Determination of the orientational distribution and orientation factor for transfer between membrane-bound fluorophores using a confocal microscope
Determination of the orientational distribution and orientation factor for transfer between membrane-bound fluorophores using a confocal microscope
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DOI：
10.1529/biophysj.106.080713
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发表时间：
2006-08-01
影响因子：
3.4
通讯作者：
Rigby, Paul
中科院分区：
文献类型：
--
作者：
Corry, Ben;Jayatilaka, Dylan;Rigby, Paul
定向荧光团已经成为物理化学、生物化学和最近的结构生物学中的有用工具，这是由于它们发射的光的偏振性质以及能量可以在它们之间转移的事实。我们提出了一个实用的方案，其中发射荧光的强度的测量可以用来确定膜结合的吸收跃迁矩的取向的平均值和分布的限制。荧光团。我们演示了如何信息的方向。荧光团可用于计算FRET光谱学中所需的取向因子k（2）。我们使用AlexaFluor探针结合到球形脂质体中的MscL机械敏感跨膜通道蛋白的图像来说明该方法。
Orientational fluorophores have been a useful tool in physical chemistry, biochemistry, and more recently structural biology due to the polarized nature of the light they emit and that fact that energy can be transferred between them. We present a practical scheme in which measurements of the intensity of emitted fluorescence can be used to determine limits on the mean and distribution of orientation of the absorption transition moment of membrane-bound. uorophores. We demonstrate how information about the orientation of. uorophores can be used to calculate the orientation factor k(2) required for use in FRET spectroscopy. We illustrate the method using images of AlexaFluor probes bound to MscL mechanosensitive transmembrane channel proteins in spherical liposomes.