Expressions and purification of a mature form of recombinant human Chemerin in Escherichia coli.
Chemerin is a novel chemokine that binds to the G protein-coupled receptor (GPCR) ChemR23, also known as chemokine-like receptor 1 (CMKLR1). It is secreted as a precursor and executes pro-inflammatory functions when the last six amino acids are removed from its C-terminus by serine proteases. After maturation, Chemerin attracts dendritic cells and macrophages through binding to ChemR23. We report a new method for expression and purification of mature recombinant human Chemerin (rhChemerin) using a prokaryotic system. After being expressed in bacteria, rhChemerin in inclusion bodies was denatured using 6M guanidine chloride. Soluble rhChemerin was prepared by the protein-specific renaturation solution under defined conditions. It was subsequently purified using ion-exchange columns to more than 95% purity with endotoxin level <1.0 EU/microg. We further demonstrated its biological activities for attracting migration of human dendritic cells and murine macrophages in vitro using established chemotaxis assays.
Xiang
Di
D
Laboratory of Regeneromics, School of Pharmacy, Shanghai Jiaotong University, Shanghai 200240, PR China.
Zhang
Jing
J
Chen
Yizhe
Y
Guo
Yiping
Y
Schalow
Adrian
A
Zhang
Zhonghui
Z
Hu
Xiaojia
X
Yu
Hongjing
H
Zhao
Mei
M
Zhu
Shunying
S
Lu
Huili
H
Wu
Mingyuan
M
Yu
Yan
Y
Moldenhauer
Anja
A
Han
Wei
W
D000818:Animals; D001483:Base Sequence; D018925:Chemokines / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D002633:Chemotaxis / Q000502:physiology; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins; D017737:Macrophages, Peritoneal / Q000166:cytology / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D011487:Protein Conformation; D017510:Protein Folding; D019707:Receptors, Chemokine / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*
C102741:CMKLR1 protein, human; D018925:Chemokines; D036341:Intercellular Signaling Peptides and Proteins; C478630:RARRES2 protein, human; D019707:Receptors, Chemokine; D011994:Recombinant Proteins