Auld Douglas S, Southall Noel T, Jadhav Ajit, Johnson Ronald L, Diller David J, Simeonov Anton, Austin Christopher P, Inglese James
NIH Chemical Genomics Center, National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland 20892-3370, USA.
J Med Chem. 2008 Apr 24;51(8):2372-86. doi: 10.1021/jm701302v. Epub 2008 Mar 26.
To aid in the interpretation of high-throughput screening (HTS) results derived from luciferase-based assays, we used quantitative HTS, an approach that defines the concentration-response behavior of each library sample, to profile the ATP-dependent luciferase from Photinus pyralis against more than 70,000 samples. We found that approximately 3% of the library was active, containing only compounds with inhibitory concentration-responses, of which 681 (0.9%) exhibited IC 50 < 10 microM. Representative compounds were shown to inhibit purified P. pyralis as well as several commercial luciferase-based detection reagents but were found to be largely inactive against Renilla reniformis luciferase. Light attenuation by the samples was also examined and found to be more prominent in the blue-shifted bioluminescence produced by R. reniformis luciferase than in the bioluminescence produced by P. pyralis luciferase. We describe the structure-activity relationship of the luciferase inhibitors and discuss the use of this data in the interpretation of HTS results and configuration of luciferase-based assays.
为了辅助解读基于荧光素酶的检测所获得的高通量筛选（HTS）结果，我们采用了定量HTS方法，该方法可定义每个文库样品的浓度-反应行为，以对超过70,000个样品分析萤火虫荧光素酶的ATP依赖性。我们发现约3%的文库具有活性，仅包含具有抑制性浓度-反应的化合物，其中681种（0.9%）的半数抑制浓度（IC50）<10微摩尔。代表性化合物被证明可抑制纯化的萤火虫荧光素酶以及几种基于荧光素酶的商业检测试剂，但发现它们对海肾荧光素酶基本无活性。还检测了样品对光的衰减，发现其在海肾荧光素酶产生的蓝移生物发光中比在萤火虫荧光素酶产生的生物发光中更为显著。我们描述了荧光素酶抑制剂的构效关系，并讨论了这些数据在解读HTS结果和基于荧光素酶的检测配置中的应用。
