Characterization of 25-hydroxyvitamin D binding protein from intestinal cells

We have previously purified a cytosolic vitamin D metabolite binding protei n (cDBP) from rat enterocytes, which has characteristics distinct from othe r vitamin D binding proteins. In these studies, we demonstrate that cDBP in a semi-purified fraction from human intestinal cells (Caco-2 cells) binds 25-hydroxy-vitamin D (25OHD) with at least a 1000-fold greater affinity tha n 1,25-dihydroxyvitamin D (1,25(OH)(2)D) or 24,25-dihydroxyvitamin D. Treat ment of cells with 1,25(OH)(2)D reduced 25OHD binding to approximately one third that of the untreated cells (0.42 CPM/mg total protein vs 1.34 CPM/mg total protein, respectively). Finally, the cDBP is not immunoreactive to a ntibodies prepared against the C-terminus of the nuclear vitamin D receptor (VDR). In summary, cDBP bound 25OHD with greater affinity than either 1,25 (OH)(2)D or 24,25 dihydroxyvitamin D, the cytosolic binding activity was do wn-regulated by 1,25(OH)(2)D and cBDP is distinct from the nuclear VDR. (C) 2000 Academic Press.