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Structure-Functional Characterization of Cytochrome P450 Sterol 14α-Demethylase (CYP51B) fromAspergillus fumigatusand Molecular Basis for the Development of Antifungal Drugs / Tatiana Y. Hargrove; Zdzislaw Wawrzak; David Lamb; F. Peter Guengerich; Galina I. Lepesheva
Journal of Biological Chemistry, Volume: 290, Issue: 39, Pages: 23916 - 23934
Swansea University Author: David, Lamb
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DOI (Published version): 10.1074/jbc.M115.677310
Aspergillus fumigatus is the opportunistic fungal pathogen that predominantly affects the immunocompromised population and causes 600,000 deaths/year. The cytochrome P450 51 (CYP51) inhibitor voriconazole is currently the drug of choice, yet the treatment efficiency remains low, calling for rational...
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Aspergillus fumigatus is the opportunistic fungal pathogen that predominantly affects the immunocompromised population and causes 600,000 deaths/year. The cytochrome P450 51 (CYP51) inhibitor voriconazole is currently the drug of choice, yet the treatment efficiency remains low, calling for rational development of more efficient agents. A. fumigatus has two CYP51 genes, CYP51A and CYP51B, which share 59% amino acid sequence identity. CYP51B is expressed constitutively, whereas gene CYP51A is reported to be inducible. We expressed, purified, and characterized A. fumigatus CYP51B, including determination of its substrate preferences, catalytic parameters, inhibition, and x-ray structure in complexes with voriconazole and the experimental inhibitor (R)-N-(1-(2,4-dichlorophenyl)-2-(1H-imidazol-1-yl)ethyl)-4-(5-phenyl-1,3,4-oxadiazol-2-yl)benzamide (VNI). The enzyme demethylated its natural substrate eburicol and the plant CYP51 substrate obtusifoliol at steady-state rates of 17 and 16 min(-1), respectively, but did not metabolize lanosterol, and the topical antifungal drug miconazole was the strongest inhibitor that we identified. The x-ray crystal structures displayed high overall similarity of A. fumigatus CYP51B to CYP51 orthologs from other biological kingdoms but revealed phylum-specific differences relevant to enzyme catalysis and inhibition. The complex with voriconazole provides an explanation for the potency of this small molecule, whereas the complex with VNI outlines a direction for further enhancement of the efficiency of this new inhibitory scaffold to treat humans afflicted with filamentous fungal infections.
cytochrome P450, azole antifungal, crystal structure
Swansea University Medical School