Purification of chicken intestinal receptor for 1,25-dihydroxyvitamin D.
The 3.3S chicken intestinal receptor for 1,25-dihydroxyvitamin D [1,25-(OH)2D] has been purified approximately 86,000-fold from the cytosolic fraction. The receptor was selectively precipitated with Polymin P from high-speed supernatants derived from 800 g of intestinal mucosa and then sequentially chromatographed on DNA-cellulose. Sephacryl, blue dextran-Sepharose, DNA-cellulose, and heparin-Sepharose. Polyacrylamide gel electrophoresis of 8-10 microgram of the purified receptor in sodium dodecyl sulfate indicated the presence of one major and three minor protein bands of molecular weights 50,000-65,000. Sucrose gradient analysis of the purified material in 0.3 M KCl suggested that a fraction of the receptor remained complexed to the 1,25-(OH)2D and that its sedimentation property of 3.3S remained unchanged. These results represent a major purification of the chick intestinal receptor for 1,25-(OH)2D, an extremely rare and labile protein whose isolation is estimated to require a 200,000-fold purification. Of primary importance is the observation tht affinity ligands such as DNA and blue dextran can effect major purification of this protein, lending credence to the hypothesis that the 1,25-(OH)2D receptor functions within the cell nucleus by altering the expression of specific genes.
D000818:Animals; D002499:Centrifugation, Density Gradient; D002645:Chickens; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D004100:Dihydroxycholecalciferols / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D006887:Hydroxycholecalciferols / Q000378:metabolism*; D007413:Intestinal Mucosa / Q000032:analysis*; D008970:Molecular Weight; D011956:Receptors, Cell Surface / Q000302:isolation & purification*; D012279:Rickets / Q000378:metabolism
D004100:Dihydroxycholecalciferols; D006887:Hydroxycholecalciferols; D011956:Receptors, Cell Surface