{"title":"基于生物层干涉仪的 SARS-COV-2 Mpro 靶向活性成分识别系统：中药配体筛选中的构建与应用","authors":"Dai Zhang, Bing Han, Xiao-Fei Chen, Shuai Zhao, Wei-Xia Li, Hui Zhang, Ming-Liang Zhang, Meng-Qi Huo, Yong-Sheng Qiu, Ying-Jie Ren, Yao-Dong Zhang, Xian-Qing Ren, Wei Wang, Jin-Fa Tang","doi":"10.1002/pca.3462","DOIUrl":null,"url":null,"abstract":"<p><strong>Introduction: </strong>Drug discovery research targeting SARS-CoV-2 and other emerging pathogens remains critically important. Active compounds derived from plants frequently serve as lead compounds for further drug discovery; however, numerous unrelated chemical constituents in crude extracts may obscure the effective ingredients in LC-MS analysis.</p><p><strong>Objective: </strong>The aim of this study is to construct a biolayer interferometry (BLI)-based system for recognizing active ingredients that inhibit the main protease (Mpro) of SARS-CoV-2 and to identify the active chemical components binding to Mpro from herbal medicines.</p><p><strong>Methodology: </strong>We developed a novel FRET fluorogenic probe by linking the amino acid sequences of the fluorescent proteins Lssmorange and mKate2 (Ls-mK). The interaction between traditional Chinese medicine and Mpro was analyzed using BLI. Ultrahigh performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS) was employed to analyze the composition of herbal medicines.</p><p><strong>Results: </strong>Fluorescence detection and spectroscopy confirmed the successful construction of an Mpro inhibitor screening system. Lanqin Oral Liquid (LQL) and Gardeniae fructus exhibited strong inhibitory effects on Mpro. Ten compounds were identified from G. fructus extracts; among them, deacetyl asperulosidic acid methyl ester (DAAME) and Gardoside were found to strongly bind to Mpro, with dissociation constants (KD) of 3.41 μM and 801 nM, respectively. The half-maximal inhibitory concentrations (IC50) of DAAME and Gardoside for Mpro were 27.46 and 13.7 μM, respectively.</p><p><strong>Conclusion: </strong>This study established a functional Mpro inhibitor screening system. Among the 10 components identified from G. fructus that bind to Mpro, DAAME and Gardoside displayed strong binding and inhibitory activity, indicating their potential as lead compounds for inhibiting SARS-CoV-2 viral replication.</p>","PeriodicalId":20095,"journal":{"name":"Phytochemical Analysis","volume":" ","pages":""},"PeriodicalIF":3.0000,"publicationDate":"2024-11-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"A Biolayer Interferometry-Based SARS-COV-2 Mpro-Targeted Active Ingredients Recognition System: Construction and Application in Ligand Screening From Herbal Medicines.\",\"authors\":\"Dai Zhang, Bing Han, Xiao-Fei Chen, Shuai Zhao, Wei-Xia Li, Hui Zhang, Ming-Liang Zhang, Meng-Qi Huo, Yong-Sheng Qiu, Ying-Jie Ren, Yao-Dong Zhang, Xian-Qing Ren, Wei Wang, Jin-Fa Tang\",\"doi\":\"10.1002/pca.3462\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><strong>Introduction: </strong>Drug discovery research targeting SARS-CoV-2 and other emerging pathogens remains critically important. Active compounds derived from plants frequently serve as lead compounds for further drug discovery; however, numerous unrelated chemical constituents in crude extracts may obscure the effective ingredients in LC-MS analysis.</p><p><strong>Objective: </strong>The aim of this study is to construct a biolayer interferometry (BLI)-based system for recognizing active ingredients that inhibit the main protease (Mpro) of SARS-CoV-2 and to identify the active chemical components binding to Mpro from herbal medicines.</p><p><strong>Methodology: </strong>We developed a novel FRET fluorogenic probe by linking the amino acid sequences of the fluorescent proteins Lssmorange and mKate2 (Ls-mK). The interaction between traditional Chinese medicine and Mpro was analyzed using BLI. Ultrahigh performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS) was employed to analyze the composition of herbal medicines.</p><p><strong>Results: </strong>Fluorescence detection and spectroscopy confirmed the successful construction of an Mpro inhibitor screening system. Lanqin Oral Liquid (LQL) and Gardeniae fructus exhibited strong inhibitory effects on Mpro. Ten compounds were identified from G. fructus extracts; among them, deacetyl asperulosidic acid methyl ester (DAAME) and Gardoside were found to strongly bind to Mpro, with dissociation constants (KD) of 3.41 μM and 801 nM, respectively. The half-maximal inhibitory concentrations (IC50) of DAAME and Gardoside for Mpro were 27.46 and 13.7 μM, respectively.</p><p><strong>Conclusion: </strong>This study established a functional Mpro inhibitor screening system. Among the 10 components identified from G. fructus that bind to Mpro, DAAME and Gardoside displayed strong binding and inhibitory activity, indicating their potential as lead compounds for inhibiting SARS-CoV-2 viral replication.</p>\",\"PeriodicalId\":20095,\"journal\":{\"name\":\"Phytochemical Analysis\",\"volume\":\" \",\"pages\":\"\"},\"PeriodicalIF\":3.0000,\"publicationDate\":\"2024-11-20\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Phytochemical Analysis\",\"FirstCategoryId\":\"99\",\"ListUrlMain\":\"https://doi.org/10.1002/pca.3462\",\"RegionNum\":3,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q2\",\"JCRName\":\"BIOCHEMICAL RESEARCH METHODS\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Phytochemical Analysis","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1002/pca.3462","RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"BIOCHEMICAL RESEARCH METHODS","Score":null,"Total":0}
A Biolayer Interferometry-Based SARS-COV-2 Mpro-Targeted Active Ingredients Recognition System: Construction and Application in Ligand Screening From Herbal Medicines.
Introduction: Drug discovery research targeting SARS-CoV-2 and other emerging pathogens remains critically important. Active compounds derived from plants frequently serve as lead compounds for further drug discovery; however, numerous unrelated chemical constituents in crude extracts may obscure the effective ingredients in LC-MS analysis.
Objective: The aim of this study is to construct a biolayer interferometry (BLI)-based system for recognizing active ingredients that inhibit the main protease (Mpro) of SARS-CoV-2 and to identify the active chemical components binding to Mpro from herbal medicines.
Methodology: We developed a novel FRET fluorogenic probe by linking the amino acid sequences of the fluorescent proteins Lssmorange and mKate2 (Ls-mK). The interaction between traditional Chinese medicine and Mpro was analyzed using BLI. Ultrahigh performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS) was employed to analyze the composition of herbal medicines.
Results: Fluorescence detection and spectroscopy confirmed the successful construction of an Mpro inhibitor screening system. Lanqin Oral Liquid (LQL) and Gardeniae fructus exhibited strong inhibitory effects on Mpro. Ten compounds were identified from G. fructus extracts; among them, deacetyl asperulosidic acid methyl ester (DAAME) and Gardoside were found to strongly bind to Mpro, with dissociation constants (KD) of 3.41 μM and 801 nM, respectively. The half-maximal inhibitory concentrations (IC50) of DAAME and Gardoside for Mpro were 27.46 and 13.7 μM, respectively.
Conclusion: This study established a functional Mpro inhibitor screening system. Among the 10 components identified from G. fructus that bind to Mpro, DAAME and Gardoside displayed strong binding and inhibitory activity, indicating their potential as lead compounds for inhibiting SARS-CoV-2 viral replication.
期刊介绍：
Phytochemical Analysis is devoted to the publication of original articles concerning the development, improvement, validation and/or extension of application of analytical methodology in the plant sciences. The spectrum of coverage is broad, encompassing methods and techniques relevant to the detection (including bio-screening), extraction, separation, purification, identification and quantification of compounds in plant biochemistry, plant cellular and molecular biology, plant biotechnology, the food sciences, agriculture and horticulture. The Journal publishes papers describing significant novelty in the analysis of whole plants (including algae), plant cells, tissues and organs, plant-derived extracts and plant products (including those which have been partially or completely refined for use in the food, agrochemical, pharmaceutical and related industries). All forms of physical, chemical, biochemical, spectroscopic, radiometric, electrometric, chromatographic, metabolomic and chemometric investigations of plant products (monomeric species as well as polymeric molecules such as nucleic acids, proteins, lipids and carbohydrates) are included within the remit of the Journal. Papers dealing with novel methods relating to areas such as data handling/ data mining in plant sciences will also be welcomed.
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