Osmond Ronald I W, Sheehan Antony, Borowicz Romana, Barnett Emma, Harvey Georgina, Turner Cheryl, Brown Andrea, Crouch Michael F, Dyer Anthony R
TGR BioSciences Pty Ltd, 31 Dalgleish Street, Thebarton, SA 5031, Australia.
J Biomol Screen. 2005 Oct;10(7):730-7. doi: 10.1177/1087057105277968. Epub 2005 Aug 29.
Discovery of novel agonists and antagonists for G protein-coupled receptors (GPCRs) relies heavily on cell-based assays because determination of functional consequences of receptor engagement is often desirable. Currently, there are several key parameters measured to achieve this, including mobilization of intracellular Ca2+ and formation of cyclic adenosine monophosphate or inositol triphosphate. However, no single assay platform is suitable for all situations, and all of the assays have limitations. The authors have developed a new high-throughput homogeneous assay platform for GPCR discovery as an alternative to current assays, which employs detection of phosphorylation of the key signaling molecule p42/44 MAP kinase (ERK 1/2). The authors show that ERK 1/2 is consistently activated in cells stimulated by Gq-coupled GPCRs and provides a new high-throughput platform for screening GPCR drug candidates. The activation of ERK 1/2 in Gq-coupled GPCR systems generates comparable pharmacological data for receptor agonist and antagonist data obtained by other GPCR activation measurement techniques.
G蛋白偶联受体（GPCR）新型激动剂和拮抗剂的发现很大程度上依赖基于细胞的检测方法，因为通常需要确定受体结合的功能后果。目前，为实现这一目标需测量几个关键参数，包括细胞内Ca2+的动员以及环磷酸腺苷或肌醇三磷酸的形成。然而，没有单一的检测平台适用于所有情况，并且所有检测方法都有局限性。作者开发了一种用于GPCR发现的新型高通量均相检测平台，作为当前检测方法的替代方法，该平台采用检测关键信号分子p42/44丝裂原活化蛋白激酶（ERK 1/2）的磷酸化。作者表明，ERK 1/2在由Gq偶联的GPCR刺激的细胞中持续被激活，并为筛选GPCR候选药物提供了一个新的高通量平台。在Gq偶联的GPCR系统中ERK 1/2的激活产生了与通过其他GPCR激活测量技术获得的受体激动剂和拮抗剂数据相当的药理学数据。
