{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,29]],"date-time":"2026-09-29T20:06:26Z","timestamp":1790712386949,"version":"4.1.0"},"reference-count":69,"publisher":"MDPI AG","issue":"1","license":[{"start":{"date-parts":[[2022,1,10]],"date-time":"2022-01-10T00:00:00Z","timestamp":1641772800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001505","name":"Health Research Council of New Zealand","doi-asserted-by":"publisher","award":["13\/263, 16\/232 and 19\/397"],"award-info":[{"award-number":["13\/263, 16\/232 and 19\/397"]}],"id":[{"id":"10.13039\/501100001505","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["JoF"],"abstract":"<jats:p>The fungal cytochrome P450 lanosterol 14\u03b1-demethylase (CYP51) is required for the biosynthesis of fungal-specific ergosterol and is the target of azole antifungal drugs. Despite proven success as a clinical target for azole antifungals, there is an urgent need to develop next-generation antifungals that target CYP51 to overcome the resistance of pathogenic fungi to existing azole drugs, toxic adverse reactions and drug interactions due to human drug-metabolizing CYPs. Candida parapsilosis is a readily transmitted opportunistic fungal pathogen that causes candidiasis in health care environments. In this study, we have characterised wild type C. parapsilosis CYP51 and its clinically significant, resistance-causing point mutation Y132F by expressing these enzymes in a Saccharomyces cerevisiae host system. In some cases, the enzymes were co-expressed with their cognate NADPH-cytochrome P450 reductase (CPR). Constitutive expression of CpCYP51 Y132F conferred a 10- to 12-fold resistance to fluconazole and voriconazole, reduced to ~6-fold resistance for the tetrazoles VT-1161 and VT-1129, but did not confer resistance to the long-tailed triazoles. Susceptibilities were unchanged in the case of CpCPR co-expression. Type II binding spectra showed tight triazole and tetrazole binding by affinity-purified recombinant CpCYP51. We report the X-ray crystal structure of ScCYP51 in complex with VT-1129 obtained at a resolution of 2.1 \u00c5. Structural analysis of azole\u2014enzyme interactions and functional studies of recombinant CYP51 from C. parapsilosis have improved understanding of their susceptibility to azole drugs and will help advance structure-directed antifungal discovery.<\/jats:p>","DOI":"10.3390\/jof8010069","type":"journal-article","created":{"date-parts":[[2022,1,10]],"date-time":"2022-01-10T17:42:25Z","timestamp":1641836545000},"page":"69","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":19,"title":["Characterisation of Candida parapsilosis CYP51 as a Drug Target Using Saccharomyces cerevisiae as Host"],"prefix":"10.3390","volume":"8","author":[{"given":"Yasmeen N.","family":"Ruma","sequence":"first","affiliation":[{"name":"Sir John Walsh Research Institute, Faculty of Dentistry, University of Otago, Dunedin 9016, New Zealand"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mikhail V.","family":"Keniya","sequence":"additional","affiliation":[{"name":"Sir John Walsh Research Institute, Faculty of Dentistry, University of Otago, Dunedin 9016, New Zealand"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0783-1635","authenticated-orcid":false,"given":"Joel D. A.","family":"Tyndall","sequence":"additional","affiliation":[{"name":"School of Pharmacy, University of Otago, Dunedin 9016, New Zealand"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2264-3480","authenticated-orcid":false,"given":"Brian C.","family":"Monk","sequence":"additional","affiliation":[{"name":"Sir John Walsh Research Institute, Faculty of Dentistry, University of Otago, Dunedin 9016, New Zealand"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2022,1,10]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"e00111-18","DOI":"10.1128\/CMR.00111-18","article-title":"Candida parapsilosis: From Genes to the Bedside","volume":"32","author":"Toth","year":"2019","journal-title":"Clin. Microbiol. Rev."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"5","DOI":"10.1111\/1469-0691.12539","article-title":"Global trends in the distribution of Candida species causing candidemia","volume":"20","author":"Guinea","year":"2014","journal-title":"Clin. Microbiol. Infect."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"606","DOI":"10.1128\/CMR.00013-08","article-title":"Candida parapsilosis, an emerging fungal pathogen","volume":"21","author":"Trofa","year":"2008","journal-title":"Clin. Microbiol. Rev."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"e206","DOI":"10.1097\/INF.0b013e3182863a1c","article-title":"Candida parapsilosis is a significant neonatal pathogen: A systematic review and meta-analysis","volume":"32","author":"Pammi","year":"2013","journal-title":"Pediatr. Infect. Dis. J."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"159","DOI":"10.1016\/j.jhin.2004.06.033","article-title":"High frequency of Candida parapsilosis on the hands of healthy hosts","volume":"59","author":"Bonassoli","year":"2005","journal-title":"J. Hosp. Infect."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"fov043","DOI":"10.1093\/femsyr\/fov043","article-title":"Influence of glucose concentration on the structure and quantity of biofilms formed by Candida parapsilosis","volume":"15","author":"Pereira","year":"2015","journal-title":"FEMS Yeast Res."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"1119","DOI":"10.1097\/00006454-200112000-00005","article-title":"Risk factors for Candida species colonization of neonatal intensive care unit patients","volume":"20","author":"Saiman","year":"2001","journal-title":"Pediatr. Infect. Dis. J."},{"key":"ref_8","doi-asserted-by":"crossref","unstructured":"Pinhati, H.M., Casulari, L.A., Souza, A.C., Siqueira, R.A., Damasceno, C.M., and Colombo, A.L. (2016). Outbreak of candidemia caused by fluconazole resistant Candida parapsilosis strains in an intensive care unit. BMC Infect. Dis., 16.","DOI":"10.1186\/s12879-016-1767-9"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"e246","DOI":"10.1016\/j.jhin.2018.06.009","article-title":"Nosocomial outbreak of Candida parapsilosis sensu stricto fungaemia in a neonatal intensive care unit in China","volume":"100","author":"Qi","year":"2018","journal-title":"J. Hosp. Infect."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"792","DOI":"10.1016\/j.cmi.2019.03.028","article-title":"Resistance of Candida to azoles and echinocandins worldwide","volume":"25","author":"Pristov","year":"2019","journal-title":"Clin. Microbiol. Infect."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"232","DOI":"10.3389\/fcimb.2020.00232","article-title":"Prevalence and Clonal Distribution of Azole-Resistant Candida parapsilosis Isolates Causing Bloodstream Infections in a Large Italian Hospital","volume":"10","author":"Martini","year":"2020","journal-title":"Front. Cell Infect. Microbiol."},{"key":"ref_12","doi-asserted-by":"crossref","unstructured":"Pfaller, M.A., Andes, D.R., Diekema, D.J., Horn, D.L., Reboli, A.C., Rotstein, C., Franks, B., and Azie, N.E. (2014). Epidemiology and outcomes of invasive candidiasis due to non-albicans species of Candida in 2496 patients: Data from the Prospective Antifungal Therapy (PATH) registry 2004-2008. PLoS ONE, 9.","DOI":"10.1371\/journal.pone.0101510"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"349","DOI":"10.1046\/j.1439-0507.2000.00582.x","article-title":"The frequency of Candida parapsilosis in onychomycosis. An epidemiological survey in Israel","volume":"43","author":"Segal","year":"2000","journal-title":"Mycoses"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"37","DOI":"10.1155\/2005\/401530","article-title":"Vaginal Candida parapsilosis: Pathogen or bystander?","volume":"13","author":"Nyirjesy","year":"2005","journal-title":"Infect. Dis. Obstet. Gynecol."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"925","DOI":"10.1590\/S0074-02762005000800016","article-title":"Candida colonization in intensive care unit patients\u2019 urine","volume":"100","author":"Passos","year":"2005","journal-title":"Mem. Inst. Oswaldo Cruz"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"241","DOI":"10.1080\/08860220600580431","article-title":"Candida parapsilosis peritonitis has more complications than other Candida peritonitis in peritoneal dialysis patients","volume":"28","author":"Chen","year":"2006","journal-title":"Ren. Fail."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"283","DOI":"10.3109\/10408410903213393","article-title":"Candida parapsilosis: A review of its epidemiology, pathogenesis, clinical aspects, typing and antimicrobial susceptibility","volume":"35","author":"Clemons","year":"2009","journal-title":"Crit. Rev. Microbiol."},{"key":"ref_18","first-page":"e3140","article-title":"Device-associated Central Nervous System Infection Caused by Candida parapsilosis","volume":"10","author":"Bhalla","year":"2018","journal-title":"Cureus"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"e1","DOI":"10.1093\/cid\/civ933","article-title":"Clinical Practice Guideline for the Management of Candidiasis: 2016 Update by the Infectious Diseases Society of America","volume":"62","author":"Pappas","year":"2016","journal-title":"Clin. Infect. Dis."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"396","DOI":"10.1128\/JCM.01398-10","article-title":"Geographic variations in species distribution and echinocandin and azole antifungal resistance rates among Candida bloodstream infection isolates: Report from the SENTRY Antimicrobial Surveillance Program (2008 to 2009)","volume":"49","author":"Pfaller","year":"2011","journal-title":"J. Clin. Microbiol."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"663","DOI":"10.1007\/s11046-016-0028-1","article-title":"Exploring the In Vitro Resistance of Candida parapsilosis to Echinocandins","volume":"181","author":"Chassot","year":"2016","journal-title":"Mycopathologia"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"713687","DOI":"10.1155\/2012\/713687","article-title":"Antifungal resistance and new strategies to control fungal infections","volume":"2012","author":"Vandeputte","year":"2012","journal-title":"Int. J. Microbiol."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"649","DOI":"10.1080\/21505594.2016.1188236","article-title":"The synthesis, regulation, and functions of sterols in Candida albicans: Well-known but still lots to learn","volume":"7","author":"Lv","year":"2016","journal-title":"Virulence"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"1994","DOI":"10.1093\/jac\/dkw091","article-title":"Emergence of azole-resistant Candida parapsilosis causing bloodstream infection: Results from laboratory-based sentinel surveillance in South Africa","volume":"71","author":"Govender","year":"2016","journal-title":"J. Antimicrob. Chemother."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"320","DOI":"10.1111\/myc.12596","article-title":"Detection of neonatal unit clusters of Candida parapsilosis fungaemia by microsatellite genotyping: Results from laboratory-based sentinel surveillance, South Africa, 2009\u20132010","volume":"60","author":"Magobo","year":"2017","journal-title":"Mycoses"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"2997","DOI":"10.3389\/fmicb.2018.02997","article-title":"An Azole-Resistant Candida parapsilosis Outbreak: Clonal Persistence in the Intensive Care Unit of a Brazilian Teaching Hospital","volume":"9","author":"Thomaz","year":"2018","journal-title":"Front. Microbiol."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"1735","DOI":"10.1128\/JCM.00409-07","article-title":"Results from the ARTEMIS DISK Global Antifungal Surveillance study, 1997 to 2005: An 8.5-year analysis of susceptibilities of Candida species and other yeast species to fluconazole and voriconazole determined by CLSI standardized disk diffusion testing","volume":"45","author":"Pfaller","year":"2007","journal-title":"J. Clin. Microbiol."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"6581","DOI":"10.1128\/AAC.01177-15","article-title":"Candida parapsilosis resistance to fluconazole: Molecular mechanisms and in vivo impact in infected galleria mellonella larvae","volume":"59","author":"Souza","year":"2015","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"1030","DOI":"10.1128\/AAC.04613-14","article-title":"Molecular mechanisms of fluconazole resistance in Candida parapsilosis isolates from a U.S. surveillance system","volume":"59","author":"Grossman","year":"2015","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"373","DOI":"10.1016\/j.diagmicrobio.2009.11.006","article-title":"Screening for amino acid substitutions in the Candida albicans Erg11 protein of azole-susceptible and azole-resistant clinical isolates: New substitutions and a review of the literature","volume":"66","author":"Morio","year":"2010","journal-title":"Diagn. Microbiol. Infect. Dis."},{"key":"ref_31","first-page":"2173","article-title":"Azole Antifungal Resistance in Candida albicans and Emerging Non-albicans Candida Species","volume":"7","author":"Whaley","year":"2016","journal-title":"Front. Microbiol."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"891","DOI":"10.1093\/jac\/dkx480","article-title":"A multicentre study of antifungal susceptibility patterns among 350 Candida auris isolates (2009-17) in India: Role of the ERG11 and FKS1 genes in azole and echinocandin resistance","volume":"73","author":"Chowdhary","year":"2018","journal-title":"J. Antimicrob. Chemother."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"778","DOI":"10.1093\/jac\/dks481","article-title":"Mechanisms of azole resistance in 52 clinical isolates of Candida tropicalis in China","volume":"68","author":"Jiang","year":"2013","journal-title":"J. Antimicrob. Chemother."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"105799","DOI":"10.1016\/j.ijantimicag.2019.09.003","article-title":"Analysis of global antifungal surveillance results reveals predominance of Erg11 Y132F alteration among azole-resistant Candida parapsilosis and Candida tropicalis and country-specific isolate dissemination","volume":"55","author":"Castanheira","year":"2020","journal-title":"Int. J. Antimicrob. Agents"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"1162","DOI":"10.1128\/AAC.05502-11","article-title":"Identification of a Cryptococcus neoformans cytochrome P450 lanosterol 14alpha-demethylase (Erg11) residue critical for differential susceptibility between fluconazole\/voriconazole and itraconazole\/posaconazole","volume":"56","author":"Sionov","year":"2012","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"3244","DOI":"10.1093\/jac\/dku316","article-title":"Aspergillus fumigatus harbouring the sole Y121F mutation shows decreased susceptibility to voriconazole but maintained susceptibility to itraconazole and posaconazole","volume":"69","author":"Lescar","year":"2014","journal-title":"J. Antimicrob. Chemother."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"669","DOI":"10.1007\/s10658-017-1216-7","article-title":"Resistance of wheat pathogen Zymoseptoria tritici to DMI and QoI fungicides in the Nordic-Baltic region\u2014A status","volume":"149","author":"Heick","year":"2017","journal-title":"Eur. J. Plant Pathol."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"15898","DOI":"10.1038\/s41598-017-16123-9","article-title":"Intrinsic short-tailed azole resistance in mucormycetes is due to an evolutionary conserved aminoacid substitution of the lanosterol 14\u03b1-demethylase","volume":"7","author":"Caramalho","year":"2017","journal-title":"Sci. Rep."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"3455","DOI":"10.1016\/j.bmcl.2014.05.068","article-title":"Design and optimization of highly-selective fungal CYP51 inhibitors","volume":"24","author":"Hoekstra","year":"2014","journal-title":"Bioorg. Med. Chem. Lett."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"e02114","DOI":"10.1128\/AAC.02114-18","article-title":"Azole Resistance Reduces Susceptibility to the Tetrazole Antifungal VT-1161","volume":"63","author":"Monk","year":"2019","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"6728","DOI":"10.1074\/jbc.M117.778308","article-title":"Structural analyses of Candida albicans sterol 14alpha-demethylase complexed with azole drugs address the molecular basis of azole-mediated inhibition of fungal sterol biosynthesis","volume":"292","author":"Hargrove","year":"2017","journal-title":"J. Biol. Chem."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"e00570-17","DOI":"10.1128\/AAC.00570-17","article-title":"Crystal Structure of the New Investigational Drug Candidate VT-1598 in Complex with Aspergillus fumigatus Sterol 14alpha-Demethylase Provides Insights into Its Broad-Spectrum Antifungal Activity","volume":"61","author":"Hargrove","year":"2017","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"e01131-18","DOI":"10.1128\/AAC.01131-18","article-title":"Heterologous Expression of Full-Length Lanosterol 14alpha-Demethylases of Prominent Fungal Pathogens Candida albicans and Candida glabrata Provides Tools for Antifungal Discovery","volume":"62","author":"Keniya","year":"2018","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"4982","DOI":"10.1128\/AAC.00925-15","article-title":"Structural Insights into Binding of the Antifungal Drug Fluconazole to Saccharomyces cerevisiae Lanosterol 14alpha-Demethylase","volume":"59","author":"Sagatova","year":"2015","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"e02242-17","DOI":"10.1128\/AAC.02242-17","article-title":"Impact of Homologous Resistance Mutations from Pathogenic Yeast on Saccharomyces cerevisiae Lanosterol 14alpha-Demethylase","volume":"62","author":"Sagatova","year":"2018","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"1150","DOI":"10.1128\/EC.00091-07","article-title":"Characterization of three classes of membrane proteins involved in fungal azole resistance by functional hyperexpression in Saccharomyces cerevisiae","volume":"6","author":"Lamping","year":"2007","journal-title":"Eukaryot. Cell"},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"99","DOI":"10.1002\/(SICI)1526-968X(200002)26:2<99::AID-GENE1>3.0.CO;2-B","article-title":"Cre recombinase: The universal reagent for genome tailoring","volume":"26","author":"Nagy","year":"2000","journal-title":"Genesis"},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"2519","DOI":"10.1093\/nar\/24.13.2519","article-title":"A new efficient gene disruption cassette for repeated use in budding yeast","volume":"24","author":"Guldener","year":"1996","journal-title":"Nucleic Acids Res."},{"key":"ref_49","doi-asserted-by":"crossref","unstructured":"Keniya, M.V., Fleischer, E., Klinger, A., Cannon, R.D., and Monk, B.C. (2015). Inhibitors of the Candida albicans major facilitator superfamily transporter Mdr1p responsible for fluconazole resistance. PLoS ONE, 10.","DOI":"10.1371\/journal.pone.0126350"},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"26213","DOI":"10.1038\/srep26213","article-title":"Triazole resistance mediated by mutations of a conserved active site tyrosine in fungal lanosterol 14alpha-demethylase","volume":"6","author":"Sagatova","year":"2016","journal-title":"Sci. Rep."},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"3865","DOI":"10.1073\/pnas.1324245111","article-title":"Architecture of a single membrane spanning cytochrome P450 suggests constraints that orient the catalytic domain relative to a bilayer","volume":"111","author":"Monk","year":"2014","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"265","DOI":"10.1016\/S0021-9258(19)52451-6","article-title":"Protein measurement with the Folin phenol reagent","volume":"193","author":"Lowry","year":"1951","journal-title":"J. Biol. Chem."},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"680","DOI":"10.1038\/227680a0","article-title":"Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4","volume":"227","author":"Laemmli","year":"1970","journal-title":"Nature"},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"e01134-18","DOI":"10.1128\/AAC.01134-18","article-title":"Crystal Structures of Full-Length Lanosterol 14alpha-Demethylases of Prominent Fungal Pathogens Candida albicans and Candida glabrata Provide Tools for Antifungal Discovery","volume":"62","author":"Keniya","year":"2018","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"26838","DOI":"10.1074\/jbc.M111.237099","article-title":"Substrate preferences and catalytic parameters determined by structural characteristics of sterol 14alpha-demethylase (CYP51) from Leishmania infantum","volume":"286","author":"Hargrove","year":"2011","journal-title":"J. Biol. Chem."},{"key":"ref_56","doi-asserted-by":"crossref","first-page":"31","DOI":"10.1016\/S0167-4838(99)00156-9","article-title":"Conformational dynamics and molecular interaction reactions of recombinant cytochrome P450scc (CYP11A1) detected by fluorescence energy transfer","volume":"1434","author":"Lepesheva","year":"1999","journal-title":"Biochim. Biophys. Acta"},{"key":"ref_57","doi-asserted-by":"crossref","first-page":"4530","DOI":"10.1128\/AAC.00349-16","article-title":"The Investigational Drug VT-1129 Is a Highly Potent Inhibitor of Cryptococcus Species CYP51 but Only Weakly Inhibits the Human Enzyme","volume":"60","author":"Warrilow","year":"2016","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_58","doi-asserted-by":"crossref","first-page":"125","DOI":"10.1107\/S0907444909047337","article-title":"Xds","volume":"66","author":"Kabsch","year":"2010","journal-title":"Acta Crystallogr. D Biol. Crystallogr."},{"key":"ref_59","doi-asserted-by":"crossref","first-page":"760","DOI":"10.1107\/S0907444994003112","article-title":"The CCP4 suite: Programs for protein crystallography","volume":"50","year":"1994","journal-title":"Acta Crystallogr. D Biol. Crystallogr."},{"key":"ref_60","doi-asserted-by":"crossref","first-page":"658","DOI":"10.1107\/S0021889807021206","article-title":"Phaser crystallographic software","volume":"40","author":"McCoy","year":"2007","journal-title":"J. Appl. Crystallogr."},{"key":"ref_61","doi-asserted-by":"crossref","first-page":"213","DOI":"10.1107\/S0907444909052925","article-title":"PHENIX: A comprehensive Python-based system for macromolecular structure solution","volume":"66","author":"Adams","year":"2010","journal-title":"Acta Crystallogr. D Biol. Crystallogr."},{"key":"ref_62","doi-asserted-by":"crossref","first-page":"486","DOI":"10.1107\/S0907444910007493","article-title":"Features and development of Coot","volume":"66","author":"Emsley","year":"2010","journal-title":"Acta Crystallogr. D Biol. Crystallogr."},{"key":"ref_63","unstructured":"(2021, March 23). Available online: http:\/\/grade.globalphasing.org."},{"key":"ref_64","doi-asserted-by":"crossref","first-page":"779","DOI":"10.1006\/jmbi.1993.1626","article-title":"Comparative protein modelling by satisfaction of spatial restraints","volume":"234","author":"Sali","year":"1993","journal-title":"J. Mol. Biol."},{"key":"ref_65","doi-asserted-by":"crossref","first-page":"205","DOI":"10.1006\/jmbi.2000.4042","article-title":"T-Coffee: A novel method for fast and accurate multiple sequence alignment","volume":"302","author":"Notredame","year":"2000","journal-title":"J. Mol. Biol."},{"key":"ref_66","doi-asserted-by":"crossref","first-page":"269","DOI":"10.1016\/0005-2744(69)90420-3","article-title":"Kinetics of the reversible inhibition of enzyme-catalysed reactions by tight-binding inhibitors","volume":"185","author":"Morrison","year":"1969","journal-title":"Biochim. Biophys. Acta"},{"key":"ref_67","doi-asserted-by":"crossref","unstructured":"Tyndall, J.D., Sabherwal, M., Sagatova, A.A., Keniya, M.V., Negroni, J., Wilson, R.K., Woods, M.A., Tietjen, K., and Monk, B.C. (2016). Structural and Functional Elucidation of Yeast Lanosterol 14alpha-Demethylase in Complex with Agrochemical Antifungals. PLoS ONE, 11.","DOI":"10.1371\/journal.pone.0167485"},{"key":"ref_68","doi-asserted-by":"crossref","first-page":"140206","DOI":"10.1016\/j.bbapap.2019.02.008","article-title":"Fungal Lanosterol 14alpha-demethylase: A target for next-generation antifungal design","volume":"1868","author":"Monk","year":"2020","journal-title":"Biochim. Biophys. Acta Proteins Proteom."},{"key":"ref_69","doi-asserted-by":"crossref","unstructured":"Riley, J., Brand, S., Voice, M., Caballero, I., Calvo, D., and Read, K.D. (2015). Development of a Fluorescence-based Trypanosoma cruzi CYP51 Inhibition Assay for Effective Compound Triaging in Drug Discovery Programmes for Chagas Disease. PLoS Negl. Trop. Dis., 9.","DOI":"10.1371\/journal.pntd.0004014"}],"container-title":["Journal of Fungi"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2309-608X\/8\/1\/69\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,13]],"date-time":"2025-10-13T14:14:46Z","timestamp":1760364886000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2309-608X\/8\/1\/69"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2022,1,10]]},"references-count":69,"journal-issue":{"issue":"1","published-online":{"date-parts":[[2022,1]]}},"alternative-id":["jof8010069"],"URL":"https:\/\/doi.org\/10.3390\/jof8010069","relation":{},"ISSN":["2309-608X"],"issn-type":[{"value":"2309-608X","type":"electronic"}],"subject":[],"published":{"date-parts":[[2022,1,10]]}}}