{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T01:25:02Z","timestamp":1760059502625,"version":"build-2065373602"},"reference-count":46,"publisher":"MDPI AG","issue":"12","license":[{"start":{"date-parts":[[2025,6,18]],"date-time":"2025-06-18T00:00:00Z","timestamp":1750204800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100012550","name":"Ministry of Culture and Innovation of Hungary from the National Research, Development and Innovation Fund","doi-asserted-by":"publisher","award":["TKP2021-EGA-20","TKP2021-NKTA-34","TKP2021-NKTA","150532","BO\/00110\/23\/5"],"award-info":[{"award-number":["TKP2021-EGA-20","TKP2021-NKTA-34","TKP2021-NKTA","150532","BO\/00110\/23\/5"]}],"id":[{"id":"10.13039\/501100012550","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100012550","name":"National Research, Development, and Innovation Fund of Hungary","doi-asserted-by":"publisher","award":["TKP2021-EGA-20","TKP2021-NKTA-34","TKP2021-NKTA","150532","BO\/00110\/23\/5"],"award-info":[{"award-number":["TKP2021-EGA-20","TKP2021-NKTA-34","TKP2021-NKTA","150532","BO\/00110\/23\/5"]}],"id":[{"id":"10.13039\/501100012550","id-type":"DOI","asserted-by":"publisher"}]},{"name":"NKFIH","award":["TKP2021-EGA-20","TKP2021-NKTA-34","TKP2021-NKTA","150532","BO\/00110\/23\/5"],"award-info":[{"award-number":["TKP2021-EGA-20","TKP2021-NKTA-34","TKP2021-NKTA","150532","BO\/00110\/23\/5"]}]},{"DOI":"10.13039\/501100003825","name":"J\u00e1nos Bolyai Research Scholarship of the Hungarian Academy of Sciences","doi-asserted-by":"publisher","award":["TKP2021-EGA-20","TKP2021-NKTA-34","TKP2021-NKTA","150532","BO\/00110\/23\/5"],"award-info":[{"award-number":["TKP2021-EGA-20","TKP2021-NKTA-34","TKP2021-NKTA","150532","BO\/00110\/23\/5"]}],"id":[{"id":"10.13039\/501100003825","id-type":"DOI","asserted-by":"publisher"}]},{"name":"University of Debrecen (UD) Program for Scientific Publication","award":["TKP2021-EGA-20","TKP2021-NKTA-34","TKP2021-NKTA","150532","BO\/00110\/23\/5"],"award-info":[{"award-number":["TKP2021-EGA-20","TKP2021-NKTA-34","TKP2021-NKTA","150532","BO\/00110\/23\/5"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the coronavirus disease of 2019 (COVID-19) and has persistently caused infections since its emergence in late 2019. The main protease (Mpro) of SARS-CoV-2 plays a crucial role in its life-cycle; thus, it is an important target for drug development. One of the first virus-specific drugs that has been approved for the treatment of COVID-19 patients is Paxlovid, which contains nirmatrelvir, a covalent inhibitor of Mpro. Screening of inhibitor candidates and specificity studies also rely on efficient substrates and activity assays. Casein is one of the most commonly applied universal substrates that can be used to study a wide range of proteases, including SARS-CoV-2 Mpro. Casein is a known substrate for Mpro in vitro, but the specific casein isoform cleaved by Mpro remained unidentified, and the cleavage sites have yet to be determined. This work studied cleavage of \u03b1-, \u03b2- and \u03ba-isoforms of bovine casein by SARS-CoV-2 Mpro, using in vitro and in silico approaches. The candidate cleavage sites were predicted in silico based on the protein sequences, and the cleavage positions were identified based on mass spectrometric analysis of cleavage fragments. Based on our results, only \u03b2-casein contains cleavage sites for Mpro and thus can be used as its substrate in vitro. The newly identified cleavage site sequences further widen the knowledge about the specificity of SARS-CoV-2 Mpro.<\/jats:p>","DOI":"10.3390\/ijms26125829","type":"journal-article","created":{"date-parts":[[2025,6,18]],"date-time":"2025-06-18T05:30:42Z","timestamp":1750224642000},"page":"5829","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Identification of SARS-CoV-2 Main Protease Cleavage Sites in Bovine \u03b2-Casein"],"prefix":"10.3390","volume":"26","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-6079-5621","authenticated-orcid":false,"given":"J\u00e1nos Andr\u00e1s","family":"M\u00f3ty\u00e1n","sequence":"first","affiliation":[{"name":"Department of Biochemistry and Molecular Biology, Faculty of Medicine, University of Debrecen, 4032 Debrecen, Hungary"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8568-914X","authenticated-orcid":false,"given":"Tibor","family":"Nagy","sequence":"additional","affiliation":[{"name":"Department of Applied Chemistry, Faculty of Sciences and Technology, University of Debrecen, 4032 Debrecen, Hungary"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"\u00c1gota","family":"Nagyn\u00e9 Veres","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Molecular Biology, Faculty of Medicine, University of Debrecen, 4032 Debrecen, Hungary"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6452-9937","authenticated-orcid":false,"given":"M\u00e1ria","family":"Golda","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Molecular Biology, Faculty of Medicine, University of Debrecen, 4032 Debrecen, Hungary"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7959-5996","authenticated-orcid":false,"given":"Mohamed","family":"Mahdi","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Molecular Biology, Faculty of Medicine, University of Debrecen, 4032 Debrecen, Hungary"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5076-8729","authenticated-orcid":false,"given":"J\u00f3zsef","family":"T\u0151zs\u00e9r","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Molecular Biology, Faculty of Medicine, University of Debrecen, 4032 Debrecen, Hungary"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2025,6,18]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"155","DOI":"10.1038\/s41579-020-00468-6","article-title":"Coronavirus biology and replication: Implications for SARS-CoV-2","volume":"19","author":"Kratzel","year":"2021","journal-title":"Nat. Rev. Microbiol."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"115491","DOI":"10.1016\/j.ejmech.2023.115491","article-title":"The research progress of SARS-CoV-2 main protease inhibitors from 2020 to 2022","volume":"257","author":"Pang","year":"2023","journal-title":"Eur. J. Med. Chem."},{"key":"ref_3","doi-asserted-by":"crossref","unstructured":"M\u00f3ty\u00e1n, J.A., Mahdi, M., Hoffka, G., and T\u0151zs\u00e9r, J. (2022). Potential Resistance of SARS-CoV-2 Main Protease (Mpro) against Protease Inhibitors: Lessons Learned from HIV-1 Protease. Int. J. Mol. Sci., 23.","DOI":"10.3390\/ijms23073507"},{"key":"ref_4","doi-asserted-by":"crossref","unstructured":"Denaro, M., Ferro, E., Barrano, G., Meli, S., Busacca, M., Corallo, D., Capici, A., Zisa, A., Cucuzza, L., and Gradante, S. (2023). Monitoring of SARS-CoV-2 Infection in Ragusa Area: Next Generation Sequencing and Serological Analysis. Int. J. Mol. 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Viruses, 17.","DOI":"10.3390\/v17050691"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"100793","DOI":"10.1016\/j.xpro.2021.100793","article-title":"Fluorogenic in vitro activity assay for the main protease Mpro from SARS-CoV-2 and its adaptation to the identification of inhibitors","volume":"2","author":"Ihssen","year":"2021","journal-title":"STAR Protoc."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"1189","DOI":"10.1177\/24725552211020681","article-title":"Efficiency Improvements and Discovery of New Substrates for a SARS-CoV-2 Main Protease FRET Assay","volume":"26","author":"Leuthold","year":"2021","journal-title":"SLAS Discov."},{"key":"ref_8","doi-asserted-by":"crossref","unstructured":"Legare, S., Heide, F., Bailey-Elkin, B.A., and Stetefeld, J. (2022). Improved SARS-CoV-2 main protease high-throughput screening assay using a 5-carboxyfluorescein substrate. J. Biol. Chem., 298.","DOI":"10.1016\/j.jbc.2022.101739"},{"key":"ref_9","doi-asserted-by":"crossref","unstructured":"Suresh, V., Sheik, D.A., Detomasi, T.C., Zhao, T., Zepeda, T., Saladi, S., Rajesh, U.C., Byers, K., Craik, C.S., and Davisson, V.J. (2023). A Prototype Assay Multiplexing SARS-CoV-2 3CL-Protease and Angiotensin-Converting Enzyme 2 for Saliva-Based Diagnostics in COVID-19. Biosensors, 13.","DOI":"10.3390\/bios13070682"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"4138","DOI":"10.1021\/acs.jcim.3c00324","article-title":"Targeting SARS-CoV-2 Main Protease (MPro) with Kinase Inhibitors: A Promising Approach for Discovering Antiviral and Anti-inflammatory Molecules against SARS-CoV-2","volume":"63","author":"Anton","year":"2023","journal-title":"J. Chem. Inf. Model."},{"key":"ref_11","doi-asserted-by":"crossref","unstructured":"Miczi, M., Golda, M., Kunkli, B., Nagy, T., T\u0151zs\u00e9r, J., and M\u00f3ty\u00e1n, J.A. (2020). Identification of Host Cellular Protein Substrates of SARS-COV-2 Main Protease. Int. J. Mol. Sci., 21.","DOI":"10.3390\/ijms21249523"},{"key":"ref_12","doi-asserted-by":"crossref","unstructured":"Miltner, N., Kall\u00f3, G., Cs\u0151sz, \u00c9., Miczi, M., Nagy, T., Mahdi, M., M\u00f3ty\u00e1n, J.A., and T\u0151zs\u00e9r, J. (2023). Identification of SARS-CoV-2 Main Protease (Mpro) Cleavage Sites Using Two-Dimensional Electrophoresis and In Silico Cleavage Site Prediction. Int. J. Mol. Sci., 24.","DOI":"10.3390\/ijms24043236"},{"key":"ref_13","first-page":"899","article-title":"Sigma\u2019s Non-specific Protease Activity Assay\u2014Casein as a Substrate","volume":"19","year":"2008","journal-title":"J. Vis. Exp."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"1867","DOI":"10.1039\/C6AN02647H","article-title":"Recent developments in protease activity assays and sensors","volume":"142","author":"Ong","year":"2017","journal-title":"Analyst"},{"key":"ref_15","first-page":"e60421","article-title":"Analysis of Group IV Viral SSHHPS Using In Vitro and In Silico Methods","volume":"154","author":"Hu","year":"2019","journal-title":"J. Vis. Exp."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"106","DOI":"10.1016\/j.antiviral.2019.02.001","article-title":"Proteolytic cleavage of host proteins by the Group IV viral proteases of Venezuelan equine encephalitis virus and Zika virus","volume":"164","author":"Morazzani","year":"2019","journal-title":"Antivir. Res."},{"key":"ref_17","doi-asserted-by":"crossref","unstructured":"Doctor, K.Z., Gilmour, E., Recarte, M., Beatty, T.R., Shifa, I., Stangel, M., Schwisow, J., Leary, D.H., and Legler, P.M. (2023). Automated SSHHPS Analysis Predicts a Potential Host Protein Target Common to Several Neuroinvasive (+)ssRNA Viruses. Viruses, 15.","DOI":"10.3390\/v15020542"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"1483","DOI":"10.1021\/acsinfecdis.0c00866","article-title":"The SARS-CoV-2 SSHHPS Recognized by the Papain-like Protease","volume":"7","author":"Reynolds","year":"2021","journal-title":"ACS Infect. Dis."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"131","DOI":"10.1016\/j.febslet.2004.08.017","article-title":"Characterization of trans- and cis-cleavage activity of the SARS coronavirus 3CLpro protease: Basis for the in vitro screening of anti-SARS drugs","volume":"574","author":"Lin","year":"2004","journal-title":"FEBS Lett."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"2622","DOI":"10.1016\/j.ijbiomac.2020.08.166","article-title":"Screening and evaluation of approved drugs as inhibitors of main protease of SARS-CoV-2","volume":"164","author":"Tripathi","year":"2020","journal-title":"Int. J. Biol. Macromol."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"33151","DOI":"10.1021\/acsomega.0c04808","article-title":"Structure-Based Virtual Screening and Biochemical Validation to Discover a Potential Inhibitor of the SARS-CoV-2 Main Protease","volume":"5","author":"Gupta","year":"2020","journal-title":"ACS Omega"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"3411","DOI":"10.1002\/ptr.6802","article-title":"Evaluation of medicinal herbs as a potential therapeutic option against SARS-CoV-2 targeting its main protease","volume":"34","author":"Upadhyay","year":"2020","journal-title":"Phytother. Res."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"5496","DOI":"10.1111\/febs.16947","article-title":"Identification of a novel inhibitor of SARS-CoV-2 main protease: An in silico, biochemical, and cell-based approach","volume":"290","author":"Trivedi","year":"2023","journal-title":"FEBS J."},{"key":"ref_24","doi-asserted-by":"crossref","unstructured":"Kiemer, L., Lund, O., Brunak, S., and Blom, N. (2004). Coronavirus 3CLpro proteinase cleavage sites: Possible relevance to SARS virus pathology. BMC Bioinform., 5.","DOI":"10.1186\/1471-2105-5-72"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"437","DOI":"10.1016\/j.virs.2022.04.006","article-title":"Prediction of coronavirus 3C-like protease cleavage sites using machine-learning algorithms","volume":"37","author":"Chen","year":"2022","journal-title":"Virol. Sin."},{"key":"ref_26","doi-asserted-by":"crossref","unstructured":"Scott, B.M., Lacasse, V., Blom, D.G., Tonner, P.D., and Blom, N.S. (2022). Predicted coronavirus Nsp5 protease cleavage sites in the human proteome. BMC Genom. Data, 23.","DOI":"10.1186\/s12863-022-01044-y"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"8744","DOI":"10.1021\/bi0621415","article-title":"Substrate specificity profiling and identification of a new class of inhibitor for the major protease of the SARS coronavirus","volume":"46","author":"Goetz","year":"2007","journal-title":"Biochemistry"},{"key":"ref_28","doi-asserted-by":"crossref","unstructured":"Chuck, C.P., Chow, H.F., Wan, D.C., and Wong, K.B. (2011). Profiling of substrate specificities of 3C-like proteases from group 1, 2a, 2b, and 3 coronaviruses. PLoS ONE, 6.","DOI":"10.1371\/journal.pone.0027228"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"e2000246","DOI":"10.1002\/pmic.202000246","article-title":"N-Terminomics for the Identification of In Vitro Substrates and Cleavage Site Specificity of the SARS-CoV-2 Main Protease","volume":"21","author":"Koudelka","year":"2021","journal-title":"Proteomics"},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"109892","DOI":"10.1016\/j.celrep.2021.109892","article-title":"Mechanistic insights into COVID-19 by global analysis of the SARS-CoV-2 3CLpro substrate degradome","volume":"37","author":"Pablos","year":"2021","journal-title":"Cell Rep."},{"key":"ref_31","doi-asserted-by":"crossref","unstructured":"Taylor, E.W., and Radding, W. (2020). Understanding Selenium and Glutathione as Antiviral Factors in COVID-19: Does the Viral Mpro Protease Target Host Selenoproteins and Glutathione Synthesis?. Front. Nutr., 7.","DOI":"10.3389\/fnut.2020.00143"},{"key":"ref_32","doi-asserted-by":"crossref","unstructured":"Gallardo, I.A., Todd, D.A., Lima, S.T., Chekan, J.R., Chiu, N.H., and Taylor, E.W. (2023). SARS-CoV-2 Main Protease Targets Host Selenoproteins and Glutathione Biosynthesis for Knockdown via Proteolysis, Potentially Disrupting the Thioredoxin and Glutaredoxin Redox Cycles. Antioxidants, 12.","DOI":"10.3390\/antiox12030559"},{"key":"ref_33","doi-asserted-by":"crossref","unstructured":"Gazda, L.D., Jo\u00f3n\u00e9 Mat\u00faz, K., Nagy, T., M\u00f3ty\u00e1n, J.A., and T\u0151zs\u00e9r, J. (2020). Biochemical characterization of Ty1 retrotransposon protease. PLoS ONE, 15.","DOI":"10.1371\/journal.pone.0227062"},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"189","DOI":"10.1007\/BF03195213","article-title":"Polymorphism of bovine beta-casein and its potential effect on human health","volume":"48","author":"Kostyra","year":"2007","journal-title":"J. Appl. Genet."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"15","DOI":"10.3168\/jds.2011-4543","article-title":"Determination of molecular weights of caprine milk proteins by matrix-assisted laser desorption\/ionization mass spectrometry","volume":"95","author":"Ham","year":"2012","journal-title":"J. Dairy Sci."},{"key":"ref_36","doi-asserted-by":"crossref","unstructured":"Vincent, D., Elkins, A., Condina, M.R., Ezernieks, V., and Rochfort, S. (2016). Quantitation and Identification of Intact Major Milk Proteins for High-Throughput LC-ESI-Q-TOF MS Analyses. PLoS ONE, 11.","DOI":"10.1371\/journal.pone.0163471"},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"5553","DOI":"10.1038\/s41467-021-25796-w","article-title":"Characterising proteolysis during SARS-CoV-2 infection identifies viral cleavage sites and cellular targets with therapeutic potential","volume":"12","author":"Meyer","year":"2021","journal-title":"Nat. Commun."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"190","DOI":"10.1186\/s12985-020-01457-0","article-title":"Analysis of the efficacy of HIV protease inhibitors against SARS-CoV-2\u2032s main protease","volume":"17","author":"Mahdi","year":"2020","journal-title":"Virol. J."},{"key":"ref_39","unstructured":"UniProt Consortium (2021). UniProt: The universal protein knowledgebase in 2021. Nucleic Acids Res., 49, D480\u2013D489."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"10423","DOI":"10.1002\/anie.202016961","article-title":"Targeting the Main Protease of SARS-CoV-2: From the Establishment of High Throughput Screening to the Design of Tailored Inhibitors","volume":"60","author":"Breidenbach","year":"2021","journal-title":"Angew. Chem. Int. Ed. Engl."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1007\/s00394-021-02551-x","article-title":"A1- and A2 beta-casein on health-related outcomes: A scoping review of animal studies","volume":"61","author":"Lohner","year":"2022","journal-title":"Eur. J. Nutr."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"127765","DOI":"10.1016\/j.foodchem.2020.127765","article-title":"\u03b2-Casomorphin: A complete health perspective","volume":"337","author":"Thiruvengadam","year":"2021","journal-title":"Food Chem."},{"key":"ref_43","doi-asserted-by":"crossref","unstructured":"Bolat, E., Eker, F., Y\u0131lmaz, S., Karav, S., Oz, E., Brennan, C., Proestos, C., Zeng, M., and Oz, F. (2024). BCM-7: Opioid-like Peptide with Potential Role in Disease Mechanisms. Molecules, 29.","DOI":"10.3390\/molecules29092161"},{"key":"ref_44","doi-asserted-by":"crossref","unstructured":"de Vasconcelos, M.L., Oliveira, L.M.F.S., Hill, J.P., and Vidal, A.M.C. (2023). Difficulties in Establishing the Adverse Effects of \u03b2-Casomorphin-7 Released from \u03b2-Casein Variants-A Review. Foods, 12.","DOI":"10.3390\/foods12173151"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"1474","DOI":"10.1021\/acschembio.4c00096","article-title":"High-Resolution Substrate Specificity Profiling of SARS-CoV-2 Mpro; Comparison to SARS-CoV Mpro","volume":"19","author":"Yaghi","year":"2024","journal-title":"ACS Chem. Biol."},{"key":"ref_46","doi-asserted-by":"crossref","unstructured":"van Vliet, V.J.E., Huynh, N., Pal\u00e0, J., Patel, A., Singer, A., Slater, C., Chung, J., van Huizen, M., Teyra, J., and Miersch, S. (2022). Ubiquitin variants potently inhibit SARS-CoV-2 PLpro and viral replication via a novel site distal to the protease active site. PLoS Pathog., 18.","DOI":"10.1371\/journal.ppat.1011065"}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/26\/12\/5829\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,9]],"date-time":"2025-10-09T17:53:59Z","timestamp":1760032439000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/26\/12\/5829"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,6,18]]},"references-count":46,"journal-issue":{"issue":"12","published-online":{"date-parts":[[2025,6]]}},"alternative-id":["ijms26125829"],"URL":"https:\/\/doi.org\/10.3390\/ijms26125829","relation":{},"ISSN":["1422-0067"],"issn-type":[{"type":"electronic","value":"1422-0067"}],"subject":[],"published":{"date-parts":[[2025,6,18]]}}}