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Recombinant occlusion bodies of baculovirus as carriers of a non-structural protein of foot-and-mouth disease virus.

ABSTRACT: Here, we developed a diagnostic ELISA for foot-and-mouth disease using recombinant occlusion bodies (rOBs) of baculovirus. We fused ?3AB1-3, a polypeptide derived from non-structural proteins of foot-and-mouth disease virus, to polyhedrin (POLH), the major constituent of OBs, under polh promoter. To further assess the most convenient strategy to improve yields, we designed two recombinant baculoviruses, vPOLH and vPOLHE44G. These carried the sequence of the fusion protein POLH-?3AB1-3 with an additional copy in cis of polh or polh E44G , respectively, under p10 promoter. Our results show that both viruses expressed POLH-?3AB1-3, which was detected by western blot in purified rOBs with anti-POLH and anti-3AB1 antibodies. We also found that vPOLHE44G produced larger polyhedra and a significant increase of antigen yield (p?

SUBMITTER: Diez M

PROVIDER: S-EPMC6195870 | biostudies-literature | 2018 Nov

REPOSITORIES: biostudies-literature

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Recombinant occlusion bodies of baculovirus as carriers of a non-structural protein of foot-and-mouth disease virus.

Diez Michay M Trotta Myrian M Alfonso Victoria V Taboga Oscar O López María Gabriela MG

3 Biotech 20181020 11

Here, we developed a diagnostic ELISA for foot-and-mouth disease using recombinant occlusion bodies (rOBs) of baculovirus. We fused Δ3AB1-3, a polypeptide derived from non-structural proteins of foot-and-mouth disease virus, to polyhedrin (POLH), the major constituent of OBs, under polh promoter. To further assess the most convenient strategy to improve yields, we designed two recombinant baculoviruses, vPOLH and vPOLHE44G. These carried the sequence of the fusion pr ...[more]

PMID: 30370198

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Project description:The relatively low infectivity of baculoviruses to their host larvae limits their use as insecticidal agents on a larger scale. In the present study, a novel strategy was developed to efficiently embed foreign proteins into Autographa californica multiple nucleopolyhedrovirus (AcMNPV) occlusion bodies (OBs) to achieve stable expression of foreign proteins and to improve viral infectivity. A recombinant AcMNPV bacmid was constructed by expressing the 150-amino-acid (aa) N-terminal segment of polyhedrin under the control of the p10 promoter and the remaining C-terminal 95-aa segment under the control of the polyhedrin promoter. The recombinant virus formed OBs in Spodoptera frugiperda 9 cells, in which the occlusion-derived viruses were embedded in a manner similar to that for wild-type AcMNPV. Next, the 95-aa polyhedrin C terminus was fused to enhanced green fluorescent protein, and the recombinant AcMNPV formed fluorescent green OBs and was stably passaged in vitro and in vivo The AcMNPV recombinants were further modified by fusing truncated Agrotis segetum granulovirus enhancin or truncated Cydia pomonella granulovirus ORF13 (GP37) to the C-terminal 95 aa of polyhedrin, and both recombinants were able to form normal OBs. Bioactivity assays indicated that the median lethal concentrations of these two AcMNPV recombinants were 3- to 5-fold lower than that of the control virus. These results suggest that embedding enhancing factors in baculovirus OBs by use of this novel technique may promote efficient and stable foreign protein expression and significantly improve baculovirus infectivity.IMPORTANCE Baculoviruses have been used as bioinsecticides for over 40 years, but their relatively low infectivity to their host larvae limits their use on a larger scale. It has been reported that it is possible to improve baculovirus infectivity by packaging enhancing factors within baculovirus occlusion bodies (OBs); however, so far, the packaging efficiency has been low. In this article, we describe a novel strategy for efficiently embedding foreign proteins into AcMNPV OBs by expressing N- and C-terminal (dimidiate) polyhedrin fragments (150 and 95 amino acids, respectively) as fusions to foreign proteins under the control of the p10 and polyhedrin promoters, respectively. When this strategy was used to embed an enhancing factor (enhancin or GP37) into the baculovirus OBs, 3- to 5-fold increases in baculoviral infectivity were observed. This novel strategy has the potential to create an efficient protein expression system and a highly efficient virus-based system for insecticide production in the future.

Dataset Information

Recombinant occlusion bodies of baculovirus as carriers of a non-structural protein of foot-and-mouth disease virus.

Publications

Recombinant occlusion bodies of baculovirus as carriers of a non-structural protein of foot-and-mouth disease virus.

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