Crystal Structure Determination of Ubiquitin by Fusion to a Protein That Forms a Highly Porous Crystal Lattice
Crystal Structure Determination of Ubiquitin by Fusion to a Protein That Forms a Highly Porous Crystal Lattice
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DOI：
10.1021/jacs.8b07512
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发表时间：
2018-10-24
影响因子：
15
通讯作者：
Maita, Nobuo
中科院分区：
文献类型：
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作者：
Maita, Nobuo
蛋白质结晶过程需要使用大量高纯度蛋白质来筛选大量条件，这使得晶体结构分析困难。因此，开发简单和通用的蛋白质结晶技术是非常可取的，也是极具挑战性的。在这里，我展示了遗传融合的高度多孔蜂窝状晶格的R1 EN的泛素的结晶和结构测定。我成功地从三个R1 EN-泛素结构中结晶并收集了X射线数据，这些结构在与原始R1 EN相同的条件下具有不同的接头长度。晶体衍射至1.7-2.4埃分辨率，并且确定了泛素结构，结果与先前公布的结构几乎相同。此外，泛素结构可以通过单独使用R1 EN的分子置换来解决。该方法可以减少结晶筛选所需的工作量，适用于从头蛋白质结构测定。
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 angstrom 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.