Molecular basis of P450 inhibition and activation: implications for drug development and drug therapy.
Molecular basis of P450 inhibition and activation: implications for drug development and drug therapy.
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发表时间：
1998-12
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通讯作者：
G. Szklarz;J. Halpert
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作者：
G. Szklarz;J. Halpert
细胞色素 P450 (P450) 2B1 和 P450 3A4 的三维同源模型已与定点诱变一起用于阐明底物特异性的分子决定因素。 2B 酶中鉴定的大多数关键残基都位于五个底物识别位点 (SRS) 内，并且在调节底物结合或进入的细菌 P450 残基中具有对应物。将抑制剂对接到 2B 模型中，为伴随特定氨基酸侧链替换的基于机制的失活的敏感性变化提供了合理的解释。这些研究为预测 P450 抑制引起的药物相互作用和合理的抑制剂设计提供了基础。此外，已经确定了能够影响底物的同向刺激和类黄酮的异向刺激的 P450 3A4 残基的位置。野生型酶的类固醇羟基化表现出 S 形动力学，表明正协同作用。基于 3A4 模型和单位点突变体，构建了 SRS-2 中的双突变体，该双突变体表现出正常的 Michaelis-Menten 动力学。建模和诱变研究的结果表明，底物和效应子在 P450 3A4 的单个大空腔内的相邻位点结合。彻底了解细胞色素 P450 3A4 中底物结合和效应位点的位置和结构要求对于合理化和预测与酶结合的多种药物和其他化合物之间的相互作用是有价值的。
Three-dimensional homology models of cytochromes P450 (P450) 2B1 and P450 3A4 have been utilized along with site-directed mutagenesis to elucidate the molecular determinants of substrate specificity. Most of the key residues identified in 2B enzymes fall within five substrate recognition sites (SRSs) and have counterparts in bacterial P450 residues that regulate substrate binding or access. Docking of inhibitors into 2B models has provided a plausible explanation for changes in susceptibility to mechanism-based inactivation that accompany particular amino acid side-chain replacements. These studies provide a basis for predicting drug interactions due to P450 inhibition and for rational inhibitor design. In addition, the location of P450 3A4 residues capable of influencing homotropic stimulation by substrates and heterotropic stimulation by flavonoids has been identified. Steroid hydroxylation by the wild-type enzyme exhibits sigmoidal kinetics, indicative of positive cooperativity. Based on the 3A4 model and single-site mutants, a double mutant in SRS-2 has been constructed that exhibits normal Michaelis-Menten kinetics. Results of modeling and mutagenesis studies suggest that the substrate and effector bind at adjacent sites within a single large cavity in P450 3A4. A thorough understanding of the location and structural requirements of the substrate-binding and effector sites in cytochrome P450 3A4 should prove valuable in rationalizing and predicting interactions among the multitude of drugs and other compounds that bind to the enzyme.