title: Soluble Expression and Purification of the Catalytic Domain of Human Vascular Endothelial Growth Factor Receptor 2 in Escherichia coli.
abstract: Vascular endothelial growth factor (VEGF) plays a key role in angiogenesis through binding to its specific receptors, which mainly occurs to VEGF receptor 2 (VEGFR-2), a kinase insert domain-containing receptor. Therefore, the disruption of VEGFR-2 signaling provides a promising therapeutic approach for the treatment of cancer by inhibiting abnormal or tumorinduced angiogenesis. To explore this potential, we expressed the catalytic domain of VEGFR- 2 (VEGFR-2-CD) as a soluble active kinase in Escherichia coli. The recombinant protein was purified and the VEGFR-2-CD activity was investigated. The obtained VEGFR-2-CD showed autophosphorylation activity and phosphate transfer activity comparable to the commercial enzyme. Furthermore, the IC50 value of known VEGFR-2 inhibitor was determined using the purified VEGFR-2-CD. These results indicated a possibility for functional and economical VEGFR-2-CD expression in E. coli to use for inhibitor screening.
authors.affiliation: State Key Laboratory of Bioreactor Engineering, Biomedical Nanotechnology Center, Shanghai Institute of Biomanufacturing Technology, School of Biotechnology, East China University of Science and Technology, Shanghai 200237, P.R. China.
authors.forename: Jia
authors.initials: J
authors.lastname: Wei
authors.affiliation: State Key Laboratory of Bioreactor Engineering, Biomedical Nanotechnology Center, Shanghai Institute of Biomanufacturing Technology, School of Biotechnology, East China University of Science and Technology, Shanghai 200237, P.R. China.
authors.forename: Xiaodan
authors.initials: X
authors.lastname: Cao
authors.affiliation: State Key Laboratory of Bioreactor Engineering, Biomedical Nanotechnology Center, Shanghai Institute of Biomanufacturing Technology, School of Biotechnology, East China University of Science and Technology, Shanghai 200237, P.R. China.
authors.forename: Shengmin
authors.initials: S
authors.lastname: Zhou
authors.affiliation: State Key Laboratory of Bioreactor Engineering, Biomedical Nanotechnology Center, Shanghai Institute of Biomanufacturing Technology, School of Biotechnology, East China University of Science and Technology, Shanghai 200237, P.R. China.
authors.forename: Chao
authors.initials: C
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Bioreactor Engineering, Biomedical Nanotechnology Center, Shanghai Institute of Biomanufacturing Technology, School of Biotechnology, East China University of Science and Technology, Shanghai 200237, P.R. China.
authors.forename: Haijun
authors.initials: H
authors.lastname: Yu
authors.affiliation: State Key Laboratory of Bioreactor Engineering, Biomedical Nanotechnology Center, Shanghai Institute of Biomanufacturing Technology, School of Biotechnology, East China University of Science and Technology, Shanghai 200237, P.R. China.
authors.forename: Yao
authors.initials: Y
authors.lastname: Zhou
authors.affiliation: State Key Laboratory of Bioreactor Engineering, Biomedical Nanotechnology Center, Shanghai Institute of Biomanufacturing Technology, School of Biotechnology, East China University of Science and Technology, Shanghai 200237, P.R. China.
authors.forename: Ping
authors.initials: P
authors.lastname: Wang
mesh_terms: D020134:Catalytic Domain / Q000235:genetics; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism*; D015870:Gene Expression; D006801:Humans; D020128:Inhibitory Concentration 50; D010710:Phosphates / Q000378:metabolism; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification*; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification*
chemical_list: D004791:Enzyme Inhibitors; D010710:Phosphates; D011994:Recombinant Proteins; C000588866:KDR protein, human; D040301:Vascular Endothelial Growth Factor Receptor-2
keywords: E. coli expression system; anticancer drug screening; catalytic domain; inhibitors; soluble expression; vascular endothelial growth factor receptor 2