Development of a tag-free plant-made interferon gamma production system with improved therapeutic efficacy against viruses

Min-Chao Jiang; Wei-Li Hsu; Ching-Yu Tseng; Na-Sheng Lin; Yau-Heiu Hsu; Chung-Chi Hu

doi:10.3389/fbioe.2023.1341340

Development of a tag-free plant-made interferon gamma production system with improved therapeutic efficacy against viruses

Front Bioeng Biotechnol. 2024 Jan 11:11:1341340. doi: 10.3389/fbioe.2023.1341340. eCollection 2023.

Authors

Min-Chao Jiang¹, Wei-Li Hsu², Ching-Yu Tseng², Na-Sheng Lin³, Yau-Heiu Hsu^{4

5}, Chung-Chi Hu^{4

5}

Affiliations

¹ PhD Program in Microbial Genomics, National Chung Hsing University and Academia Sinica, Taichung, Taiwan.
² Graduate Institute of Microbiology and Public Health, College of Veterinary Medicine, National Chung Hsing University, Taichung, Taiwan.
³ Institute of Plant and Microbial Biology, Academia Sinica, Taipei, Taiwan.
⁴ Graduate Institute of Biotechnology, National Chung Hsing University, Taichung, Taiwan.
⁵ Advanced Plant Biotechnology Center, National Chung Hsing University, Taichung, Taiwan.

Abstract

Plants offer a promising platform for cost-effective production of biologically active therapeutic glycoproteins. In previous studies, we have developed a plant expression system based on Bamboo mosaic virus (BaMV) by incorporating secretory signals and an affinity tag, which resulted in notably enhanced yields of soluble and secreted fusion glycoproteins (FGs) in Nicotiana benthamiana. However, the presence of fusion tags on recombinant glycoproteins is undesirable for biomedical applications. This study aimed to develop a refined expression system that can efficiently produce tag-free glycoproteins in plants, with enhanced efficacy of mature interferon gamma (mIFNγ) against viruses. To accommodate the specific requirement of different target proteins, three enzymatically or chemically cleavable linkers were provided in this renovated BaMV-based expression system. We demonstrated that Tobacco etch virus (TEV) protease could process the specific cleavage site (L_TEV) of the fusion protein, designated as SS^ExtHis(SP)₁₀L_TEV-mIFNγ, with optimal efficiency under biocompatible conditions to generate tag-free mIFNγ glycoproteins. The TEV protease and secretory-affinity tag could be effectively removed from the target mIFNγ glycoproteins through Ni²⁺-NTA chromatography. In addition, the result of an antiviral assay showed that the tag-free mIFNγ glycoproteins exhibited enhanced biological properties against Sindbis virus, with comparable antiviral activity of the commercialized HEK293-expressed hIFNγ. Thus, the improved BaMV-based expression system developed in this study may provide an alternative strategy for producing tag-free therapeutic glycoproteins intended for biomedical applications.

Keywords: Bamboo mosaic virus (BaMV); antivirus activity; cleavable peptide; tag-free IFNγ; therapeutic protein; viral vector.

Grants and funding

The author(s) declare that no financial support was received for the research, authorship, and/or publication of this article. This work was financially supported in part by the Advanced Plant and Food Crop Biotechnology Center from The Featured Areas Research Center Program within the framework of the Higher Education Sprout Project by the Ministry of Education (MOE) in Taiwan and by the Ministry of Science and Technology, Taiwan (MOST 111-2313-B-005-051 and MOST 111-2313-B-005-004).