Trimerization of the N-terminal Domain of ACE2 for Bifunctional Trapping of Future SARS-CoV-2 Variants

摘要 由SARS-CoV-2引发的新冠肺炎大流行已导致188人感染3,830万人 到2020年10月14日死亡人数超过108万的国家和地区。虽然各种疫苗都是 在加快开发的情况下，人们严重担心SARS-COV-2的未来变种将进化和 在不久的将来感染人类。我们建议开发一种新型的通用分子阻滞剂来对抗 基于hACE2 N端三螺旋束(3HB，残基20-99)的SARS-CoV-2，这是公认的 通过SARS-CoV和SARS-CoV-2作为细胞表面受体感染人细胞。基于我们之前的工作 在三螺旋束结构域的三聚化中，我们假设hACE2的N-末端3HB结构域可以是 三聚体，与SARS-CoV-2所有变种的RBD可能具有高度的效力和特异性结合。二 这个项目将追求具体的目标。在具体目标1中，我们将开发基于N-的三聚体陷阱。 高特异性与SARS-CoV-2 RBD结合的hACE2末端三螺旋束(3HB) 和力量。在特定的目标2中，我们将使用定向分子进化来从一个 并将其与hACE2的N端3HB融合，构建双功能SARS-CoV-2陷阱 高度特异地结合病毒RBD，同时抑制其由Furin介导的预激活。由此产生的新型SARS-CoV- 2捕获分子将有可能被广泛应用于阻断hACE2介导的感染。 SARS-CoV-2未来变种的大部分，如果不是全部的话。

Abstract The Covid-19 pandemic caused by SARS-Cov-2 has resulted in 38.3 million infection cases across 188 countries and territories with more than 1.08 million deaths by October 14 2020. While various vaccines are under expedite development, there is a serious concern that future variants of SARS-COV-2 will evolve and infect humans in the near future. We propose to develop a novel class of universal molecular blockers against SARS-CoV-2 based on the N-terminal three-helix bundle (3HB, residues 20-99) of hACE2, which is recognized by both SARS-CoV and SARS-CoV-2 as cell surface receptor to infect human cells. Based on our previous work in trimerizing three-helical bundle domains, we hypothesize that the N-terminal 3HB domain of hACE2 can be trimerized for highly potent and specific binding with presumably the RBD of all variants of SARS-CoV-2. Two specific aims will be pursued in this project. In specific aim 1, we will develop a trimeric trap based on the N- terminal three-helix bundle (3HB) of hACE2 that trivalently binds to the RBD of SARS-CoV-2 with high specificity and potency. In specific aim 2, we will use directed molecular evolution to identify a human furin inhibitor from a protein domain library and fuse it with the N-terminal 3HB of hACE2 for a bifunctional SARS-CoV-2 trap that highly specifically binds viral RBD while inhibits its furin-mediated preactivation. The resulting novel SARS-CoV- 2 trapping molecules will have the potential to be universally applied to block hACE2-mediated infection by vast majority of, if not all, future variants of SARS-CoV-2.