Establishment of a large-scale purification procedure for purified recombinant bovine interferon-tau produced by a silkworm-baculovirus gene expression system.
We developed a procedure for the large-scale purification of bovine interferon-tau (boIFN-tau) by means of a silkworm-baculovirus gene expression system. Recombinant boIFN-tau (rboIFN-tau) was efficiently produced in the silkworm infected with boIFN-tau cDNA recombinant baculovirus and accumulated in the haemolymph. To establish a purification method suitable for mass production, we tried three crude purification methods, namely, an acidification and neutralization treatment (ANT), silica gel column chromatography (SGCC), and Blue sepharose column chromatography (BSCC) with a combination of Q-sepharose (QSC) and chelating sepharose column chromatographies (CSCC). As a result, the acidification and neutralization treatment was found to be the most efficient and cost effective. With this combination, we obtained 91% pure products. To confirm the applicability of the procedure for mass production, we inoculated 100 silkworms with the recombinant virus, and recovered about 4.55 mg (1.26 x 10(8)U/mg) of 91% pure rboIFN-tau by means of a combination of the ANT, followed by QSC and CSCC.
Nagaya
Hidekazu
H
Katakura Industries Co., Ltd., Saitama, Japan
Kanaya
Toshimichi
T
Kaki
Hiroki
H
Tobita
Yoneko
Y
Takahashi
Masashi
M
Takahashi
Hitomi
H
Yokomizo
Yuichi
Y
Inumaru
Shigeki
S
D000818:Animals; D016367:Baculoviridae / Q000235:genetics*; D001709:Biotechnology; D012831:Bombyx / Q000378:metabolism* / Q000821:virology; D002417:Cattle; D002845:Chromatography / Q000379:methods; D015870:Gene Expression; D005822:Genetic Vectors; D006863:Hydrogen-Ion Concentration; D007370:Interferon Type I / Q000096:biosynthesis* / Q000302:isolation & purification*; D007814:Larva / Q000378:metabolism / Q000821:virology; D011257:Pregnancy Proteins / Q000096:biosynthesis* / Q000302:isolation & purification*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification*
D007370:Interferon Type I; D011257:Pregnancy Proteins; D011994:Recombinant Proteins; C020947:interferon tau