Proluciferin Acetals as Bioluminogenic Substrates for Cytochrome P450 Activity and Probes for CYP3A Inhibition
Proluciferin Acetals as Bioluminogenic Substrates for Cytochrome P450 Activity and Probes for CYP3A Inhibition
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DOI：
10.1124/dmd.111.041541
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发表时间：
2011-12-01
影响因子：
3.9
通讯作者：
Cali, James J.
中科院分区：
文献类型：
--
作者：
Meisenheimer, Poncho L.;Uyeda, H. Tetsuo;Cali, James J.
细胞色素P450 （P450）测定使用探针底物来询问新的化学实体对P450酶的影响。我们报道了一个家族的生物致光荧光素缩醛底物的合成和研究，被P450酶氧化形成荧光素酶底物。荧光素缩醛通过一组纯化的P450酶进行筛选。其中一种产荧光素缩醛在cyp3a4催化下表现出对荧光素酯的敏感和选择性氧化作用，其代谢产物k(m)和k（cat）分别为2.88 μ m和5.87 pmol。分钟(1)。Pmol酶（-1）。用缩醛荧光素作为探针底物，测定已知的CYP3A4或人肝微粒体抑制剂的IC（50）值。已知抑制剂的IC（50）值与传统的基于高效液相色谱的探针底物睾酮计算的IC（50）值密切相关。荧光素缩醛被CYP3A迅速氧化为不稳定的半正位酯，产生荧光素酯，因此有利于简单快速的CYP3A生物发光测定。
Cytochrome P450 (P450) assays use probe substrates to interrogate the influence of new chemical entities toward P450 enzymes. We report the synthesis and study of a family of bioluminogenic luciferin acetal substrates that are oxidized by P450 enzymes to form luciferase substrates. The luciferin acetals were screened against a panel of purified P450 enzymes. In particular, one proluciferin acetal has demonstrated sensitive and selective CYP3A4-catalyzed oxidation to a luciferin ester-K(m) and k(cat) are 2.88 mu M and 5.87 pmol metabolite . min(-1) . pmol enzyme(-1), respectively. The proluciferin acetal was used as a probe substrate to measure IC(50) values of known inhibitors against recombinant CYP3A4 or human liver microsomes. IC(50) values for the known inhibitors correlate strongly with IC(50) values calculated from the traditional high-performance liquid chromatography-based probe substrate testosterone. Luciferin acetals are rapidly oxidized to unstable hemi-orthoesters by CYP3A resulting in luciferin esters and, therefore, are conducive to simple rapid CYP3A bioluminescent assays.