New insights into Vitamin D sterol-VDR proteolysis, allostery, structure-function from the perspective of a conformational ensemble model.
Recently, we have developed a Vitamin D sterol (VDS)-VDR conformational ensemble model. This model can be broken down into three individual, yet interlinked parts: (a) the conformationally flexible VDS, (b) the apo/holo-VDR helix-12 (H12) conformational ensemble, and (c) the presence of two VDR ligand binding pockets (LBPs); one thermodynamically favored (the genomic pocket, G-pocket) and the other kinetically favored by VDSs (the alternative pocket, A-pocket). One focus of this study is to use directed VDR mutagenesis to (1) demonstrate H12 is stabilized in the transcriptionally active closed conformation (hVDR-c1) by three salt-bridges that span the length of H12 (cationic residues R154, K264 and R402), (2) to elucidate the VDR trypsin sites [R173 (hVDR-c1), K413 (hVDR-c2) and R402 (hVDR-c3)] and (3) demonstrate the apo-VDR H12 equilibrium can be shifted. The other focus of this study is to apply the model to generate a mechanistic understanding to discrepancies observed in structure-function data obtained with a variety of 1alpha,25(OH)(2)-Vitamin D(3) (1,25D) A-ring and side-chain analogs, and side-chain metabolites. We will demonstrate that these structure-function conundrums can be rationalized, for the most part by focusing on alterations in the VDS conformational flexibility and the elementary interaction between the VDS and the VDR A- and G-pockets, relative to the control, 1,25D.
Mizwicki
Mathew T
MT
Department of Biochemistry, University of California-Riverside, Riverside, CA 92521, USA.
Bula
Craig M
CM
Bishop
June E
JE
Norman
Anthony W
AW
D000495:Allosteric Site; D002460:Cell Line; D006801:Humans; D008024:Ligands; D008958:Models, Molecular; D009154:Mutation / Q000235:genetics; D017434:Protein Structure, Tertiary; D018167:Receptors, Calcitriol / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D055672:Static Electricity; D014357:Trypsin / Q000378:metabolism*; D014807:Vitamin D / Q000737:chemistry* / Q000378:metabolism*
D008024:Ligands; D018167:Receptors, Calcitriol; D014807:Vitamin D; D014357:Trypsin