Strikingly Different Roles of SARS-CoV-2 Fusion Peptides Uncovered by Neutron Scattering

Armando Maestro1,2 and Nathan Zaccai3

1 Centro de Fı́sica de Materiales (CSIC, UPV/EHU) – Materials Physics Center MPC, Paseo Manuel de Lardizabal 5, E-20018 San Sebastián, Spain.
2 IKERBASQUE—Basque Foundation for Science, Plaza Euskadi 5, Bilbao, 48009 Spain
3 Cambridge Institute for Medical Research, University of Cambridge, Cambridge CB22 7QQ, United Kingdom.

En raison de la pandémie du COVID-19, il est devenu important de comprendre les mécanismes moléculaires à la base de l’infection par les coronavirus. Une étape critique dans la pénétration du virus SARS-CoV-2 dans la cellule se produit lorsque la protéine virale Spike sert de médiateur de la fusion entre les membranes du virus et de son hôte. Notre récente publication dans le Journal of the American Chemical Society présente une étude du rôle joué par certaines régions de la protéine Spike et de l’influence du calcium et du cholestérol dans le processus de fusion par diffusion des neutrons. Nos résultats ont révélé que des fonctions étonnamment différentes sont codées dans cette protéine.
Due to the COVID-19 pandemic, a thorough understanding of the molecular mechanisms of cellular infection by coronaviruses has become imperative. A critical stage in cell entry by the SARS-CoV-2 virus occurs when its Spike protein mediates fusion between viral and host membranes. Recently published in the Journal of the American Chemical Society, we presented a detailed investigation of the role of selected regions of the Spike protein, and the influence of calcium and cholesterol, in this fusion process.

Importantly, in order to gain insights about how the fusion proceeds in vivo, we recreated important elements of the fusion mechanism by simplifying the system down to its core elements, amenable to experimental analysis by neutron scattering. In our in vitro models, cellular membranes were mimicked with hydrogenated and deuterated lipid monolayers and bilayers, produced in a recently set-up facility within the Partnership for Soft Condensed Matter at the ILL (www.ill.eu/L-Lab), while different sections of the Spike protein’s unstructured region, crucial for viral fusion, were synthesised as (fusion) peptides.
Structural information from specular neutron reflectometry1 and small angle neutron scattering2, complemented by dynamical information from quasi-elastic and spin–echo neutron spectroscopy3, was therefore employed to study the interaction of fusion peptides with model membrane. Neutrons are particularly well suited for the study of soft and biological matter since they allow measurements at room-temperature with better than nanometer resolution and at energies corresponding to thermal fluctuations. They are non-destructive and highly penetrating, thus allowing work in physiological conditions. Furthermore, as neutrons interact very differently with hydrogen (1H) and deuterium (2H), it is possible through isotopic substitution, for example in lipids, to observe hydrogen atoms and water molecules in biological samples, and therefore highlight structural and chemical differences in specific regions of interest.
Our experiments revealed strikingly different functions encoded in the Spike fusion domain. Calcium drives the N-terminal of the Spike fusion domain to fully cross the host plasma membrane. Removing calcium, however, reorients the peptide back to the lipid leaflet closest to the virus, leading to significant changes in lipid fluidity and rigidity. In conjunction with other regions of the fusion domain, which are also positioned to bridge and dehydrate viral and host membranes, the molecular events leading to cell entry by SARS-CoV-2 are proposed.
The data are of interest not only in the context of the current COVID-19 pandemic, but they also provide a powerful interdisciplinary framework for future investigations of eukaryotic and viral fusion mechanisms. For example, it remains to be discovered if other viral fusion peptides can also harpoon through cellular membranes during infection. Our comprehensive study will also interest a wider audience, since this scientific approach, combining structural and dynamics characterization by neutrons under physiological conditions, can be readily applied to many biological questions.

fait marquant mai 2022

Schematic view of how during SARS-Cov-2 infection, the critically important fusion of human and viral membranes is induced by the viral Spike protein. The presence of calcium (Ca+) drives the fusion peptides of the Spike protein to harpoon through the host cell membrane’s lipid bilayer, while other regions of the Spike protein help bridge viral and host membranes, thereby facilitating fusion.

Reference : Santamaria A, Batchu KC, Matsarskaia O, Prévost SF, Russo D, Natali F, Seydel T, Hoffmann I, Laux V, Haertlein M, Darwish TA, Russell RA, Corucci G, Fragneto G, Maestro A, Zaccai NR. Strikingly Different Roles of SARS-CoV-2 Fusion Peptides Uncovered by Neutron Scattering. J Am Chem Soc. (2022) 144(7):2968-2979. DOI : 10.1021/jacs.1c09856.