Yano Hideaki, Cai Ning Sheng, Javitch Jonathan A, Ferré Sergi
National Institute on Drug Abuse, National Institutes of Health, Baltimore, MD, USA.
Departments of Psychiatry & Pharmacology, College of Physicians & Surgeons, Columbia University, New York, NY, USA.
Biotechniques. 2018 Jul;65(1):9-14. doi: 10.2144/btn-2018-0039.
Protein complementation assays (PCA) are used as pharmacological tools, enabling a wide array of applications, ranging from studies of protein-protein interactions to second messenger effects. Methods to detect activities of G protein-coupled receptors (GPCRs) have particular relevance for drug screening. Recent development of an engineered luciferase NanoLuc created the possibility of generating a novel PCA, which in turn could open a new avenue for developing drug screening assays. Here we identified a novel split position for NanoLuc and demonstrated its use in a series of fusion constructs to detect the activity of GPCRs. The split construct can be applied to a variety of pharmacological screening systems.
蛋白质互补分析（PCA）被用作药理学工具，可实现广泛的应用，从蛋白质-蛋白质相互作用的研究到第二信使效应。检测G蛋白偶联受体（GPCR）活性的方法在药物筛选中具有特殊意义。一种工程化荧光素酶NanoLuc的最新进展为产生一种新型PCA创造了可能性，这反过来又可能为开发药物筛选分析开辟一条新途径。在这里，我们确定了NanoLuc的一个新的分裂位置，并证明了其在一系列融合构建体中用于检测GPCR活性的用途。这种分裂构建体可应用于多种药理学筛选系统。
