Martin Hagemann ^1* Eva-Maria Brouwer ^1* Hitesh K R Medipally ² Saskia Schwab ¹ Shanshan Song ¹ Marc M Nowaczyk ¹

• ¹ Institute of Biosciences, University of Rostock, Rostock, Germany
• ² School of Chemistry, Biotechnology and Health, Royal Institute of Technology, Stockholm, Stockholm, Sweden

Fixation of CO2 into the organic compound formate by formate dehydrogenase (Fdh) is regarded as the oldest autotrophic process on Earth. It has been proposed that an Fdh-dependent CO2 fixation module could support CO2 assimilation even in photoautotrophic organisms. In the present study we characterized the Fdh from Clostridium carboxidivorans (ccFdh), because it has the capability to reduce CO2 under aerobic conditions. During the production of recombinant ccFdh, in which the selenocysteine codon was exchanged by Cys, we could replace the W as transition metal in the ccFdh metal cofactor by Mo, which resulted in two-fold higher enzyme activity of 6 µmol formate min -1 . Then, we generated ccFdh variants, in which the strict NADH preference of the enzyme was changed to NADPH, because this reducing agent is produced in high amounts in the photosynthetic light process.Finally, we showed that the native ccFdh can also directly use ferredoxin as reducing agent, which is produced by the photosynthetic light reaction at the photosystem I. Collectively, these data suggest that ccFdh and particularly optimized variants can be regarded as a suitable enzyme to couple formate production to photosynthesis in photoautotroph organism, which could potentially support CO2 assimilation via the Calvin-Benson-Bassham cycle and minimize CO2 losses due to photorespiration.