Carbon dioxide is an important carbon feedstock for a future green economy. This requires efficient strategies for its conversion into multicarbon compounds. We developed a synthetic cycle for the continuous fixation of CO2 in vitro. The crotonyl–CoA/ethylmalonyl-CoA/hydroxybutyryl-CoA (CETCH) cycle is a reaction network of 17 enzymes that converts CO2 into organic molecules. The CETCH cycle was drafted by metabolic retrosynthesis and established with enzymes originating from nine different organisms of all three domains of life. This cycle adds a seventh, synthetic alternative to the six naturally evolved CO2 fixation pathways, thereby opening the way for in vitro and in vivo applications.