{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,24]],"date-time":"2026-07-24T04:35:05Z","timestamp":1784867705810,"version":"3.55.0"},"reference-count":61,"publisher":"MDPI AG","issue":"12","license":[{"start":{"date-parts":[[2023,6,7]],"date-time":"2023-06-07T00:00:00Z","timestamp":1686096000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","award":["NSFC 21927814"],"award-info":[{"award-number":["NSFC 21927814"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Molecules"],"abstract":"<jats:p>The main protease (Mpro) is a promising drug target for inhibiting the coronavirus due to its conserved properties and lack of homologous genes in humans. However, previous studies on Mpro\u2019s kinetic parameters have been confusing, hindering the selection of accurate inhibitors. Therefore, obtaining a clear view of Mpro\u2019s kinetic parameters is necessary. In our study, we investigated the kinetic behaviors of Mpro from SARS-CoV-2 and SARS-CoV using both FRET-based cleavage assay and the LC-MS method, respectively. Our findings indicate that the FRET-based cleavage assay could be used for preliminary screening of Mpro inhibitors, while the LC-MS method should be applied to select the effective inhibitors with higher reliability. Furthermore, we constructed the active site mutants (H41A and C145A) and measured the kinetic parameters to gain a deeper understanding of the atomic-level enzyme efficiency reduction compared to the wild type. Overall, our study provides valuable insights for inhibitor screening and design by offering a comprehensive understanding of Mpro\u2019s kinetic behaviors.<\/jats:p>","DOI":"10.3390\/molecules28124605","type":"journal-article","created":{"date-parts":[[2023,6,8]],"date-time":"2023-06-08T02:02:28Z","timestamp":1686189748000},"page":"4605","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":7,"title":["Comprehensive Understanding of the Kinetic Behaviors of Main Protease from SARS-CoV-2 and SARS-CoV: New Data and Comparison to Published Parameters"],"prefix":"10.3390","volume":"28","author":[{"given":"Fangya","family":"Li","sequence":"first","affiliation":[{"name":"School of Pharmaceutical Science and Technology, Tianjin University, Tianjin 300072, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tingting","family":"Fang","sequence":"additional","affiliation":[{"name":"School of Pharmaceutical Science and Technology, Tianjin University, Tianjin 300072, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Feng","family":"Guo","sequence":"additional","affiliation":[{"name":"School of Pharmaceutical Science and Technology, Tianjin University, Tianjin 300072, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zipeng","family":"Zhao","sequence":"additional","affiliation":[{"name":"School of Pharmaceutical Science and Technology, Tianjin University, Tianjin 300072, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jianyu","family":"Zhang","sequence":"additional","affiliation":[{"name":"School of Pharmaceutical Science and Technology, Tianjin University, Tianjin 300072, China"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2023,6,7]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"24765","DOI":"10.1074\/jbc.M311744200","article-title":"Dissection study on the severe acute respiratory syndrome 3C-like protease reveals the critical role of the extra domain in dimerization of the enzyme: Defining the extra domain as a new target for design of highly specific protease inhibitors","volume":"279","author":"Shi","year":"2004","journal-title":"J. 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