Heparin inhibits cellular invasion by SARS-CoV-2: structural dependence of the interaction of the surface protein (spike) S1 receptor binding domain with heparin

Abstract

The dependence of the host on the interaction of hundreds of extracellular proteins with the cell surface glycosaminoglycan heparan sulphate (HS) for the regulation of homeostasis is exploited by many microbial pathogens as a means of adherence and invasion. The closely related polysaccharide heparin, the widely used anticoagulant drug, which is structurally similar to HS and is a common experimental proxy, can be expected to mimic the properties of HS. Heparin prevents infection by a range of viruses if added exogenously, including S-associated coronavirus strain HSR1 and here, we show that the addition of heparin (100 μg.ml -1 ) to vero cells inhibits invasion by SARS-CoV-2 by 70%. We also demonstrate that heparin binds to the Spike (S1) protein receptor binding domain and induces a conformational change, illustrated by surface plasmon resonance and circular dichroism spectroscopy studies. The structural features of heparin on which this interaction depends were investigated using a library of heparin derivatives and size-defined fragments. Binding is more strongly dependent on the presence of 2-O or 6-O sulphation, and the consequent conformational consequences in the heparin structure, than on N-sulphation. A hexasaccharide is required for conformational changes to be induced in the secondary structure that are comparable to those that arise from heparin binding. Enoxaparin, a low molecular weight clinical anticoagulant, also binds the S1 RBD protein and induces conformational change. These findings have implications for the rapid development of a first-line therapeutic by repurposing heparin as well as for next-generation, tailor-made, GAG-based antiviral agents against SARS-CoV-2 and other members of the Coronaviridae.

Purification of recombinant SARS-CoV-2 spike, its receptor binding domain, and CR3022 mAb for serological assay

“…Both heparan sulfate [24] and heparin [25] were recently shown to interact with SARS-CoV-2 spike and its RBD, further strengthening our postulation that CR3022 and HSPG could compete with one another for spike binding. All these evidences in the literature, along with the experimental observation presented in this study, suggest that HSPG co-purification is not a coincidental event.…”

The utility of native MS for understanding the mechanism of action of repurposed therapeutics in COVID-19: heparin as a disruptor of the SARS-CoV-2 interaction with its host cell receptor

“…10 In particular, heparin interaction with the S-protein has been shown to induce conformational changes within the latter 11 and to have inhibitory effects on the cellular entry by the virus. 12 Combined with the well-documented anti-coagulant and anti-inflammatory 13 properties of heparin (that are highly relevant vis-à-vis the two hallmarks of COVID-19, the coagulopathy 14,15 and the cytokine storm 16 ), this led to a suggestion that heparin or related compounds may play multiple roles in both arresting the SARS-CoV-2 infection and mitigating its consequences. 17,18 In fact, heparin treatment of COVID-19 patients has been adopted by some physicians and is associated with a better prognosis.…”

Contribution of syndecans to the cellular entry of SARS-CoV-2

“…According to these studies, the conformation change induced by the attachment of spikeS1 to heparin is required for e cient SARS-CoV-2 entry into the cells 87 . Another very recent ndings of also showed heparin effectively blocking SARS-CoV-2 invasion into Vero cells 88 . It has to be noted that the primary sequence of spikeS1 (namely Pro681 -Arg685) contains the heparin-binding core motif PRRAR [33][34][35] ( Supplementary Fig.…”