2-Amino-1,3-propanediol (2-APD) is a chemical building block for the production of various value-added pharmaceuticals. However, the current manufacture of 2-APD predominantly relies on chemical processes by utilizing fossil fuel-derived and highly explosive raw materials. Herein, we established an artificial biosynthetic pathway for converting glucose to 2-APD in a metabolically engineered Escherichia coli. This artificial pathway employs an engineered heterogeneous aminotransferase RtxA for diverting dihydroxyacetone phosphate to generate 2-APD phosphate and an endogenous phosphatase for converting it into the target product 2-APD. Through fine-tuning the activity and solubility of RtxA for efficiently extending the glycolysis pathway, enhancing the metabolic recycling of amino-containing substrate supply via nitrogen-borrowing, and unlocking the dephosphorylation involved in the downstream pathway, the best metabolically engineered E. coli strain LYC-5 was constructed stepwise. Under aerobic conditions, a fed-batch fermentation of the strain LYC-5 produced 14.6 g/L 2-APD with a productivity of 0.122 g/L/h in a 6-L bioreactor, which was the highest reported titer to the best of our knowledge. This work demonstrates the great potential to provide an environmentally friendly and efficient approach for 2-APD production.