Production of recombinant ovine interferon tau using a Bombyx mori nuclear polyhedrosis baculovirus expression system.
Ovine interferon tau (oIFN-tau) is an embryonic protein of critical importance in the establishment of pregnancy in the sheep. We have produced recombinant (r) oIFN-tau using a baculovirus expression system and demonstrated the biological activity of the protein produced. Bombyx mori larvae were infected with B. mori nuclear polyhedrosis virus (BmNPV), modified by inserting a cDNA coding for oIFN-tau downstream of the strong polyhedron promoter. Following infection, antiviral activity of the haemolymph rose to a maximum of 3.6 x 10(8) u/mL (equivalent to 3 mg roIFN-tau/mL) by day 5, when haemolymph was collected and stored frozen. Control haemolymph, collected from uninfected insects at an equivalent time, contained no antiviral activity. The roIFN-tau was partially purified by gel filtration column chromatography and the presence of roIFN-tau confirmed by western blotting. The biological activity of the partially purified roIFN-tau was tested in ewes. Treatment with roIFN-tau caused a significant delay in luteolysis confirming biological potency. The results demonstrate that this system can be successfully used to produce large quantities of roIFN-tau.
Mann
S J
SJ
University of Nottingham, School of Biosciences, Division of Animal Physiology, Sutton Bonington, Loughborough, UK.
Lamming
G E
GE
Fisher
P A
PA
Maeda
S
S
Stewart
H J
HJ
Flint
A P F
AP
Mann
G E
GE
D000818:Animals; D015153:Blotting, Western / Q000662:veterinary; D012831:Bombyx / Q000821:virology*; D002850:Chromatography, Gel / Q000662:veterinary; D003001:Cloning, Molecular / Q000379:methods*; D015237:Dinoprost / Q000031:analogs & derivatives* / Q000097:blood; D005260:Female; D066298:In Vitro Techniques; D007370:Interferon Type I / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D003341:Luteolysis / Q000187:drug effects; D017924:Nucleopolyhedroviruses / Q000235:genetics* / Q000378:metabolism; D011247:Pregnancy; D011257:Pregnancy Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D011374:Progesterone / Q000097:blood; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D012756:Sheep / Q000502:physiology*
D007370:Interferon Type I; D011257:Pregnancy Proteins; D011994:Recombinant Proteins; C020947:interferon tau; C008228:15-keto-13,14-dihydroprostaglandin F2alpha; D011374:Progesterone; D015237:Dinoprost