Long Range Control of Reactivity in CO Dehydrogenases

• Applicant: Professor Dr. Holger Dobbek
• Subject Area: Biochemistry
• Term: since 2023
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 529954943

Homologous enzymes are defined by a set of highly conserved “core” residues that are required for function. The catalytic properties (catalytic rates, Michaelis constants, susceptibility to inhibitors, and even bidirectionality) are frequently determined by non-conserved residues remote from the active site. Identifying core residues defining the function, understanding the influence of the secondary sphere residues, and comprehending long-range effects are fundamental to reveal the mechanisms of metalloenzymes and understand how they have been tuned by Evolution. Our proposal concerns NiFe CO dehydrogenases (CODHs), which catalyze the reversible reduction of CO2 to CO. The partners (CNRS and HU) have contributed to elucidating the mechanisms of CODHs, separately and together. HU has recently isolated and characterized one particular enzyme, CooS-V, which is very similar in terms of sequence to prototypical CODHs but performs a distinct (yet unknown) reaction. The observed flexibility of the CooS-V active site is reminiscent of conformational changes that are functionally important in a CODH studied by CNRS. With its unique situation of being the closest enzyme to a CODH without being one, CooS-V provides opportunities to learn both about the core residues that define CODH function and about the long-range interactions that modulate CODH activity by using site-directed mutagenesis to either engineer CODH function into CooS-V or transform a CODH into an enzyme with CooS-V activity. In this project, both partners will design and produce protein variants that are intermediate between known CODHs and CooS-V, and they will join forces to characterize them using crystallography (HU) and electrochemistry (CNRS), providing structural information and in-depth functional and kinetic characterization. Combining the approaches of both partners will give unprecedented insight into how the different properties of CODHs have evolved and can be harnessed for future applications.

• DFG Programme: Research Grants
• International Connection: France
• Cooperation Partner: Professor Dr. Vincent Fourmond