Expression of human VEGF165 in silkworm (Bombyx mori L.) by using a recombinant baculovirus and its bioactivity assay.
Silkworm larva has a lot of advantages as a "biofactory" to produce recombinant protein. A recombinant baculovirus, carrying cDNA encoding the 165 amino-acid long isoform of human vascular endothelial growth factor (VEGF) was successfully constructed for the large-scale production of this protein using silkworms as an in vivo host. The fifth-instar silkworm larvae were inoculated with the recombinant virus. Time-course expression analysis indicated that the expression level was highest at around 80 h post-infection and the recombinant protein was found mainly in the haemolymph. Therefore, the hemolymph was collected from the infected larvae and the recombinant protein was purified by using Nickel affinity chromatography under native condition. The expression level was estimated to be as high as approximately 426 microg per larva. Furthermore, the recombinant protein was characterized and was found biologically active in inducing endothelial cell proliferation in vitro.
Wu
Xiaofeng
X
College of Animal Sciences, Zhejiang University, Huajiachi Campus, Hangzhou 310029, China. wuxiaofeng@zju.edu.cn
Yin
Zhaozheng
Z
Cao
Cuiping
C
Huang
Ling
L
Lu
Xingmeng
X
Liu
Jianxin
J
Cui
Weizheng
W
D000595:Amino Acid Sequence; D000818:Animals; D016367:Baculoviridae / Q000235:genetics*; D012831:Bombyx / Q000235:genetics* / Q000378:metabolism*; D002455:Cell Division / Q000187:drug effects; D003001:Cloning, Molecular / Q000379:methods; D004305:Dose-Response Relationship, Drug; D042783:Endothelial Cells / Q000166:cytology* / Q000187:drug effects; D018014:Gene Transfer Techniques; D006801:Humans; D008969:Molecular Sequence Data; D008970:Molecular Weight; D015202:Protein Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D014162:Transfection; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics* / Q000494:pharmacology
D011994:Recombinant Proteins; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A