Heparan Sulfate Proteoglycans as Attachment Factor for SARS-CoV-2.

Lin Liu,Pradeep Chopra,Kim M Bouwman,Robert P De Vries,S Mark Tompkins,Margreet A Wolfert,Geert-Jan Boons,Xiuru Li

doi:10.1021/acscentsci.1c00010

Abstract

Severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) is causing an unprecedented global pandemic demanding the urgent development of therapeutic strategies. Microarray binding experiments, using an extensive heparan sulfate (HS) oligosaccharide library, showed that the receptor binding domain (RBD) of the spike of SARS-CoV-2 can bind HS in a length- and sequence-dependent manner. A hexasaccharide composed of IdoA2S-GlcNS6S repeating units was identified as the minimal binding epitope. Surface plasmon resonance showed the SARS-CoV-2 spike protein binds with a much higher affinity to heparin (KD = 55 nM) compared to the RBD (KD = 1 μM) alone. It was also found that heparin does not interfere in angiotensin-converting enzyme 2 (ACE2) binding or proteolytic processing of the spike. However, exogenous administered heparin or a highly sulfated HS oligosaccharide inhibited RBD binding to cells. Furthermore, an enzymatic removal of HS proteoglycan from physiological relevant tissue resulted in a loss of RBD binding. The data support a model in which HS functions as the point of initial attachment allowing the virus to travel through the glycocalyx by low-affinity high-avidity interactions to reach the cell membrane, where it can engage with ACE2 for cell entry. Microarray binding experiments showed that ACE2 and HS can simultaneously engage with the RBD, and it is likely no dissociation between HS and RBD is required for binding to ACE2. The results highlight the potential of using HS oligosaccharides as a starting material for therapeutic agent development.

Highlights

Our response to the severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) pandemic demands an urgent development of therapeutic strategies
Surface plasmon resonance experiments were performed to probe whether the receptor binding domain (RBD) domain of the SARS-CoV-2 spike protein can bind with heparin
Biotinylated heparin was immobilized on a streptavidin-coated sensor chip, and binding experiments were performed by employing as analytes different concentrations of the RBD, monomeric spike protein, and trimeric spike protein of SARS-CoV-2

Summary

■ INTRODUCTION

Our response to the severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) pandemic demands an urgent development of therapeutic strategies. We prepared an unprecedented library of structurally well-defined HS oligosaccharides that differ in chain length, backbone composition, and sulfation pattern.[23,24] This collection of HS oligosaccharides was used to develop a glycan microarray for the systematic analysis of selectivity of HS-binding proteins Using this microarray platform in conjugation with detailed binding studies, we found that the RBD domain of SARS-CoV-2 spike can bind HS in a lengthand sequence-dependent manner, and the observations support a model in which the RBD confers sequence selectivity, and the affinity of binding is enhanced by additional interactions with other HS binding sites in, for example, the S1/S2 proteolytic cleavage site.[9] Identified HS oligosaccharide ligands could inhibit the binding of RBD to cells. The spike can, bind simultaneously with HS and ACE2, and no dissociation between RBD and HS is required before it can engage with ACE2 for cell entry

■ RESULTS AND DISCUSSION

■ DISCUSSION AND CONCLUSIONS

■ ACKNOWLEDGMENTS

■ REFERENCES