SARS-COV-2 spike binding to ACE2 in living cells monitored by TR-FRET.
Targeting the interaction between the SARS-CoV-2 spike protein and human ACE2, its primary cell membrane receptor, is a promising therapeutic strategy to prevent viral entry. Recent in vitro studies revealed that the receptor binding domain (RBD) of the spike protein plays a prominent role in ACE2 binding, yet a simple and ... quantitative assay for monitoring this interaction in a cellular environment is lacking. Here, we developed an RBD-ACE2 binding assay that is based on time-resolved FRET, which reliably monitors the interaction in a physiologically relevant and cellular context. Because it is modular, the assay can monitor the impact of different cellular components, such as heparan sulfate, lipids, and membrane proteins on the RBD-ACE2 interaction and it can be extended to the full-length spike protein. The assay is HTS compatible and can detect small-molecule competitive and allosteric modulators of the RBD-ACE2 interaction with high relevance for SARS-CoV-2 therapeutics.
Mesh Terms:
Angiotensin-Converting Enzyme 2, Cells, Cultured, Fluorescence Resonance Energy Transfer, HEK293 Cells, Humans, Protein Binding, Spike Glycoprotein, Coronavirus, Time Factors
Angiotensin-Converting Enzyme 2, Cells, Cultured, Fluorescence Resonance Energy Transfer, HEK293 Cells, Humans, Protein Binding, Spike Glycoprotein, Coronavirus, Time Factors
Cell Chem Biol
Date: Jan. 20, 2022
PubMed ID: 34246414
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