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Identification of the ligand‐binding domain of human vascular‐endothelial‐growth‐factor receptor Flt‐1
Author(s) -
Ma Li,
Wang Xiaoning,
Zhang Zhiqing,
Zhou Xiaoming,
Chen Aijun,
Yao Lihong
Publication year - 2001
Publication title -
biotechnology and applied biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.468
H-Index - 70
eISSN - 1470-8744
pISSN - 0885-4513
DOI - 10.1042/ba20010043
Subject(s) - umbilical vein , angiogenesis , vasculogenesis , pichia pastoris , vascular endothelial growth factor , biology , receptor , endothelial stem cell , kinase insert domain receptor , microbiology and biotechnology , extracellular , human umbilical vein endothelial cell , yeast , in vitro , biochemistry , vascular endothelial growth factor a , recombinant dna , cancer research , vegf receptors , gene
The vascular‐endothelial‐growth‐factor (VEGF) receptor Flt‐1 has been shown to be involved in vasculogenesis and angiogenesis. The receptor is characterized by seven immunoglobulin‐like loops within the extracellular domain and the first three N‐terminal immunoglobulin‐like loops are involved in high‐affinity binding of VEGF. The minimal extracellular domains of Flt‐1 to achieve VEGF binding were screened using the yeast two‐hybrid system. The result showed that the binding capacity of loop2‐3 was close to that of loop1‐3. The two truncated mutants consisting of loop2‐3 and loop1‐3 were expressed in the methylotrophic yeast Pichia pastoris at high levels (0.3 mg/litre). The corresponding proteins, named soluble (s)Flt‐1(2‐3) and sFlt‐1(1‐3), were purified. An in vitro biological activity assay showed that the binding capacity of sFlt‐1(2‐3) to human VEGF 165 and the inhibiting effect of it on human umbilical‐vein endothelial cell proliferation stimulated by human VEGF 165 were close to those of sFlt‐1(1‐3). Animal tests showed that sFlt‐1(2‐3) could significantly inhibit the formation of regenerated blood vessels stimulated by hVEGF 165 .