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,
Lisa L. Kirkemo,
Josef A. Gramespacher,
Julio Cesar Cetrulo Lorenzi,
Davide F. Robbiani,
Michel C. Nussenzweig,
Paul 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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