Method for purifying porcine mature interleukin-18 from silkworm haemolymph.
Recombinant porcine mature interleukin-18 (rPomIL-18) has been purified from the haemolymph of silkworms. After co-infection of two recombinant baculoviruses (BmAcpVL1392-IL-18-His and BmAcpVL1392-casp-1) into the silkworm, rPomIL-18 was produced and secreted into the haemolymph. Polyethylene glycol (PEG) 6000 was added to the haemolymph at 8% (v/w) to precipitate storage proteins which would otherwise bind non-specifically to the metal chelating column and the supernatant then was applied to Sepharose bonded with Ni2+. rPomIL-18 was eluted from the column using 100 mM imidazole buffer at pH 8 with a purity of 93.6%. Approximately 5.3 mg purified rPomIL-18 was obtained from 22 ml haemolymph. It could induce interferon-gamma formation from porcine peripheral blood mononuclear cells.
Katakura Industries Co., Ltd., 1548 Shimo-okutomi, Sayama, Saitama 350-1332, Japan. h.nagaya@katakura.co.jp
Hidekazu
H
Nagaya
Yoshihiro
Y
Muneta
Chiaki
C
Enomoto
Sayaka
S
Matsumoto
Yuichi
Y
Yokomizo
Yasuyuki
Y
Mori
D000818:Animals; D012831:Bombyx / Q000235:genetics* / Q000378:metabolism*; D002845:Chromatography / Q000379:methods*; D003001:Cloning, Molecular / Q000379:methods; D006458:Hemolymph / Q000378:metabolism*; D020382:Interleukin-18 / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification*; D015202:Protein Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D013552:Swine
D020382:Interleukin-18; D011994:Recombinant Proteins