Purification and refolding of recombinant human IGF II from silkworms infected with recombinant Bombyx mori nuclear polyhedrosis virus.
We have reported that insulin-like growth factor II (IGF II) was produced as a fusion protein in Bombyx mori (silkworm) larval bodies infected with recombinant B. mori nuclear polyhedrosis virus [J. Gen. Virol., 68, 2599-2606 (1987)]. In this study, the purification of IGF II from the infected silkworms is reported. The fusion protein was extracted with 6.0 M guanidine-HCl from the infected larval bodies homogenized in water. The use of organic solvents to remove the impurities, such as lipid derived from the larval bodies, was a very effective method of purification. IGF II was released from the partially purified fusion protein by treatment with CNBr, purified by HPLC, and refolded by air-oxidization. Refolded IGF II had an identical primary structure including disulfide bonds and showed identical thymidine uptake stimulation activity with human IGF II. Furthermore, protein disulfide-isomerase was shown to be able to refold scrambled IGF II rapidly.
Marumoto
Y
Y
Molecular Biology Research Laboratory, Daiichi Pharmaceutical Co., Ltd., Tokyo, Japan.
Teruuchi
T
T
Enjoh
T
T
Numata
F
F
Sakano
K
K
D000595:Amino Acid Sequence; D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D012831:Bombyx; D005822:Genetic Vectors; D006801:Humans; D007335:Insulin-Like Growth Factor II / Q000235:genetics / Q000302:isolation & purification* / Q000494:pharmacology; D008969:Molecular Sequence Data; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000302:isolation & purification / Q000494:pharmacology; D013936:Thymidine / Q000378:metabolism
D011993:Recombinant Fusion Proteins; D007335:Insulin-Like Growth Factor II; D013936:Thymidine