J. Biol. Chem., Vol. 255, Issue 9, 4081-4086, May, 1980

A specific 1,25-dihydroxyvitamin D3 binding macromolecule in chicken bone

WS Mellon and HF DeLuca

Cytosol prepared from homogenates of bone from vitamin D3-deficient chicks contains a 3.7 S macromolecule having high affinity and low capacity for 1,25-dihydroxyvitamin D3. Employing 1,25-dihydroxy-[26,27- 3H]vitamin D3 (160 Ci/mmol) an apparent Kd has been calculated to be 7.6 x 10(-11) M while the association and dissociation rate constants for the binding process at 25 degrees C were determined to be 9.5 x 10(8) M-1 min-1 and 2.3 x 10(-2) min-1, respectively. A 5.5 S molecule is also present which binds 1,25-dihydroxyvitamin D3 and 25- hydroxyvitamin D3 but appears to prefer 25-hydroxyvitamin D3 and is increased by the addition of chick serum to cytosol. The 3.7 S material is neither a serum contaminant nor a component of the 5.5 S molecular species and is likely of intracellular origin. Under low salt conditions the 3.7 S macromolecule migrates to 4.3 S and 5.5 S regions on sucrose gradients suggesting aggregation of the protein. Several vitamin D3 metabolites are capable of specifically binding to the 3.7 S macromolecule. The relative order of potency for several analogs causing displacement of specifically bound 1,25-dihydroxy-[26,27- 3H]vitamin D3 is: 1,25-dihydroxyvitamin D3 greater than 1 alpha- hydroxyvitamin D3 greater than or equal to 25-hydroxyvitamin D3 greater 24(R),25-dihydroxyvitamin D3. It is concluded that chick bone cytosol contains a macromolecule of high affinity and low capacity for 1,25- dihydroxyvitamin D3 which may function as a receptor for some physiological events in bone.
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