Lanosterol

About: Lanosterol is a research topic. Over the lifetime, 1239 publications have been published within this topic receiving 36737 citations. The topic is also known as: (3β)-lanosta-8,24-dien-3-ol & (3β,20R)-lanosta-8,24-dien-3-ol.

Structural and Functional Elucidation of Yeast Lanosterol 14α-Demethylase in Complex with Agrochemical Antifungals.

Abstract: 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.

Synthesis, specificity, and antifungal activity of inhibitors of the Candida albicans delta 24-sterol methyltransferase.

Abstract: A series of side chain modified analogues of cholesterol and lanosterol (1-10) have been synthesized and evaluated as inhibitors of the Candida albicans delta 24-sterol methyltransferase. Two sterol substrate analogues 1 and 2 which contained a 24-thia substituent were relatively modest inhibitors of the enzyme (Ki = 1.5-72 microM). Compounds which mimic the carbocation intermediates proposed for the methyltransferase reaction, including sulfonium salts 4-6, amidines 7 and 8, and imidazoles 9 and 10 were substantially more potent inhibitors (Ki = 5-500 nM). All of the sterol analogues examined displayed less than 10-fold selectivity for inhibition of the methyltransferase versus the rat liver delta 24-sterol reductase. The sterol analogues were tested for in vitro antifungal activity against C. albicans, Candida tropicalis, and Torulopsis glabrata. The minimum inhibitory concentrations versus C. albicans correlated well with the Ki values for methyltransferase inhibition, and the potency of several compounds approached that of amphotericin B, although only modest fungicidal activity was observed.

Inhibition by ketoconazole of mitogen-induced DNA synthesis and cholesterol biosynthesis in lymphocytes.

Abstract: The effects of ketoconazole on mitogen-induced DNA synthesis and cholesterol biosynthesis in human and murine lymphocytes have been examined. Ketoconazole concentrations which do not affect cell viability (0.1 to 10 micrograms/ml) in culture led to a dose-dependent inhibition of DNA synthesis, as measured by [3H]thymidine incorporation, induced by either T-cell or B-cell mitogens. At drug concentrations 5- to 10-fold lower, ketoconazole inhibited the incorporation of [14C]acetate into cholesterol, with a resultant accumulation of [14C]lanosterol. The suppressive effects of ketoconazole on DNA synthesis were reversed by increasing the concentration of human serum in the culture medium from 5 to 20%. The depletion of lipoproteins in human serum by density centrifugation reduced the cholesterol content by 90% but did not affect the ability of the serum to overcome the inhibition by ketoconazole of DNA synthesis. Unlike DNA synthesis, cholesterol biosynthesis was not restored by 20% fresh human serum or lipoprotein-depleted human serum. These results demonstrate that ketoconazole potently inhibits DNA synthesis and cholesterol synthesis in mitogen-stimulated lymphocytes at drug concentrations obtained therapeutically. Further, the uncoupling of endogenous cholesterol synthesis and DNA synthesis indicates at least two levels of action of ketoconazole in mammalian lymphocytes.

Mechanism of azole antifungal activity as determined by liquid chromatographic/mass spectrometric monitoring of ergosterol biosynthesis

Abstract: A liquid chromatography/mass spectrometry (LC/MS) method for separation and characterization of ergosterol biosynthetic precursors was developed to study the effect of Posaconazole on sterol biosynthesis in fungi. Ergosterol biosynthetic precursors were characterized from their electron ionization mass spectra acquired by a normal-phase chromatography, particle beam LC/MS method. Fragment ions resulting from cleavage across the D-ring and an abundant M - 15 fragment ion were diagnostic for methyl substitution at C-4 and C-14. Comparison of the sterol profile in control and treated Candida albicans incubations showed depletion of ergosterol and accumulation of C-4 and C-14 methyl-substituted sterols following treatment with Posaconazole. These C-4 and C-14 methyl sterols are known to be incapable of sustaining cell growth. The results demonstrate that Posaconazole exerts its antifungal activity by inhibition of ergosterol biosynthesis. Furthermore, Posaconazole appears to disrupt ergosterol biosynthesis by inhibition of lanosterol 14alpha-demethylase.