Expression and Purification of Porcine Rotavirus Structural Proteins in Silkworm Larvae as a Vaccine Candidate.
Tatsuya KatoTatsuki KakutaAmi YonezukaTomofumi SekiguchiYuki MachidaJian XuTohru SuzukiEnoch Y ParkPublished in: Molecular biotechnology (2022)
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 μg and 50 μg 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-ΔVP7). Both VP7-PA and 30k-T-ΔVP7 were expressed in the hemolymph of silkworm larvae, with respective amounts of 26 μg and 49 μg 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.