Klostermeier D, Millar D P
Department of Molecular Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, California 92037, USA.
Methods. 2001 Mar;23(3):240-54. doi: 10.1006/meth.2000.1135.
Fluorescence resonance energy transfer (FRET) results from nonradiative coupling of two fluorophores and reports on distances in the range 10-100 A. It is therefore a suitable probe to determine distances in RNA molecules and define their global structure, to follow kinetics of RNA conformational changes during folding in real time, to monitor ion binding, or to analyze conformational equilibria and assess the thermodynamic stability of tertiary structure conformers. Along with the basic principles of steady-state and time-resolved fluorescence resonance energy transfer measurements, approaches to investigate RNA conformational transitions and folding are described and illustrated with selected examples. The versatility of FRET-based techniques has recently been demonstrated by implementations of FRET in high-throughput screening of potential drugs as well as studies of energy transfer that monitor RNA conformational changes on the single-molecule level.
荧光共振能量转移（FRET）源于两个荧光团的非辐射耦合，可报告10 - 100埃范围内的距离。因此，它是一种合适的探针，可用于确定RNA分子中的距离并定义其整体结构，实时跟踪RNA折叠过程中构象变化的动力学，监测离子结合，或分析构象平衡并评估三级结构构象体的热力学稳定性。本文将结合稳态和时间分辨荧光共振能量转移测量的基本原理，描述并通过选定的示例说明研究RNA构象转变和折叠的方法。基于FRET的技术的多功能性最近已通过在潜在药物的高通量筛选中实施FRET以及在单分子水平监测RNA构象变化的能量转移研究得到证明。
