Biosynthesis of Glycine from One-Carbon Resources Using an Engineered Escherichia coli Whole-Cell Catalyst

Carbon dioxide (CO₂) is a cost-effective, abundant, and renewable carbon source, but its utilization technologies face several issues. The reductive glycine pathway (RGP) is recognized as one of the most efficient one-carbon (C1) assimilation routes in nature, with its core component-the glycine cleavage system (GCS: GcvP, GcvH, GcvT, and GcvL)-playing an essential role in C1 metabolism. To develop efficient CO₂ conversion and utilization pathways, we identified <i>Nh</i>FtfL and <i>Am</i>FchA-MtdA with high catalytic efficiency through gene mining and constructed a four-plasmid co-expression system in <i>E. coli</i> BL21(DE3) using Gibson Assembly. This system integrated GcvP-GcvH, GcvT-GcvL, <i>Nh</i>FtfL-<i>Am</i>FchA-MtdA, and <i>Rs</i>PPK2, thereby reconstituting the complete RGP while enhancing ATP supply. The engineered strain functioned as an efficient whole-cell biocatalyst, achieving a glycine space-time productivity of 0.125 mmol/L/h via one-pot conversion of formate. Furthermore, we expanded the application scope by developing a whole-cell electrocatalysis system that directly synthesized glycine from CO₂ and NH₄Cl, achieving a glycine space-time productivity of 0.135 mmol/L/h. This study demonstrates the potential of the engineered RGP system for upgrading C1 resources and supports the transition toward carbon neutrality.