High level expression and purification of bioactive bovine interleukin-18 using a baculovirus system

Bioactive recombinant bovine interleukin-18 (rboIL-18) was expressed using a baculovirus system. Normally, IL-18 is translated as a precursor form of a 24 kDa polypeptide and processed by IL-1β converting enzyme (ICE) to a mature bioactive form of 18 kDa protein. Hence, to express active form IL-18,...

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Bibliographic Details
Published in:Veterinary immunology and immunopathology 2002-08, Vol.87 (1), p.65-72
Main Authors: Nagata, Tomoshi, Ishikawa, Satoko, Shimokawa, Eiko, Kamikawa, Makiyo, Kikuma, Reiko, Muneta, Yoshihiro, Yokomizo, Yuichi, Nakamura, Masayuki, Takehara, Kazuaki
Format: Article
Language:English
Subjects:
Animals
Baculoviridae - genetics
Baculovirus
Blotting, Western
Bovine IL-12
Bovine IL-18
Caspase 1 - genetics
Cattle
Cloning, Molecular
Electrophoresis, Polyacrylamide Gel - veterinary
Expression
Interferon-gamma - analysis
Interferon-gamma - biosynthesis
Interleukin-18 - biosynthesis
Interleukin-18 - genetics
Interleukin-18 - isolation & purification
Purification
Recombinant Proteins - biosynthesis
Recombinant Proteins - genetics
Recombinant Proteins - isolation & purification
Spodoptera - genetics
Superinfection - veterinary
Synergistic effect
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Summary:Bioactive recombinant bovine interleukin-18 (rboIL-18) was expressed using a baculovirus system. Normally, IL-18 is translated as a precursor form of a 24 kDa polypeptide and processed by IL-1β converting enzyme (ICE) to a mature bioactive form of 18 kDa protein. Hence, to express active form IL-18, we constructed two recombinant baculoviruses containing boIL-18 and human ICE ( hICE) genes, respectively, and superinfected these viruses into insect cells. Superinfection of both recombinant viruses into the cells resulted in the expression of a 24 kDa precursor form and an 18 kDa mature form detectable in the supernatant by immunoblotting using anti-porcine IL-18 antibody. Culture supernatant from the superinfected cells showed a synergistic effect with recombinant boIL-12 for production of interferon-γ (IFN-γ) in bovine peripheral mononuclear cells. By addition of histidine hexamer at the C-terminal of boIL-18, the mature IL-18 was purified. Bioactivity remained after purification.
ISSN:0165-2427
1873-2534
DOI:10.1016/S0165-2427(02)00127-7