Maita Nobuo
Division of Disease Proteomics, Institute of Advanced Medical Sciences , Tokushima University , 3-18-15 Kuramoto-cho , Tokushima 770-8503 , Japan.
J Am Chem Soc. 2018 Oct 24;140(42):13546-13549. doi: 10.1021/jacs.8b07512. Epub 2018 Oct 10.
The protein crystallization process requires screening of a large number of conditions using a large quantity of high-purity protein, which makes crystal structure analysis difficult. Thus, the development of easy and versatile protein crystallization techniques is both extremely desirable and highly challenging. Here I demonstrate the crystallization and structure determination of ubiquitin by genetic fusion to the highly porous honeycomb lattice of R1EN. I successfully crystallized and collected X-ray data from three R1EN-ubiquitin constructs with various linker lengths under the same conditions as the original R1EN. The crystals diffracted to 1.7-2.4 Å resolution, and the ubiquitin structures were determined with results almost identical to the previously published structure. Moreover, the ubiquitin structure could be solved by molecular replacement using R1EN alone. This method may reduce the effort required for crystallization screening and is applicable to de novo protein structure determination.
蛋白质结晶过程需要使用大量高纯度蛋白质筛选大量条件，这使得晶体结构分析变得困难。因此，开发简便通用的蛋白质结晶技术既极为必要又极具挑战性。在此，我展示了通过与高度多孔的R1EN蜂窝晶格进行基因融合来实现泛素的结晶和结构测定。我在与原始R1EN相同的条件下，成功地对三种具有不同连接子长度的R1EN-泛素构建体进行了结晶，并收集了X射线数据。这些晶体的衍射分辨率达到1.7 - 2.4 Å，所确定的泛素结构与先前发表的结构几乎相同。此外，仅使用R1EN通过分子置换就能解析泛素结构。该方法可能会减少结晶筛选所需的工作量，并且适用于全新蛋白质结构的测定。
