Immunoaffinity purification of active rat recombinant 1,25-dihydroxyvitamin D3 receptor.
We have developed a large-scale immunoaffinity purification procedure for the recombinant vitamin D receptor. The purified receptor is homogeneous, and is bound by 1,25-dihydroxyvitamin D3 with a Kd of 5 x 10(-10) M. The isolated receptor binds to the osteocalcin vitamin D response element in the presence of porcine intestinal nuclear extract stripped of endogenous vitamin D receptor as well. However, the binding of D3 and the vitamin D3 response element does not completely assure a native conformation of the protein. The availability of large quantities of highly purified active vitamin D receptor makes possible detailed structural analysis.
Li
Z
Z
Department of Biochemistry, University of Wisconsin-Madison, College of Agricultural and Life Sciences 53706.
Prahl
J M
JM
Hellwig
W
W
DeLuca
H F
HF
D000818:Animals; D000911:Antibodies, Monoclonal; D015153:Blotting, Western; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D002846:Chromatography, Affinity / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel; D007413:Intestinal Mucosa / Q000378:metabolism; D007700:Kinetics; D009036:Moths; D015675:Osteocalcin / Q000235:genetics / Q000378:metabolism; D051381:Rats; D018167:Receptors, Calcitriol / Q000096:biosynthesis / Q000302:isolation & purification* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism; D013552:Swine; D014162:Transfection
D000911:Antibodies, Monoclonal; D018167:Receptors, Calcitriol; D011994:Recombinant Proteins; D015675:Osteocalcin