Acevedo Alejandra, Stoynova Ludmilla, Davis Karen, Solórzano Ramona, Collins Elaine D
Department of Chemistry, San Jose State University, San Jose, CA 95192-0101, USA.
J Steroid Biochem Mol Biol. 2004 May;89-90(1-5):83-7. doi: 10.1016/j.jsbmb.2004.03.051.
Most of the actions of 1,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3)] are mediated by binding to the Vitamin D nuclear receptor (VDR). The crystal structure of a deletion mutant (Delta165-215) of the VDR ligand-binding domain (LBD) bound to 1,25(OH)(2)D(3) indicates that amino acid residues tyrosine-143 and serine-278 form hydrogen bonding interactions with the 3-hydroxyl group of 1,25(OH)(2)D(3). Studies of VDR and three mutants (Y143F, S278A, and Y143F/S278A) did not indicate any differences in the binding affinity between the variant receptors and the wild-type receptor. This might indicate that the 3-hydroxyl group binds differently to the full-length VDR than the to deletion mutant. To further investigate, four deletion VDR mutants were constructed: VDR(Delta165-215), VDR(Delta165-215) (Y143F), VDR(Delta165-215) (S278A), VDR(Delta165-215) (Y143F/S278A). There were no significant differences in binding affinity between the wild-type receptor and the deletion mutants except for VDR(Delta165-215) (Y143F/S278A). In gene activation assays, VDR constructs with the single mutation Y143F and the double mutation Y143F/S278A, but not the single mutation S278A required higher doses of 1,25(OH)(2)D(3) for half-maximal response. This suggests that there are some minor structural and functional differences between the wild-type VDR and the Delta165-215 deletion mutant and that Y143 residue is more important for receptor function than residue S278.
1,25 - 二羟基维生素D(3)[1,25(OH)₂D(3)]的大多数作用是通过与维生素D核受体（VDR）结合来介导的。与1,25(OH)₂D(3)结合的VDR配体结合域（LBD）缺失突变体（Δ165 - 215）的晶体结构表明，氨基酸残基酪氨酸 - 143和丝氨酸 - 278与1,25(OH)₂D(3)的3 - 羟基形成氢键相互作用。对VDR及其三个突变体（Y143F、S278A和Y143F/S278A）的研究表明，变体受体与野生型受体之间的结合亲和力没有任何差异。这可能表明3 - 羟基与全长VDR的结合方式与缺失突变体不同。为了进一步研究，构建了四个VDR缺失突变体：VDR(Δ165 - 215)、VDR(Δ165 - 215)(Y143F)、VDR(Δ165 - 215)(S278A)、VDR(Δ165 - 215)(Y143F/S278A)。除了VDR(Δ165 - 215)(Y143F/S278A)外，野生型受体与缺失突变体之间的结合亲和力没有显著差异。在基因激活试验中，具有单突变Y143F和双突变Y143F/S278A的VDR构建体，但不是单突变S278A的构建体，需要更高剂量的1,25(OH)₂D(3)才能达到半最大反应。这表明野生型VDR与Δ165 - 215缺失突变体之间存在一些微小的结构和功能差异，并且Y143残基对受体功能比S278残基更重要。
