TIME ULTRA HIGH RESOLUTION OF GUANINE NUCLEOTIDE DISSOCIATION INHIBITOR

鸟嘌呤核苷酸解离抑制剂（GDI）调节 在囊泡运输中恢复Rab GTP酶，同时抑制GDP 与Rabs分离 可溶性复合物，GDI-Rab，作为 用于将Rab递送至新形成的囊泡的胞质储库， 其中Rab蛋白被激活为GTP形式。 晶体 该55 kDa蛋白质的结构先前已确定为1.8 E分辨率。 SSRL光束线9-1在波长 0.79这使我们能够收集超高分辨率的数据集 在-176 ℃下达到1.04 E。高分辨率和中等分辨率（1.4 E）数据集， 从单晶中收集的数据进行了合并和处理 使用总体Rmerge为7%的MOLSFLM（所有数据）。 锻 使用SHELXL-97进行，在该决议中， 执行单独的各向异性热位移参数 精致。 发现许多残留物具有多个 构象和一些不正确的旋转异构体被鉴定， 特别是对于亮氨酸残基。 此外，His标记的 用于纯化目的的N末端残基 在电子密度图中可以清楚地识别出来。 400多 在精制过程中逐步加入水分子， 氢位于接近精制结束的位置。 当前 Rcryst和Rfree值分别为15.6%和19.0%。

The guanine nucleotide dissociation inhibitor (GDI) regulates the retrieval of Rab GTPases in vesicular traffic, while inhibiting GDP dissociation from Rabs. The soluble complex, GDI-Rab, serves as a cytosolic reservoir for the delivery of Rab to newly formed vesicles, where the Rab protein is activated to the GTP form. The crystal structure of this 55 kDa protein had been previously determined to 1.8 E resolution. The SSRL beamline 9-1 was optimized at a wavelength 0.79 E, which allowed us to collect an ultra high resolution data set to 1.04 E at -176 C. High and medium (1.4 E) resolution data sets were collected from a single crystal and the data were merged and processed using MOLSFLM with an overall Rmerge of 7% (all data). Refinement was carried out using SHELXL-97 which, at this resolution, allowed us to perform individual anisotropic thermal displacement parameter refinement. A number of residues were found to possess multiple conformations and some incorrect rotomers were identified, particularly for leucine residues. In addition, the His-tagged residues at the N terminus that were used for purification purpose could now be clearly identified in the electron density map. Over 400 water molecules were added stepwise during the refinement and hydrogens were positioned towards the end of refinement. The current Rcryst and Rfree values are 15.6% and 19.0%, respectively.