Biology subjects

Dronsella, B. B.

Publications and source records attributed to Dronsella, B. B..

3 recordsLinked to original sources

bioRxiv2026

Combining evolution and machine learning-guided pathway optimization to engineer a novel methylsuccinate module for synthetic C1 metabolism in vivo

De novo metabolic pathways open possibilities for sustainable biotransformations in microbes. However, the in vivo-implementation of such new-to-nature pathways is highly challenging and heavily relies on adaptive laboratory evolution (ALE) of the hosts native metabolic network. Here, we assess how much this need for host-centric ALE can be overcome and/or complemented through the informed design of the newly introduced pathway. Exemplifying for a synthetic CO2-fixation module via methyl-succinate, we established methylsuccinate-dependent growth of Escherichia coli over six months by ALE of E. colis native metabolism. In parallel, we developed a machine-learning guided workflow (MEVIS) for the automated engineering of the synthetic pathway, resulting in methylsuccinate-dependent growth within three weeks. Critically, performing MEVIS in the background of the ALE-evolved strain is necessary to further approach wild-type like growth, demonstrating how ALE in combination with machine-learning-guided lab automation holds great potential to accelerate and improve design-build-test-learn cycles in contemporary metabolic engineering.

bioengineering↗

bioRxiv2023

Engineering the biological conversion of formate into crotonate in Cupriavidus necator

To advance the sustainability of the biobased economy, our society needs to develop novel bioprocesses based on truly renewable resources. The C1-molecule formate is increasingly proposed as carbon and energy source for microbial fermentations, as it can be efficiently generated electrochemically from CO2 and renewable energy. Yet, its biotechnological conversion into value-added compounds has been limited to a handful of examples. In this work, we engineered the natural formatotrophic bacterium C. necator as cell factory to enable biological conversion of formate into crotonate, a platform short-chain unsaturated carboxylic acid of biotechnological relevance. First, we developed a small-scale (150-mL working volume) cultivation setup for growing C. necator in minimal medium using formate as only carbon and energy source. By using a fed-batch strategy with automatic feeding of formic acid, we could increase final biomass concentrations 15-fold compared to batch cultivations in flasks. Then, we engineered a heterologous crotonate pathway in the bacterium via a modular approach, where each pathway section was assessed using multiple candidates. The best performing modules included a malonyl-CoA bypass for increasing the thermodynamic drive towards the intermediate acetoacetyl-CoA and subsequent conversion to crotonyl-CoA through partial reverse {beta}-oxidation. This pathway architecture was then tested for formate-based biosynthesis in our fed-batch setup, resulting in a two-fold higher titer, three-fold higher productivity, and five-fold higher yield compared to the strain not harboring the bypass. Eventually, we reached a maximum product titer of 148.0 {+/-} 6.8 mg/L. Altogether, this work consists in a proof-of-principle integrating bioprocess and metabolic engineering approaches for the biological upgrading of formate into a value-added platform chemical.

synthetic biology↗

bioRxiv2022

Engineered synthetic one-carbon fixation exceeds yield of the Calvin Cycle

One-carbon (C1) feedstocks derived from CO2 and renewable electricity, such as formate, are promising substrates for sustainable production of chemicals, food and fuels. Energetically more efficient, engineered C1-fixation pathways were proposed to increase biomass yields above their natural counterparts, but have so far not been shown to achieve this. Here, we replace the native energy-inefficient Calvin-Benson-Bassham (CBB) cycle in Cupriavidus necator by genomic integration of the synthetic reductive glycine pathway for growth on formate. Our final engineered strain reaches a higher biomass yield than the CBB-cycle-utilizing wild type, showing for the first time that efficiencies found in natural metabolism can be exceeded via a synthetic pathway. This yield increase demonstrates the potential of synthetic metabolism and is an important step towards realizing truly sustainable, economically feasible bio-based production.

synthetic biology↗