Hexamethylene Amiloride Binds the SARS-CoV-2 Envelope Protein at the Protein-Lipid Interface.
Noah H SombergJoão Medeiros-SilvaHyunil JoJun WangWilliam F DeGradoMei HongPublished in: Protein science : a publication of the Protein Society (2023)
The SARS-CoV-2 envelope (E) protein forms a five-helix bundle in lipid bilayers whose cation-conducting activity is associated with the inflammatory response and respiratory distress symptoms of COVID-19. E channel activity is inhibited by the drug 5-(N,N-hexamethylene) amiloride (HMA). However, the binding site of HMA in E has not been determined. Here we use solid-state NMR to measure distances between HMA and the E transmembrane domain (ETM) in lipid bilayers. 13 C, 15 N-labeled HMA is combined with fluorinated or 13 C-labeled ETM. Conversely, fluorinated HMA is combined with 13 C, 15 N-labeled ETM. These orthogonal isotopic labeling patterns allow us to conduct dipolar recoupling NMR experiments to determine the HMA binding stoichiometry to ETM as well as HMA-protein distances. We find that HMA binds ETM with a stoichiometry of one drug per pentamer. Unexpectedly, the bound HMA is not centrally located within the channel pore, but lies on the lipid-facing surface in the middle of the TM domain. This result suggests that HMA may inhibit the E channel activity by interfering with the gating function of an aromatic network. These distance data are obtained under much lower drug concentrations than in previous chemical shift data, which showed the largest perturbation for N-terminal residues. This difference suggests that HMA has higher affinity for the protein-lipid interface than the channel pore, which gives insight into the inhibition mechanism of HMA for SARS-CoV-2 E. This article is protected by copyright. All rights reserved.
Keyphrases
- sars cov
- solid state
- inflammatory response
- protein protein
- fatty acid
- respiratory syndrome coronavirus
- amino acid
- magnetic resonance
- binding protein
- coronavirus disease
- emergency department
- high resolution
- electronic health record
- pet imaging
- small molecule
- physical activity
- mass spectrometry
- ionic liquid
- transcription factor
- data analysis
- sleep quality