Thesis presented December 07, 2021

Abstract:
CO₂ metabolism plays a fundamental role in biology, as it represents a “building block” for the synthesis of organic molecules in living organisms. On the other hand, the excessive CO₂ emissions due to human activity in recent decades requires today the search for effective solutions dedicated to the closure of the carbon cycle. Thus, it is interesting to develop some alternatives inspired by nature. In particular, carbon monoxide dehydrogenase (CODH) catalyses the reversible reduction of CO₂ to CO and represents an attractive biocatalyst in terms of activity, selectivity, ability to operate in mild conditions with low overpotentials. In addition, it does not require high production costs and uses earth abundant-metals. For these reasons, several studies have been conducted on [NiFe]-CODHs for applications in electrocatalytic CO₂ reduction reaction (CO₂RR). [NiFe]-CODHs consist of a complex multimetallic NiFe₄S₄ active site, unique in biology, and additional FeS clusters involved in the electron transfer from the buried active site to the protein surface. Here, we present a novel and efficient method to overproduce the fully active recombinant [NiFe]-CODH from Rhodospirillum rubrum in the easy-to-handle bacteria E. coli. We also developed the stable and favourable immobilization of the enzyme on non-covalently functionalized carbon nanotubes (CNTs), subsequently integrated in a gas-diffusion electrode device This device has comparable performance to molecular and metal catalysts for CO₂RR. To go further, the understanding of the intramolecular electron transfer within the enzyme has been undertaken, in order to optimize its capacity to work in direct electron transfer with the electrode. Other optimizations are considered, such as the direct evolution of the enzyme to provide a better resistance towards O₂. All together, the results of this work have paved the way to study and use this powerful biocatalyst in the near future for biotechnological applications.

Keywords:
CO₂ reduction, CO oxidation, Biocatalysis, Carbon Monoxide Dehydrogenase, Carbon Nanotubes

On-line thesis.