| Autor: |
Warrilow AG; Centre for Cytochrome P450 Biodiversity, Institute of Life Science, Swansea University Medical School, Swansea, Wales SA2 8PP, United Kingdom., Price CL; Centre for Cytochrome P450 Biodiversity, Institute of Life Science, Swansea University Medical School, Swansea, Wales SA2 8PP, United Kingdom., Parker JE; Centre for Cytochrome P450 Biodiversity, Institute of Life Science, Swansea University Medical School, Swansea, Wales SA2 8PP, United Kingdom., Rolley NJ; Centre for Cytochrome P450 Biodiversity, Institute of Life Science, Swansea University Medical School, Swansea, Wales SA2 8PP, United Kingdom., Smyrniotis CJ; Chemistry and Biochemistry Department, University of California, Santa Cruz, CA 95064 USA., Hughes DD; Plant Chemistry Group, School of Chemistry, Bangor University, Bangor, Gwynedd, Wales, LL57 2UW, United Kingdom., Thoss V; Plant Chemistry Group, School of Chemistry, Bangor University, Bangor, Gwynedd, Wales, LL57 2UW, United Kingdom., Nes WD; Center for Chemical Biology, Department of Chemistry and Biochemistry, Texas Tech University, Lubbock, Texas 79409-1061., Kelly DE; Centre for Cytochrome P450 Biodiversity, Institute of Life Science, Swansea University Medical School, Swansea, Wales SA2 8PP, United Kingdom., Holman TR; Chemistry and Biochemistry Department, University of California, Santa Cruz, CA 95064 USA., Kelly SL; Centre for Cytochrome P450 Biodiversity, Institute of Life Science, Swansea University Medical School, Swansea, Wales SA2 8PP, United Kingdom. |
| Abstrakt: |
Malassezia globosa cytochromes P450 CYP51 and CYP5218 are sterol 14α-demethylase (the target of azole antifungals) and a putative fatty acid metabolism protein (and a potential azole drug target), respectively. Lanosterol, eburicol and obtusifoliol bound to CYP51 with Kd values of 32, 23 and 28 μM, respectively, catalyzing sterol 14α-demethylation with respective turnover numbers of 1.7 min(-1), 5.6 min(-1) and 3.4 min(-1). CYP5218 bound a range of fatty acids with linoleic acid binding strongest (Kd 36 μM), although no metabolism could be detected in reconstitution assays or role in growth on lipids. Clotrimazole, fluconazole, itraconazole, ketoconazole, voriconazole and ketaminazole bound tightly to CYP51 (Kd ≤ 2 to 11 nM). In contrast, fluconazole did not bind to CYP5218, voriconazole and ketaminazole bound weakly (Kd ~107 and ~12 μM), whereas ketoconazole, clotrimazole and itraconazole bound strongest to CYP5218 (Kd ~1.6, 0.5 and 0.4 μM) indicating CYP5218 to be only a secondary target of azole antifungals. IC50 determinations confirmed M. globosa CYP51 was strongly inhibited by azole antifungals (0.15 to 0.35 μM). MIC100 studies showed itraconazole should be considered as an alternative to ketoconazole given the potency and safety profiles and the CYP51 assay system can be used in structure-activity studies in drug development. |
