Abstract
PurposeThe thermodynamically favored complex between the nuclear vitamin D receptor (VDR) and 1α,25(OH)2-vitamin D3 (1,25D3) triggers a shift in equilibrium to favor VDR binding to DNA, heterodimerization with the nuclear retinoid x receptor (RXR) and subsequent regulation of gene transcription. The key amino acids and structural requirements governing VDR binding to nuclear coactivators (NCoA) are well defined. Yet very little is understood about the internal changes in amino acid flexibility underpinning the control of ligand affinity, helix 12 conformation and function. Herein, we use molecular dynamics (MD) to study how the backbone and side-chain flexibility of the VDR differs when a) complexed to 1α,25(OH)2-vitamin D3 (1,25D3, agonist) and (23S),25-dehydro-1α(OH)-vitamin D3-26,23-lactone (MK, antagonist); b) residues that form hydrogen bonds with the C25-OH (H305 and H397) of 1,25D3 are mutated to phenylalanine; c) helix 12 conformation is changed and ligand is removed; and d) x-ray water near the C1- and C3-OH groups of 1,25D3 are present or replaced with explicit solvent.MethodsWe performed molecular dynamic simulations on the apo- and holo-VDRs and used T-Analyst to monitor the changes in the backbone and side-chain flexibility of residues that form regions of the VDR ligand binding pocket (LBP), NCoA surface and control helix 12 conformation.ResultsThe VDR-1,25D3 and VDR-MK MD simulations demonstrate that 1,25D3 and MK induce highly similar changes in backbone and side-chain flexibility in residues that form the LBP. MK however did increase the backbone and side-chain flexibility of L404 and R274 respectively. MK also induced expansion of the VDR charge clamp (i.e. NCoA surface) and weakened the intramolecular interaction between H305---V418 (helix 12) and TYR401 (helix 11). In VDR_FF, MK induced a generally more rigid LBP and stronger interaction between F397 and F422 than 1,25D3, and reduced the flexibility of the R274 side-chain. Lastly the VDR MD simulations indicate that R274 can sample multiple conformations in the presence of ligand. When the R274 is extended, the β-OH group of 1,25D3 lies proximal to the backbone carbonyl oxygen of R274 and the side-chain forms H-bonds with hinge domain residues. This differs from the x-ray, kinked geometry, where the side-chain forms an H-bond with the 1α-OH group. Furthermore, 1,25D3, but not MK was observed to stabilize the x-ray geometry of R274 during the > 30 ns MD runs.ConclusionsThe MD methodology applied herein provides an in silico foundation to be expanded upon to better understand the intrinsic flexibility of the VDR and better understand key side-chain and backbone movements involved in the bimolecular interaction between the VDR and its’ ligands.Electronic supplementary materialThe online version of this article (doi:10.1186/2193-9616-1-2) contains supplementary material, which is available to authorized users.
Highlights
The nuclear vitamin D receptor (VDR) is a member of the nuclear receptor (NR) transcription factor super family (Nuclear receptor nomenclature committee 1999)
The VDR-1,25D3 and VDR-MK molecular dynamics (MD) simulations demonstrate that 1,25D3 and MK induce highly similar changes in backbone and side-chain flexibility in residues that form the ligand binding pocket (LBP)
The VDR falls into this classification of NRs, given the active conformation is dramatically stabilized by binding to the seco-steroid hormone, 1α,25(OH)2-vitamin D3 (1,25D3) (Peleg et al 1995; Mizwicki et al 2009a)
Summary
The nuclear vitamin D receptor (VDR) is a member of the nuclear receptor (NR) transcription factor super family (Nuclear receptor nomenclature committee 1999). Domain E, the ligand binding domain (LBD), varies cross-family in sequence homology; the transcriptionally active conformation of NR LBDs is highly conserved (Wurtz et al 1996) and is highlighted by a closed position of the C-terminal helix of domain E (helix 12), capping the ligand binding pocket (LBP) (Renaud and Moras 2000). The VDR falls into this classification of NRs, given the active conformation is dramatically stabilized by binding to the seco-steroid hormone, 1α,25(OH)2-vitamin D3 (1,25D3) (Peleg et al 1995; Mizwicki et al 2009a). Like the other NR family members, the closure of helix 12 completes the nuclear co-activator (NCoA) binding surface (Renaud and Moras 2000). The landscape of the NCoA binding surface for the VDR can best be described as a surface of hydrophobic residues that lie between a charge clamp that is made between a conserved LYS and GLU residue, residues 246 (helix 4) and 420 (helix 12) respectively (Figure 1)
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