Fast single-molecule FRET spectroscopy: theory and experiment.
Fast single-molecule FRET spectroscopy: theory and experiment.
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DOI：
10.1039/c4cp02489c
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发表时间：
2014-09-21
期刊：
影响因子：
--
通讯作者：
Gopich IV
中科院分区：
文献类型：
--
作者：
Chung HS;Gopich IV
单分子光谱广泛用于研究大分子动力学。尽管该技术提供了在系综水平上无法获得的独特信息，但研究快速分子动力学的可能性受到每单位时间检测到的光子数量（光子计数率）的限制，该数量与照明强度成正比。然而，由于各种光物理和光化学问题，简单地增加照明强度通常没有帮助。在本视角中，我们展示了如何通过考虑每个光子并使用最大似然法来提高给定光子计数率下单分子荧光光谱的动态范围。对于记录了光子颜色和光子间时间的光子轨迹，通过最大化适当的似然函数来获得描述分子动力学的模型参数。我们讨论各种似然函数、它们的适用性以及提取参数的准确性。最大似然法已用于分析快速两态蛋白质折叠的实验并测量过渡路径时间。在二维 FRET 效率-寿命直方图的框架中讨论了荧光寿命等其他信息的利用。
Single-molecule spectroscopy is widely used to study macromolecular dynamics. Although this technique provides unique information that cannot be obtained at the ensemble level, the possibility of studying fast molecular dynamics is limited by the number of photons detected per unit time (photon count rate), which is proportional to the illumination intensity. However, simply increasing the illumination intensity often does not help because of various photophysical and photochemical problems. In this Perspective, we show how to improve the dynamic range of single-molecule fluorescence spectroscopy at a given photon count rate by considering each and every photon and using a maximum likelihood method. For a photon trajectory with recorded photon colors and inter-photon times, the parameters of a model describing molecular dynamics are obtained by maximizing the appropriate likelihood function. We discuss various likelihood functions, their applicability, and the accuracy of the extracted parameters. The maximum likelihood method has been applied to analyze the experiments on fast two-state protein folding and to measure transition path times. Utilizing other information such as fluorescence lifetimes is discussed in the framework of two-dimensional FRET efficiency-lifetime histograms.
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影响因子：
48
作者：
Gambin, Yann;VanDelinder, Virginia;Ferreon, Allan Chris M.;Lemke, Edward A.;Groisman, Alex;Deniz, Ashok A.
通讯作者：
Deniz, Ashok A.
影响因子：
3.4
作者：
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通讯作者：
Puglisi, Joseph D.
影响因子：
2.3
作者：
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通讯作者：
Gopich, Irina V.
影响因子：
2.9
作者：
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通讯作者：
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影响因子：
48
作者：
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通讯作者：
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