Elucidating coronavirus fusion mechanisms.

Abstract

Enveloped viruses initiate infection by attaching to a cellular receptor, followed by triggering and engagement of the viral fusion glycoprotein. We have developed a method by which the receptor binding is replaced by a synthetic DNA tether, permitting study of the triggering and fusion processes independent of receptor engagement. This is achieved by embedding complementary strands of DNA into the membranes of the viral particle and its target. Here, we apply this technique to SARS-CoV-2 and related coronaviruses. We find that ACE-2 binding can be effectively replaced with a synthetic tether and that fusion can be triggered using an exogenous protease. Furthermore, the rate of fusion is robust to protease concentration, suggesting that conformational transitions of the spike glycoprotein, rather than ACE-2 binding or proteolytic cleavage, are the limiting steps for SARS-CoV-2 entry. Using this same technique, we are able to compare the fusion kinetics of two SARS-CoV-2 variants with a bat coronavirus. We find that the Wuhan and BA.1 or Omicron strains have similar fusion kinetics, while WIV1 fuses slightly faster. DNA-tethering thus permits comparison of coronavirus fusion independent of receptor binding.