title: A 65-kilodalton nuclear protein binds to the human vitamin D receptor: a bacterial-expressed histidine-tagged receptor study.
abstract: We report here that the human 1,25-dihydroxyvitamin D3 receptor (hVDR) binds to a 65 kD nuclear protein in a ligand-dependent manner. Histidine-tagged full-length hVDR was overexpressed in E.coli and purified to near homogeneity using Ni-NTA and gel filtration columns without denature/renature procedures. Nuclear extract from the osteoblastic cell line MG-63 was incubated with the recombinant hVDR and Ni-NTA agar in the presence of a double-stranded DNA fragment containing vitamin D responsive element. Proteins bound to the hVDR were eluted and analyzed on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. A 65 kD protein was detected with full-length hVDR in the presence of 100 nM 1,25(OH)2D3, while this interaction was not observed in the absence of the ligand nor with carboxyl-terminally truncated hVDR, which lacks an activation function-2 domain. Therefore, this nuclear protein may be involved in the ligand-dependent transcriptional regulation via the hVDR.
authors.affiliation: Department of Environmental Medicine, Research Institute, Osaka Medical Center for Maternal and Child Health, Japan.
authors.forename: S
authors.initials: S
authors.lastname: Nakajima
authors.forename: I
authors.initials: I
authors.lastname: Yanagihara
authors.forename: K
authors.initials: K
authors.lastname: Ozono
mesh_terms: D001665:Binding Sites; D002460:Cell Line; D004926:Escherichia coli / Q000235:genetics; D006639:Histidine / Q000737:chemistry; D006801:Humans; D008970:Molecular Weight; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism*; D010006:Osteoblasts / Q000378:metabolism; D011485:Protein Binding; D018167:Receptors, Calcitriol / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D009687:Nuclear Proteins; D018167:Receptors, Calcitriol; D011994:Recombinant Proteins; D006639:Histidine