open access publication

Article, 2024

Two Receptor Binding Strategy of SARS-CoV-2 Is Mediated by Both the N-Terminal and Receptor-Binding Spike Domain

Journal of Physical Chemistry B, ISSN 1520-6106, 1089-5647, Volume 128, 2, Pages 451-464, 10.1021/acs.jpcb.3c06258


Monti M. (Corresponding author) [1] Milanetti E. 0000-0002-3046-5170 [1] [2] Frans M.T. Miotto M. [1] Di Rienzo L. [1] Baranov M.V. Gosti G. [1] [3] Somavarapu A.K. 0000-0001-6123-6875 [4] Nagaraj M. 0000-0002-1294-3810 [4] Golbek T.W. 0000-0002-3632-2701 [4] Rossing E. 0000-0002-6381-6475 [5] Moons S.J. 0000-0001-5507-3811 [5] Boltje T.J. 0000-0001-9141-8784 [5] van den Bogaart G. 0000-0003-2180-6735 Weidner T. 0000-0002-7083-7004 [4] Otzen D.E. 0000-0002-2918-8989 [4] Tartaglia G.G. 0000-0001-7524-6310 [1] Ruocco G. [1] [2] Roeters S.J. 0000-0003-3238-2181 (Corresponding author) [4] [6]


  1. [1] Istituto Italiano di Tecnologia
  2. [NORA names: Italy; Europe, EU; OECD];
  3. [2] Dipartimento di Fisica
  4. [NORA names: Italy; Europe, EU; OECD];
  5. [3] National Research Council
  6. [NORA names: Italy; Europe, EU; OECD];
  7. [4] Aarhus University
  8. [NORA names: AU Aarhus University; University; Denmark; Europe, EU; Nordic; OECD];
  9. [5] Institute for Mathematics
  10. [NORA names: Netherlands; Europe, EU; OECD];


It is not well understood why severe acute respiratory syndrome (SARS)-CoV-2 spreads much faster than other β-coronaviruses such as SARS-CoV and Middle East respiratory syndrome (MERS)-CoV. In a previous publication, we predicted the binding of the N-terminal domain (NTD) of SARS-CoV-2 spike to sialic acids (SAs). Here, we experimentally validate this interaction and present simulations that reveal a second possible interaction between SAs and the spike protein via a binding site located in the receptor-binding domain (RBD). The predictions from molecular-dynamics simulations and the previously-published 2D-Zernike binding-site recognition approach were validated through flow-induced dispersion analysis (FIDA)─which reveals the capability of the SARS-CoV-2 spike to bind to SA-containing (glyco)lipid vesicles, and flow-cytometry measurements─which show that spike binding is strongly decreased upon inhibition of SA expression on the membranes of angiotensin converting enzyme-2 (ACE2)-expressing HEK cells. Our analyses reveal that the SA binding of the NTD and RBD strongly enhances the infection-inducing ACE2 binding. Altogether, our work provides in silico, in vitro, and cellular evidence that the SARS-CoV-2 virus utilizes a two-receptor (SA and ACE2) strategy. This allows the SARS-CoV-2 spike to use SA moieties on the cell membrane as a binding anchor, which increases the residence time of the virus on the cell surface and aids in the binding of the main receptor, ACE2, via 2D diffusion.


  • Lundbeckfonden
  • HEK293 Cells
  • Horizon 2020 Framework Programme
  • National Institute of Allergy and Infectious Diseases
  • BEI Resources
  • Novo Nordisk Fonden
  • EIC-2022-PathfinderOpen
  • National Center for Gene Therapy and Drugs
  • European Research Council
  • NextGeneration EU
  • National Institutes of Health
  • Innovation Foundation Denmark

Data Provider: Elsevier