Astbury Centre for Structural Molecular Biology , University of Leeds , Leeds LS2 9JT , United Kingdom.
School of Chemistry , University of Leeds , Leeds LS2 9JT , United Kingdom.
J Am Chem Soc. 2019 Apr 3;141(13):5211-5219. doi: 10.1021/jacs.8b11480. Epub 2019 Mar 21.
The self-assembly of proteins into higher order structures is ubiquitous in living systems. It is also an essential process for the bottom-up creation of novel molecular architectures and devices for synthetic biology. However, the complexity of protein-protein interaction surfaces makes it challenging to mimic natural assembly processes in artificial systems. Indeed, many successful computationally designed protein assemblies are prescreened for "designability", limiting the choice of components. Here, we report a simple and pragmatic strategy to assemble chosen multisubunit proteins into more complex structures. A coiled-coil domain appended to one face of the pentameric cholera toxin B-subunit (CTB) enabled the ordered assembly of tubular supra-molecular complexes. Analysis of a tubular structure determined by X-ray crystallography has revealed a hierarchical assembly process that displays features reminiscent of the polymorphic assembly of polyomavirus proteins. The approach provides a simple and straightforward method to direct the assembly of protein building blocks which present either termini on a single face of an oligomer. This scaffolding approach can be used to generate bespoke supramolecular assemblies of functional proteins. Additionally, structural resolution of the scaffolded assemblies highlight "native-state" forced protein-protein interfaces, which may prove useful as starting conformations for future computational design.
蛋白质自行组装成更高阶结构在生命系统中普遍存在。这也是自下而上创造新型分子结构和合成生物学器件的必要过程。然而，蛋白质-蛋白质相互作用表面的复杂性使得在人工系统中模拟自然组装过程具有挑战性。事实上，许多成功的计算设计蛋白质组装体都经过了“可设计性”的预先筛选，限制了组件的选择。在这里，我们报告了一种简单而实用的策略，可将选定的多亚基蛋白质组装成更复杂的结构。卷曲螺旋结构域添加到五聚体霍乱毒素 B 亚基（CTB）的一个面上，可实现管状超分子复合物的有序组装。X 射线晶体学分析确定的管状结构揭示了一个分层组装过程，其特征类似于多瘤病毒蛋白的多态性组装。该方法提供了一种简单直接的方法来指导呈现于寡聚体单一面上的蛋白质构建块的组装。这种支架方法可用于生成功能蛋白的定制超分子组装体。此外，支架组装体的结构分辨率突出了“天然状态”强制的蛋白质-蛋白质界面，这可能作为未来计算设计的起始构象有用。
