Milligram Production and Biological Activity Characterization of the Human Chemokine Receptor CCR3

被引：17

作者：

Wang, Mingqing ^{[1
,2
]}

Ge, Baosheng ^{[1
,2
]}

Li, Renmin ^{[3
]}

Wang, Xiaoqiang ^{[1
,2
]}

Lao, Jun ^{[1
,2
]}

Huang, Fang ^{[1
,2
]}

机构：

[1] China Univ Petr Huadong, State Key Lab Heavy Oil Proc, Qingdao, Shandong, Peoples R China

[2] China Univ Petr Huadong, Ctr Bioengn & Biotechnol, Qingdao, Shandong, Peoples R China

[3] Chinese Acad Sci, Qingdao Inst Bioenergy & Bioproc Technol, Qingdao, Shandong, Peoples R China

来源：

PLOS ONE | 2013年 / 8卷 / 06期

基金：

中国国家自然科学基金;

关键词：

PROTEIN-COUPLED RECEPTORS; EOSINOPHIL EOTAXIN RECEPTOR; HETEROLOGOUS EXPRESSION; MOLECULAR-CLONING; FUNCTIONAL EXPRESSION; PURIFICATION; SOLUBILIZATION; GPCR; TRAFFICKING; RECRUITMENT;

D O I：

10.1371/journal.pone.0065500

中图分类号：

O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];

学科分类号：

07 ; 0710 ; 09 ;

摘要：

Human chemokine receptor CCR3 (hCCR3) belongs to the G protein-coupled receptors (GPCRs) superfamily of membrane proteins and plays major roles in allergic diseases and angiogenesis. In order to study the structural and functional mechanism of hCCR3, it is essential to produce pure protein with biological functions on a milligram scale. Here we report the expression of hCCR3 gene in a tetracycline-inducible stable mammalian cell line. A cell clone with high hCCR3 expression was selected from 46 stably transfected cell clones and from this cell line pure hCCR3 on a milligram scale was obtained after two-step purification. Circular dichroism spectrum with a characteristic shape and magnitude for alpha-helix indicated proper folding of hCCR3 after purification. The biological activity of purified hCCR3 was verified by its high binding affinity with its endogenous ligands CCL11 and CCL24, with K-D in the range of 10(-8) M to 10(-6) M.

引用

页数：10

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