Single-Molecule FRET Imaging of Virus Spike–Host Interactions

Abstract

As a major surface glycoprotein of enveloped viruses, the virus spike protein is a primary target for vaccines and anti-viral treatments. Current vaccines aiming at controlling the COVID-19 pandemic are mostly directed against the SARS-CoV-2 spike protein. To promote virus entry and facilitate immune evasion, spikes must be dynamic. Interactions with host receptors and coreceptors trigger a cascade of conformational changes/structural rearrangements in spikes, which bring virus and host membranes in proximity for membrane fusion required for virus entry. Spike-mediated viral membrane fusion is a dynamic, multi-step process, and understanding the structure–function-dynamics paradigm of virus spikes is essential to elucidate viral membrane fusion, with the ultimate goal of interventions. However, our understanding of this process primarily relies on individual structural snapshots of endpoints. How these endpoints are connected in a time-resolved manner, and the order and frequency of conformational events underlying virus entry, remain largely elusive. Single-molecule Förster resonance energy transfer (smFRET) has provided a powerful platform to connect structure–function in motion, revealing dynamic aspects of spikes for several viruses: SARS-CoV-2, HIV-1, influenza, and Ebola. This review focuses on how smFRET imaging has advanced our understanding of virus spikes’ dynamic nature, receptor-binding events, and mechanism of antibody neutralization, thereby informing therapeutic interventions.

Hypericin Inhibit Alpha-Coronavirus Replication by Targeting 3CL Protease

“…However, we found this catalytic site (Cys144) was next to our solution 2 ( Figure 3 B). To further determine whether hypericin could inhibit the enzymatic activity of 3CLpro, we took advantage of an established FRET assay [ 37 , 38 , 39 ] by using the reporter FRET substrate Dabcyl-YNSTLQ↓AGLRKM-E-Edans. As a start, we expressed the recombinant PEDV 3CLpro with conventional methods of prokaryotic expression, followed by purification with Glutathione Sepharose (GSH) 4B column (GE Healthcare, Chicago, IL, USA) ( Figure 4 A).…”

Understanding viruses and viral infections by biophotonic methods

“…For deeper insight into cell-virion interaction more advanced fluorescence imaging methods, such as FRET, fluorescence recovery after photobleaching (FRAP) and fluorescence lifetime imaging (FLIM) are being used to visualize the virus-cell protein interactions, detection and quantification of the virus within the cell, surface receptor- binding, and so on [142,143]. For example, Gordon et al investigated SARS and MERS viral-human protein interaction using 3-Å cryoelectron microscopy, immunofluorescence, functional genetic screening and proteomics combined with cell biology [144].…”

Monitoring COVID‐19 through SARS‐CoV‐2 quantification in wastewater: progress, challenges and prospects

“…To improve the feasibility and practicability of WBE, research efforts are also being focused on developing faster and cost‐effective methods such as biosensors, which have the potential to be miniaturized, can be easily operated by non‐experts, are portable and disposable and can allow on‐site measurements (Bhalla et al ., 2020; Mao et al ., 2020; Lu, 2021). In this sense, paper‐based devices have been strongly recommended for in situ analysis of SARS‐CoV‐2 in water environments, although its practical application in WW has to be validated (Mao et al ., 2020; Tran et al ., 2021).…”