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Updated: Jun 12, 2025

Analysis of Group IV Viral SSHHPS Using In Vitro and In Silico Methods
Published on: December 21, 2019
Insights into the Main Protease of SARS-CoV-2: Thermodynamic Analysis, Structural Characterization, and the Impact of
Jamie P Butalewicz1, Sarah N Sipe1, Kyle J Juetten1
1Department of Chemistry, The University of Texas at Austin, Austin, Texas 78712, United States.
Abstract:
The main protease (Mpro) of SARS-CoV-2 is an essential enzyme for coronaviral maturation and is the target of Paxlovid, which is currently the standard-of-care treatment for COVID-19. There remains a need to identify new inhibitors of Mpro as viral resistance to Paxlovid emerges. Here, we report the use of native mass spectrometry coupled with 193 nm ultraviolet photodissociation (UVPD) and integrated with other biophysical tools to structurally characterize Mpro and its interactions with potential covalent inhibitors. The overall energy landscape was obtained using variable temperature nanoelectrospray ionization (vT-nESI), thus providing quantitative evaluation of inhibitor binding on the stability of Mpro. Thermodynamic parameters extracted from van't Hoff plots revealed that the dimeric complexes containing each inhibitor showed enhanced stability through increased melting temperatures as well as overall lower average charge states, giving insight into the basis for inhibition mechanisms.
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