title: Genetic basis for differential activities of fluconazole and voriconazole against Candida krusei.
abstract: Invasive infections caused by Candida krusei are a significant concern because this organism is intrinsically resistant to fluconazole. Voriconazole is more active than fluconazole against C. krusei in vitro. One mechanism of fluconazole resistance in C. krusei is diminished sensitivity of the target enzyme, cytochrome P450 sterol 14alpha-demethylase (CYP51), to inhibition by this drug. We investigated the interactions of fluconazole and voriconazole with the CYP51s of C. krusei (ckCYP51) and fluconazole-susceptible Candida albicans (caCYP51). We found that voriconazole was a more potent inhibitor of both ckCYP51 and caCYP51 in cell extracts than was fluconazole. Also, the ckCYP51 was less sensitive to inhibition by both drugs than was caCYP51. These results were confirmed by expressing the CYP51 genes from C. krusei and C. albicans in Saccharomyces cerevisiae and determining the susceptibility of the transformants to voriconazole and fluconazole. We constructed homology models of the CYP51s of C. albicans and C. krusei based on the crystal structure of CYP51 from Mycobacterium tuberculosis. These models predicted that voriconazole is a more potent inhibitor of both caCYP51 and ckCYP51 than is fluconazole, because the extra methyl group of voriconazole results in a stronger hydrophobic interaction with the aromatic amino acids in the substrate binding site and more extensive filling of this site. Although there are multiple differences in the predicted amino acid sequence of caCYP51 and ckCYP51, the models of the two enzymes were quite similar and the mechanism for the relative resistance of ckCYP51 to the azoles was not apparent.
authors.affiliation: Division of Infectious Diseases, Department of Medicine, Harbor-UCLA Research and Education Institute, Torrance, California 90502, USA.
authors.forename: Takashi
authors.initials: T
authors.lastname: Fukuoka
authors.forename: Douglas A
authors.initials: DA
authors.lastname: Johnston
authors.forename: Carol A
authors.initials: CA
authors.lastname: Winslow
authors.forename: Marcel J
authors.initials: MJ
authors.lastname: de Groot
authors.forename: Catherine
authors.initials: C
authors.lastname: Burt
authors.forename: Christopher A
authors.initials: CA
authors.lastname: Hitchcock
authors.forename: Scott G
authors.initials: SG
authors.lastname: Filler
mesh_terms: D000595:Amino Acid Sequence; D000935:Antifungal Agents / Q000494:pharmacology*; D002175:Candida / Q000187:drug effects* / Q000201:enzymology / Q000235:genetics; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry / Q000235:genetics; D025141:Drug Resistance, Fungal; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D015725:Fluconazole / Q000494:pharmacology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010088:Oxidoreductases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics; D017433:Protein Structure, Secondary; D011743:Pyrimidines / Q000494:pharmacology*; D058886:Sterol 14-Demethylase; D014230:Triazoles / Q000494:pharmacology*; D065819:Voriconazole
chemical_list: D000935:Antifungal Agents; D065607:Cytochrome P-450 Enzyme Inhibitors; D004791:Enzyme Inhibitors; D011743:Pyrimidines; D014230:Triazoles; D015725:Fluconazole; D003577:Cytochrome P-450 Enzyme System; D010088:Oxidoreductases; D058886:Sterol 14-Demethylase; D065819:Voriconazole