Cherezov V, Fersi H, Caffrey M
Biochemistry, Biophysics, and Chemistry, The Ohio State University, Columbus, Ohio 43210, USA.
Biophys J. 2001 Jul;81(1):225-42. doi: 10.1016/S0006-3495(01)75694-9.
The in meso method for growing crystals of membrane proteins uses a spontaneously forming lipidic cubic mesophase. The detergent-solubilized protein is dispersed with lipid, typically monoolein, and in so doing the cubic phase self-assembles. A precipitant is added to trigger crystal nucleation and growth. The commercial screen solution series are convenient for use in crystallization trials. The aim of this study was to determine which of the Hampton Screen and Screen 2 series of solutions are compatible with the in meso method. These screens contain components any of which could destroy the cubic phase. X-ray diffraction was used for phase identification and for microstructure characterization. The study was done at 4 degrees C and at 20 degrees C. Two types of sample preparations were examined. One used an excess of half-strength screen solution (Prep. 1). The other used a limiting quantity of undiluted screen solution (Prep. 2). At 20 degrees C, over 90% of the screen solutions produced the cubic phase with Prep. 1. This figure dropped to 50% with Prep. 2. In contrast, 50 to 60% of the screens were cubic phase compatible at 4 degrees C under Prep. 1 conditions. The figure fell to 25% with Prep. 2. The mode of action of the diverse screen components are explained on the basis of the phase properties of the monoolein/water system.
用于生长膜蛋白晶体的“in meso”方法使用自发形成的脂质立方相。去污剂增溶的蛋白质与脂质（通常是单油酸甘油酯）混合，在此过程中立方相自组装。加入沉淀剂以触发晶体成核和生长。商业筛选溶液系列便于用于结晶试验。本研究的目的是确定汉普顿筛选溶液和筛选2系列溶液中哪些与“in meso”方法兼容。这些筛选溶液包含任何一种都可能破坏立方相的成分。使用X射线衍射进行相鉴定和微观结构表征。该研究在4℃和20℃下进行。检查了两种类型的样品制备方法。一种使用过量的半强度筛选溶液（制备方法1）。另一种使用限量的未稀释筛选溶液（制备方法2）。在20℃下，超过90%的筛选溶液在制备方法1下产生立方相。在制备方法2下，这一数字降至50%。相比之下，在4℃下，制备方法1条件下50%至60%的筛选溶液与立方相兼容。在制备方法2下，这一数字降至25%。根据单油酸甘油酯/水体系的相性质解释了各种筛选成分的作用方式。
