[Fusion expression, purification and bioassay of IFN-gamma inducible protein-10 and thioredoxin gene in E. coli]

Sheng Wu Yi Xue Gong Cheng Xue Za Zhi. 2005 Jun;22(3):535-9.
[Article in Chinese]

Abstract

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.

MeSH terms

  • Chemokine CXCL10 / biosynthesis*
  • Escherichia coli / metabolism
  • Genetic Vectors
  • Humans
  • Recombinant Proteins / biosynthesis
  • T-Lymphocytes / cytology
  • Thioredoxins / biosynthesis

Substances

  • CXCL10 protein, human
  • Chemokine CXCL10
  • Recombinant Proteins
  • Thioredoxins