Deep Mutational Scanning of SARS-CoV-2 Receptor Binding Domain Reveals Constraints on Folding and ACE2 Binding

By Tyler N. Starr, Allison J. Greaney, Sarah K. Hilton, Daniel Ellis, K. H. Crawford, A. Dingens, M. Navarro, John E. Bowen, M. A. Tortorici, A. Walls and 3 more. 2020

968 citations recorded by the source corpus.

The receptor binding domain (RBD) of the SARS-CoV-2 spike glycoprotein mediates viral attachment to ACE2 receptor and is a major determinant of host range and a dominant target of neutralizing antibodies. Here, we experimentally measure how all amino acid mutations to the RBD affect expression of folded protein and its affinity for ACE2. Most mutations are deleterious for RBD expression and ACE2 binding, and we identify constrained regions on the RBD’s surface that may be desirable targets for vaccines and antibody-based therapeutics. But a substantial number of mutations are well tolerated or even enhance ACE2 binding, including at ACE2 interface residues that vary across SARS-related coronaviruses. However, we find no evidence that these ACE2-affinity-enhancing mutations have been selected in current SARS-CoV-2 pandemic isolates. We present an interactive visualization and open analysis pipeline to facilitate use of our dataset for vaccine design and functional annotation of mutations observed during viral surveillance.

Subfields: Infectious Diseases, Molecular Biology, Radiology, Nuclear Medicine and Imaging

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Citations in the corpus

Learning inverse folding from millions of predicted structures (2022, methodology): “We assess whether inverse folding models can predict mutational effects on binding by leveraging a dataset generated by Starr et al. (2020) in which all single amino acid substitutions to the SARS-CoV-2 receptor binding domain (RBD) were experimentally measured for binding affinity to human ACE2.”
Elicitation of Potent Neutralizing Antibody Responses by Designed Protein Nanoparticle Vaccines for SARS-CoV-2 (2020, positive): “Structures of the SARS-CoV-2 S receptor-binding domain (RBD) in complex with ACE2 defined key residues involved in recognition and guide surveillance studies aiming to detect the emergence of mutants with altered binding affinity for ACE2 or distinct antigenicity (Lan et al., 2020; Shang et al., 202”
Evidence of escape of SARS-CoV-2 variant B.1.351 from natural and vaccine-induced sera (2021, positive): “In vitro evolution experiments have recently been reported (Starr et al., 2020) in which live virus has been induced to evolve in the face of immune pressure from either mAbs or polyclonal serum.”
Nanobody cocktails potently neutralize SARS-CoV-2 D614G N501Y variant and protect mice (2021, neutral): “N501 is one of six key contact residues within the RBD, and the N501Y mutation increases binding affinity to human and mouse ACE2 (22, 23).”
SARS-CoV-2 Omicron-B.1.1.529 leads to widespread escape from neutralizing antibody responses (2022, positive): “The standard non-cooperative Hill equation was ﬁtted by nonlinear least-squares regression with two additional parameters using Python 3.7 (Starr et al., 2020; Zahradnik et al., 2021a).”
Title: Mosaic nanoparticles elicit cross-reactive immune responses to zoonotic coronaviruses in mice (2020, neutral): “After incubation with 293TACE2 target cells for 48 hours at 37˚C, cells were washed twice with phosphate-buffered saline (PBS) and lysed with Luciferase Cell Culture Lysis 5x reagent (Promega).”
Estimated transmissibility and impact of SARS-CoV-2 lineage B.1.1.7 in England (2021, background): “Mutation N501Y is a key contact residue in the receptor binding domain and enhances virus binding affinity to human angiotensin-converting enzyme 2 (ACE2) ( 3 , 4 ).”