Isolation of a 110 000 molecular weight protein in the purification of the nuclear chick intestinal 1,25-dihydroxyvitamin D-3 receptor.
A single protein band of molecular weight 110 000 has been obtained after sodium dodecyl sulfate polyacrylamide gel electrophoresis of purified 1,25-dihydroxyvitamin D-3 (1,25-(OH)2D-3) receptor from crude nuclear extracts of chick intestinal mucosa, prepared in the presence of the protease inhibitors phenylmethylsulfonyl fluoride and epsilon-aminocaproic acid. The nuclear extract was subjected to a six-step purification scheme, involving polymin P and ammonium sulfate fractionation, DNA-cellulose affinity chromatography, Sephacryl S-200 gel filtration, blue dextran-Sepharose and a final DNA-cellulose chromatographic step. The receptor was obtained in about 1% yield and was purified approx. 3700-fold from the nuclear extract, as assessed by specific activity. Single peaks were observed with 3H-1,25-(OH)2D-3-labeled crude nuclear extracts on Sephacryl S-200 gel filtration (Stokes' radius = 35.5 A) and sucrose density gradient centrifugation (3.5 S). Although the identity of the Mr 110 000 protein will remain inconclusive until methods for further characterization are available, it may represent evidence for a higher molecular weight form of the 1,25-(OH)2D-3 receptor than that observed previously.
D000818:Animals; D002645:Chickens; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D007413:Intestinal Mucosa / Q000032:analysis*; D008297:Male; D008970:Molecular Weight; D011506:Proteins / Q000302:isolation & purification*; D018167:Receptors, Calcitriol; D011987:Receptors, Steroid / Q000302:isolation & purification*
D011506:Proteins; D018167:Receptors, Calcitriol; D011987:Receptors, Steroid