Print Email Facebook Twitter Optimizing the balance between heterologous acetate- and CO2-reduction pathways in anaerobic cultures of Saccharomyces cerevisiae strains engineered for low-glycerol production Title Optimizing the balance between heterologous acetate- and CO2-reduction pathways in anaerobic cultures of Saccharomyces cerevisiae strains engineered for low-glycerol production Author van Aalst, Aafke C.A. (Student TU Delft) Geraats, Ellen H. (Student TU Delft) Martini, M.L.A. (DSM) Mans, R. (TU Delft BT/Industriele Microbiologie) Pronk, J.T. (TU Delft BT/Biotechnologie) Department BT/Biotechnologie Date 2023 Abstract In anaerobic Saccharomyces cerevisiae cultures, NADH (reduced form of nicotinamide adenine dinucleotide)-cofactor balancing by glycerol formation constrains ethanol yields. Introduction of an acetate-to-ethanol reduction pathway based on heterologous acetylating acetaldehyde dehydrogenase (A-ALD) can replace glycerol formation as 'redox-sink' and improve ethanol yields in acetate-containing media. Acetate concentrations in feedstock for first-generation bioethanol production are, however, insufficient to completely replace glycerol formation. An alternative glycerol-reduction strategy bypasses the oxidative reaction in glycolysis by introducing phosphoribulokinase (PRK) and ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO). For optimal performance in industrial settings, yeast strains should ideally first fully convert acetate and, subsequently, continue low-glycerol fermentation via the PRK-RuBisCO pathway. However, anaerobic batch cultures of a strain carrying both pathways showed inferior acetate reduction relative to a strain expressing only the A-ALD pathway. Complete A-ALD-mediated acetate reduction by a dual-pathway strain, grown anaerobically on 50 g L-1 glucose and 5 mmol L-1 acetate, was achieved upon reducing PRK abundance by a C-terminal extension of its amino acid sequence. Yields of glycerol and ethanol on glucose were 55% lower and 6% higher, respectively, than those of a nonengineered reference strain. The negative impact of the PRK-RuBisCO pathway on acetate reduction was attributed to sensitivity of the reversible A-ALD reaction to intracellular acetaldehyde concentrations. Subject acetatebiofuelsfermentationNADHredox cofactor balance To reference this document use: http://resolver.tudelft.nl/uuid:e139a76b-9fcf-4527-b49d-2a1b4fcc61ba DOI https://doi.org/10.1093/femsyr/foad048 Source FEMS Yeast Research, 23 Part of collection Institutional Repository Document type journal article Rights © 2023 Aafke C.A. van Aalst, Ellen H. Geraats, M.L.A. Martini, R. Mans, J.T. Pronk Files PDF foad048.pdf 1.17 MB Close viewer /islandora/object/uuid:e139a76b-9fcf-4527-b49d-2a1b4fcc61ba/datastream/OBJ/view