{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,21]],"date-time":"2026-07-21T05:05:22Z","timestamp":1784610322414,"version":"3.55.0"},"reference-count":72,"publisher":"MDPI AG","issue":"7","license":[{"start":{"date-parts":[[2026,7,17]],"date-time":"2026-07-17T00:00:00Z","timestamp":1784246400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Pharmaceuticals"],"abstract":"<jats:p>Background\/Objectives: Cryptosporidiosis, caused by Cryptosporidium parvum, is a significant cause of diarrheal disease, particularly affecting young children and immunocompromised individuals. With current treatments offering limited efficacy, there is an urgent need for novel therapeutic targets. Methods: In this study, we report the cloning, expression, and functional characterization of a glutathione transferase (GST) from C. parvum (CpGST). Results: Biocomputing analysis revealed a single gene encoding a cytosolic enzyme with distinct structural features, compared to human cytosolic homologs. Structural modeling indicated a non-canonical thioredoxin fold and a truncated C-terminal domain, suggesting functional divergence. CpGST was expressed in Escherichia coli, and its enzymatic properties were characterized. Although the enzyme displayed a narrow substrate spectrum, it showed a distinct substrate preference, retaining catalytic activity toward the standard GST substrates 1-chloro-2,4-dinitrobenzene (CDNB) and cumene hydroperoxide (CuOOH). Steady-state kinetic analysis revealed limited affinity for both reduced glutathione (GSH) and CDNB. Inhibition analysis identified several polyphenols and synthetic curcumin analogues as potent inhibitors, with IC50 values in the low micromolar range. Kinetic analysis with the most potent inhibitor revealed a mixed-type inhibition mechanism. Conclusions: These findings support the classification of CpGST as a structurally and functionally distinct member of the GST family, likely adapted to the parasite\u2019s physiology and metabolism. The enzyme\u2019s divergence from human GSTs, along with its favorable druggability profile, underscores its potential as a target for anti-cryptosporidial drug development, particularly in strategies aimed at disrupting stress response and detoxification pathways.<\/jats:p>","DOI":"10.3390\/ph19071106","type":"journal-article","created":{"date-parts":[[2026,7,20]],"date-time":"2026-07-20T00:47:32Z","timestamp":1784508452000},"page":"1106","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Functional Characterization and Inhibition Analysis of a Glutathione Transferase from Cryptosporidium parvum: A Potential Target for Antiparasitic Drug Development"],"prefix":"10.3390","volume":"19","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-4675-0435","authenticated-orcid":false,"given":"Panagiota D.","family":"Pantiora","sequence":"first","affiliation":[{"name":"Laboratory of Enzyme Technology, Department of Biotechnology, School of Applied Biology and Biotechnology, Agricultural University of Athens, 75 Iera Odos Street, 11855 Athens, Greece"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2437-0546","authenticated-orcid":false,"given":"Nikolaos D.","family":"Georgakis","sequence":"additional","affiliation":[{"name":"Laboratory of Enzyme Technology, Department of Biotechnology, School of Applied Biology and Biotechnology, Agricultural University of Athens, 75 Iera Odos Street, 11855 Athens, Greece"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0059-952X","authenticated-orcid":false,"given":"Dimitris","family":"Matiadis","sequence":"additional","affiliation":[{"name":"Laboratory of Chemistry, Department of Food Science & Human Nutrition, School of Food and Nutritional Sciences, Agricultural University of Athens, Iera Odos 75, 11855 Athens, Greece"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4231-6658","authenticated-orcid":false,"given":"Marina","family":"Sagnou","sequence":"additional","affiliation":[{"name":"Institute of Biosciences & Applications, National Centre for Scientific Research \u201cDemokritos\u201d, 15310 Athens, Greece"},{"name":"Chemical Biology Laboratory, RTIN and Department of Science and Mathematics\u2014Deree, The American College of Greece, 15342 Athens, Greece"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1925-9867","authenticated-orcid":false,"given":"Nikolaos E.","family":"Labrou","sequence":"additional","affiliation":[{"name":"Laboratory of Enzyme Technology, Department of Biotechnology, School of Applied Biology and Biotechnology, Agricultural University of Athens, 75 Iera Odos Street, 11855 Athens, Greece"},{"name":"Institute of Nanobiotechnology, Sustainable Development and Artificial Intelligence, Research and Innovation Center, Agricultural University of Athens, Iera Odos 75, 11855 Athens, Greece"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,7,17]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"988","DOI":"10.15212\/ZOONOSES-2022-0011","article-title":"Zoonotic Cryptosporidium parasites possess a unique carbohydrate-binding protein (malectin) that is absent in other apicomplexan lineages","volume":"2","author":"Wu","year":"2022","journal-title":"Zoonoses"},{"key":"ref_2","doi-asserted-by":"crossref","unstructured":"Helmy, Y.A., El-Adawy, H., and Abdelwhab, E.M. (2017). A comprehensive review of common bacterial, parasitic and viral zoonoses at the human\u2013animal interface in Egypt. Pathogens, 6.","DOI":"10.3390\/pathogens6030033"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"568","DOI":"10.15585\/mmwr.mm6825a3","article-title":"Cryptosporidiosis outbreaks\u2014United States, 2009\u20132017","volume":"68","author":"Gharpure","year":"2019","journal-title":"Morb. Mortal. Wkly. Rep."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"1","DOI":"10.2166\/wst.2020.515","article-title":"A review of Cryptosporidium spp. and their detection in water","volume":"83","author":"Hassan","year":"2021","journal-title":"Water Sci. Technol."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"3395","DOI":"10.5433\/1679-0359.2017v38n5p3395","article-title":"Waterborne Giardia and Cryptosporidium: Contamination of human drinking water by sewage and cattle feces","volume":"38","author":"Martins","year":"2017","journal-title":"Semin. Ci\u00eanc. Agr\u00e1r."},{"key":"ref_6","doi-asserted-by":"crossref","unstructured":"Khan, S.M., and Witola, W.H. (2023). Past, current, and potential treatments for cryptosporidiosis in humans and farm animals: A comprehensive review. Front. Cell. Infect. Microbiol., 13.","DOI":"10.3389\/fcimb.2023.1115522"},{"key":"ref_7","first-page":"2300733","article-title":"Marked increase in cryptosporidiosis cases, Spain, 2023","volume":"29","author":"Martinez","year":"2024","journal-title":"Eurosurveillance"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"62","DOI":"10.1007\/s40588-021-00159-7","article-title":"Cryptosporidium: Host\u2013parasite interactions and pathogenesis","volume":"8","author":"Pinto","year":"2021","journal-title":"Curr. Clin. Microbiol. Rep."},{"key":"ref_9","unstructured":"European Centre for Disease Prevention and Control (2021). Cryptosporidiosis\u2014Annual Epidemiological Report for 2021, ECDC."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"549","DOI":"10.1097\/QAD.0b013e3283437e88","article-title":"Cryptosporidiosis in patients with HIV\/AIDS","volume":"25","author":"Shaffie","year":"2011","journal-title":"AIDS"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"48","DOI":"10.1111\/tid.12336","article-title":"Cryptosporidium infection after renal transplantation in an endemic area","volume":"17","author":"Bhadauria","year":"2015","journal-title":"Transpl. Infect. Dis."},{"key":"ref_12","doi-asserted-by":"crossref","unstructured":"Helmy, Y.A., and Hafez, H.M. (2022). Cryptosporidiosis: From prevention to treatment, a narrative review. Microorganisms, 10.","DOI":"10.3390\/microorganisms10122456"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"72","DOI":"10.1128\/CMR.17.1.72-97.2004","article-title":"Cryptosporidium taxonomy: Recent advances and implications for public health","volume":"17","author":"Xiao","year":"2004","journal-title":"Clin. Microbiol. Rev."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"1667","DOI":"10.1017\/S0031182014001085","article-title":"Cryptosporidium species in humans and animals: Current understanding and research needs","volume":"141","author":"Ryan","year":"2014","journal-title":"Parasitology"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"455","DOI":"10.1097\/QCO.0000000000000761","article-title":"Emerging treatment options for cryptosporidiosis","volume":"34","author":"Love","year":"2021","journal-title":"Curr. Opin. Infect. Dis."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"1375","DOI":"10.1016\/S0140-6736(02)11401-2","article-title":"Effect of nitazoxanide on morbidity and mortality in Zambian children with cryptosporidiosis: A randomised controlled trial","volume":"360","author":"Amadi","year":"2002","journal-title":"Lancet"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"45","DOI":"10.1016\/j.exppara.2009.07.005","article-title":"Cryptosporidium and Giardia: Treatment options and prospects for new drugs","volume":"124","author":"Rossignol","year":"2010","journal-title":"Exp. Parasitol."},{"key":"ref_18","unstructured":"Fayer, R. (1997). Prophylaxis and chemotherapy: Human and animal. Cryptosporidium and Cryptosporidiosis, CRC Press."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"213","DOI":"10.1007\/s100960050461","article-title":"Effect of antiretroviral therapy on cryptosporidiosis and microsporidiosis in patients infected with human immunodeficiency virus type 1","volume":"19","author":"Maggi","year":"2000","journal-title":"Eur. J. Clin. Microbiol. Infect. Dis."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"193","DOI":"10.1159\/000086920","article-title":"Treatment of cryptosporidiosis in immunocompromised hosts","volume":"51","author":"Zardi","year":"2005","journal-title":"Chemotherapy"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"453","DOI":"10.1093\/jac\/45.4.453","article-title":"Activity of nitazoxanide alone and in combination with azithromycin and rifabutin against Cryptosporidium parvum in cell culture","volume":"45","author":"Giacometti","year":"2000","journal-title":"J. Antimicrob. Chemother."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"85","DOI":"10.1016\/S1473-3099(14)70772-8","article-title":"A review of the global burden, novel diagnostics, therapeutics, and vaccine targets for Cryptosporidium","volume":"15","author":"Checkley","year":"2015","journal-title":"Lancet Infect. Dis."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"915","DOI":"10.1080\/14728222.2020.1785432","article-title":"Novel drug targets for treatment of cryptosporidiosis","volume":"24","author":"Wang","year":"2020","journal-title":"Expert Opin. Ther. Targets"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"331","DOI":"10.1021\/ml100096t","article-title":"Discovery of potent and selective inhibitors of CDPK1 from C. parvum and T. gondii","volume":"1","author":"Murphy","year":"2010","journal-title":"ACS Med. Chem. Lett."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"602","DOI":"10.1038\/nsmb.1818","article-title":"Toxoplasma gondii calcium-dependent protein kinase 1 is a target for selective kinase inhibitors","volume":"17","author":"Ojo","year":"2010","journal-title":"Nat. Struct. Mol. Biol."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"376","DOI":"10.1038\/nature22337","article-title":"A Cryptosporidium PI(4)K inhibitor is a drug candidate for cryptosporidiosis","volume":"546","author":"Manjunatha","year":"2017","journal-title":"Nature"},{"key":"ref_27","doi-asserted-by":"crossref","unstructured":"Mead, J.R., and Arrowood, M.J. (2020). Cryptosporidium: Methods and Protocols; Methods in Molecular Biology, Humana Press.","DOI":"10.1007\/978-1-4939-9748-0"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"638","DOI":"10.2307\/3283792","article-title":"Efficacy of serine protease inhibitors against Cryptosporidium parvum infection in a bovine fallopian tube epithelial cell culture system","volume":"82","author":"Forney","year":"1996","journal-title":"J. Parasitol."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"233","DOI":"10.1111\/jeu.12272","article-title":"Differential gene expression and protein localization of Cryptosporidium parvum fatty acyl-CoA synthetase isoforms","volume":"63","author":"Guo","year":"2016","journal-title":"J. Eukaryot. Microbiol."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"186","DOI":"10.1016\/j.copbio.2014.12.024","article-title":"Plant GSTome: Structure and functional role in xenome network and plant stress response","volume":"32","author":"Labrou","year":"2015","journal-title":"Curr. Opin. Biotechnol."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"63","DOI":"10.3390\/antiox12010063","article-title":"Monocarbonyl curcumin analogues as potent inhibitors against human glutathione transferase P1-1","volume":"12","author":"Pantiora","year":"2023","journal-title":"Antioxidants"},{"key":"ref_32","doi-asserted-by":"crossref","unstructured":"Sabt, A., Kitsos, S., Ebaid, M.S., Furlan, V., Pantiora, P.D., Tsolka, M., Elkaeed, E.B., Hamissa, M.F., Angelis, N., and Tsitsilonis, O.E. (2024). Novel coumarin-6-sulfonamide-chalcone hybrids as glutathione transferase P1-1 inhibitors. PLoS ONE, 19.","DOI":"10.1371\/journal.pone.0306124"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"1373","DOI":"10.1038\/cdd.2010.80","article-title":"Glutathione transferases as mediators of signaling pathways involved in cell proliferation and cell death","volume":"17","author":"Laborde","year":"2010","journal-title":"Cell Death Differ."},{"key":"ref_34","doi-asserted-by":"crossref","unstructured":"Karpusas, M., Axarli, I., Chiniadis, L., Papakyriakou, A., Bethanis, K., Scopelitou, K., Clonis, Y.D., and Labrou, N.E. (2013). The interaction of the chemotherapeutic drug chlorambucil with human glutathione transferase A1-1: Kinetic and structural analysis. PLoS ONE, 8.","DOI":"10.1371\/journal.pone.0056337"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"511","DOI":"10.1080\/07388551.2017.1375890","article-title":"Recent advances in protein engineering and biotechnological applications of glutathione transferases","volume":"38","author":"Perperopoulou","year":"2018","journal-title":"Crit. Rev. Biotechnol."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"24876","DOI":"10.1074\/jbc.270.42.24876","article-title":"Heme degradation in the presence of glutathione: A proposed mechanism to account for the high levels of non-heme iron found in the membranes of hemoglobinopathic red blood cells","volume":"270","author":"Atamna","year":"1995","journal-title":"J. Biol. Chem."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"365","DOI":"10.1016\/j.jsb.2015.06.008","article-title":"High resolution structures of Plasmodium falciparum GST complexes provide novel insights into the dimer\u2013tetramer transition and a novel ligand-binding site","volume":"191","author":"Perbandt","year":"2015","journal-title":"J. Struct. Biol."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"e21352","DOI":"10.1096\/fj.202001987RR","article-title":"Toxoplasma gondii glutathione S-transferase 2 plays an important role in partial secretory protein transport","volume":"35","author":"Li","year":"2021","journal-title":"FASEB J."},{"key":"ref_39","first-page":"8486021","article-title":"Exploring the infectious drug target glutathione S-transferase in Plasmodium falciparum with the inhibitory potential of Azadirachta indica phytocompounds","volume":"2024","year":"2024","journal-title":"Adv. Public Health"},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"441","DOI":"10.1126\/science.1094786","article-title":"Complete genome sequence of the apicomplexan, Cryptosporidium parvum","volume":"304","author":"Abrahamsen","year":"2004","journal-title":"Science"},{"key":"ref_41","doi-asserted-by":"crossref","unstructured":"Mauzy, M.J., Enomoto, S., Lancto, C.A., Abrahamsen, M.S., and Rutherford, M.S. (2012). The Cryptosporidium parvum transcriptome during in vitro development. PLoS ONE, 7.","DOI":"10.1371\/journal.pone.0031715"},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"20370","DOI":"10.1038\/s41598-020-77233-5","article-title":"Comparative analyses and structural insights of new class glutathione transferases in Cryptosporidium species","volume":"10","author":"Mfeka","year":"2020","journal-title":"Sci. Rep."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"148","DOI":"10.1016\/j.ijpddr.2016.07.003","article-title":"Evaluation of Giardia lamblia thioredoxin reductase as drug activating enzyme and as drug target","volume":"6","author":"Leitsch","year":"2016","journal-title":"Int. J. Parasitol. Drugs Drug Resist."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"15631","DOI":"10.1021\/bi991179x","article-title":"Role of the C-terminal helix 9 in the stability and ligandin function of class \u03b1 glutathione transferase A1-1","volume":"38","author":"Dirr","year":"1999","journal-title":"Biochemistry"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"5639","DOI":"10.1074\/jbc.M109.074757","article-title":"Minor modifications of the C-terminal helix reschedule the favored chemical reactions catalyzed by theta class glutathione transferase T1-1","volume":"285","author":"Shokeer","year":"2010","journal-title":"J. Biol. Chem."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"857","DOI":"10.1006\/jmbi.1997.1338","article-title":"Crystallization, structural determination and analysis of a novel parasite vaccine candidate: Fasciola hepatica glutathione S-transferase","volume":"273","author":"Rossjohn","year":"1997","journal-title":"J. Mol. Biol."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"e21838","DOI":"10.1002\/jbt.21838","article-title":"Crystal structure of a class-mu glutathione S-transferase from whiteleg shrimp Litopenaeus vannamei: Structural changes in the xenobiotic binding H-site may alter the spectra of molecules bound","volume":"31","year":"2017","journal-title":"J. Biochem. Mol. Toxicol."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"453","DOI":"10.1042\/bj3110453","article-title":"Isothiocyanates as substrates for human glutathione transferases: Structure\u2013activity studies","volume":"311","author":"Kolm","year":"1995","journal-title":"Biochem. J."},{"key":"ref_49","doi-asserted-by":"crossref","unstructured":"Morgenstern, R. (2024). Kinetic behavior of glutathione transferases: Understanding cellular protection from reactive intermediates. Biomolecules, 14.","DOI":"10.3390\/biom14060641"},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1042\/bj3600001","article-title":"Structure, function and evolution of glutathione transferases: Implications for classification of non-mammalian members of an ancient enzyme superfamily","volume":"360","author":"Sheehan","year":"2001","journal-title":"Biochem. J."},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"325","DOI":"10.2174\/138920308785132677","article-title":"Mammalian cytosolic glutathione transferases","volume":"9","author":"Dourado","year":"2008","journal-title":"Curr. Protein Pept. Sci."},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"1155","DOI":"10.1007\/s00436-009-1535-5","article-title":"Effects of curcumin on Cryptosporidium parvum in vitro","volume":"105","author":"Shahiduzzaman","year":"2009","journal-title":"Parasitol. Res."},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"7","DOI":"10.1016\/j.vetpar.2017.12.008","article-title":"Comparative efficacy of curcumin and paromomycin against Cryptosporidium parvum infection in a BALB\/c model","volume":"250","author":"Asadpour","year":"2018","journal-title":"Vet. Parasitol."},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"5885","DOI":"10.1128\/IAI.67.11.5885-5891.1999","article-title":"Reactive nitrogen and oxygen species ameliorate experimental cryptosporidiosis in the neonatal BALB\/c mouse model","volume":"67","author":"Leitch","year":"1999","journal-title":"Infect. Immun."},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"13","DOI":"10.1017\/S0031182096008037","article-title":"Anti-oxidant enzymes in Cryptosporidium parvum oocysts","volume":"114","author":"Entrala","year":"1997","journal-title":"Parasitology"},{"key":"ref_56","doi-asserted-by":"crossref","first-page":"747","DOI":"10.1016\/j.ijpara.2008.11.012","article-title":"An apicomplexan ankyrin-repeat histone deacetylase with relatives in photosynthetic eukaryotes","volume":"39","author":"Rider","year":"2009","journal-title":"Int. J. Parasitol."},{"key":"ref_57","doi-asserted-by":"crossref","first-page":"181","DOI":"10.1007\/s00436-003-0831-8","article-title":"The role of Cryptosporidium parvum-derived phospholipase in intestinal epithelial cell invasion","volume":"90","author":"Pollok","year":"2003","journal-title":"Parasitol. Res."},{"key":"ref_58","doi-asserted-by":"crossref","unstructured":"Tsouri, S., Tselo, E., Premetis, G.E., Furlan, V., Pantiora, P.D., Mavroidi, B., Matiadis, D., Pelecanou, M., Papageorgiou, A.C., and Bren, U. (2024). A monocarbonyl curcuminoid derivative inhibits the activity of human glutathione transferase A4-4 and chemosensitizes glioblastoma cells to temozolomide. Pharmaceuticals, 17.","DOI":"10.3390\/ph17030365"},{"key":"ref_59","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1016\/j.jgar.2019.01.023","article-title":"Drug resistance in protozoan parasites: An incessant wrestle for survival","volume":"18","author":"Pramanik","year":"2019","journal-title":"J. Glob. Antimicrob. Resist."},{"key":"ref_60","doi-asserted-by":"crossref","first-page":"2212","DOI":"10.1021\/acs.biochem.5c00059","article-title":"Targeting apicomplexan parasites: Structural and functional characterization of Cryptosporidium thioredoxin reductase as a novel drug target","volume":"64","author":"Gabriele","year":"2025","journal-title":"Biochemistry"},{"key":"ref_61","doi-asserted-by":"crossref","first-page":"W174","DOI":"10.1093\/nar\/gkv342","article-title":"I-TASSER server: New development for protein structure and function predictions","volume":"43","author":"Yang","year":"2015","journal-title":"Nucleic Acids Res."},{"key":"ref_62","doi-asserted-by":"crossref","first-page":"W291","DOI":"10.1093\/nar\/gkx366","article-title":"COFACTOR: Improved protein function prediction by combining structure, sequence and protein\u2013protein interaction information","volume":"45","author":"Zhang","year":"2017","journal-title":"Nucleic Acids Res."},{"key":"ref_63","doi-asserted-by":"crossref","first-page":"D344","DOI":"10.1093\/nar\/gkaa977","article-title":"The InterPro protein families and domains database: 20 years on","volume":"49","author":"Blum","year":"2021","journal-title":"Nucleic Acids Res."},{"key":"ref_64","doi-asserted-by":"crossref","first-page":"2922","DOI":"10.1093\/molbev\/msz185","article-title":"Sequenceserver: A modern graphical user interface for custom BLAST databases","volume":"36","author":"Priyam","year":"2019","journal-title":"Mol. Biol. Evol."},{"key":"ref_65","doi-asserted-by":"crossref","first-page":"W293","DOI":"10.1093\/nar\/gkab301","article-title":"Interactive Tree Of Life (iTOL) v5: An online tool for phylogenetic tree display and annotation","volume":"49","author":"Letunic","year":"2021","journal-title":"Nucleic Acids Res."},{"key":"ref_66","doi-asserted-by":"crossref","first-page":"539","DOI":"10.1038\/msb.2011.75","article-title":"Fast, scalable generation of high-quality protein multiple sequence alignments using Clustal Omega","volume":"7","author":"Sievers","year":"2011","journal-title":"Mol. Syst. Biol."},{"key":"ref_67","doi-asserted-by":"crossref","first-page":"W636","DOI":"10.1093\/nar\/gkz268","article-title":"The EMBL-EBI search and sequence analysis tools APIs in 2019","volume":"47","author":"Madeira","year":"2019","journal-title":"Nucleic Acids Res."},{"key":"ref_68","doi-asserted-by":"crossref","first-page":"W320","DOI":"10.1093\/nar\/gku316","article-title":"Deciphering key features in protein structures with the new ENDscript server","volume":"42","author":"Robert","year":"2014","journal-title":"Nucleic Acids Res."},{"key":"ref_69","doi-asserted-by":"crossref","first-page":"583","DOI":"10.1038\/s41586-021-03819-2","article-title":"Highly accurate protein structure prediction with AlphaFold","volume":"596","author":"Jumper","year":"2021","journal-title":"Nature"},{"key":"ref_70","doi-asserted-by":"crossref","first-page":"590","DOI":"10.1038\/s41586-021-03828-1","article-title":"Highly accurate protein structure prediction for the human proteome","volume":"596","author":"Tunyasuvunakool","year":"2021","journal-title":"Nature"},{"key":"ref_71","unstructured":"Corso, G., Deng, A., Fry, B., Polizzi, N., Barzilay, R., and Jaakkola, T. (2024). Deep Confident Steps to New Pockets: Strategies for Docking Generalization. arXiv."},{"key":"ref_72","doi-asserted-by":"crossref","first-page":"853","DOI":"10.1002\/cmdc.200700010","article-title":"From Modeling to Medicinal Chemistry: Automatic Generation of Two-Dimensional Complex Diagrams","volume":"2","author":"Stierand","year":"2007","journal-title":"ChemMedChem"}],"container-title":["Pharmaceuticals"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1424-8247\/19\/7\/1106\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,7,21]],"date-time":"2026-07-21T04:29:25Z","timestamp":1784608165000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1424-8247\/19\/7\/1106"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,7,17]]},"references-count":72,"journal-issue":{"issue":"7","published-online":{"date-parts":[[2026,7]]}},"alternative-id":["ph19071106"],"URL":"https:\/\/doi.org\/10.3390\/ph19071106","relation":{},"ISSN":["1424-8247"],"issn-type":[{"value":"1424-8247","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,7,17]]}}}