Edmiston P L, Saavedra S S
Department of Chemistry, University of Arizona, Tucson 85721, USA.
Biophys J. 1998 Feb;74(2 Pt 1):999-1006. doi: 10.1016/S0006-3495(98)74024-X.
Molecular orientation in a hydrated monolayer film of yeast cytochrome c, immobilized via disulfide bonding between Cys-102 and a pyridyl disulfide-capped phospholipid bilayer deposited from an air-water interface onto glass substrates, was investigated. The orientation distribution of the heme groups in the protein film was determined using a combination of absorption linear dichroism, measured in a planarintegrated optical waveguide-attenuated total reflection geometry- and fluorescence anisotropy, measured in a total internal reflection geometry. A gaussian model for the orientation distribution was used to recover the mean heme tilt angle and angular distribution about the mean, which were 40 and 11 degrees, respectively. Additional experiments showed that a large fraction of the cytochrome c was disulfide bonded to the bilayer, which correlates with the high degree of macroscopic order in the protein film. However, a subpopulation of yeast cytochrome c molecules in the film (approximately 30% of the total) appeared to be nonspecifically adsorbed. The orientation distribution of this subpopulation was found to be much broader than the specifically bound fraction.
研究了通过半胱氨酸-102与从气-水界面沉积到玻璃基板上的吡啶二硫键封端的磷脂双层之间的二硫键固定的酵母细胞色素c水合单层膜中的分子取向。使用吸收线性二色性（在平面集成光波导衰减全反射几何结构中测量）和荧光各向异性（在全内反射几何结构中测量）相结合的方法，确定蛋白质膜中血红素基团的取向分布。使用取向分布的高斯模型来恢复平均血红素倾斜角和围绕平均值的角度分布，分别为40度和11度。额外的实验表明，大部分细胞色素c通过二硫键与双层结合，这与蛋白质膜中的高度宏观有序性相关。然而，膜中的酵母细胞色素c分子亚群（约占总数的30%）似乎是非特异性吸附的。发现该亚群的取向分布比特异性结合部分宽得多。
