Two models of Smad4 and Hoxa9 complex are proposed: structural and interactional perspective

Transforming growth factor-beta superfamily growth factors (TGF-β) regulate a diverse range of cellular functions, including proliferation, differentiation, extracellular matrix secretion and cell adhesion. TGF-β is also one of the most abundant of the known growth factors. Osteopontin (OPN), the major non-collagenous bone matrix protein is a secreted, arginine-glycine-aspertate containing phosphorylated glycoprotein. Analysis of the OPN promoter sequence reveals both Hoxc8 and Hoxa9 (mouse homeotic gene) recognize and utilize the same consensus TAAT motif in the binding sequence to mediate the repression. Hoxa9 functions as a strong transcriptional repressor, similar to Hoxc-8 (X. Shi, X. Yang, D. Chen, Z. Chang, and X. Cao, J Biol Chem 274, 13711-13717, 1999). The DNA-binding protein Hoxa9 interacts with Smad4 (X. Shi, S. Bai, L. Li and X, and Cao X, J Biol Chem 276, 850-855, 2001), but not with Smad3 (which binds to OPN promoter), and the interaction between Smad4 and Hoxa9 results in the transcriptional activation of OPN in response to TGF- stimulation. In this paper we have proposed two possible model structures of Hoxa9 and Smad4 complex. These have been modeled based on homology modeling and a new method has been used to model the flexible loop part. Manual docking has been used to achieve the final model involving the Hoxa9 -Smad4 complex which tallies with the experimental results. We have mutated some selective important residues and looked at their effect in terms of interaction energy in complex formation in both the models.