Isolation and molecular characterisation of the gene encoding eburicol 14α-demethylase (CYP51) from Penicillium italicum
article
The CYP51 gene encoding eburicol 14α-demethylase (P450(14DM)) was cloned from a genomic library of the filamentous fungal plant pathogen Penicillium italicum, by heterologous hybridisation with the corresponding gene encoding lanosterol 14α-demethylase from the yeast Candida tropicalis. The nucleotide sequence of a 1739-bp genomic fragment and the corresponding cDNA clone comprises an open reading frame (ORE) of 1545 bp, encoding a protein of 515 amino acids with a predicted molecular mass of 57.3 kDa. The ORF is interrupted by three introns of 60, 72 and 62 bp. The C-terminal part of the protein includes a characteristic haem-binding domain, HR2, common to all P450 genes. The deduced P. italicum P450(14DM) protein and the P450(14DM) proteins from Candida albicans, C. tropicalis and Saccharomyces cerevisiae share 47.2, 47.0 and 45.8% amino acid sequence identity. Therefore, the cloned gene is classified as a member of the CYP51 family. Multiple copies of a genomic DNA fragment of P. italicum containing the cloned P450 gene were introduced into Aspergillus niger by transformation. Transformants were significantly less sensitive to fungicides which inhibit P450(14DM) activity, indicating that the cloned gene encodes a functional eburicol 14α-demethylase.
Topics
14α-demethylaseCytochrome P450FungicidesPenicillium italicumSterol biosynthesis inhibitorsCytochrome p450 isoenzymeEnilconazoleEtaconazoleFenarimolFungicideUnclassified drugArticleControlled studyGene isolationGenetic analysisMolecular cloningMolecular geneticsNonhumanPenicilliumPriority journalSterol synthesisYeastAmino Acid SequenceBase SequenceBlotting, NorthernCloning, MolecularCytochrome P-450 Enzyme SystemDNA PrimersFungal ProteinsFungicides, IndustrialGenes, FungalLanosterolMolecular Sequence DataOxidoreductasesPenicilliumPyrimidinesRestriction MappingSequence Analysis, DNASequence Homology, Amino Acid
TNO Identifier
233310
ISSN
00268925
Source
Molecular and General Genetics, 250(6), pp. 725-733.
Pages
725-733
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