Munshi S, Liljas L, Johnson J E
Department of Molecular Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, LaJolla, California 92037, USA.
Acta Crystallogr D Biol Crystallogr. 1998 Nov 1;54(Pt 6 Pt 2):1295-305. doi: 10.1107/s0907444998004454.
The structure of Nudaurelia capensis omega virus (NomegaV), a single-stranded RNA virus, was determined to 2.8 A resolution. Triclinic crystals (a = 413.6, b = 410.2, c = 419.7 A, alpha = 59.13, beta = 58.9, gamma = 64.0 degrees ) diffracted X-rays beyond 2.7 A resolution. The unit cell contained one icosahedral virus particle, providing 60-fold non-crystallographic symmetry (n.c.s.) and structural redundancy. The particle orientation in the unit cell was determined by self-rotation function analyses. Initial phases to 18 A resolution were derived from a hollow spherical model of 192 A outer radius and 139 A inner radius, filled with uniform electron density. Radii of the model were determined by maximizing the correlation of the model-based calculated data with the low-resolution X-ray diffraction and solution-scattering data. Phases were refined by 60-fold non-crystallographic electron-density averaging and extended in small steps to a resolution of 5 A. The phases obtained represented a mixture of four different phase sets, each consistent with the icosahedral symmetry constraints. The resulting electron density was not interpretable. A difference Fourier map computed with the native and an isomorphous heavy-atom derivative data sets and phases refined by real-space averaging was interpretable only if data within the 10 A resolution shell were used. Maps calculated with data significantly higher than 10 A resolution failed to display a constellation of heavy-atom sites consistent with the T = 4 icosahedral symmetry. Attempts to extend the phases beyond 10 A resolution, starting with either phases based on a model or single isomorphous replacement, were unsuccessful. Successful phase extension was achieved by computing the phases for the higher resolution reflections from a partial atomic model (poly gly) built into the averaged 10 A electron-density map. Phases from this model served as the starting point for n.c.s. phase refinement and extension to slightly higher resolution. The atomic model was improved at each extension interval and these phases were used for the subsequent phase calculation and extension. The entire polypeptide backbone corresponding to the NomegaV structure was built into the map at 4 A. The same procedure for phase refinement was used to extend the phases to 2.8 A in small increments of resolution. The overall molecular averaging R factor and correlation coefficient at 2.8 A resolution were 18.4% and 0.87, respectively.
卡普斯金眼大蚕蛾欧米伽病毒（NomegaV）是一种单链RNA病毒，其结构已确定至2.8埃分辨率。三斜晶体（a = 413.6，b = 410.2，c = 419.7埃，α = 59.13，β = 58.9，γ = 64.0度）的X射线衍射超过2.7埃分辨率。晶胞包含一个二十面体病毒粒子，提供60重非晶体学对称性（n.c.s.）和结构冗余。通过自旋转函数分析确定了晶胞中粒子的取向。18埃分辨率的初始相位源自一个外半径为192埃、内半径为139埃的空心球形模型，内部填充均匀电子密度。通过使基于模型计算的数据与低分辨率X射线衍射和溶液散射数据的相关性最大化来确定模型的半径。通过60重非晶体学电子密度平均对相位进行精修，并以小步长扩展至5埃分辨率。获得的相位代表四种不同相位集的混合，每种都符合二十面体对称约束。所得电子密度无法解释。只有使用10埃分辨率壳层内的数据时，用天然数据集和同晶型重原子衍生物数据集以及通过实空间平均精修的相位计算的差分傅里叶图才是可解释的。用分辨率显著高于10埃的数据计算的图谱未能显示与T = 4二十面体对称一致的重原子位点群。从基于模型的相位或单同晶置换开始将相位扩展至10埃分辨率以上的尝试均未成功。通过从构建到平均10埃电子密度图中的部分原子模型（聚甘氨酸）计算更高分辨率反射的相位，成功实现了相位扩展。该模型的相位作为n.c.s.相位精修和扩展至稍高分辨率的起点。在每个扩展间隔改进原子模型，并将这些相位用于后续的相位计算和扩展。在4埃分辨率下将与NomegaV结构相对应的整个多肽主链构建到图谱中。使用相同的相位精修程序以小分辨率增量将相位扩展至2.8埃。2.8埃分辨率下的总体分子平均R因子和相关系数分别为18.4%和0.87。
