A 65-kilodalton nuclear protein binds to the human vitamin D receptor: a bacterial-expressed histidine-tagged receptor study.

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.