Creation of Fluorescent RXR Antagonists Based on CBTF-EE and Application to a Fluorescence Polarization Binding Assay.
Creation of Fluorescent RXR Antagonists Based on CBTF-EE and Application to a Fluorescence Polarization Binding Assay.
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DOI：
10.1021/acsmedchemlett.1c00201
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发表时间：
2021-05
影响因子：
4.2
通讯作者：
Maho Takioku;Y. Takamura;Michiko Fujihara;Masaki Watanabe;S. Yamada;M. Kawasaki;S. Ito;S. Nakano;H. Kakuta
中科院分区：
文献类型：
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作者：
Maho Takioku;Y. Takamura;Michiko Fujihara;Masaki Watanabe;S. Yamada;M. Kawasaki;S. Ito;S. Nakano;H. Kakuta
类维生素A X受体（RXR）配体通常以羧基在配体结合口袋（LBP）内形成氢键的模式结合。然而，我们先前报道的RXR拮抗剂CBTF-EE（4a）以其羧基指向LBP外部并且其烷氧基侧链位于LBP内部的方式结合。在这里，我们研究了结合模式的4 b和4c轴承硝基苯并恶二唑（NBD）或硼-二吡咯亚甲基（BODIPY）荧光团，分别在4a的烷氧基链的末端。这两种化合物均作为RXR拮抗剂发挥作用。4c而不是4 b可用于荧光偏振结合测定，表明BODIPY而不是NBD的旋转在结合状态下受到限制。通过对接模拟支持的荧光发现表明，荧光团位于LBP之外，因此4 b和4c的结合模式与4a的结合模式不同。测定结果与[3 H]9-顺式维甲酸测定结果高度相关。
Retinoid X receptor (RXR) ligands often bind in modes in which the carboxy group forms a hydrogen bond inside the ligand-binding pocket (LBP). However, our previously reported RXR antagonist, CBTF-EE (4a), binds with its carboxy group directed outside the LBP and its alkoxy side chain located inside the LBP. Here, we examined the binding modes of 4b and 4c bearing a nitrobenzoxadiazole (NBD) or boron-dipyrromethene (BODIPY) fluorophore, respectively, at the end of the alkoxy chain of 4a. Both compounds function as RXR antagonists. 4c, but not 4b, was available for a fluorescence polarization binding assay, indicating that rotation of BODIPY, but not NBD, is restricted in the bound state. The fluorescence findings, supported by docking simulations, suggest the fluorophores are located outside the LBP, so that the binding mode of 4b and 4c is different from that of 4a. The assay results were highly correlated with those of a [3H]9-cis-retinoic acid assay.