Quantitative Microscopy: Protein Dynamics and Membrane Organisation
Quantitative Microscopy: Protein Dynamics and Membrane Organisation
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DOI：
10.1111/j.1600-0854.2009.00908.x
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发表时间：
2009-08-01
期刊：
影响因子：
4.5
通讯作者：
Gaus, Katharina
中科院分区：
文献类型：
--
作者：
Owen, Dylan M.;Williamson, David;Gaus, Katharina
膜蛋白的移动性是其相互作用能力和蛋白质功能的关键决定因素。细胞膜的异质性赋予蛋白质不同类型的运动；不动，随机布朗运动，反常亚扩散，跳跃或受限扩散，或定向流动。因此，定量蛋白质的运动可以深入了解细胞膜的横向组织，特别是具有高粘度的膜微域，如脂筏。在这篇综述中，我们研究了三种主要的蛋白质动力学分析技术的假设和发现：光漂白后的荧光恢复，单粒子跟踪和荧光相关光谱。这些技术，以及数据分析中使用的物理模型，已经变得越来越复杂，并提供了蛋白质动力学和细胞膜结构域的生物物理特性的前所未有的细节。然而，尽管取得了这些进展，在胆固醇依赖性脂质微结构域、蛋白质-蛋白质相互作用以及潜在细胞骨架的影响之间的关系仍然存在重大的未知因素。新的多维显微镜方法可以提供更高的时间和空间分辨率，从而更准确地量化活细胞中的蛋白质和膜动力学。
The mobility of membrane proteins is a critical determinant of their interaction capabilities and protein functions. The heterogeneity of cell membranes imparts different types of motion onto proteins; immobility, random Brownian motion, anomalous sub-diffusion, 'hop' or confined diffusion, or directed flow. Quantifying the motion of proteins therefore enables insights into the lateral organisation of cell membranes, particularly membrane microdomains with high viscosity such as lipid rafts. In this review, we examine the hypotheses and findings of three main techniques for analysing protein dynamics: fluorescence recovery after photobleaching, single particle tracking and fluorescence correlation spectroscopy. These techniques, and the physical models employed in data analysis, have become increasingly sophisticated and provide unprecedented details of the biophysical properties of protein dynamics and membrane domains in cell membranes. Yet despite these advances, there remain significant unknowns in the relationships between cholesterol-dependent lipid microdomains, protein-protein interactions, and the effect of the underlying cytoskeleton. New multi-dimensional microscopy approaches may afford greater temporal and spatial resolution resulting in more accurate quantification of protein and membrane dynamics in live cells.