Gryczynski I, Johnson M L, Lakowicz J R
University of Maryland School of Medicine, Department of Biological Chemistry, Baltimore 21201.
Biophys Chem. 1994 Sep;52(1):1-13. doi: 10.1016/0301-4622(94)00031-x.
We describe the theory and practical aspects of analyzing fluorescence anisotropy decays in terms of correlation times distributions. In our model the rotational motions of the fluorophores were described using Gaussian or Lorentzian distributions of the correlation times. The theory is presented both for time and frequency-domain measurements, although the simulations and measurements are focused on the frequency-domain measurements of the anisotropy decays. Analysis of simulated data is presented to illustrate the nature of the data and the resolution which can be expected with presently available frequency-domain measurements. Additionally, we describe experimental data for samples where one can reasonably expect a single exponential and/or discrete multi-exponential correlation time distributions, and for samples where the anisotropy decay might be expected to display a distribution of correlation times. These samples include small single tryptophan peptides in propylene glycol, the single tryptophan residue in S. Nuclease, and the single tryptophan residue in the native and partially unfolded states of ribonuclease T1.
我们描述了根据相关时间分布分析荧光各向异性衰减的理论和实际方面。在我们的模型中，使用相关时间的高斯或洛伦兹分布来描述荧光团的旋转运动。尽管模拟和测量集中在各向异性衰减的频域测量上，但该理论同时适用于时域和频域测量。给出了模拟数据分析，以说明数据的性质以及目前可用的频域测量所能达到的分辨率。此外，我们描述了一些样品的实验数据，在这些样品中可以合理预期单一指数和/或离散多指数相关时间分布，以及在各向异性衰减可能呈现相关时间分布的样品。这些样品包括丙二醇中的小单色氨酸肽、核酸酶S中的单个色氨酸残基，以及核糖核酸酶T1天然和部分未折叠状态下的单个色氨酸残基。
