biostudies-literatureUnknown13Li JThis study engineered the thermotolerant yeast Kluyveromyces marxianus to produce 3-hydroxypropionic acid (3-HP), a key precursor for biodegradable plastics, via the malonyl-CoA pathway using non-food feedstocks. The 3-HP titer was further increased through deleting Adh2A and Ach1, which prevents the synthesis of byproducts ethanol and acetic acid. Using Jerusalem artichoke tuber powder, engineered strain produced 27.32 and 32.31 g/L of 3-HP at 37 °C and 42 °C through fed-batch fermentation. Metabolic reconstruction replaced the native FADH2-dependent glycerol pathway (GUT1/GUT2) with an NADH-generating GDH1/DAK1 pathway, significantly enhancing glycerol utilization and increasing intracellular NADH supply by 62 %. Overexpression of Utr1 can further enhance the NADPH supply. Combined with heterologous expression of a codon-optimized, high-activity malonyl-CoA reductase (MCR) mutant (MCRN940V/K1106W/S1114R), the engineered strain achieved 3-HP titers of 33.15 g/L in fed-batch fermentation using pure glycerol at 42 °C. Crucially, it also produced 26.57 g/L 3-HP directly from crude glycerol at 42 °C. The thermotolerant fermentation at 42 °C, unprecedented for yeast-based 3-HP synthesis, reduces cooling water consumption by approximately 60 %, translating to an estimated annual CO2 reduction of 27.1 tons per 1000-ton fermenter. This work establishes a cost-effective, industrially scalable bioprocess for valorizing Jerusalem artichoke tubers and crude glycerol into a key platform chemical for biodegradable plastics and green chemicals, leveraging the strain's substrate flexibility, process robustness, and significant environmental advantages.Synthetic and systems biotechnology1-13https://www.ebi.ac.uk/biostudies/studies/S-EPMC12856427biostudies-literatureEngineering Kluyveromyces marxianus for 3-hydroxypropionic acid production at elevated temperature from Jerusalem artichoke tubers and crude glycerol.PMC12856427Li YZhao MLi JXie YRen LHu ZZhang BZha HfalseEngineering Kluyveromyces marxianus for 3-hydroxypropionic acid production at elevated temperature from Jerusalem artichoke tubers and crude glycerol.This study engineered the thermotolerant yeast Kluyveromyces marxianus to produce 3-hydroxypropionic acid (3-HP), a key precursor for biodegradable plastics, via the malonyl-CoA pathway using non-food feedstocks. The 3-HP titer was further increased through deleting Adh2A and Ach1, which prevents the synthesis of byproducts ethanol and acetic acid. Using Jerusalem artichoke tuber powder, engineered strain produced 27.32 and 32.31 g/L of 3-HP at 37 °C and 42 °C through fed-batch fermentation. Metabolic reconstruction replaced the native FADH2-dependent glycerol pathway (GUT1/GUT2) with an NADH-generating GDH1/DAK1 pathway, significantly enhancing glycerol utilization and increasing intracellular NADH supply by 62 %. Overexpression of Utr1 can further enhance the NADPH supply. Combined with heterologous expression of a codon-optimized, high-activity malonyl-CoA reductase (MCR) mutant (MCRN940V/K1106W/S1114R), the engineered strain achieved 3-HP titers of 33.15 g/L in fed-batch fermentation using pure glycerol at 42 °C. Crucially, it also produced 26.57 g/L 3-HP directly from crude glycerol at 42 °C. The thermotolerant fermentation at 42 °C, unprecedented for yeast-based 3-HP synthesis, reduces cooling water consumption by approximately 60 %, translating to an estimated annual CO2 reduction of 27.1 tons per 1000-ton fermenter. This work establishes a cost-effective, industrially scalable bioprocess for valorizing Jerusalem artichoke tubers and crude glycerol into a key platform chemical for biodegradable plastics and green chemicals, leveraging the strain's substrate flexibility, process robustness, and significant environmental advantages.2026-01-01T00:00:00Z2026 Sep2026-06-18T05:50:21.13Z2026-06-18T03:07:49.033ZS-EPMC128564274162498410.1016/j.synbio.2026.01.008