Exploring Biosynthetic Pathways Through Genome Mining: Case Studies of Two Penicillium Species
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Philipps-Universität Marburg
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Abstract
Nature, as a vast reservoir of resources closely connected to human life, not only provides the material foundation for daily living but also serves as a vital source for health protection. Among these resources are numerous natural products that have been developed into medicines. However, with the rise of drug resistance and the accelerating environmental changes, the demand for new drugs continues to grow. Microorganisms, as a key component of the natural world, offer promising solutions to this challenge. Having evolved a diverse array of adaptive strategies in response to various environmental pressures, microorganisms engage in processes such as communication and competition, often mediated by secondary metabolites. These molecules, in turn, represent a valuable resource for further drug discovery.
The research on the biosynthesis of natural products has laid a valuable foundation for the production and utilization of medicinal compounds. With the rapid advancements in sequencing and bioinformatics technologies, an enormous amount of microbial genomic data has been become available. This surge in data has significantly increased the number of predicted biosynthetic gene clusters, highlighting the vast potential of microbial systems for synthesizing secondary metabolites. However, many of these biosynthetic gene clusters remain unlinked to the secondary metabolites they produce, creating an urgent need for research focused on predicting and elucidating biosynthetic pathways through genome mining strategies.
In collaboration with Zheng-Xi Zhang, we identified two biosynthetic pathways in Penicillium palitans NRRL 2033 by employing a top-down genome mining strategy with the isolated compounds as the starting point. This led to the identification of a gene cluster, named vdo cluster, which shows notable similarity to the pen and asq clusters involved in the biosynthesis of viridicatin and 4'-methoxyviridicatin. Heterologous expression in Aspergillus nidulans LO8030 and feeding experiments confirmed that four genes, vdoABCD, within this newly identified cluster are responsible for viridicatol biosynthesis. In particular, the cytochrome P450 enzyme VdoD catalyzes a key reaction by converting cyclopenin to cyclopenol, shedding light on a previously uncharacterized meta-hydroxylation process on a monoalkylated benzene ring.
Similarly, another gene cluster, termed spe, was identified in the genome of P. palitans, displaying high similarity to the cpa clusters involved in cyclopiazonic acid (CPA) and speradine A biosynthesis. Disruption of the backbone gene speA in the native producer, coupled with the heterologous expression of speABCDE in A. nidulans LO8030, confirmed the role of these five genes in the biosynthesis of both CPA and speradines including speradine F. Additional heterologous expression and precursor feeding experiments revealed that the P450 enzyme SpeD catalyzes an oxidation reaction at C-2 of the indole ring, while the methyltransferase SpeE is responsible for N-1 methylation. Intriguingly, the conversion of speradine A to speradine F involves multiple spontaneous hydroxylations, leading to the formation of a cyclic ketal ring. This discovery highlights an example of less-studied multiple non-enzymatic hydroxylations and provides insight into the formation of similar chemical skeletons.
In a third project, in collaboration with Yu Dai and Dr. Jing Zhou, a bottom-up approach focused on targeted gene cluster mining was employed to investigate a gene cluster in Penicillium roqueforti FM164, which is responsible for the biosynthesis of alkyl salicylaldehyde derivatives. The products of this biosynthetic gene cluster were identified through heterologous expression in A. nidulans LO8030 combined with promoter replacement. These products were determined to be novel alkyl salicylaldehyde derivatives containing an ethanolamine moiety. Heterologous expression and feeding experiments revealed that their biosynthesis requires a cytochrome P450 enzyme and two flavin-containing monooxygenases, which operate in a coordinated sequence of oxidation steps both prior to and following ethanolamine incorporation. This finding enhances our understanding of the diverse modification potential in alkyl salicylaldehyde scaffolds. As the second gene cluster identified in P. roqueforti responsible for producing alkyl salicylaldehyde derivatives, these results further emphasize the role of gene cluster duplication in diversifying biosynthetic pathways and expanded the structural variety of secondary metabolites.
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Dates
Created: 2024Issued: 2025-08-06Updated: 2025-08-06
Faculty
Fachbereich Pharmazie
Publisher
Philipps-Universität Marburg
Language
eng
Data types
DoctoralThesis
Keywords
natürliche ProduktePenicilliumsecondary metabolitebiosynthesisfilamentöse Pilzegenome mining
DFG-subjects
SekundärmetaboliteGenome-MiningPharmakologieBiosynthese
DDC-Numbers
615
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Li, Zhang-Hai (0000-0001-9729-1972): Exploring Biosynthetic Pathways Through Genome Mining: Case Studies of Two Penicillium Species. : Philipps-Universität Marburg 2025-08-06. DOI: https://doi.org/10.17192/z2024.0506.