High level expression of C-terminal truncated recombinant chicken interferon-gamma in baculovirus vector system.
Chicken interferon-gamma (ChIFN-gamma) was expressed by baculovirus in a C-terminal truncated form, namely ChIFN-gammaT, to accelerate the secretion of the expressed protein. It is also expressed as ChIFN-gammaT bearing poly His tag, ChIFN-gammaTHis, for easy purification. The expressed proteins were detected by SDS-PAGE analysis with Coomassie brilliant blue staining. The purified ChIFN-gammaTHis with nickel chelated column showed anti-viral activity in vitro and stimulation of the secretion of nitrogen intermediates such as nitric oxide in chicken peripheral blood mononuclear cells. Antiserum against ChIFN-gammaTHis recognized the 15 kDa, 16 kDa, and 32 kDa bands that seemed to be an unglycosylated monomer, a glycosylated monomer, and a homodimer of ChIFN-gammaTHis in the culture supernatant, respectively. The anti-serum also recognized around 14 kDa and 28 kDa bands in the sera of chickens or concanavalin A stimulated spleen cell culture supernatants that seemed to be monomeric and dimeric forms of a natural ChIFN-gamma, respectively.
Takehara
Kazuaki
K
Laboratory of Poultry Diseases, School of Veterinary Medicine and Animal Sciences, Kitasato University, Towada, Aomori, Japan.
Kamikawa
Makiyo
M
Ohnuki
Naomi
N
Nagata
Tomoshi
T
Nakano
Ayako
A
Yamaguchi
Daisuke
D
Yokomizo
Yuichi
Y
Nakamura
Masayuki
M
D000595:Amino Acid Sequence; D000818:Animals; D016367:Baculoviridae / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D002478:Cells, Cultured; D002645:Chickens; D003001:Cloning, Molecular; D003208:Concanavalin A / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel / Q000662:veterinary; D015967:Gene Expression Regulation, Viral; D005822:Genetic Vectors; D007371:Interferon-gamma / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D007963:Leukocytes, Mononuclear / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009569:Nitric Oxide / Q000378:metabolism; D011994:Recombinant Proteins; D013154:Spleen / Q000378:metabolism
D011994:Recombinant Proteins; D003208:Concanavalin A; D009569:Nitric Oxide; D007371:Interferon-gamma