The steady increase of CO2 levels in the atmosphere is one of the main reasons for the current climate and environmental crisis, which is also fueled by the carbon-intensive industrial production of chemicals and organic compounds. Autotrophic microorganisms, which consume CO2 instead of glucose, sucrose or starch, are an exciting opportunity to sustainably produce organic chemical products. However, the rates of natural CO2-fixing pathways and energy-harvesting efficiency in photoautotrophs, such as microalgae, is generally low (around 1–2%). One approach to bypass this limitation is to genetically engineer bacteria and yeasts to improve their CO2 assimilation efficiency. Now, writing in the Proceedings of the National Academy of Sciences, Diethard Mattanovich and colleagues integrated pathways in the synthetic autotrophic yeast Komagataella phaffii to produce lactic and itaconic acid (two important feedstocks for polymer production) based on CO2 as the only carbon source.

The team had previously converted the methanol-assimilation pathway of methylotrophic yeast Pichia pastoris into a CO2-fixating pathway resembling the Calvin–Benson–Bessham (CBB) cycle. Building on this work, the team replaced the formaldehyde-assimilating dihydroxyacetone synthase (DAS) enzyme of K. phaffi with a bacterial ribulose 1,5-bisphosphate carboxylase/oxygenase (RuBisCO) enzyme and added phosphoribulose kinase (PRK) from spinach to enable the yeast strain to directly assimilate CO2 instead of methanol. The researchers went on to show the feasibility of using the same platform to produce organic acids from CO2. “Thus far, CO2 assimilation in yeast has only been achieved by co-assimilation with sugars, such as glucose or xylose, increasing the yield of ethanol production,” explains Mattanovich. “Our approach enables the growth of yeast biomass, and the production of different chemicals, with CO2 as the only carbon source. In other words, everything that is produced is made of CO2.”

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Specifically, the expression of cytosolic cis-aconitate decarboxylase (CadA), a gene encoding the enzyme for the decarboxylation reaction of cis-aconitate to itaconic acid, and a bacterial lactate dehydrogenase gene, induced yeast-mediated production of up to 2,000 and 600 mg l−1 itaconic and lactic acid, respectively. Interestingly, balancing the co-expression of CadA with the mitochondrial tricarboxylic acid transporter (mttA), which shuttles the substrate cis-aconitate to CadA in the cytosol, improved the production of itaconic acid. “The technical difference in our approach is that we built the strains using a natural pathway and methanol (also a single carbon molecule, such as CO2),” says Mattanovich. “This pathway is present in some yeast species, it is separated from the rest of the yeast metabolism in specialized compartments, and it is highly optimized by evolution, promising a high success rate.”

“Everything that is produced is made of CO2

However, the titres obtained in shake-flask cultivations are not yet industrially relevant. “We need to boost the rate of CO2 assimilation, using enzymes from different sources and balancing the activities of the CO2 assimilation pathway,” explains Mattanovich. The team will also address the energy efficiency of CO2 assimilation (exploring alternative pathways) and engage in bioprocess development using model-based process design and experimental optimization. If industrially feasible yields can be achieved, this approach could enable the sustainable production of chemicals and other products.