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Protocetraric and Salazinic Acids as Potential Inhibitors of SARS-CoV-2 3CL Protease: Biochemical, Cytotoxic, and Computational Characterization of Depsidones as Slow-Binding Inactivators

The study investigated the inhibitory activity of protocetraric and salazinic acids against SARS-CoV-2 3CL(pro). The kinetic parameters were determined by microtiter plate-reading fluorimeter using a fluorogenic substrate. The cytotoxic activity was tested on murine Sertoli TM4 cells. In silico anal...

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Autores principales: Fagnani, Lorenza, Nazzicone, Lisaurora, Bellio, Pierangelo, Franceschini, Nicola, Tondi, Donatella, Verri, Andrea, Petricca, Sabrina, Iorio, Roberto, Amicosante, Gianfranco, Perilli, Mariagrazia, Celenza, Giuseppe
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9227325/
https://www.ncbi.nlm.nih.gov/pubmed/35745633
http://dx.doi.org/10.3390/ph15060714
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author Fagnani, Lorenza
Nazzicone, Lisaurora
Bellio, Pierangelo
Franceschini, Nicola
Tondi, Donatella
Verri, Andrea
Petricca, Sabrina
Iorio, Roberto
Amicosante, Gianfranco
Perilli, Mariagrazia
Celenza, Giuseppe
author_facet Fagnani, Lorenza
Nazzicone, Lisaurora
Bellio, Pierangelo
Franceschini, Nicola
Tondi, Donatella
Verri, Andrea
Petricca, Sabrina
Iorio, Roberto
Amicosante, Gianfranco
Perilli, Mariagrazia
Celenza, Giuseppe
author_sort Fagnani, Lorenza
collection PubMed
description The study investigated the inhibitory activity of protocetraric and salazinic acids against SARS-CoV-2 3CL(pro). The kinetic parameters were determined by microtiter plate-reading fluorimeter using a fluorogenic substrate. The cytotoxic activity was tested on murine Sertoli TM4 cells. In silico analysis was performed to ascertain the nature of the binding with the 3CL(pro). The compounds are slow-binding inactivators of 3CL(pro) with a K(i) of 3.95 μM and 3.77 μM for protocetraric and salazinic acid, respectively, and inhibitory efficiency k(inact)/K(i) at about 3 × 10(−5) s(−1)µM(−1). The mechanism of inhibition shows that both compounds act as competitive inhibitors with the formation of a stable covalent adduct. The viability assay on epithelial cells revealed that none of them shows cytotoxicity up to 80 μM, which is well below the K(i) values. By molecular modelling, we predicted that the catalytic Cys145 makes a nucleophilic attack on the carbonyl carbon of the cyclic ester common to both inhibitors, forming a stably acyl-enzyme complex. The computational and kinetic analyses confirm the formation of a stable acyl-enzyme complex with 3CL(pro). The results obtained enrich the knowledge of the already numerous biological activities exhibited by lichen secondary metabolites, paving the way for developing promising scaffolds for the design of cysteine enzyme inhibitors.
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spelling pubmed-92273252022-06-25 Protocetraric and Salazinic Acids as Potential Inhibitors of SARS-CoV-2 3CL Protease: Biochemical, Cytotoxic, and Computational Characterization of Depsidones as Slow-Binding Inactivators Fagnani, Lorenza Nazzicone, Lisaurora Bellio, Pierangelo Franceschini, Nicola Tondi, Donatella Verri, Andrea Petricca, Sabrina Iorio, Roberto Amicosante, Gianfranco Perilli, Mariagrazia Celenza, Giuseppe Pharmaceuticals (Basel) Article The study investigated the inhibitory activity of protocetraric and salazinic acids against SARS-CoV-2 3CL(pro). The kinetic parameters were determined by microtiter plate-reading fluorimeter using a fluorogenic substrate. The cytotoxic activity was tested on murine Sertoli TM4 cells. In silico analysis was performed to ascertain the nature of the binding with the 3CL(pro). The compounds are slow-binding inactivators of 3CL(pro) with a K(i) of 3.95 μM and 3.77 μM for protocetraric and salazinic acid, respectively, and inhibitory efficiency k(inact)/K(i) at about 3 × 10(−5) s(−1)µM(−1). The mechanism of inhibition shows that both compounds act as competitive inhibitors with the formation of a stable covalent adduct. The viability assay on epithelial cells revealed that none of them shows cytotoxicity up to 80 μM, which is well below the K(i) values. By molecular modelling, we predicted that the catalytic Cys145 makes a nucleophilic attack on the carbonyl carbon of the cyclic ester common to both inhibitors, forming a stably acyl-enzyme complex. The computational and kinetic analyses confirm the formation of a stable acyl-enzyme complex with 3CL(pro). The results obtained enrich the knowledge of the already numerous biological activities exhibited by lichen secondary metabolites, paving the way for developing promising scaffolds for the design of cysteine enzyme inhibitors. MDPI 2022-06-04 /pmc/articles/PMC9227325/ /pubmed/35745633 http://dx.doi.org/10.3390/ph15060714 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Fagnani, Lorenza
Nazzicone, Lisaurora
Bellio, Pierangelo
Franceschini, Nicola
Tondi, Donatella
Verri, Andrea
Petricca, Sabrina
Iorio, Roberto
Amicosante, Gianfranco
Perilli, Mariagrazia
Celenza, Giuseppe
Protocetraric and Salazinic Acids as Potential Inhibitors of SARS-CoV-2 3CL Protease: Biochemical, Cytotoxic, and Computational Characterization of Depsidones as Slow-Binding Inactivators
title Protocetraric and Salazinic Acids as Potential Inhibitors of SARS-CoV-2 3CL Protease: Biochemical, Cytotoxic, and Computational Characterization of Depsidones as Slow-Binding Inactivators
title_full Protocetraric and Salazinic Acids as Potential Inhibitors of SARS-CoV-2 3CL Protease: Biochemical, Cytotoxic, and Computational Characterization of Depsidones as Slow-Binding Inactivators
title_fullStr Protocetraric and Salazinic Acids as Potential Inhibitors of SARS-CoV-2 3CL Protease: Biochemical, Cytotoxic, and Computational Characterization of Depsidones as Slow-Binding Inactivators
title_full_unstemmed Protocetraric and Salazinic Acids as Potential Inhibitors of SARS-CoV-2 3CL Protease: Biochemical, Cytotoxic, and Computational Characterization of Depsidones as Slow-Binding Inactivators
title_short Protocetraric and Salazinic Acids as Potential Inhibitors of SARS-CoV-2 3CL Protease: Biochemical, Cytotoxic, and Computational Characterization of Depsidones as Slow-Binding Inactivators
title_sort protocetraric and salazinic acids as potential inhibitors of sars-cov-2 3cl protease: biochemical, cytotoxic, and computational characterization of depsidones as slow-binding inactivators
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9227325/
https://www.ncbi.nlm.nih.gov/pubmed/35745633
http://dx.doi.org/10.3390/ph15060714
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