Abstract

The Spike (S) protein is the main handle for SARS-CoV-2 to enter host cells via surface ACE2 receptors. How ACE2 binding activates proteolysis of S protein is unknown. Here, using amide hydrogen-deuterium exchange mass spectrometry and molecular dynamics simulations, we have mapped the S:ACE2 interaction interface and uncovered long-range allosteric propagation of ACE2 binding to sites necessary for host-mediated proteolysis of S protein, critical for viral host entry. Unexpectedly, ACE2 binding enhances dynamics at a distal S1/S2 cleavage site and flanking protease docking site ~27 Å away while dampening dynamics of the stalk hinge (central helix and heptad repeat) regions ~130 Å away. This highlights that the stalk and proteolysis sites of the S protein are dynamic hotspots in the pre-fusion state. Our findings provide a dynamics map of the S:ACE2 interface in solution and also offer mechanistic insights into how ACE2 binding is allosterically coupled to distal proteolytic processing sites and viral-host membrane fusion. Our findings highlight protease docking sites flanking the S1/S2 cleavage site, fusion peptide and heptad repeat 1 (HR1) as alternate allosteric hotspot targets for potential therapeutic development.

Data availability

All data generated or analysed during this study are included in the manuscript and supporting files. Source data files have been provided for Figures 2, 3, 4 and 5.

Article and author information

Author details

  1. Palur V Raghuvamsi

    Biological Sciences, National University of Singapore, Singapore, Singapore
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-0897-6935
  2. Nikhil Kumar Tulsian

    Biological Sciences, National University of Singapore, Singapore, Singapore
    Competing interests
    The authors declare that no competing interests exist.
  3. Firdaus Samsudin

    Bioinformatics Institute, A*STAR, Singapore, Singapore
    Competing interests
    The authors declare that no competing interests exist.
  4. Xinlei Qian

    Life Sciences Institute, National University of Singapore, Singapore, Singapore
    Competing interests
    The authors declare that no competing interests exist.
  5. Kiren Purushotorman

    Microbiology and Immunology, National University of Singapore, Singapore, Singapore
    Competing interests
    The authors declare that no competing interests exist.
  6. Gu Yue

    Microbiology and Immunology, National University of Singapore, Singapore, Singapore
    Competing interests
    The authors declare that no competing interests exist.
  7. Mary M Kozma

    Life Sciences Institute, National University of Singapore, Singapore, Singapore
    Competing interests
    The authors declare that no competing interests exist.
  8. Wong Yee Hwa

    School of Biological Sciences, National Technological University, Singapore, Singapore
    Competing interests
    The authors declare that no competing interests exist.
  9. Julien Lescar

    School of Biological Sciences, National Technological University, Singapore, Singapore
    Competing interests
    The authors declare that no competing interests exist.
  10. Peter J Bond

    Microbiology and Immunology, National University of Singapore, Singapore, Singapore
    For correspondence
    peterjb@bii.a-star.edu.sg
    Competing interests
    The authors declare that no competing interests exist.
  11. Paul Anthony MacAry

    Microbiology and Immunology, National University of Singapore, Singapore, Singapore
    For correspondence
    micpam@nus.edu.sg
    Competing interests
    The authors declare that no competing interests exist.
  12. Ganesh Srinivasan Anand

    Biological Sciences, National University of Singapore, Singapore, Singapore
    For correspondence
    gsa5089@psu.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8995-3067

Funding

Ministry of Education - Singapore (Research Fellowship)

  • Ganesh Srinivasan Anand

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Donald Hamelberg, Georgia State University, United States

Version history

  1. Received: October 1, 2020
  2. Accepted: February 5, 2021
  3. Accepted Manuscript published: February 8, 2021 (version 1)
  4. Version of Record published: March 4, 2021 (version 2)

Copyright

© 2021, Raghuvamsi et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 9,304
    views
  • 1,161
    downloads
  • 99
    citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Palur V Raghuvamsi
  2. Nikhil Kumar Tulsian
  3. Firdaus Samsudin
  4. Xinlei Qian
  5. Kiren Purushotorman
  6. Gu Yue
  7. Mary M Kozma
  8. Wong Yee Hwa
  9. Julien Lescar
  10. Peter J Bond
  11. Paul Anthony MacAry
  12. Ganesh Srinivasan Anand
(2021)
SARS-CoV-2 S protein:ACE2 interaction reveals novel allosteric targets
eLife 10:e63646.
https://doi.org/10.7554/eLife.63646

Share this article

https://doi.org/10.7554/eLife.63646

Further reading

    1. Biochemistry and Chemical Biology
    2. Cell Biology
    Christopher TA Lewis, Elise G Melhedegaard ... Julien Ochala
    Research Article

    Hibernation is a period of metabolic suppression utilized by many small and large mammal species to survive during winter periods. As the underlying cellular and molecular mechanisms remain incompletely understood, our study aimed to determine whether skeletal muscle myosin and its metabolic efficiency undergo alterations during hibernation to optimize energy utilization. We isolated muscle fibers from small hibernators, Ictidomys tridecemlineatus and Eliomys quercinus and larger hibernators, Ursus arctos and Ursus americanus. We then conducted loaded Mant-ATP chase experiments alongside X-ray diffraction to measure resting myosin dynamics and its ATP demand. In parallel, we performed multiple proteomics analyses. Our results showed a preservation of myosin structure in U. arctos and U. americanus during hibernation, whilst in I. tridecemlineatus and E. quercinus, changes in myosin metabolic states during torpor unexpectedly led to higher levels in energy expenditure of type II, fast-twitch muscle fibers at ambient lab temperatures (20 °C). Upon repeating loaded Mant-ATP chase experiments at 8 °C (near the body temperature of torpid animals), we found that myosin ATP consumption in type II muscle fibers was reduced by 77–107% during torpor compared to active periods. Additionally, we observed Myh2 hyper-phosphorylation during torpor in I. tridecemilineatus, which was predicted to stabilize the myosin molecule. This may act as a potential molecular mechanism mitigating myosin-associated increases in skeletal muscle energy expenditure during periods of torpor in response to cold exposure. Altogether, we demonstrate that resting myosin is altered in hibernating mammals, contributing to significant changes to the ATP consumption of skeletal muscle. Additionally, we observe that it is further altered in response to cold exposure and highlight myosin as a potentially contributor to skeletal muscle non-shivering thermogenesis.

    1. Biochemistry and Chemical Biology
    2. Neuroscience
    Maximilian Nagel, Marco Niestroj ... Marc Spehr
    Research Article

    In most mammals, conspecific chemosensory communication relies on semiochemical release within complex bodily secretions and subsequent stimulus detection by the vomeronasal organ (VNO). Urine, a rich source of ethologically relevant chemosignals, conveys detailed information about sex, social hierarchy, health, and reproductive state, which becomes accessible to a conspecific via vomeronasal sampling. So far, however, numerous aspects of social chemosignaling along the vomeronasal pathway remain unclear. Moreover, since virtually all research on vomeronasal physiology is based on secretions derived from inbred laboratory mice, it remains uncertain whether such stimuli provide a true representation of potentially more relevant cues found in the wild. Here, we combine a robust low-noise VNO activity assay with comparative molecular profiling of sex- and strain-specific mouse urine samples from two inbred laboratory strains as well as from wild mice. With comprehensive molecular portraits of these secretions, VNO activity analysis now enables us to (i) assess whether and, if so, how much sex/strain-selective ‘raw’ chemical information in urine is accessible via vomeronasal sampling; (ii) identify which chemicals exhibit sufficient discriminatory power to signal an animal’s sex, strain, or both; (iii) determine the extent to which wild mouse secretions are unique; and (iv) analyze whether vomeronasal response profiles differ between strains. We report both sex- and, in particular, strain-selective VNO representations of chemical information. Within the urinary ‘secretome’, both volatile compounds and proteins exhibit sufficient discriminative power to provide sex- and strain-specific molecular fingerprints. While total protein amount is substantially enriched in male urine, females secrete a larger variety at overall comparatively low concentrations. Surprisingly, the molecular spectrum of wild mouse urine does not dramatically exceed that of inbred strains. Finally, vomeronasal response profiles differ between C57BL/6 and BALB/c animals, with particularly disparate representations of female semiochemicals.