Spectral fluctuation of a single fluorophore conjugated to a protein molecule.
Spectral fluctuation of a single fluorophore conjugated to a protein molecule.
复制标题
与蛋白质分子结合的单个荧光团的光谱波动。
DOI：
10.1016/s0006-3495(00)76708-7
复制
发表时间：
2000
影响因子：
3.4
通讯作者：
T. Yanagida
中科院分区：
文献类型：
--
作者：
T. Wazawa;Y. Ishii;T. Funatsu;T. Yanagida
我们使用全内反射荧光显微镜测量了附着在水溶液中单个蛋白质分子上的单个荧光团的荧光光谱。肌球蛋白亚片段 1 (S1) 最具反应性的半胱氨酸残基用四甲基罗丹明标记。在单分子和批量测量中，由溶剂从水性缓冲液变为甲醇引起的光谱变化相似，这表明即使在单分子水平上，荧光光谱对荧光团的微环境变化也很敏感。附着在 S1 分子上的荧光团的荧光光谱的时间依赖性仅显示出在秒的时间尺度上随时间波动。由于直接缀合到玻璃表面的相同荧光团的荧光光谱保持恒定，因此观察到连接到 S1 的荧光团的光谱波动很可能是由于 S1 分子中缓慢的自发构象变化所致。因此，单分子荧光光谱似乎是研究单个生物分子动态行为的有力工具。
We have measured the fluorescence spectra of a single fluorophore attached to a single protein molecule in aqueous solution using a total internal reflection fluorescence microscope. The most reactive cysteine residue of myosin subfragment-1 (S1) was labeled with tetramethylrhodamine. The spectral shift induced by a change in solvent from aqueous buffer to methanol in both single-molecule and bulk measurements were similar, indicating that, even at the single molecule level, the fluorescence spectrum is sensitive to microenvironmental changes of fluorophores. The time dependence of the fluorescence spectra of fluorophores attached to S1 molecules solely showed a fluctuation with time over a time scale of seconds. Because the fluorescence spectra of the same fluorophores directly conjugated to a glass surface remained constant, the spectral fluctuation observed for the fluorophores attached to S1 is most likely due to slow spontaneous conformational changes in the S1 molecule. Thus, single-molecule fluorescence spectroscopy appears to be a powerful tool to study the dynamic behavior of single biomolecules.
DOI：
10.1073/pnas.96.3.893
发表时间：
1999-02-02
影响因子：
11.1
作者：
Ha, TJ;Ting, AY;Weiss, S
通讯作者：
Weiss, S