potent and proven vaccine candidates require appropriate adjuvants to trigger better immune response. Virus-like particles carrying repetitive copies of foreign antigens can induce both T and B cell-mediated immunity required for conferring protection against intracellular pathogens. In this study, we developed hybrid potyvirus-like particles (PVLPs) displaying mycobacterial antigens on their surface by translationally fusing the coat protein (CP) gene derived from Johnson grass mosaic virus with CFP 10 or/and ESAT 6 gene(s). The recombinant plasmids carrying fusion constructs were transformed into Escherichia coli, the fusion proteins, viz. ESAT6-CP, CP-CFP10 and ESAT6-CP-CFP10, were expressed and purified using Ni-NTA2+ affinity chromatography under denaturing conditions. The chimeric CP fusion proteins were self-assembled in vitro into PVLPs by the gradual removal of denaturing conditions. The purified hybrid PVLPs carrying Mtb antigens when injected into mice showed enhanced immunogenicity for both ESAT6 and CFP10 antigens compared to the same antigens immunized without any adjuvant. In vitro stimulation of splenocytes derived from mice immunized with chimeric PVLPs upregulates the expression of cytokines involved in TB immune response.
Keywords: Tuberculosis · CFP10 · ESAT6 · Johnson grass mosaic virus · Potyvirus-like particles

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