NMR study on the structure and interaction of human HOXC9-homeodomain

Abstract

Abstract

NMR study on the structure and interaction Of human HOXC9-homeodomain

HOX genes are a family of transcription factors, which share a common 61 amino acid DNA-binding domain, the homeobox. This contains a helix-turn-helix motif for DNA-protein interaction. HOX genes are important regulators of embryogenesis and envelopment along the anterior–posterior axis. They are involved in formation of body patterns, organogenesis, cell and organ differentiation, cell adhesion, migration and cell cycle control. Some HOX genes can act as oncogenes, playing a role in tumor development. HOXC9 is also Known that associated with angiogenesis and neuroblastoma. The purpose of this study is to determine three dimensional structures of proteins from Homeodomain, and identify the function of the transcription factor by DNA-binding activity. To achieve the goal, Homeodomain of HOXC9 was selected and cloned through the known genome sequence. They were expressed using E.coli expression system and codon optimization process was performed to obtain large quantities of proteins. The expressed proteins were carried out solubility test to optimize the best soluble condition of the proteins. After purification, the secondary structures of the proteins were predicted via Circular Dichroism (CD). The final samples were predicted their three dimensional structure through NMR spectroscopy, and the part of the structure was analyzed. S As the results, HOXC9 is consist of three α –helixes and it has monomer form. The secondary structure and 3D structure of HP0268 were predicted by TALOS+ and MODELLER programs respectively. From this study, we established technical purification methods, optimized conditions for NMR experiments. We complete 95% of the backbone assignments, determinate secondary structure of HOXC9 and confirm HOXC9-DNA interaction by EMSA so we identify function as transcription factor of HOXC9. Additional experiment about binding of Smad4-MH1 with HOXC9 is on-going. This experiment provide the fundamental information for the drug development target to human HOXC9 protein and help to search the lead compound for structure based drug design like high throughput screening.