Molecular Aspects of Vitamin D Hormone Action Dr. Elaine Collins San Jose State University, California The hormonally active form of vitamin D is 1,25-dihydroxy vitamin D3 [1,25(OH)2D3]. Biological actions of 1,25(OH)2D3 include maintaining calcium homeostasis and affecting differentiation and proliferation of a variety of tissues including skin, immune cells, and many cancer cells. Most of these actions are mediated through the actions of its nuclear receptor (VDR). VDR is a member of a superfamily of nuclear receptors which are ligand-dependent transcription factors. Other members of the family include receptors for steroid hormones, retinoids, thyroid hormone and several orphan receptors. Hormone binding to VDR is an early step in the nuclear signaling pathway. Binding results in conformational changes in the ligand binding domain (LBD) of the receptor. We are investigating the molecular details of ligand binding and the relationship of binding to conformational changes in LBD. Recently, the x-ray crystal structure of the LBD was published. However, 50 amino acid residues were deleted from the protein due to problems with the crystalization process. We are studying binding characteristics of the native receptor and the deletion mutant to determine the importance of the deleted residues in hormone binding. Our experimental approach includes: 1) Construction of deletion mutants and point mutations. 2) Binding assays to determine binding affinity of the mutant receptors. These experiments should help define the role of 1,25(OH)2D3 binding in the vitamin D signal transduction pathway.
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