Heterologous expression of pyruvate formate lyase enhances cell growth of Clostridium ljungdahlii during microbial electrosynthesis

Slow cell growth and low biomass yields are hurdles for gas fermentation by acetogens. Microbial electrosynthesis (MES) utilizes acetogens as biocatalysts to reduce CO ₂ and produce commodity chemicals using electricity. However, limited electron supply in a bioelectrochemical system (BES) aggravates the poor cell growth of acetogens resulting in low productivities. Formate is a soluble C1 feedstock that can be produced by CO ₂ reduction (1). Thus, assimilation of formate can unburden the amount of electrons required for MES. Acetobacterium woodii posseses multiple pyruvate formate lyase (PFL) genes that are upregulated during formatotrophic growth. In this study, Clostridium ljugdahlii was engineered to heterologously express a pfl geneset of A. woodii to increase the cell growth of C. ljungdahlii during microbial electrosynthesis. Different combinations of pfl A and pfl B from A. woodii were tested in C. ljungdahlii to find the best working combination under control of P _fdx -riboswitch expression system. Expression of pfl B1 and pfl A of A. woodii showed higher maximum OD of C. ljugdahlii under H ₂ :CO ₂ condition with supplementation of 80 mM sodium formate. More than 40 mM of sodium formate caused significantly longer lag phase but the lag phase could be shortened after adaptation in 80 mM of sodium formate. At the end, the engineered strain showed improved cell growth (OD _max 0.22 ± 0.05) and acetate production (21.8 ± 5.6 mM) during microbial electrosynthesis compared to the control strain (OD _max 0.10 ±0.06 and 10.2 ±2.5 mM acetate). These results will be useful for strain development for microbial electrosynthesis, as well as gas fermentation.