An Efficient CO2-Upcycling Platform Based on Engineered Halomonas TD with Enhanced Acetate-Utilizing Capacity via Adaptive Laboratory Evolution.

Abstract

Biohybrid conversion of carbon dioxide (CO₂) into value-added bioproducts via engineered microbes using CO₂-derived electrolytes (CDE) addresses global CO₂ emissions, but most recombinants have poor saline CDE tolerance and low carbon conversion rate (CCR). Herein, Halomonas TD (salt-resistant) was adaptively evolved into TD80, which efficiently uses acetate; its aceE gene mutation (encoding pyruvate dehydrogenase) drives acetate utilization. Subsequently, different biosynthesis pathways in TD80 enabled high yields of poly-3-hydroxybutyrate (PHB), poly-3-hydroxybutyrate-co-4-hydroxybutyrate (P34HB), 3-hydroxybutyrate (3HB), violacein, ectoine, 1,3-diaminopropane (1,3-DAP) and superoxide dismutase (SOD), respectively. Moreover, 26.0 g L^-1 ectoine and 29.6 g L^-1 PHB can be achieved by recombinant TD80 strains during fed-batch studies. Finally, a non-canonical pathway was designed to recycle the excess malonyl-CoA into PHB. The resultant PHB content in fed-batch study was increased from 60 wt% to 80 wt%. Moreover, co-producing ectoine and PHB could further boost the CCR of CDE-to-product up to 53.7 mol%, which exemplified promising potential for biohybrid CO₂ upcycling involved in carbon capture and utilization system. Furthermore, TD80 was engineered to grow on formate only aiming to achieve the full use of CDE. The establishment of technology and economy assessment (TEA) confirmed the Halomonas-based platform's efficiency and economic viability for carbon footprint reduction.