Synthesis of complex-type N-glycans on recombinant proteins produced by silkworm-baculovirus expression system

2016 ◽  
Author(s):  
Daisuke Morokuma
Keyword(s):  
Recombinant Proteins ◽  
Expression System ◽  
Baculovirus Expression System ◽  
Complex Type ◽  
Baculovirus Expression

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

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.

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

PeerJ ◽  
2016 ◽  
Vol 4 ◽  
pp. e2183 ◽  
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.

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

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.

scholarly journals Identification of Drosophila melanogaster yellow-f and yellow-f2 proteins as dopachrome-conversion enzymes

2002 ◽  
Vol 368 (1) ◽  
pp. 333-340 ◽  
Author(s):  
Qian HAN ◽  
Jianmin FANG ◽  
Haizhen DING ◽  
Jody K. JOHNSON ◽  
Bruce M. CHRISTENSEN ◽  
...  
Keyword(s):  
Enzyme Activity ◽  
Drosophila Melanogaster ◽  
Recombinant Proteins ◽  
Adult Development ◽  
Expression System ◽  
Baculovirus Expression System ◽  
Structural Rearrangement ◽  
High Abundance ◽  
Northern Hybridization ◽  
Baculovirus Expression

This study describes the identification of Drosophila yellow-f and yellow-f2 as dopachrome-conversion enzymes responsible for catalysing the conversion of dopachrome into 5,6-dihydroxyindole in the melanization pathway. Drosophilayellow-y gene and yellow-b, -c, -f and -f2 genes were expressed in an insect cell/baculovirus expression system and their corresponding recombinant proteins were screened for dopachrome-conversion enzyme activity. Among the yellow and yellow-related genes, the yellow-f and yellow-f2 genes were identified as the genes coding for Drosophila dopachrome-conversion enzyme based on the high activity of their recombinant proteins in catalysing the production of 5,6-dihydroxyindole from dopachrome. Both yellow-f and yellow-f2 are capable of mediating a decarboxylative structural rearrangement of dopachrome, as well as an isomerization/tautomerization of dopamine chrome and dopa methyl ester chrome. Northern hybridization revealed the transcription of yellow-f in larvae and pupae, but a high abundance of mRNA was observed in later larval and early pupal stages. In contrast, yellow-f2 transcripts were present at all stages, but high abundance of its mRNA was observed in later-stage pupae and adults. These data indicate that yellow-f and yellow-f2 complement each other during Drosophila development and that the yellow-f is involved in larval and pupal melanization, and yellow-f2 plays a major role in melanization reactions in Drosophila during later pupal and adult development. Results from this study provide the groundwork towards a better understanding of the physiological roles of the Drosophilayellow gene family.

scholarly journals Expression of functional G protein beta gamma dimers of defined subunit composition using a baculovirus expression system.

1992 ◽  
Vol 267 (19) ◽  
pp. 13123-13126 ◽  
Author(s):  
S.G. Graber ◽  
R.A. Figler ◽  
V.K. Kalman-Maltese ◽  
J.D. Robishaw ◽  
J.C. Garrison
Keyword(s):  
G Protein ◽  
Expression System ◽  
Baculovirus Expression System ◽  
Subunit Composition ◽  
Baculovirus Expression
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