Interaction of azole antifungal antibiotics with cytochrome P-450-dependent 14α-sterol demethylase purified from Candida albicans

Abstract

The interaction of azole antifungal antibiotics with purified Candida albicans cytochrome P-450-dependent 14 alpha-sterol demethylase (P-450DM) was measured spectrophotometrically and by inhibition of enzyme activity. Ketoconazole and ICI 153066 (a triazole derivative) formed low-spin complexes with the ferric cytochrome and induced type II difference spectra. These spectra are indicative of an interaction between the azole moiety and the sixth co-ordination position of P-450DM haem. Both azoles inhibited the binding of CO to the sodium dithionite-reduced ferrous cytochrome, and inhibited reconstituted P-450DM activity by binding to the cytochrome with a one-to-one stoichiometry. Similarly, total inhibition of enzyme activity occurred when equimolar amounts of clotrimazole, miconazole or fluconazole were added to reconstituted P-450DM. These results correlated with the inhibition of P-450DM in broken cell preparations, confirming that all five azoles are potent inhibitors of ergosterol biosynthesis in C. albicans.

Molecular diversity of sterol 14α‐demethylase substrates in plants, fungi and humans

“…All other chemicals used were purchased from Sigma, Poole, UK. The 14K-demethylase activity of the plant, C. albicans and human enzymes consisting of CYP51 and its respective reductase was assayed according to previous methods [12,13]. Each sterol was dispersed in dilaurylphosphatidylcholine (DLPC) and added to the reaction mixture at varying concentrations.…”

First Step of Glycosylphosphatidylinositol (GPI) Biosynthesis Cross-talks with Ergosterol Biosynthesis and Ras Signaling in Candida albicans

“…To understand this, we first examined the response of GPI2 heterozygote to azoles, which act by specifically targeting Erg11 in the ergosterol biosynthetic pathway (19). We found it to be azole-resistant (Fig.…”

Transient Receptor Potential Channels in Sensory Neurons Are Targets of the Antimycotic Agent Clotrimazole

“…CLT exerts its antifungal actions by inhibiting P450-dependent enzymes (Hitchcock et al, 1990). Other well known molecular targets of CLT are intermediate conductance Ca 2ϩ -activated potassium (IK Ca ) channels, including the erythrocyte Gardos channel (KCNN4) (Alvarez et al, 1992); CLT-induced inhibition of the Gardos channel, leading to reduced erythrocyte dehydratation, is a promising therapy in sickle cell disease (Brugnara et al, 1996).…”