RESUMEN
Entomopathogenic fungus Metarhizium majus contains the nine-gene PPZ cluster, with ppzA, encoding a peramine-producing nonribosomal peptide synthetase, as the central component. In this work, the roles of two α-ketoglutarate, iron-dependent oxygenases encoded by the PPZ genes ppzC and ppzD were elucidated. PpzD was found to produce both trans-4-hydroxy-l-proline and trans-3-hydroxy-l-proline in a 13.1:1 ratio, yielding a key precursor for peramine biosynthesis. PpzC was found to act directly on peramine, yielding the novel analogue 8-hydroxyperamine.
Asunto(s)
Compuestos Heterocíclicos con 2 Anillos , Hierro , Ácidos Cetoglutáricos , Metarhizium , Poliaminas , Familia de Multigenes , Compuestos FerrososRESUMEN
Pyrazomycin is a rare C-nucleoside antibiotic containing a naturally occurring pyrazole ring, the biosynthetic origin of which has remained obscure for decades. In this study we report the identification of the gene cluster responsible for pyrazomycin biosynthesis in Streptomyces candidus NRRLâ 3601, revealing that the StrR-family regulator PyrR is the cluster-situated transcriptional activator governing pyrazomycin biosynthesis. Furthermore, our results from in vivo reconstitution and stable-isotope feeding experiments provide support for the hypothesis that PyrN is a new nitrogen-nitrogen bond-forming enzyme that catalyzes the linkage of the ϵ-NH2 nitrogen atom of l-N6 -OH-lysine and the α-NH2 nitrogen atom of l-glutamic acid. This study lays the foundation for further genetic and biochemical characterization of pyrazomycin pathway enzymes involved in constructing the characteristic pyrazole ring.
Asunto(s)
Antibacterianos/biosíntesis , Ribosa/biosíntesis , Streptomyces/metabolismo , Amidas , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Familia de Multigenes , Pentosiltransferasa/genética , Pentosiltransferasa/metabolismo , Pirazoles , Proteínas Represoras/genética , Proteínas Represoras/metabolismo , Streptomyces/genéticaRESUMEN
Microorganisms produce a wide range of natural products (NPs) with clinically and agriculturally relevant biological activities. In bacteria and fungi, genes encoding successive steps in a biosynthetic pathway tend to be clustered on the chromosome as biosynthetic gene clusters (BGCs). Historically, "activity-guided" approaches to NP discovery have focused on bioactivity screening of NPs produced by culturable microbes. In contrast, recent "genome mining" approaches first identify candidate BGCs, express these biosynthetic genes using synthetic biology methods, and finally test for the production of NPs. Fungal genome mining efforts and the exploration of novel sequence and NP space are limited, however, by the lack of a comprehensive catalog of BGCs encoding experimentally-validated products. In this study, we generated a comprehensive reference set of fungal NPs whose biosynthetic gene clusters are described in the published literature. To generate this dataset, we first identified NCBI records that included both a peer-reviewed article and an associated nucleotide record. We filtered these records by text and homology criteria to identify putative NP-related articles and BGCs. Next, we manually curated the resulting articles, chemical structures, and protein sequences. The resulting catalog contains 197 unique NP compounds covering several major classes of fungal NPs, including polyketides, non-ribosomal peptides, terpenoids, and alkaloids. The distribution of articles published per compound shows a bias toward the study of certain popular compounds, such as the aflatoxins. Phylogenetic analysis of biosynthetic genes suggests that much chemical and enzymatic diversity remains to be discovered in fungi. Our catalog was incorporated into the recently launched Minimum Information about Biosynthetic Gene cluster (MIBiG) repository to create the largest known set of fungal BGCs and associated NPs, a resource that we anticipate will guide future genome mining and synthetic biology efforts toward discovering novel fungal enzymes and metabolites.
Asunto(s)
Productos Biológicos , Vías Biosintéticas/genética , Genes Fúngicos , Genoma Fúngico , Familia de Multigenes , Alcaloides , Secuencia de Aminoácidos , Biología Computacional , Curaduría de Datos , Hongos/genética , Filogenia , Policétidos , TerpenosRESUMEN
Pyridoxal 5'-phosphate (PLP) is an organic cofactor employed by ~ 4% of enzymes. The structure of the PLP cofactor allows for the stabilization of carbanions through resonance. A small number of PLP-dependent enzymes employ molecular oxygen as a cosubstrate. Here, we review the biological roles and possible mechanisms of these enzymes, and we observe that these enzymes are found in multiple protein families, suggesting that reaction with oxygen might have emerged de novo in several protein families and thus could be directed to emerge again through laboratory evolution experiments.