Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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Purification of Caudal-Related Homeodomain Transcription Factor and Its Binding Characterization

  • Jeong, Mi-Suk (Department of Molecular Biology, College of Natural Sciences, Pusan National University) ;
  • Hwang, Eun-Young (Department of Molecular Biology, College of Natural Sciences, Pusan National University) ;
  • Kim, Hyun-Tae (Department of Molecular Biology, College of Natural Sciences, Pusan National University) ;
  • Yoo, Mi-Ae (Department of Molecular Biology, College of Natural Sciences, Pusan National University) ;
  • Jang, Se-Bok (Department of Molecular Biology, College of Natural Sciences, Pusan National University)
  • Published : 2009.12.31
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Abstract

Human CDX2 is known as a caudal-related homeodomain transcription factor that is expressed in the intestinal epithelium and is important in differentiation and maintenance of the intestinal epithelial cells. The caudal-related homeobox proteins bind DNA according to a helix-turn-helix structure, thereby increasing the structural stability of DNA. A cancer-tumor suppressor role for Cdx2 has been shown by a decrease in the level of the expression of Cdx2 in colorectal cancer, but the mechanism of transcriptional regulation has not been examined at the molecular level. We developed a large-scale system for expression of the recombinant, novel CDX2, in Escherichia coli. A highly purified and soluble CDX2 protein was obtained in E. coli strain BL21(DE3)RIL and a hexahistidine fusion system using Ni-NTA affinity column, anion exchange, and gel filtration chromatographies. The identity and secondary structure of the purified CDX2 protein were confirmed by MALDI-TOF MS, Western blot, and a circular dichroism analyses. In addition, we studied the DNA-binding activity of recombinant CDX2 by ELISA experiment and isolated human CDX2-binding proteins derived from rat cells by an immobilized GST-fusion method. Three CDX2-binding proteins were found in the gastric tissue, and those proteins were identified to the homeobox protein Hox-D8, LIM homeobox protein 6, and SMC1L1 protein.

Keywords

References

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