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活细胞中GPCR二聚体的smFRET成像
作者:小柯机器人 发布时间:2021/3/10 16:08:25

美国哥伦比亚大学Jonathan A. Javitch和美国圣裘德儿童研究医院Scott C. Blanchard研究组合作,开发了活细胞中G蛋白偶联受体(GPCR)二聚体的单分子荧光共振能量转移(smFRET)成像技术。该项研究成果发表在2021年3月8日出版的《自然-方法学》上。

他们报告使用smFRET来检测和追踪哺乳动物细胞质膜内扩散的跨膜蛋白的普适方法。他们利用这种细胞内smFRET方法来显示激动剂诱导的个体代谢型谷氨酸受体二聚体内的结构动力学。他们将这些方法应用于代表性的A、B和C类受体,分别找到受体单体,密度依赖二聚体和组成型二聚体的证据。

据了解,已知C类GPCR形成稳定同型二聚体或异二聚体对功能至关重要,但是构成所有GPCR的> 90%的A和B类受体的低聚状态仍是研究重点。SmFRET是一种强大的方法,有可能揭示有关GPCR组织的宝贵见解,但很少在活细胞中用于研究蛋白质系统。

附:英文原文

Title: Single-molecule FRET imaging of GPCR dimers in living cells

Author: Wesley B. Asher, Peter Geggier, Michael D. Holsey, Grant T. Gilmore, Avik K. Pati, Jozsef Meszaros, Daniel S. Terry, Signe Mathiasen, Megan J. Kaliszewski, Mitchell D. McCauley, Alekhya Govindaraju, Zhou Zhou, Kaleeckal G. Harikumar, Khuloud Jaqaman, Laurence J. Miller, Adam W. Smith, Scott C. Blanchard, Jonathan A. Javitch

Issue&Volume: 2021-03-08

Abstract: Class C G protein-coupled receptors (GPCRs) are known to form stable homodimers or heterodimers critical for function, but the oligomeric status of class A and B receptors, which constitute >90% of all GPCRs, remains hotly debated. Single-molecule fluorescence resonance energy transfer (smFRET) is a powerful approach with the potential to reveal valuable insights into GPCR organization but has rarely been used in living cells to study protein systems. Here, we report generally applicable methods for using smFRET to detect and track transmembrane proteins diffusing within the plasma membrane of mammalian cells. We leverage this in-cell smFRET approach to show agonist-induced structural dynamics within individual metabotropic glutamate receptor dimers. We apply these methods to representative class A, B and C receptors, finding evidence for receptor monomers, density-dependent dimers and constitutive dimers, respectively. 

DOI: 10.1038/s41592-021-01081-y

Source: https://www.nature.com/articles/s41592-021-01081-y

期刊信息

Nature Methods:《自然—方法学》,创刊于2004年。隶属于施普林格·自然出版集团,最新IF:28.467
官方网址:https://www.nature.com/nmeth/
投稿链接:https://mts-nmeth.nature.com/cgi-bin/main.plex