Purified Human Vitamin D Receptor Overexpressed in Escherichia coli and Baculovirus Systems Does Not Bind 1,25-Dihydroxyvitamin D₃ Hormone Efficiently Unless Supplemented with a Rat Liver Nuclear Extract

We report here that highly purified human vitamin D receptor (hVDR) derived from E. coli or baculovirus expression systems does not exhibit saturable, high affinity 1,25-dihydroxyvitamin D₃ (1,25(OH)₂D₃) ligand binding when these preparations alone are analyzed. Inclusion of rat liver nuclear extract, which does not itself contain detectable 1,25(OH)₂D₃ binding activity, is required to endow hVDR isolated from bacterial or insect cells with the property of high affinity hormone binding (K_d 0.13-0.22 nM). This observation should facilitate the valid assay of 1,25(OH)₂D₃ binding activity and kinetics in samples of overexpressed hVDR. Moreover, since rat liver nuclear extract contains retinoid X receptors and possibly other auxiliary factors capable of forming heterodimers with hVDR that in turn associate with vitamin D responsive elements, we hypothesize that like DNA binding, 1,25(OH)₂D₃ binding to hVDR requires the cooperation of a co-receptor or some uncharacterized receptor activating/stabilizing factor.