An integrated metabolomic and proteomic approach for the identification of covalent inhibitors of the main protease (Mpro) of SARS-COV-2 from crude natural extracts

Screening of Mpro Protease (SARS-CoV-2) Covalent Inhibitors from an Anthocyanin-Rich Blueberry Extract Using an HRMS-Based Analytical Platform

“…At this point, isorhamnetin-3-glucoside/galactoside, malvidin-3-arabinoside, malvidin-3-glucoside/galactoside, peonidin-3-arabinoside and peonidin-3-glucoside/galactoside were excluded from the panel of electrophilic compounds since their structural formulae do not contain an available catechol moiety that promotes the formation of the highly reactive quinone intermediate by oxidative activation; this is the basic mechanism through which many catechol-containing natural products react with M pro , as recently demonstrated [ 12 ]. All the hypothesized structural formulae for the covalent adducts resulting from the reaction of each phytocomponent with cysteine were confirmed, except for petunidin-3-glucoside/galactoside, which exhibits a reduced reactivity due to O-methylation on the 3-hydroxy group of pyrogallol (methylated derivative of delphinidin-3-glucoside/galactoside).…”

“…In an early published paper, it was demonstrated how this methodological approach can achieve high levels of analytical performance in terms of sensitivity while operating in untargeted mode, thus enabling the complete characterization of covalent adducts between phytoconstituents and nucleophilic residues of M pro without the need to make a priori assumptions [ 12 ]. The results obtained confirmed the formation of covalent adducts between the molecules used as a model of inhibition and three of the nucleophilic residues of M pro exposed on the binding pocket: Cys145 and His41 constituting the catalytic dyad, and the vicinal histidine residues His163/His164.…”

“…Against this background, screening strategies aimed at identifying bioactive natural compounds as potential M pro inhibitors could have a substantial impact in the fight against SARS-CoV-2. In this scenario, we proposed an innovative strategy, based on high-resolution mass spectrometry (HR-MS) and integrating advanced methods for metabolomics and protein structure analysis to elucidate the identity and the binding mode of covalent M pro binders [ 12 ]. Furthermore, thanks to the state-of-the-art equipment used, which enables high-sensitivity analytical performance levels, it has been possible to work in untargeted mode, thus identifying any unknown or non-commercially available compounds by testing a wide range of natural extracts and expanding the current knowledge base.…”

Discovery of the covalent SARS‐CoV‐2 Mpro inhibitors from antiviral herbs via integrating target‐based high‐throughput screening and chemoproteomic approaches

“…Notably, some flavonoids (e.g., quercetin and luteolin) and catechins (e.g., gallic acid and caffeic acid) in LJF contain one or more catechol groups in their structures, which are susceptible to oxidation and form o ‐quinone. These o ‐quinones have the potential to form covalent bonds with the cysteines of M pro 16,29,30 …”

Investigation on Hydrazonobenzenesulfonamides as Human Carbonic Anhydrase I, II, IX and XII Inhibitors

“…Molecular docking simulations were carried out by using the crystal structures of CAII (PDB ID 3HS4) [50], CAIX (PDB ID 3IAI) [51] and CAXII (PDB ID 1JD0) [52] in complex with the inhibitor AAZ that was used as a reference compound in this study. The protein structures were prepared as described elsewhere [53]. Ligand structures were submitted to a preliminary conjugate gradient minimization by using AMMP tool implemented in VEGA ZZ software [54].…”