Construction and extension of the SM cycle for synthetic one‑carbon compounds utilization

One‑carbon (C1) compounds are important carbon feedstocks for achieving a sustainable bio-economy. However, it remains challenging to tap the full potential of C1 compounds utilization, due to the inefficiency of co-utilization of different C1 compounds. Here, a new-to-nature C1-fixing pathway, the serine aldolase/malic enzyme (SM) cycle, was developed to convert C1 compounds to acetyl-CoA by programming CO 2 -fixing carboxylation reaction, formaldehyde-assimilating aldolization reaction and acetyl-CoA-producing cleavage reaction. The SM cycle was drafted by metabolic retrosynthesis, constructed with enzymes originating from six different organisms, and optimized by engineering key enzymes and adjusting reaction system, and thus its C1 utilization rate was up to 30.2 nmol min −1  mg −1  cycle proteins. Sequentially, the SM cycle was programmed in a plug-and-play fashion by installing carbon chain extension modules for converting C1 compounds to various organic molecules such as C 2 -compounds ( e.g. , acetaldehyde and acetyl-CoA), C 3 -compounds ( e.g. , pyruvate, 3-hydroxypropionate and propionyl-CoA), C 4 -compounds ( e.g. , succinate, fumarate and succinyl-CoA). Finally, the applicability of the SM cycle was ascertained in a 5 L fermenter for the production of diverse chemicals, such as pyruvate and succinate, with C1 utilization rate up to 34.5 and 35.8 mg/gDCW/h, respectively, surpassing that of microalgae and cyanobacteria. This study presented here provides a carbon-negative flexible platform for widening the versatility of C1 utilization and may boost C1-based biomanufacturing in a sustainable future.