McMillan, LKCarr, RLYoung, CAAstin, JWLowe, RGTParker, EJJameson, GBFinch, SCMiles, COMcManus, OBSchmalhofer, WAGarcia, MLKaczorowski, GJGoetz, MTkacz, JSScott, B2025-01-082025-01-082003-10(2003). Molecular Genetics and Genomics, 270(1), 9-23.1617-4615https://hdl.handle.net/2292/70985The gene cluster required for paxilline biosynthesis in Penicillium paxilli contains two cytochrome P450 monooxygenase genes, paxP and paxQ. The primary sequences of both proteins are very similar to those of proposed cytochrome P450 monooxygenases from other filamentous fungi, and contain several conserved motifs, including that for a haem-binding site. Alignment of these sequences with mammalian and bacterial P450 enzymes of known 3-D structure predicts that there is also considerable conservation at the level of secondary structure. Deletion of paxP and paxQ results in mutant strains that accumulate paspaline and 13-desoxypaxilline, respectively. These results confirm that paxP and paxQ are essential for paxilline biosynthesis and that paspaline and 13-desoxypaxilline are the most likely substrates for the corresponding enzymes. Chemical complementation of paxilline biosynthesis in paxG (geranygeranyl diphosphate synthase) and paxP, but not paxQ, mutants by the external addition of 13-desoxypaxilline confirms that PaxG and PaxP precede PaxQ, and are functionally part of the same biosynthetic pathway. A pathway for the biosynthesis of paxilline is proposed on the basis of these and earlier results. Electrophysiological experiments demonstrated that 13-desoxypaxilline is a weak inhibitor of mammalian maxi-K channels (Ki=730 nM) compared to paxilline (Ki=30 nM), indicating that the C-13 OH group of paxilline is crucial for the biological activity of this tremorgenic mycotoxin. Paspaline is essentially inactive as a channel blocker, causing only slight inhibition at concentrations up to 1 microM.Print-ElectronicItems in ResearchSpace are protected by copyright, with all rights reserved, unless otherwise indicated. Previously published items are made available in accordance with the copyright policy of the publisher.https://researchspace.auckland.ac.nz/docs/uoa-docs/rights.htmAnimalsMammalsPenicilliumIndolesCytochrome P-450 Enzyme SystemPotassium Channels, Calcium-ActivatedRecombinant ProteinsDNA, ComplementaryRestriction MappingGenetic Complementation TestReverse Transcriptase Polymerase Chain ReactionSequence AlignmentMutagenesisGene DeletionAmino Acid SequenceConserved SequenceSequence Homology, Amino AcidGenes, BacterialMultigene FamilyMolecular Sequence DataLarge-Conductance Calcium-Activated Potassium Channels3101 Biochemistry and Cell Biology3102 Bioinformatics and Computational Biology31 Biological SciencesGeneticsBiotechnologyScience & TechnologyLife Sciences & BiomedicineBiochemistry & Molecular BiologyGenetics & HeredityPenicillium paxilliindole-diterpenespaxillinecytochrome P450monooxygenasesmaxi-K channelsPROTEIN SECONDARY STRUCTURELANOSTEROL 14-ALPHA-METHYL DEMETHYLASEGIBBERELLA-FUJIKUROI ENCODESACTIVATED POTASSIUM CHANNELSAMINO-ACIDALBOPHOMA-YAMANASHIENSISINDOLE-DITERPENOIDSNODULISPORIC ACIDLOLIUM-PERENNEBETA-SUBUNIT0601 Biochemistry and Cell Biology0604 GeneticsBiomedicalBasic ScienceGeneric Health Relevance0607 Plant Biology3105 Genetics3108 Plant biologyJournal Article10.1007/s00438-003-0887-2Copyright: Springer Nature12884010 (pubmed)1617-4623