Trimeric SARS-CoV-2 Spike interacts with dimeric ACE2 with limited intra-Spike avidity
Xin X. Zhou,
Shion A Lim,
Susanna K. Elledge,
Nicholas J. Rettko,
Beth Shoshana Zha,
Lisa L. Kirkemo,
Josef A. Gramespacher,
Julio Cesar Cetrulo Lorenzi,
Davide F. Robbiani,
Michel C. Nussenzweig,
Paul D D Bieniasz,
Oren S. Rosenburg,
Kevin K. Leung,
James A Wells
Posted 21 May 2020
bioRxiv DOI: 10.1101/2020.05.21.109157
Posted 21 May 2020
A serious public health crisis is currently unfolding due to the SARS-CoV-2 pandemic. SARS-CoV-2 viral entry depends on an interaction between the receptor binding domain of the trimeric viral Spike protein (Spike-RBD) and the dimeric human angiotensin converting enzyme 2 (ACE2) receptor. While it is clear that strategies to block the Spike/ACE2 interaction are promising as anti-SARS-CoV-2 therapeutics, our current understanding is insufficient for the rational design of maximally effective therapeutic molecules. Here, we investigated the mechanism of Spike/ACE2 interaction by characterizing the binding affinity and kinetics of different multimeric forms of recombinant ACE2 and Spike-RBD domain. We also engineered ACE2 into a split Nanoluciferase-based reporter system to probe the conformational landscape of Spike-RBDs in the context of the Spike trimer. Interestingly, a dimeric form of ACE2, but not monomeric ACE2, binds with high affinity to Spike and blocks viral entry in pseudotyped virus and live SARS-CoV-2 virus neutralization assays. We show that dimeric ACE2 interacts with an RBD on Spike with limited intra-Spike avidity, which nonetheless contributes to the affinity of this interaction. Additionally, we demonstrate that a proportion of Spike can simultaneously interact with multiple ACE2 dimers, indicating that more than one RBD domain in a Spike trimer can adopt an ACE2-accessible 'up' conformation. Our findings have significant implications on the design strategies of therapeutic molecules that block the Spike/ACE2 interaction. The constructs we describe are freely available to the research community as molecular tools to further our understanding of SARS-CoV-2 biology. ### Competing Interest Statement The authors have declared no competing interest.
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