Expression of biologically active monomeric form of human M-CSF in baculovirus infected silkworm, Bombyx mori.
The human macrophage colony stimulating factor (hM-CSF) in its monomeric form has been over-produced in BmN cells and in silkworm larvae infected with the recombinant baculovirus Bm284M-CSF. The recombinant monomeric M-CSF (rhM-CSF) exhibited the activity of 8-14 x 10(4) units/ml of cell culture medium. When the insect larvae were infected with the recombinant virus, the maximum rhM-CSF was expressed 4-5 days post infection with an activity of 3 x 10(6) units/ml hemolymph. The monomeric rhM-CSF was purified to homogeneity through three steps of purification. A pilot purification yielded 1 mg of homogeneous monomeric rhM-CSF from 10 larvae. The purified rhM-CSF monomers gradually dimerized in vitro. In contrast, the crude or the semi-purified monomers did not dimerize in vitro, indicating that the presence of an unknown moiety in the rhM-CSF preparations obtained from hemolymph interfered with dimerization.
Qiu
P
P
Department of Biochemistry, Nanjing University, P.R. China.
Ding
Y
Y
Qin
J
J
Han
K K
KK
Zhu
D
D
D000478:Alkylation; D000818:Animals; D016367:Baculoviridae / Q000378:metabolism*; D015153:Blotting, Western; D012831:Bombyx / Q000378:metabolism* / Q000821:virology; D001853:Bone Marrow / Q000187:drug effects; D001854:Bone Marrow Cells; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D018076:DNA, Complementary / Q000096:biosynthesis; D006458:Hemolymph / Q000737:chemistry; D006801:Humans; D007814:Larva / Q000378:metabolism; D016173:Macrophage Colony-Stimulating Factor / Q000096:biosynthesis* / Q000302:isolation & purification; D051379:Mice; D008810:Mice, Inbred C57BL; D016186:Receptor, Macrophage Colony-Stimulating Factor; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D013438:Sulfhydryl Compounds / Q000032:analysis
D018076:DNA, Complementary; D011994:Recombinant Proteins; D013438:Sulfhydryl Compounds; D016173:Macrophage Colony-Stimulating Factor; D016186:Receptor, Macrophage Colony-Stimulating Factor