Natural and Recombinant SARS-CoV-2 Isolates Rapidly Evolve <i>In Vitro</i> to Higher Infectivity through More Efficient Binding to Heparan Sulfate and Reduced S1/S2 Cleavage
Nikita Shiliaev, Tetyana Lukash, Oksana Palchevska, David K. Crossman, Todd J. Green, Michael R. Crowley, Elena I. Frolova, Ilya Frolov
Abstract
The spike protein of SARS-CoV-2 is a major determinant of viral pathogenesis. It mediates binding to the ACE2 receptor and, later, fusion of viral envelope and cellular membranes. The results of our study demonstrate that SARS-CoV-2 rapidly evolves during propagation in cultured cells. Its spike protein acquires mutations in the NTD and in the P1' position of the furin cleavage site (FCS). The amino acid substitutions or insertions of short peptides in NTD are closely located on the protein surface and increase its positive charge. They strongly increase affinity of the virus to heparan sulfate, make it dramatically more infectious for the cultured cells, and decrease the genome equivalent to PFU (GE/PFU) ratio by orders of magnitude. The S686G mutation also transforms the FCS into the heparin-binding peptide. Thus, the evolved SARS-CoV-2 variants efficiently use glycosaminoglycans on the cell surface for primary attachment before the high-affinity interaction of the spikes with the ACE2 receptor.