Identification of amino acids in rat pregnane X receptor that determine species-specific activation.
Identification of amino acids in rat pregnane X receptor that determine species-specific activation.
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DOI：
10.1124/mol.65.1.36
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发表时间：
2004
影响因子：
3.6
通讯作者：
R. Tirona;B. Leake;L. Podust;R. Kim
中科院分区：
文献类型：
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作者：
R. Tirona;B. Leake;L. Podust;R. Kim
甾烷X受体（PXR）是一种核受体，与药物代谢酶和转运蛋白的转录调控密切相关。有趣的是，某些PXR配体如利福平已被证明容易诱导人类和兔，但不是啮齿动物细胞色素P450 3A的成员。由于不同化学结构的药物似乎受到类似的影响，我们假设大鼠PXR中的特定氨基酸残基或结构域影响某些人PXR配体的受体活化。为了鉴定这样的结构域，使用随机嵌合体生成方法产生串联头-尾构型的人-大鼠和大鼠-人嵌合PXR cDNA阵列。这些嵌合体的药理学表征揭示了大鼠和人PXR的配体结合结构域内的一个离散片段对利福平作用至关重要。在这个区域内，相应的残基Leu 308和Phe 305的人和大鼠PXR，分别被发现是重要的利福平激活。最近确定的人类PXR晶体结构的同源性建模表明，这些氨基酸位于或邻近的柔性环，形成孔的一部分，配体结合腔。利福平，紫杉醇，贯叶金丝桃素的敏感性被赋予大鼠PXR时，Phe 305被转化为亮氨酸，而衰减的敏感性时，观察到人PXR的Leu 308被苯丙氨酸取代。因此，我们的数据提供了令人信服的新见解的重要性，氨基酸组成的孔配体结合腔作为一个关键的调制器PXR反应。
The pregnane X receptor (PXR) is a nuclear receptor significantly involved in the transcriptional regulation of drug-metabolizing enzymes and transporters. Interestingly, certain PXR ligands such as rifampin have been shown to readily induce human and rabbit but not rodent members of the cytochrome P450 3A. Because drugs of divergent chemical structures seem to be similarly affected, we hypothesized that specific amino acid residue(s) or domains in rat PXR affect receptor activation by certain human PXR ligands. To identify such a domain(s), an array of human-rat and rat-human chimeric PXR cDNAs in a tandem head-to-tail configuration were created using a random chimeragenesis method. Pharmacological characterization of these chimeras revealed a discreet segment within the ligand-binding domain of rat and human PXR to be essential for the rifampin effect. Within this region, the corresponding residues Leu308 and Phe305 of human and rat PXR, respectively, were found to be important for rifampin activation. Homology modeling derived from the recently determined crystal structure of human PXR indicates that these amino acids are located within or neighboring the flexible loop that forms part of the pore to the ligand-binding cavity. Rifampin, paclitaxel, and hyperforin sensitivity was conferred to rat PXR when Phe305 was converted to leucine, whereas attenuation of sensitivity was observed when Leu308 of human PXR was replaced with phenylalanine. Accordingly, our data provide compelling new insight into the importance of the amino acids comprising the pore to the ligand-binding cavity as a critical modulator of PXR response.