Variants of the Ebola virus matrix protein VP40 have differential effects on oligomerization and plasma membrane interactions.

The Ebola virus (EBOV) has been responsible for several outbreaks in Africa over the last decade causing severe hemorrhagic fever with limited treatment options. EBOV is a lipid-enveloped filamentous negative-sense RNA virus with a genome encoding seven proteins. This includes the matrix protein VP40, which regulates assembly and budding of new virions from the inner leaflet of the host cell plasma membrane (PM). VP40 is a multifaceted protein that has different oligomeric states to regulate different parts of the virus life cycle. We investigated how two patient-derived mutations of VP40 samples (R204H and H269R) altered VP40 electrostatics, VP40 oligomeric state, interactions with PM anionic lipids and how these changes affect virus-like particle formation using both laboratory and computational approaches. The R204H mutation induced exclusive octamer formation destabilizing the dimer structure needed for proper lipid binding, PM localization, and virus-like particle formation. In contrast, H269R altered VP40 C-terminal domain (CTD) electrostatics and had similar dimer formation to WT VP40. H269R led to an increase in PM localization compared to WT VP40 and had a three-fold increase in affinity for PM anionic lipids (phosphatidylserine and PI(4,5)P2), consistent with the increased PM localization. Molecular dynamics simulations revealed increased interactions of H269R with phosphatidylserine by several CTD residues. These results highlight the properties of two electrostatic changes found in nature on VP40 structure and function. Understanding the effects of amino acid substitutions on VP40 biophysical properties will be helpful in explaining changes in EBOV variants that occur in nature.