Determination of Intermediate State Structures in the Opening Pathway of SARS-CoV-2 Spike Using Cryo-Electron Microscopy

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the cause of COVID19, a highly infectious disease that is severely affecting our society and welfare systems. In order to develop therapeutic interventions against this condition, one promising strategy is to target spike, the trimeric transmembrane glycoprotein that the virus uses to recognise and bind its host cells. Here we use a metainference cryo-electron microscopy approach to determine the opening pathway that brings spike from its inactive (closed) conformation to its active (open) one. The knowledge of the structures of the intermediate states of spike along these opening pathways enables us to identify a cryptic pocket that is not exposed in the open and closed states. We then identify compounds that bind the cryptic pocket by screening a library of repurposed drugs. These results underline the opportunities offered by the determination of the structures of the intermediate states populated during the dynamics of proteins to allow the therapeutic targeting of otherwise invisible cryptic binding sites.

Determination of Intermediate State Structures in the Opening Pathway of SARS-CoV-2 Spike Using Cryo-Electron Microscopy

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the cause of COVID19, a highly infectious disease that is severely affecting our society and welfare systems. In order to develop therapeutic interventions against this condition, one promising strategy is to target spike, the trimeric transmembrane glycoprotein that the virus uses to recognise and bind its host cells. Here we use a metainference cryo-electron microscopy approach to determine the opening pathway that brings spike from its inactive (closed) conformation to its active (open) one. The knowledge of the structures of the intermediate states of spike along these opening pathways enables us to identify a cryptic pocket that is not exposed in the open and closed states. We then identify compounds that bind the cryptic pocket by screening a library of repurposed drugs. These results underline the opportunities offered by the determination of the structures of the intermediate states populated during the dynamics of proteins to allow the therapeutic targeting of otherwise invisible cryptic binding sites.

Weak catch bonds make strong networks

Molecular catch bonds are ubiquitous in biology and well-studied in the context of leukocyte extravasion1, cellular mechanosensing2,3, and urinary tract infection4. Unlike normal (slip) bonds, catch bonds strengthen under tension. The current paradigm is that this remarkable ability enables cells to increase their adhesion in fast fluid flows1,4, and hence provides ‘strength-on-demand’. Recently, cytoskeletal crosslinkers have been discovered that also display catch bonding5–8. It has been suggested that they strengthen cells, following the strength-on-demand paradigm9,10. However, catch bonds tend to be weaker compared to regular (slip) bonds because they have cryptic binding sites that are often inactive11–13. Therefore, the role of catch bonding in the cytoskeleton remains unclear. Here we reconstitute cytoskeletal actin networks to show that catch bonds render them both stronger and more deformable than slip bonds, even though the bonds themselves are weaker. We develop a model to show that weak binding allows the catch bonds to mitigate crack initiation by moving from low- to high-tension areas in response to mechanical loading. By contrast, slip bonds remain trapped in stress-free areas. We therefore propose that the mechanism of catch bonding is typified by dissociation-on-demand rather than strength-on-demand. Dissociation-on-demand can explain how both cytolinkers5–8,10,14,15 and adhesins1,2,4,12,16–20 exploit continuous redistribution to combine mechanical strength with the adaptability required for movement and proliferation21. Our findings provide a mechanistic understanding of diseases where catch bonding is compromised11,12 such as kidney focal segmental glomerulosclerosis22,23, caused by the α-actinin-4 mutant studied here. Moreover, catch bonds provide a route towards creating life-like materials that combine strength with deformability24.