Division of Basic Sciences, Fred Hutchinson Cancer Research Center, 1100 Fairview Ave. North, Seattle, WA, 98109, USA.
Lumen Bioscience Inc., 1441 North 34th Street, Seattle, WA, 98103, USA.
Commun Biol. 2021 Oct 29;4(1):1240. doi: 10.1038/s42003-021-02766-y.
Circular tandem repeat proteins ('cTRPs') are de novo designed protein scaffolds (in this and prior studies, based on antiparallel two-helix bundles) that contain repeated protein sequences and structural motifs and form closed circular structures. They can display significant stability and solubility, a wide range of sizes, and are useful as protein display particles for biotechnology applications. However, cTRPs also demonstrate inefficient self-assembly from smaller subunits. In this study, we describe a new generation of cTRPs, with longer repeats and increased interaction surfaces, which enhanced the self-assembly of two significantly different sizes of homotrimeric constructs. Finally, we demonstrated functionalization of these constructs with (1) a hexameric array of peptide-binding SH2 domains, and (2) a trimeric array of anti-SARS CoV-2 VHH domains. The latter proved capable of sub-nanomolar binding affinities towards the viral receptor binding domain and potent viral neutralization function.
环形串联重复蛋白（“cTRPs”）是从头设计的蛋白质支架（在本研究和之前的研究中，基于反平行双螺旋束），包含重复的蛋白质序列和结构基序，并形成封闭的环形结构。它们可以表现出显著的稳定性和可溶性，具有广泛的大小，并可用作生物技术应用的蛋白质展示颗粒。然而，cTRPs 也表现出从小的亚基组装效率低下。在这项研究中，我们描述了新一代的 cTRPs，具有更长的重复序列和增加的相互作用表面，这增强了两种明显不同大小的同源三聚体构建体的自组装。最后，我们证明了这些构建体的功能化，（1）具有六聚体肽结合 SH2 结构域的阵列，和（2）具有抗 SARS CoV-2 VHH 结构域的三聚体阵列。后者被证明能够对病毒受体结合域具有亚纳摩尔结合亲和力和有效的病毒中和功能。
