About: Lanosterol is a(n) research topic. Over the lifetime, 1239 publication(s) have been published within this topic receiving 36737 citation(s). The topic is also known as: (3β)-lanosta-8,24-dien-3-ol & (3β,20R)-lanosta-8,24-dien-3-ol.
Papers published on a yearly basis
TL;DR: Mapping mutations identified in Candida albicans azole-resistant isolates indicates that azole resistance in fungi develops in protein regions involved in orchestrating passage of CYP51 through different conformational stages along the catalytic cycle rather than in residues directly contacting fluconazole.
Abstract: Cytochrome P450 14α-sterol demethylases (CYP51) are essential enzymes in sterol biosynthesis in eukaryotes. CYP51 removes the 14α-methyl group from sterol precursors such as lanosterol, obtusifoliol, dihydrolanosterol, and 24(28)-methylene-24,25-dihydrolanosterol. Inhibitors of CYP51 include triazole antifungal agents fluconazole and itraconazole, drugs used in treatment of topical and systemic mycoses. The 2.1- and 2.2-Å crystal structures reported here for 4-phenylimidazole- and fluconazole-bound CYP51 from Mycobacterium tuberculosis (MTCYP51) are the first structures of an authentic P450 drug target. MTCYP51 exhibits the P450 fold with the exception of two striking differences—a bent I helix and an open conformation of BC loop—that define an active site-access channel running along the heme plane perpendicular to the direction observed for the substrate entry in P450BM3. Although a channel analogous to that in P450BM3 is evident also in MTCYP51, it is not open at the surface. The presence of two different channels, with one being open to the surface, suggests the possibility of conformationally regulated substrate-in/product-out openings in CYP51. Mapping mutations identified in Candida albicans azole-resistant isolates indicates that azole resistance in fungi develops in protein regions involved in orchestrating passage of CYP51 through different conformational stages along the catalytic cycle rather than in residues directly contacting fluconazole. These new structures provide a basis for rational design of new, more efficacious antifungal agents as well as insight into the molecular mechanism of P450 catalysis.
TL;DR: These studies showed that erg11/erg11 mutants of a C. albicans strain harboring a defective erg11 allele can be obtained in vitro in the presence of amphotericin B, and could therefore be selected by similar mechanisms during antifungal therapy.
Abstract: The role of sterol mutations in the resistance of Candida albicans to antifungal agents has not been thoroughly investigated. Previous work reported that clinical C. albicans strains resistant to both azole antifungals and amphotericin B were defective in ERG3, a gene encoding sterol Δ5,6-desaturase. It is also believed that a deletion of the lanosterol 14α-demethylase gene, ERG11, is possible only under aerobic conditions when ERG3 is not functional. We tested these hypotheses by creating mutants by targeted deletion of the ERG3 and ERG11 genes and subjecting those mutants to antifungal susceptibility testing and sterol analysis. The homozygous erg3/erg3 mutant created, DSY1751, was resistant to azole derivatives, as expected. This mutant was, however, slightly more susceptible to amphotericin B than the parent wild type. It was possible to generate erg11/erg11 mutants in the DSY1751 background but also, surprisingly, in the background of a wild-type isolate with functional ERG3 alleles under aerobic conditions. This mutant (DSY1769) was obtained by exposure of an ERG11/erg11 heterozygous strain in a medium containing 10 μg of amphotericin B per ml. Amphotericin B-resistant strains were obtained only from ERG11/erg11 heterozygotes at a frequency of approximately 5 × 10−5 to 7 × 10−5, which was consistent with mitotic recombination between the first disrupted erg11 allele and the other remaining functional ERG11 allele. DSY1769 was also resistant to azole derivatives. The main sterol fraction in DSY1769 contained lanosterol and eburicol. These studies showed that erg11/erg11 mutants of a C. albicans strain harboring a defective erg11 allele can be obtained in vitro in the presence of amphotericin B. Amphotericin B-resistant strains could therefore be selected by similar mechanisms during antifungal therapy.
TL;DR: Biochemical analysis demonstrates that Erg16p became less susceptible to fluconazole in isolate 13 in a series of isolates from an HIV-infected patient, demonstrating azole resistance in the pathogenic yeast Candida albicans.
Abstract: Azole resistance in the pathogenic yeast Candida albicans is an emerging problem in the human immunodeficiency virus (HIV)-infected population. The target enzyme of the azole drugs is lanosterol 14alpha demethylase (Erg16p), a cytochrome P-450 enzyme in the biosynthetic pathway of ergosterol. Biochemical analysis demonstrates that Erg16p became less susceptible to fluconazole in isolate 13 in a series of isolates from an HIV-infected patient. PCR-single-strand conformation polymorphism (PCR-SSCP) analysis was used to scan for genomic alterations of ERG16 in the isolates that would cause this change in the enzyme in isolate 13. Alterations near the 3' end of the gene that were identified by PCR-SSCP were confirmed by DNA sequencing. A single amino acid substitution (R467K) that occurred in isolate 13 was identified in both alleles of ERG16. Allelic differences within the ERG16 gene, in the ERG16 promoter, and in the downstream THR1 gene were eliminated in isolate 13. The loss of allelic variation in this region of the genome is most likely the result of mitotic recombination or gene conversion. The R467K mutation and loss of allelic variation that occur in isolate 13 are likely responsible for the azole-resistant enzyme activity seen in this and subsequent isolates. The description of R467K represents the first point mutation to be identified within ERG16 of a clinical isolate of C. albicans that alters the fluconazole sensitivity of the enzyme.
TL;DR: It is demonstrated that host-induced gene silencing (HIGS) targeting the fungal sterol 14α-demethylase (CYP51) genes restricts Fusarium infection in plants, demonstrating that HIGS is a powerful tool, which could revolutionize crop plant protection.
Abstract: Head blight, which is caused by mycotoxin-producing fungi of the genus Fusarium, is an economically important crop disease. We assessed the potential of host-induced gene silencing targeting the fungal cytochrome P450 lanosterol C-14α-demethylase (CYP51) genes, which are essential for ergosterol biosynthesis, to restrict fungal infection. In axenic cultures of Fusarium graminearum, in vitro feeding of CYP3RNA, a 791-nt double-stranded (ds)RNA complementary to CYP51A, CYP51B, and CYP51C, resulted in growth inhibition [half-maximum growth inhibition (IC50) = 1.2 nM] as well as altered fungal morphology, similar to that observed after treatment with the azole fungicide tebuconazole, for which the CYP51 enzyme is a target. Expression of the same dsRNA in Arabidopsis and barley rendered susceptible plants highly resistant to fungal infection. Microscopic analysis revealed that mycelium formation on CYP3RNA-expressing leaves was restricted to the inoculation sites, and that inoculated barley caryopses were virtually free of fungal hyphae. This inhibition of fungal growth correlated with in planta production of siRNAs corresponding to the targeted CYP51 sequences, as well as highly efficient silencing of the fungal CYP51 genes. The high efficiency of fungal inhibition suggests that host-induced gene-silencing targeting of the CYP51 genes is an alternative to chemical treatments for the control of devastating fungal diseases.
TL;DR: The target protein with an inhibitor that showed cholesterol lowering in vivo opens the way for the structure-based design of new OSC inhibitors, and the complex with the reaction product lanosterol gives a clear picture of the way in which the enzyme achieves product specificity in this highly exothermic cyclization reaction.
Abstract: In higher organisms the formation of the steroid scaffold is catalysed exclusively by the membrane-bound oxidosqualene cyclase (OSC; lanosterol synthase). In a highly selective cyclization reaction OSC forms lanosterol with seven chiral centres starting from the linear substrate 2,3-oxidosqualene. Valuable data on the mechanism of the complex cyclization cascade have been collected during the past 50 years using suicide inhibitors, mutagenesis studies and homology modelling. Nevertheless it is still not fully understood how the enzyme catalyses the reaction. Because of the decisive role of OSC in cholesterol biosynthesis it represents a target for the discovery of novel anticholesteraemic drugs that could complement the widely used statins. Here we present two crystal structures of the human membrane protein OSC: the target protein with an inhibitor that showed cholesterol lowering in vivo opens the way for the structure-based design of new OSC inhibitors. The complex with the reaction product lanosterol gives a clear picture of the way in which the enzyme achieves product specificity in this highly exothermic cyclization reaction.
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