Natural product synthesis by non-ribosomal peptide synthetases (NRPS) is greatly defined by the substrate selectivity of the adenylation (A) domains. Previous assays for specificity determination were mainly performed in vitro and were requiring protein purification. In this work, we developed - based on NRPS engineering - a novel in vivo assay suitable for high-throughput application named ASCR (A domain screening). Using the recently described XUT fusion sites, A domains and their upstream condensation domains were assembled as di-domains to characterized NRPS model system, which allowed detection of defined tripeptide products via mass spectrometry directly after cell culture extraction. We evaluated the assay by screening in total 54 A domains from five known and seven uncharacterized NRPS, covering a broad range organism taxonomy and GC content of the investigated NRPS-encoding genes. Additionally, we applied the assay to elucidate and confirm the structures of novel cyclic pentapeptides derived from three novel NRPS from Photorhabdus temperata K122.

The evolutionary emergence of complex multicellularity in bacteria and fungi, such as the ability for mycelial growth, is strongly associated with increased genomic carriage of biosynthetic machinery ...

Microbial Natural Products Meet Data Science. The Zdouc research lab at the University of Vienna investigates naturally occurring, specialized molecules called natural products. These small molecules ...

Scientific Reports - Development of a bioassay-guided genome mining approach for antifungal natural product discovery from pseudomonads

This work presents DeepSeMS, a transformer-based framework that translates biosynthetic gene clusters into candidate secondary metabolite structures, enabling scalable exploration of chemical diversit...

Abstract. The root microbiome is important for plant development. The impact of the root microbiome is the result of multiple complex interactions among mi

Automated detection of metallophore biosynthesis reveals that metal-chelating non-ribosomal peptides are widespread, chemically diverse, and deeply rooted in bacterial evolution.

Automated detection of metallophore biosynthesis reveals that metal-chelating non-ribosomal peptides are widespread, chemically diverse, and deeply rooted in bacterial evolution.

Abstract. Biocuration is essential to transform molecular sequence data into standardized, machine-readable resources. Such curated datasets enable compara

An improved mechanistic understanding of microbial interactions can guide targeted interventions or inform the rational design of microbial communities to optimize them for applications such as pathog...

The tree microbiome is essential for host health and pathogen suppression. Synthetic microbial communities (SynComs) are emerging as important tools to understand microbiome dynamics and engineer micr...

The tree microbiome is essential for host health and pathogen suppression. Synthetic microbial communities (SynComs) are emerging as important tools to understand microbiome dynamics and engineer micr...

Standard European Vector Architecture 4.0 Welcome to the CNB-hosted database and material repository of standard and modular plasmid vectors for (de/re) constructing complex bacterial phenotypes WHAT…

Background High-throughput sequencing technologies have greatly advanced our understanding of microbiomes, but resolving microbial communities at species and strain levels remains challenging. Results...

Polyunsaturated fatty acid (PUFA) synthase-like enzymes are best known for their role in membrane lipid biosynthesis in marine bacteria, but have also been repurposed for the assembly of specialized l...

TL;DR: I created a freely available template to facilitate setting up “modern” Python projects.

Microbial metabolic gene clusters encode the biosynthesis or catabolism of metabolites that facilitate ecological specialization, mediate microbiome interactions and constitute a major source of medic...

The N-acyl-cyclolysine system regulates the expression of co-localized operons encoding diverse secreted molecules in Bacteroidota.

Crop protection against fungal pathogens is essential to prevent crop losses and maintain food security. Current crop protection relies heavily on che…

An abundance of microbial genomes have been sequenced in the past two decades. For fundamental comparative genomic investigations, where the goal is to determine the major gain and loss events shaping...

The validation of inhibitors targeting PurF, a novel drug target for tuberculosis drug discovery, is described.

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But turning what they find into drugs isn’t so easy

Zymoseptoria tritici causes Septoria Leaf Blotch disease of wheat and has evolved to overcome most chemical and genetic control methods. As such, new tools are required for future disease control. We...

🚨 Preprint (update) alert 🚨 --> Prioritizing Metabolite Features with FERMO Very proud of Mitja Zdouc, PhD, Hannah Augustijn 😊👏, and all colleagues and collaborators involved: the FERMO preprint...

Abstract. Accurate and accessible phylogenetic analysis is essential for understanding microbial taxonomy and evolution, which are integral to microbiology

Diversity within lower taxonomic units in microbial communities is a key trait, giving rise to important ecological functions. In the rhizosphere, the…

Microbial secondary metabolites are compounds produced by bacteria and fungi that are not required for their replication and unconditional survival (1, 2). While they are thus not expected to be unive...

Covering: 2019 to 2024Secondary metabolites (SMs) are naturally occurring defense or signaling molecules that are also utilized as human and animal drugs, crop protection agents, and fine chemicals. C...

Abstract. Bacteroidota species are enriched in the plant microbiome and provide several beneficial functions for their host, including disease suppression.

Rapid advances in mass spectrometry (MS) data analysis have accelerated the identification of natural products from complex mixtures such as natural product extracts. However, limitations in MS data in metabolite libraries and dereplication strategies are still lacking for assigning structures to known compounds and searching for unidentified compounds. To overcome these limitations, we present an approach that combines molecular networking with MS database-derived mass defect analysis to preferentially discover new compounds with high structural novelty in the initial stage of a discovery workflow. Specifically, unknown metabolites or clusters generated from molecular networking are assigned to a compound class based on their relative mass defects (RMDs) calculated using open-source databases. If ancillary data such as ultraviolet and MS/MS spectra of the unknown clusters are incongruent with the RMD-assigned compound class, metabolites are considered to have a new skeleton that exhibits a large difference in RMD value due to structural changes. Here, we applied this RMD-assisted method to a desert-derived bacterial strain library and validated it through the discovery of brasiliencin A (1), a new 18-membered macrolide from Nocardia brasiliensis. A putative biosynthetic pathway of brasiliencin A was proposed through whole-genome sequence analysis, and an additional 29 analogs were detected using absolute mass defect filtering (AMDF) based on plausible biosynthetic products. This led to the isolation of three additional macrolides, brasiliencins B–D (2–4). The structures of the brasiliencins (1–4) were fully elucidated through spectroscopic data analysis and quantum chemical calculations including ROE distance and 13C NMR chemical shift calculations, and experimental and theoretical electronic circular dichroism (ECD). Brasiliencin A showed strong activity against Mycobacterium smegmatis and Streptococcus australis (MIC = 31.3 nM and 7.81 μM, respectively) compared to brasiliencin B (MIC = 1000 nM and 62.5 μM, respectively) that differs at a single stereocenter.