[Fusion expression, purification and bioassay of IFN-gamma inducible protein-10 and thioredoxin gene in E. coli].
Interferon gamma-inducible protein 10, a member of the family of CXC chemokines, is secreted by interferon gamma-stimulated, monocytes, endothelial cells and keratinocytes. Interferon gamma-inducible protein 10 plays an important role in recruiting activated T cells into sites of tissue inflammation. In this experiment, PCR products of Interferon gamma-inducible protein 10 were cloned into prokaryote expression vector pET 32(a) to generate recombinant pET-IP10 with S-Tag at the N-terminus, and expressed successfully in E. coli BL21 (DE3). The total expressed products amounted to 25.3% in all bacterion proteins. pET-IP10 mainly formed inclusion body in E. coli. Soluble recombinant protein accounted for 20% among IP-10 fusion protein. The soluble recombinant proteins were purified by using S-Tag affinity chromatography effectively with purity of over 90%. The chemotaxis biological activity of purified Interferon gamma-inducible protein 10 could specifically exhibit the directional migration of stimulated T cells at concentration of 100 ng/ml. The results indicated that the strategy we used in this experiment was effective for recombinant Interferon gamma-inducible protein 10 production with biological activity.
Li
Gang
G
Key Laboratory of Biotherapy of Human Diseases of Ministry of Education and Cancer Center, West China Hospital, Sichuan University, Chengdu 610041, China.
Tian
Ling
L
Wei
Yuquan
Y
Wen
Yanjun
Y
Xiao
Fei
F
Yao
Bing
B
Zhang
Ling
L
Zhang
Ru
R
Mei
Kai
K
D054357:Chemokine CXCL10 / Q000096:biosynthesis*; D004926:Escherichia coli / Q000378:metabolism; D005822:Genetic Vectors; D006801:Humans; D011994:Recombinant Proteins / Q000096:biosynthesis; D013601:T-Lymphocytes / Q000166:cytology; D013879:Thioredoxins / Q000096:biosynthesis
C114753:CXCL10 protein, human; D054357:Chemokine CXCL10; D011994:Recombinant Proteins; D013879:Thioredoxins