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Characterisation of Candida parapsilosis CYP51 as a Drug Target Using Saccharomyces cerevisiae as Host

The fungal cytochrome P450 lanosterol 14α-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...

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Autores principales: Ruma, Yasmeen N., Keniya, Mikhail V., Tyndall, Joel D. A., Monk, Brian C.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8781857/
https://www.ncbi.nlm.nih.gov/pubmed/35050009
http://dx.doi.org/10.3390/jof8010069
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author Ruma, Yasmeen N.
Keniya, Mikhail V.
Tyndall, Joel D. A.
Monk, Brian C.
author_facet Ruma, Yasmeen N.
Keniya, Mikhail V.
Tyndall, Joel D. A.
Monk, Brian C.
author_sort Ruma, Yasmeen N.
collection PubMed
description The fungal cytochrome P450 lanosterol 14α-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 Å. Structural analysis of azole—enzyme 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.
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spelling pubmed-87818572022-01-22 Characterisation of Candida parapsilosis CYP51 as a Drug Target Using Saccharomyces cerevisiae as Host Ruma, Yasmeen N. Keniya, Mikhail V. Tyndall, Joel D. A. Monk, Brian C. J Fungi (Basel) Article The fungal cytochrome P450 lanosterol 14α-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 Å. Structural analysis of azole—enzyme 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. MDPI 2022-01-10 /pmc/articles/PMC8781857/ /pubmed/35050009 http://dx.doi.org/10.3390/jof8010069 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Ruma, Yasmeen N.
Keniya, Mikhail V.
Tyndall, Joel D. A.
Monk, Brian C.
Characterisation of Candida parapsilosis CYP51 as a Drug Target Using Saccharomyces cerevisiae as Host
title Characterisation of Candida parapsilosis CYP51 as a Drug Target Using Saccharomyces cerevisiae as Host
title_full Characterisation of Candida parapsilosis CYP51 as a Drug Target Using Saccharomyces cerevisiae as Host
title_fullStr Characterisation of Candida parapsilosis CYP51 as a Drug Target Using Saccharomyces cerevisiae as Host
title_full_unstemmed Characterisation of Candida parapsilosis CYP51 as a Drug Target Using Saccharomyces cerevisiae as Host
title_short Characterisation of Candida parapsilosis CYP51 as a Drug Target Using Saccharomyces cerevisiae as Host
title_sort characterisation of candida parapsilosis cyp51 as a drug target using saccharomyces cerevisiae as host
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8781857/
https://www.ncbi.nlm.nih.gov/pubmed/35050009
http://dx.doi.org/10.3390/jof8010069
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