Ratto Timothy V, Longo Marjorie L
Biophysics Graduate Group, Division of Biological Sciences, University of California-Davis, One Shields Avenue, Davis, CA 95616, USA.
Biophys J. 2002 Dec;83(6):3380-92. doi: 10.1016/S0006-3495(02)75338-1.
Proteins and other macromolecules are believed to hinder molecular lateral diffusion in cellular membranes. We have constructed a well-characterized model system to better understand how obstacles in lipid bilayers obstruct diffusion. Fluorescence recovery after photobleaching was used to measure the lateral diffusion coefficient in single supported bilayers composed of mixtures of 1,2-dilauroylphosphotidylcholine (DLPC) and 1,2-distearoylphosphotidylcholine (DSPC). Because these lipids are immiscible and phase separate at room temperature, a novel quenching technique allowed us to construct fluid DLPC bilayers containing small disk-shaped gel-phase DSPC domains that acted as obstacles to lateral diffusion. Our experimental setup enabled us to analyze the same samples with atomic force microscopy and exactly characterize the size, shape, and number of gel-phase domains before measuring the obstacle-dependent diffusion coefficient. Lateral obstructed diffusion was found to be dependent on obstacle area fraction, size, and geometry. Analysis of our results using a free area diffusion model shows the possibility of unexpected long-range ordering of fluid-phase lipids around the gel-phase obstacles. This lipid ordering has implications for lipid-mediated protein interactions in cellular membranes.
蛋白质和其他大分子被认为会阻碍细胞膜中的分子横向扩散。我们构建了一个特征明确的模型系统，以更好地理解脂质双层中的障碍物是如何阻碍扩散的。采用光漂白后荧光恢复技术来测量由1,2 - 二月桂酰磷脂酰胆碱（DLPC）和1,2 - 二硬脂酰磷脂酰胆碱（DSPC）混合物组成的单支撑双层膜中的横向扩散系数。由于这些脂质在室温下互不相溶且会发生相分离，一种新颖的猝灭技术使我们能够构建含有小圆盘状凝胶相DSPC结构域的流体DLPC双层膜，这些结构域对横向扩散起到障碍物的作用。我们的实验装置使我们能够在测量依赖于障碍物的扩散系数之前，用原子力显微镜分析相同的样品，并准确表征凝胶相结构域的大小、形状和数量。发现横向受阻扩散取决于障碍物的面积分数、大小和几何形状。使用自由面积扩散模型对我们的结果进行分析表明，在凝胶相障碍物周围的流体相脂质可能存在意想不到的长程有序排列。这种脂质有序排列对细胞膜中脂质介导的蛋白质相互作用具有重要意义。
