Biology subjects

van Ede, J. M.

Publications and source records attributed to van Ede, J. M..

4 recordsLinked to original sources

bioRxiv2026

A protocol for lab-scale production of 13C yeast extract as internal standard for metabolomics and quantification of intracellular metabolites

Metabolomics enables the profiling of small-molecule metabolites and thereby captures the biochemical state of a living organism at a given moment and enables to monitor its cellular responses to stimuli. This technique has become a powerful tool in pharmaceutical research, the food industry, and microbial research. Metabolomics aims to obtain an unbiased metabolic profile; however, this is complicated by compound instability, complex and often extensive sample processing, and nonlinear responses in mass spectrometry. Therefore, correcting for metabolite loss and mass spectrometry-related artifacts is essential, typically achieved through relative quantification against an isotopically labelled internal standard for each metabolite of interest. This article describes how to produce 13C-labelled yeast extract and its use as internal standard for metabolomics. More specifically, it provides step-by-step protocols for the fed-batch fermentation, quenching, metabolite extraction, and LC-MS and GC-MS characterization of the internal standard. It also includes a protocol explaining how to use the internal standard for the quantification of metabolites in yeast samples.

biochemistry↗

bioRxiv2026

SugarBase: mapping glycomolecule precursors in microbes

Glycan biosynthesis relies on nucleotide-activated sugars, essential metabolites across all domains of life, yet their usage in microbes is poorly understood. Here we present SugarBase, a mass spectrometry and bioinformatic pipeline for untargeted exploration of microbial nucleotide sugar networks. SugarBase resolves the chemical complexity of microbial metabolism by combining narrow-window DIA fragmentation with a chemistry-informed parent ion identification algorithm. Applying SugarBase across a broad phylogenetic range of microbes revealed extensive, species-specific nucleotide sugar profiles, including many candidates with no existing annotation, generating the most comprehensive inventory of nucleotide sugars to date. SugarBase guided identification of gene clusters and allowed discrimination between pseudaminic- and legionaminic acid-producing strains, where genomic and proteomic data provided only ambiguous information. We resolved distinct nonulosonic acid profiles in several Campylobacter jejuni strains, sugars which may alter susceptibility towards distinct flagellotropic phages. We further identify previously undescribed CMP-activated higher-carbon ulosonic acids in Magnetospirillum, expanding the known chemical space in glycan biosynthesis. In summary, SugarBase supports scalable discovery of microbial nucleotide sugar pathways and enzymes, expanding access to chemically complex glycans and providing new targets for antimicrobial development.

systems biology↗

bioRxiv2025

Measuring Temporal Variations of Nucleotide Pools in Microbial Granular Biofilm Performing Enhanced Biological Phosphorous Removal

Microbial communities often face environmental fluctuations that occur on timescales much shorter than their growth rate or proteome turnover. In such cases, cellular responses are likely driven by rapid changes in metabolite pools, particularly energy nucleotides including ATP, ADP, and AMP. However, robust methods to quantify these metabolites in biofilm-forming microbial communities are lacking. Here, we developed and systematically evaluated a metabolomics workflow for a granular biofilm enrichment, which performs Enhanced Biological Phosphorous Removal (EBPR). We combined fast quenching in liquid nitrogen and a boiling water extraction, followed by high resolution mass spectrometry, using porous graphitic carbon chromatography and 13C-labeled internal reference standards. Among tested procedures, a boiling water extraction was most suitable for extraction of nucleotides, as indicated by stable adenylate energy charge (AEC) and isotopic ratios. Applied to an anaerobic-aerobic cycle of a lab scale EBPR system, the method revealed dynamic changes in AEC and uridylate energy charge (UEC) during acetate uptake and polyphosphate degradation. These results demonstrate that energy pool imbalances underlie rapid metabolic switching observed in EBPR systems. Moreover, the established method provides a foundation for performing metabolomic studies of microbial biofilms in general.

systems biology↗

bioRxiv2025

Discovery of microbial glycoside hydrolases via enrichment and metaproteomics

The immense microbial diversity on Earth represents a vast genomic resource, yet discovering novel enzymes from complex environments remains challenging. Here, we combine microbial enrichment with metagenomics and metaproteomics to facilitate the identification of microbial glycoside hydrolases that operate under defined conditions. We enriched microbial communities using the carbohydrate polymer pullulan at elevated temperatures and acidic conditions. Pullulan is a natural polysaccharide composed of maltotriose units linked by -1,6 glycosidic bonds. Along with its hydrolyzing enzymes, it has broad applications across various industries. The enrichment inocula were sampled from thermophilic compost and soil from the bank of a pond. In both cases, Alicyclobacillus emerged as the dominant microorganism. Metaproteomic analysis of the enrichment biomass and secretome identified several pullulan-degrading enzymes from this organism. Notably, these enzymes were absent in the metagenomic analysis of the initial inoculum, underscoring the effectiveness of combining microbial enrichment with multi-omics for uncovering novel enzymes from complex microbial environments.

ecology↗