A Basis for Reduced Chemical Library Inhibition of Firefly Luciferase Obtained from Directed Evolution

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• 作者：Douglas S. Auld* ; Ya-Qin Zhang ; Noel T. Southall ; Ganesha Rai ; Marc Landsman ; Jennifer MacLure ; Daniel Langevin ; Craig J. Thomas ; Christopher P. Austin ; James Inglese
• 刊名：Journal of Medicinal Chemistry
• 出版年：2009
• 出版时间：March 12, 2009
• 年：2009
• 卷：52
• 期：5
• 页码：1450-1458
• 全文大小：385K
• 年卷期：v.52,no.5(March 12, 2009)
• ISSN：1520-4804

文摘

We measured the “druggability” of the ATP-dependent luciferase derived from the firefly Photuris pennsylvanica that was optimized using directed evolution (Ultra-Glo, Promega). Quantitative high-throughput screening (qHTS) was used to determine IC₅₀s of 198899 samples against a formulation of Ultra-Glo luciferase (Kinase-Glo). We found that only 0.1% of the Kinase-Glo inhibitors showed an IC₅₀ < 10 μM compared to 0.9% found from a previous qHTS against the firefly luciferase from Photinus pyralis (lucPpy). Further, the maximum affinity identified in the lucPpy qHTS was 50 nM, while for Kinase-Glo this value increased to 600 nM. Compounds with interactions stretching outside the luciferin binding pocket were largely lost with Ultra-Glo luciferase. Therefore, Ultra-Glo luciferase will show less compound interference when used as an ATP sensor compared to lucPpy. This study demonstrates the power of large-scale quantitative analysis of structure−activity relationships (>100K compounds) in addressing important questions such as a target’s druggability.