Structural and Functional Elucidation of Yeast Lanosterol 14α-Demethylase in Complex with Agrochemical Antifungals
| Autor: | Matthew A. Woods, Klaus Tietjen, Mikhail V. Keniya, Jacopo Negroni, Brian C. Monk, Manya Sabherwal, Joel D. A. Tyndall, Alia A. Sagatova, Rajni K. Wilson |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2016 |
| Předmět: |
Azoles
0301 basic medicine Antifungal Agents Molecular Conformation lcsh:Medicine Yeast and Fungal Models Plant Science Pathology and Laboratory Medicine Biochemistry Substrate Specificity Isomers Sterol 14-Demethylase chemistry.chemical_compound Heterocyclic Compounds Stereochemistry Catalytic Domain Yeasts Medicine and Health Sciences Stereoisomers Post-Translational Modification Candida albicans lcsh:Science Fungal Pathogens chemistry.chemical_classification Ergosterol Crystallography Multidisciplinary Molecular Structure biology Organic Compounds Antimicrobials Physics Lanosterol Drugs Condensed Matter Physics Chemistry 14-alpha Demethylase Inhibitors Medical Microbiology Physical Sciences Crystal Structure Saccharomyces Cerevisiae Pathogens Agrochemicals Protein Binding Research Article 030106 microbiology Plant Pathogens Microbial Sensitivity Tests Heme Mycology Research and Analysis Methods Microbiology Structure-Activity Relationship Saccharomyces 03 medical and health sciences Model Organisms Isomerism Microbial Control Solid State Physics Binding site Microbial Pathogens Pharmacology Antifungals Binding Sites Candida glabrata Organic Chemistry lcsh:R Chemical Compounds Organisms Fungi Biology and Life Sciences Proteins Plant Pathology biology.organism_classification Yeast 030104 developmental biology Enantiomers chemistry biology.protein Demethylase Azole lcsh:Q |
| Zdroj: | PLoS ONE, Vol 11, Iss 12, p e0167485 (2016) PLoS ONE |
| ISSN: | 1932-6203 |
| Popis: | Azole antifungals, known as demethylase inhibitors (DMIs), target sterol 14α-demethylase (CYP51) in the ergosterol biosynthetic pathway of fungal pathogens of both plants and humans. DMIs remain the treatment of choice in crop protection against a wide range of fungal phytopathogens that have the potential to reduce crop yields and threaten food security. We used a yeast membrane protein expression system to overexpress recombinant hexahistidine-tagged S. cerevisiae lanosterol 14α-demethylase and the Y140F or Y140H mutants of this enzyme as surrogates in order characterize interactions with DMIs. The whole-cell antifungal activity (MIC50 values) of both the R- and S-enantiomers of tebuconazole, prothioconazole (PTZ), prothioconazole-desthio, and oxo-prothioconazole (oxo-PTZ) as well as for fluquinconazole, prochloraz and a racemic mixture of difenoconazole were determined. In vitro binding studies with the affinity purified enzyme were used to show tight type II binding to the yeast enzyme for all compounds tested except PTZ and oxo-PTZ. High resolution X-ray crystal structures of ScErg11p6×His in complex with seven DMIs, including four enantiomers, reveal triazole-mediated coordination of all compounds and the specific orientation of compounds within the relatively hydrophobic binding site. Comparison with CYP51 structures from fungal pathogens including Candida albicans, Candida glabrata and Aspergillus fumigatus provides strong evidence for a highly conserved CYP51 structure including the drug binding site. The structures obtained using S. cerevisiae lanosterol 14α-demethylase in complex with these agrochemicals provide the basis for understanding the impact of mutations on azole susceptibility and a platform for the structure-directed design of the next-generation of DMIs. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|