Department of Synthetic Biology and Immunology, National Institute of Chemistry, Ljubljana, Slovenia.
Graduate School of Biomedicine, University of Ljubljana, Ljubljana, Slovenia.
Nat Biotechnol. 2017 Nov;35(11):1094-1101. doi: 10.1038/nbt.3994. Epub 2017 Oct 16.
Polypeptides and polynucleotides are natural programmable biopolymers that can self-assemble into complex tertiary structures. We describe a system analogous to designed DNA nanostructures in which protein coiled-coil (CC) dimers serve as building blocks for modular de novo design of polyhedral protein cages that efficiently self-assemble in vitro and in vivo. We produced and characterized >20 single-chain protein cages in three shapes-tetrahedron, four-sided pyramid, and triangular prism-with the largest containing >700 amino-acid residues and measuring 11 nm in diameter. Their stability and folding kinetics were similar to those of natural proteins. Solution small-angle X-ray scattering (SAXS), electron microscopy (EM), and biophysical analysis confirmed agreement of the expressed structures with the designs. We also demonstrated self-assembly of a tetrahedral structure in bacteria, mammalian cells, and mice without evidence of inflammation. A semi-automated computational design platform and a toolbox of CC building modules are provided to enable the design of protein cages in any polyhedral shape.
多肽和多核苷酸是天然可编程的生物聚合物，可以自组装成复杂的三级结构。我们描述了一个类似于设计 DNA 纳米结构的系统，其中蛋白质卷曲螺旋（CC）二聚体作为模块从头设计多面蛋白笼的构建块，这些蛋白笼能够在体外和体内有效地自我组装。我们生产并表征了 >20 种单链蛋白笼，形状为四面体、四面金字塔和三棱柱，其中最大的含有 >700 个氨基酸残基，直径为 11nm。它们的稳定性和折叠动力学与天然蛋白质相似。溶液小角 X 射线散射（SAXS）、电子显微镜（EM）和生物物理分析证实了表达结构与设计的一致性。我们还证明了在细菌、哺乳动物细胞和小鼠中四面体结构的自组装，没有炎症的证据。提供了一个半自动的计算设计平台和一个 CC 构建模块工具箱，以实现任何多面体形的蛋白笼设计。
