Efficient silkworm expression of human GPCR (nociceptin receptor) by a Bombyx mori bacmid DNA system

Guanine nucleotide-binding protein (G protein) coupled receptors (GPCRs) are frequently expressed by a baculovirus expression vector system (BEVS). We recently established a novel BEVS using the bacmid system of Bombyx mori nucleopolyhedrovirus (BmNPV), which is directly applicable for protein expre...

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Bibliographic Details
Published in:Biochemical and biophysical research communications 2009-07, Vol.385 (3), p.375-379
Main Authors: Kajikawa, Mizuho, Sasaki, Kaori, Wakimoto, Yoshitaro, Toyooka, Masaru, Motohashi, Tomoko, Shimojima, Tsukasa, Takeda, Shigeki, Park, Enoch Y., Maenaka, Katsumi
Format: Article
Language:English
Subjects:
60 APPLIED LIFE SCIENCES
Animals
Bacmid
Baculovirus
BmNPV
Bombyx - genetics
Bombyx - metabolism
Bombyx - virology
Bombyx mori
Bombyx mori NPV
CENTRIFUGATION
DNA
DNA - genetics
Genetic Vectors - genetics
GPCR
GTP-ASES
GUANINE
Humans
INJECTION
LARVAE
Membrane protein
Nucleopolyhedrovirus - genetics
NUCLEOTIDES
Protein Biosynthesis
Protein expression
RECEPTORS
Receptors, G-Protein-Coupled - biosynthesis
Receptors, G-Protein-Coupled - genetics
Receptors, Opioid - biosynthesis
Receptors, Opioid - genetics
Recombinant Fusion Proteins - biosynthesis
Recombinant Fusion Proteins - genetics
SILKWORM
STIMULATION
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Summary:Guanine nucleotide-binding protein (G protein) coupled receptors (GPCRs) are frequently expressed by a baculovirus expression vector system (BEVS). We recently established a novel BEVS using the bacmid system of Bombyx mori nucleopolyhedrovirus (BmNPV), which is directly applicable for protein expression in silkworms. Here, we report the first example of GPCR expression in silkworms by the simple injection of BmNPV bacmid DNA. Human nociceptin receptor, an inhibitory GPCR, and its fusion protein with inhibitory G protein alpha subunit (Giα) were both successfully expressed in the fat bodies of silkworm larvae as well as in the BmNPV viral fraction. Its yield was much higher than that from Sf9 cells. The microsomal fractions including the nociceptin receptor fusion, which are easily prepared by only centrifugation steps, exhibited [35S]GTPγS-binding activity upon specific stimulation by nociceptin. Therefore, this rapid method is easy-to-use and has a high expression level, and thus will be an important tool for human GPCR production.
ISSN:0006-291X
1090-2104
DOI:10.1016/j.bbrc.2009.05.063