Expression and Purification of Porcine Rotavirus Structural Proteins in Silkworm Larvae as a Vaccine Candidate

作者：Kato, Tatsuya^*; Kakuta, Tatsuki; Yonezuka, Ami; Sekiguchi, Tomofumi; Machida, Yuki; Xu, Jian; Suzuki, Tohru; Park, Enoch Y.

来源：MOLECULAR BIOTECHNOLOGY, 2023, 65(3): 401-409.

DOI：10.1007/s12033-022-00548-3

• Porcine rotavirus
• VP6
• VP7
• Silkworm larvae
• Vaccine

In this study, silkworm larvae were used for expression of porcine rotavirus A (KS14 strain) inner capsid protein, VP6, and outer capsid protein, VP7. Initially, VP6 was fused with Strep-tag II and FLAG-tag (T-VP6), and T-VP6 was fused further with the signal peptide of Bombyx mori 30k6G protein (30k-T-VP6). T-VP6 and 30 k-T-VP6 were then expressed in the fat body and hemolymph of silkworm larvae, respectively, with respective amounts of 330 mu g and 50 mu g per larva of purified protein. Unlike T-VP6, 30k-T-VP6 was N-glycosylated due to attached signal peptide. Also, VP7 was fused with PA-tag (VP7-PA). Additionally, VP7 was fused with Strep-tag II, FLAG-tag, and the signal peptide of Bombyx mori 30k6G protein (30k-T-Delta VP7). Both VP7-PA and 30k-T-Delta VP7 were expressed in the hemolymph of silkworm larvae, with respective amounts of 26 mu g and 49 mu g per larva of purified protein, respectively. The results from our study demonstrated that T-VP6 formed nanoparticles of greater diameter compared with the ones formed by 30k-T-VP6. Also, higher amount of VP6 expressed in silkworm larvae reveal that VP6 holds the potential for its use in vaccine development against porcine rotavirus with silkworm larvae as a promising host for the production of such multi-subunit vaccines.