A novel far-red bimolecular fluorescence complementation system that allows for efficient visualization of protein interactions under physiological conditions
A novel far-red bimolecular fluorescence complementation system that allows for efficient visualization of protein interactions under physiological conditions
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一种新型远红双分子荧光互补系统，可有效可视化生理条件下蛋白质相互作用
DOI：
10.1016/j.bios.2009.06.008
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发表时间：
2009-09-15
影响因子：
12.6
通讯作者：
Luo, Qingming
中科院分区：
文献类型：
--
作者：
Chu, Jun;Zhang, Zhihong;Luo, Qingming
荧光蛋白（FP）已经能够分析活细胞中的生物分子相互作用，而双分子荧光互补（BiFC）代表了新开发的成像技术之一，可以直接可视化活细胞中的蛋白质-蛋白质相互作用。虽然已经证明了覆盖广泛光谱范围的10种不同FP支持BiFC，但只有基于Cerulean（青色FP变体）、Citrine和Venus（黄色FP变体）的BiFC系统可以在37 ℃生理温度下使用。两种基于mRFP的红色BiFC系统对更高温度（即，37摄氏度）限制了它们在大多数哺乳动物细胞研究中的应用。在这里，我们报告说，mLumin，一个新分离的mKate的远红荧光蛋白变体，发射最大值为621 nm，使BiK分析蛋白质-蛋白质相互作用在37摄氏度的活哺乳动物细胞。此外，mLumin与基于Cerulean和Venus的BiFC系统的组合允许同时可视化同一细胞中的三对蛋白质-蛋白质相互作用。基于mLumin的BiK系统将促进活细胞中多种蛋白质-蛋白质相互作用的同时可视化，并提供可视化活动物中蛋白质-蛋白质相互作用的潜力。(C)2009 Elsevier B. V.保留所有权利。
Fluorescent protein (FP) has enabled the analysis of biomolecular interactions in living cells,and bimolecular fluorescence complementation (BiFC) represents one of the newly developed imaging technologies to directly visualize protein-protein interactions in living cells. Although 10 different FPs that cover a broad range of spectra have been demonstrated to support BiFC, only Cerulean (cyan FP variant), Citrine and Venus (yellow FP variants)-based BiFC systems can be used under 37 degrees C physiological temperature. The sensitivity of two mRFP-based red BiFC systems to higher temperatures (i.e., 37 degrees C) limits their applications in most mammalian cell-based studies. Here we report that mLumin, a newly isolated far-red fluorescent protein variant of mKate with an emission maximum of 621 nm, enables BiK analysis of protein-protein interactions at 37 degrees C in living mammalian cells. Furthermore, the combination of mLumin with Cerulean- and Venus-based BiFC systems allows for simultaneous visualization of three pairs of protein-protein interactions in the same cell. The mLumin-based BiK system will facilitate simultaneous visualization of multiple protein-protein interactions in living cells and offer the potential to visualize protein-protein interactions in living animals. (C) 2009 Elsevier B.V. All rights reserved.