Biology subjects

Moses, T.

Publications and source records attributed to Moses, T..

3 recordsLinked to original sources

bioRxiv2025

An automated metabolite extraction workflow for global metabolomics analysis using the Agilent Bravo liquid handling platform

Metabolomics is the comprehensive study of small molecules that provides a snapshot of an organisms physiological state. Reflecting phenotype more closely than genes or proteins, metabolites reveal changes linked to diseases, mutations, genetic interventions, and environmental stimuli. Recent technological advancements in metabolomics analysis through the application of ion mobility mass spectrometry have enhanced the comprehensive analysis of complex metabolic mixtures. However, pre-analytical bottlenecks in throughput and consistent extraction persist. We developed an automated, sample-agnostic metabolite extraction workflow for diverse liquid samples using an Agilent Bravo liquid handling platform. Here, we provide device, protocol, and form files for efficient sample processing to extract metabolites for global metabolomics analysis using liquid chromatography - mass spectrometry techniques.

biochemistry↗

bioRxiv2025

Real-time tracking of intracellular prenyl phosphate pools in the marine diatom Phaeodactylum tricornutum with a metabolite protein-based biosensor

Metabolite-responsive, protein-based biosensors are a powerful tool for monitoring cellular metabolite dynamics in vivo and accelerating strain engineering workflows in microorganisms. In this study, we introduced a previously developed protein-based biosensor, computationally designed to detect farnesyl diphosphate (FPP), in the marine diatom Phaeodactylum tricornutum. We expressed two versions of the biosensor constitutively, under a strong promoter-terminator pair using extrachromosomal episomes, and we parameterized the capacity of both designs in detecting intracellular metabolite levels. Initial assays revealed that the two versions of the biosensor we investigated, S3-2D and S3-3A, had specificity not only for FPP but also for other exogenously supplied prenyl phosphates such as geranyl diphosphate (GPP) and geranylgeranyl diphosphate (GGPP) in a dose-dependent manner. We further demonstrated the capacity of S3-3A to track perturbations in the endogenous prenyl phosphate pools by testing it in the presence of pharmacological inhibition of the mevalonate pathway. Moreover, S3-3A generated signal "hot-spots" around the peroxisomes, suggesting their involvement in isoprenoid biosynthesis, which led us to characterize the subcellular localization of the key enzyme mevalonate kinase. These findings lay the groundwork for developing metabolite-responsive biosensors as robust tools for monitoring and investigating prenyl phosphate dynamics, providing a foundation for advanced metabolic engineering of microalgae.

synthetic biology↗

bioRxiv2023

The type of carbon source not the growth rate it supports can determine diauxie

How cells choose between potential carbon sources is a classic example of cellular decision-making, and we know that many organisms prioritise glucose. Yet there has been little investigation of whether other sugars are also preferred, blinkering our view of carbon sensing. Here we study eukaryotic budding yeast and its growth on mixtures of palatinose, an isomer of sucrose, with other sugars. We find that yeast prioritise galactose over palatinose, but not sucrose or fructose, despite all three of these sugars being able to support faster growth than palatinose. Our results therefore disfavour carbon flux-sensing as the sole mechanism. By using genetic perturbations and transcriptomics, we show that repression is active and through Gal4, the master regulator of the GAL regulon. Cells enforce their preference for galactose over palatinose by preventing runaway positive feedback in the MAL regulon, whose genes enable palatinose catabolism. They do so both by repressing MAL11, the gene encoding the palatinose transporter, and by first expressing the isomaltases, IMA1 and IMA5, which cleave palatinose and so prevent its intracellular concentration becoming enough to induce further MAL expression. Our results demonstrate that budding yeast actively maintain a preference for carbon sources other than glucose and that such preferences have been selected by more than differences in growth rates. They imply that carbon-sensing strategies even in unicellular organisms are more complex than previously thought.

systems biology↗