Expression and purification of recombinant swine interleukin-4.
The swine interleukin-4 (SwIL-4) cDNA was cloned by RT-PCR. It was expressed using an expression vector pQE30 in E. coli, a baculovirus AcNPV vector pVL1392 in insect cells, and a pCAGGS vector in mammalian cells. The rSwIL-4 proteins expressed from bacteria and insect cells were purified using a chelating affinity column and a mAb-coupled immunoaffinity column. The amount of the products and their bioactivities were compared. All recombinant cytokines were efficiently reacted with the specific antibodies and the molecular weight of rSwIL-4 was approximately 16 kDa in E. coli, 15 and 18 kDa in insect cells, and 15 and 20 kDa in mammalian cells. Variations of molecular weight observed in insect and mammalian cells were probably due to different modification ways of glycosylation. All these recombinant proteins retained their antigenicity and were biologically active in inducing human TF-1 cell proliferation in vitro. The simple purification method will make it possible to evaluate the in vitro and in vivo effects of IL-4 in pigs.
Nuntaprasert
A
A
Laboratory of Animal Research Center, Institution of Medical Science, The University of Tokyo, 4-6-1 Shirokanedai Minato-ku, Tokyo 108-8639, Japan.
Mori
Y
Y
Fujita
K
K
Yoneda
M
M
Miura
R
R
Tsukiyama-Kohara
K
K
Kai
C
C
D000818:Animals; D016367:Baculoviridae; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D003001:Cloning, Molecular; D004926:Escherichia coli; D005822:Genetic Vectors; D006801:Humans; D015847:Interleukin-4 / Q000096:biosynthesis* / Q000302:isolation & purification / Q000494:pharmacology; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000302:isolation & purification / Q000494:pharmacology; D013552:Swine
D011994:Recombinant Proteins; D015847:Interleukin-4