Expression of Muscovy Duck Parvovirus Capsid Proteins (VP2 and VP3) in a Baculovirus Expression System and Demonstration of Immunity Induced by the Recombinant Proteins

The baculovirus expression vector system (BEVS) has been widely used to produce a large number of recombinant proteins, and is becoming one of the most powerful, robust, and cost-effective systems for the production of eukaryotic proteins. Nevertheless, as in any other protein expression system, it is important to improve the production capabilities of this vector. The orf46 viral gene was identified among the most highly abundant sequences in the transcriptome of Spodoptera exigua larvae infected with its native baculovirus, the S. exigua multiple nucleopolyhedrovirus (SeMNPV). Different sequences upstream of the orf46 gene were cloned, and their promoter activities were tested by the expression of the GFP reporter gene using the Autographa californica nucleopolyhedrovirus (AcMNPV) vector system in different insect cell lines (Sf21, Se301, and Hi5) and in larvae from S. exigua and Trichoplusia ni. The strongest promoter activity was defined by a 120 nt sequence upstream of the ATG start codon for the orf46 gene. On average, GFP expression under this new promoter was more than two fold higher than the expression obtained with the standard polyhedrin (pph) promoter. Additionally, the orf46 promoter was also tested in combination with the pph promoter, revealing an additive effect over the pph promoter activity. In conclusion, this new characterized promoter represents an excellent alternative to the most commonly used baculovirus promoters for the efficient expression of recombinant proteins using the BEVS.

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Propagation ofSpodoptera frugiperda cells (Sf9) and production of recombinant proteins with the baculovirus expression system using improved spinner flasks with membrane aeration

Biotechnology Techniques ◽

10.1007/bf00160832 ◽

1995 ◽

Vol 9 (6) ◽

pp. 435-440 ◽

Cited By ~ 7

Author(s):

Volker J�ger ◽

Anja Kobold

Keyword(s):

Recombinant Proteins ◽

Expression System ◽

Baculovirus Expression System ◽

Baculovirus Expression ◽

Membrane Aeration

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Analysis of the capsid processing strategy of Thosea asigna virus using baculovirus expression of virus-like particles

Journal of General Virology ◽

10.1099/0022-1317-82-1-259 ◽

2001 ◽

Vol 82 (1) ◽

pp. 259-266 ◽

Cited By ~ 15

Author(s):

Fiona M. Pringle ◽

James Kalmakoff ◽

Vernon K. Ward

Keyword(s):

Expression System ◽

Baculovirus Expression System ◽

Terminal Region ◽

Precursor Protein ◽

Capsid Proteins ◽

Processing Strategy ◽

Structural Protein ◽

Virus Like Particles ◽

Capsid Precursor ◽

Baculovirus Expression

Thosea asigna virus (TaV), a putative member of the genus Betatetravirus of the family Tetraviridae, is predicted to have a novel capsid expression strategy compared with other characterized tetraviruses. The capsid precursor protein is cleaved twice to generate three proteins. Two of the proteins, L (58·3 kDa) and S (6·8 kDa), are incorporated into the TaV virion. The third, non-structural protein, produced from the N terminus of the precursor protein, is up to 17 kDa in size and is of unknown function. The TaV capsid precursor protein sequence without the 17 kDa N-terminal region was modelled against the solved structure from Nudaurelia ω virus (NωV) using SwissModel. The TaV model was very similar to the solved structure determined for subunit A of NωV and had features that are conserved between tetraviruses and nodaviruses, including the positioning of the cleavage site between the L and S capsid proteins. The production of virus-like particles (VLPs) using the baculovirus expression system was used to analyse the capsid processing strategy employed by TaV. VLPs were formed in both the presence and absence of the 17 kDa N-terminal region of the capsid precursor. VLPs were not formed when the L and S regions were expressed from separate promoters, indicating that cleavage between the L and S capsid proteins was an essential part of TaV capsid assembly. Expression of the TaV 17 kDa protein in bacteria did not produce intracellular tubules similar to those formed by bacterial expression of the p17 protein from Helicoverpa armigera stunt virus.

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A novel baculovirus-derived promoter with high activity in the Baculovirus Expression System

10.7287/peerj.preprints.2024 ◽

2016 ◽

Author(s):

María Martínez-Solís ◽

Silvia Gómez-Sebastián ◽

José M Escribano ◽

Agata K Jakubowska ◽

Salvador Herrero

Keyword(s):

Recombinant Proteins ◽

Trichoplusia Ni ◽

Promoter Activity ◽

Expression System ◽

Cost Effective ◽

Baculovirus Expression System ◽

Start Codon ◽

Vector System ◽

Viral Gene ◽

Baculovirus Expression

The baculovirus expression vector system (BEVS) has been widely used to produce a large number of recombinant proteins, and is becoming one of the most powerful, robust, and cost-effective systems for the production of eukaryotic proteins. Nevertheless, as in any other protein expression system, it is important to improve the production capabilities of this vector. The orf46 viral gene was identified among the most highly abundant sequences in the transcriptome of Spodoptera exigua larvae infected with its native baculovirus, the S. exigua multiple nucleopolyhedrovirus (SeMNPV). Different sequences upstream of the orf46 gene were cloned, and their promoter activities were tested by the expression of the GFP reporter gene using the Autographa californica nucleopolyhedrovirus (AcMNPV) vector system in different insect cell lines (Sf21, Se301, and Hi5) and in larvae from S. exigua and Trichoplusia ni. The strongest promoter activity was defined by a 120 nt sequence upstream of the ATG start codon for the orf46 gene. On average, GFP expression under this new promoter was more than two fold higher than the expression obtained with the standard polyhedrin (pph) promoter. Additionally, the orf46 promoter was also tested in combination with the pph promoter, revealing an additive effect over the pph promoter activity. In conclusion, this new characterized promoter represents an excellent alternative to the most commonly used baculovirus promoters for the efficient expression of recombinant proteins using the BEVS.

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A novel baculovirus-derived promoter with high activity in the baculovirus expression system

PeerJ ◽

10.7717/peerj.2183 ◽

2016 ◽

Vol 4 ◽

pp. e2183 ◽

Cited By ~ 6

Author(s):

María Martínez-Solís ◽

Silvia Gómez-Sebastián ◽

José M. Escribano ◽

Agata K. Jakubowska ◽

Salvador Herrero

Keyword(s):

Recombinant Proteins ◽

Trichoplusia Ni ◽

Promoter Activity ◽

Expression System ◽

Cost Effective ◽

Baculovirus Expression System ◽

Start Codon ◽

Vector System ◽

Viral Gene ◽

Baculovirus Expression

The baculovirus expression vector system (BEVS) has been widely used to produce a large number of recombinant proteins, and is becoming one of the most powerful, robust, and cost-effective systems for the production of eukaryotic proteins. Nevertheless, as in any other protein expression system, it is important to improve the production capabilities of this vector. Theorf46viral gene was identified among the most highly abundant sequences in the transcriptome ofSpodoptera exigualarvae infected with its native baculovirus, theS. exiguamultiple nucleopolyhedrovirus (SeMNPV). Different sequences upstream of theorf46gene were cloned, and their promoter activities were tested by the expression of the GFP reporter gene using theAutographa californicanucleopolyhedrovirus (AcMNPV) vector system in different insect cell lines (Sf21, Se301, and Hi5) and in larvae fromS. exiguaandTrichoplusia ni. The strongest promoter activity was defined by a 120 nt sequence upstream of the ATG start codon for theorf46gene. On average, GFP expression under this new promoter was more than two fold higher than the expression obtained with the standard polyhedrin (polh) promoter. Additionally, theorf46promoter was also tested in combination with the polh promoter, revealing an additive effect over the polh promoter activity. In conclusion, this new characterized promoter represents an excellent alternative to the most commonly used baculovirus promoters for the efficient expression of recombinant proteins using the BEVS.

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