Rebooting the Calvin-Benson Cycle – CalvinReboot

Autotrophic carbon fixation, the Calvin-Benson cycle (CBC), is the main limiting factor of photosynthesis. It is believed to be impossible to improve and that Evolution reached the best possible fitness in current environment. However, a different maximum fitness of CBC was never ruled out. We will assess this question through a novel synthetic biology approach, which might improve the CBC. First, we will analyze the natural sequence space and infer from it the ancestral sequences of selected CBC enzymes. Second, we shall assess the fitness of synthetic CBC in vivo by replacing endogenous CBC enzymes by their ancestral counterpart. This will be performed in parallel in two well-studied chassis, the eukaryotic green alga Chlamydomonas reinhardtii and the Cyanobacteria Synechocystis PCC 6803. Finally, we will use these ancestral sequences as a platform for directed evolution. Through this approach, we will reassess the CBC evolution through synthesis, to understand by building.