Single A326G mutation converts human CYP24A1 from 25-OH-D3-24-hydroxylase into-23-hydroxylase, generating 1α,25-(OH)2D3-26,23-lactone
Single A326G mutation converts human CYP24A1 from 25-OH-D3-24-hydroxylase into-23-hydroxylase, generating 1α,25-(OH)2D3-26,23-lactone
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DOI：
10.1073/pnas.0702093104
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发表时间：
2007-07-31
影响因子：
11.1
通讯作者：
Jones, Glenville
中科院分区：
文献类型：
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作者：
Prosser, David E.;Kaufmann, Martin;Jones, Glenville
对25-羟基维生素D-3-24-羟基酶(CyP24A1)的研究表明，它是一种双功能酶，可使1α，25(OH)(2)D-3发生24-羟基化，生成骨化柠檬酸和23-羟基化，最终生成1α，25-(OH)(2)D-3-26，23-内酯。CYP24A1进行23-或24-羟基化的程度取决于物种。在这篇文章中，我们证明了以24-羟基化底物为主的人类酶不同于负鼠的酶，负鼠的酶只在有限数量的氨基酸残基上对其进行23-羟基化。人的形式在单一底物结合残基(A326G)上的突变极大地改变了酶的区域选择性，从24-羟基酶变成23-羟基酶，而其他修饰没有影响。在CYP24A1同源模型中，Ala-326位于I-螺旋上，靠近对接的25-羟化侧链的末端，我们解释的结果表明，在326处的甘氨酸取代为底物的侧链提供了额外的空间，使其更深地进入口袋，并将其置于23-羟化的最佳立体化学位置。我们讨论了这些结果对具有A326G替代的物种的生理影响，以及对最佳维生素D类似物设计的启示。
Studies of 25-hydroxyvitamin D-3-24-hydroxylase (CYP24A1) have demonstrated that it is a bifunctional enzyme capable of the 24-hydroxylation of 1 alpha,25-(OH)(2)D-3, leading to the excretory form, calcitroic acid, and 23-hydroxylation, culminating in 1 alpha,25-(OH)(2)D-3-26,23-lactone. The degree to which CYP24A1 performs either 23- or 24-hydroxylation is species-dependent. In this paper, we show that the human enzyme that predominantly 24-hydroxylates its substrate differs from the opossum enzyme that 23-hydroxylates it at only a limited number of amino acid residues. Mutagenesis of the human form at a single substrate-binding residue (A326G) dramatically changes the regioselectivity of the enzyme from a 24-hydroxylase to a 23-hydroxylase, whereas other modifications have no effect. Ala-326 is located in the I-helix, close to the terminus of the docked 25-hydroxylated side chain in a CYP24A1 homology model, a result that we interpret indicates that substitution of a glycine at 326 provides extra space for the side chain of the substrate to move deeper into the pocket and place it in a optimal stereochemical position for 23-hydroxylation. We discuss the physiological ramifications of these results for species possessing the A326G substitution, as well as implications for optimal vitamin D analog design.