Efficient application of a baculovirus-silkworm larvae expression system for obtaining porcine circovirus type 2 virus-l
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ORIGINAL ARTICLE
Efficient application of a baculovirus‑silkworm larvae expression system for obtaining porcine circovirus type 2 virus‑like particles for a vaccine Qianhua He1 · Zhenming Cao1 · Pengwei Wang1 · Qiuyuan Lu1 · Hao Zheng1 · Jingchen Sun1 Received: 21 October 2019 / Accepted: 24 June 2020 © Springer-Verlag GmbH Austria, part of Springer Nature 2020
Abstract Porcine circovirus type 2 (PCV2) is a major pathogen associated with swine diseases. It is the smallest single-stranded DNA virus, and its genome contains four major open reading frames (ORFs). ORF2 encodes the major structural protein Cap, which can self-assemble into virus-like particles (VLPs) in vitro and contains the primary antigenic determinants. In this study, we developed a high-efficiency method for obtaining VLPs and optimized the purification conditions. In this method, we expressed the protein Cap with a 6× His tag using baculovirus-infected silkworm larvae as well as the E. coli BL21(DE3) prokaryotic expression system. The PCV2 Cap proteins produced by the silkworm larvae and E. coli BL21(DE3) were purified. Cap proteins purified from silkworm larvae self-assembled into VLPs in vitro, while the Cap proteins purified from bacteria were unable to self-assemble. Transmission electron microscopy confirmed the self-assembly of VLPs. The immunogenicity of the VLPs produced using the baculovirus system was demonstrated using an enzyme-linked immunosorbent assay (ELISA). Furthermore, the purification process was optimized. The results demonstrated that the expression system using baculovirus-infected silkworm larvae is a good choice for obtaining VLPs of PCV2 and has potential for the development of a low-cost and efficient vaccine.
Introduction Porcine circovirus type 2 (PCV2) is the smallest singlestranded DNA virus [7, 9] and it is a member of the family Circoviridae. The first porcine circovirus (PCV) was Handling Editor: Roman Pogranichniy. Qianhua He and Zhenming Cao contributed equally to this work. Electronic supplementary material The online version of this article (https://doi.org/10.1007/s00705-020-04754-9) contains supplementary material, which is available to authorized users.
reported in Canada and the associated disease was described as a PCV disease [11, 21]. There are three types of PCVs. PCV1 and PCV2 were first found in the 1970s and the 1990s, respectively, while PCV3 was first found in 2016 [11, 26, 37]. The PCV2 genome consists of only 1.7 kb, and it contains four major open reading frames (ORFs). These ORFs encode two proteins associated with replication, a structural protein that forms the viral capsid, a protein that induces apoptosis of host cells, and a protein that inhibits caspase activity and regulates CD4 and CD8T lymphocytes [13, 31]. However, PCV2 lacks transcription factors, which means that the replication of PCV2 strongly relies on host
* Hao Zheng [email protected]
Qiuyuan Lu [email protected]
* Jingchen Sun [email protected]
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Qianhua He [email protected] Zhenming Cao [email protected]
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