Abstract

We performed total alanine scanning mutational analysis (ASMA) of the residues lining the ligand binding pocket (LBP) of the human vitamin D receptor (hVDR) to investigate allosteric effects of ligands in the function of nuclear receptors (NRs). This was accomplished for the first time in the NR superfamily. The effects of ligand structure were also examined in this system (termed 2D-ASMA) using 8 representative VDR ligands. The results clearly revealed the role and importance of all amino acid residues lining the LBP and the relationships between ligand binding and transcriptional potency. 2D-ASMA indicated ligand-specific ligand-protein interactions, which are essential in determining the transactivation potency of the ligand. Taking the results as a whole, we suggest a ligand-mediated allosteric network, which allows transmission of information from ligands to the interfaces of the VDR in association with protein cofactors and was shown to be linked to a part of the network identified by statistical coupling analysis (SCA).

Keywords: ligand binding pocket (LBP), conformation, VDR agonists, 19-Norvitamin D Analogs, R391A