Thiaudière E, Soekarjo M, Kuchinka E, Kuhn A, Vogel H
Institute of Physical Chemistry, Swiss Federal Institute of Technology, Lausanne.
Biochemistry. 1993 Nov 16;32(45):12186-96. doi: 10.1021/bi00096a031.
A new, simple, and efficient purification method has been developed for the extremely hydrophobic M13 procoat, M13 coat, and Pf3 coat proteins. Homogeneous preparations were obtained in 2-propanol/0.1% TFA, where M13 coat protein is found to be dissolved in a monomeric form, and the two other proteins as dimers or trimers. The conformations of these particular proteins in different environments have been determined by circular dichroism and infrared spectroscopy. In organic solvents, the proteins adopt a conformation with an average helix content of 90%. In lipid bilayers composed of phosphatidylcholine and phosphatidylglycerol lipids, the average helix content is 50% for M13 procoat protein, 60% for M13 coat protein, and 75% for Pf3 coat protein. The orientational order parameter S alpha of the protein helices in planar lipid bilayers have been determined by polarized infrared measurements in the amide I spectral range. The helices of the three proteins are oriented preferentially parallel to the membrane normal, with S alpha = 0.63 for M13 procoat protein, S alpha = 0.58 for Pf3 coat protein, and a distinctly higher value of S alpha = 0.81 for M13 coat protein.
已开发出一种针对极疏水的M13前衣壳蛋白、M13衣壳蛋白和Pf3衣壳蛋白的新型、简单且高效的纯化方法。在2-丙醇/0.1%三氟乙酸中获得了均一制剂，其中发现M13衣壳蛋白以单体形式溶解，另外两种蛋白则以二聚体或三聚体形式存在。通过圆二色光谱和红外光谱确定了这些特定蛋白在不同环境中的构象。在有机溶剂中，这些蛋白呈现出平均螺旋含量为90%的构象。在由磷脂酰胆碱和磷脂酰甘油脂质组成的脂质双层中，M13前衣壳蛋白的平均螺旋含量为50%，M13衣壳蛋白为60%，Pf3衣壳蛋白为75%。通过在酰胺I光谱范围内的偏振红外测量确定了平面脂质双层中蛋白螺旋的取向序参数Sα。这三种蛋白的螺旋优先平行于膜法线取向，M13前衣壳蛋白的Sα = 0.63，Pf3衣壳蛋白的Sα = 0.58，而M13衣壳蛋白的Sα值明显更高，为0.81。
