Redox thermodynamics of high-spin and low-spin forms of chlorite dismutases of diverse subunit and oligomeric structure

Chlorite dismutases (Clds) are heme b containing oxidoreductases that convert chlorite to chloride and dioxygen. In this work the thermodynamics of the one-electron reduction of the ferric high-spin forms and of the six-coordinate low-spin cyanide adducts of the enzymes from Nitrobacter winogradskyi (NwCld) and Candidatus “Nitrospira defluvii” (NdCld) were determined through spectroelectrochemical experiments. The above proteins belong to two phylogenetically separated lineages that differ in subunit (21.5 kDa versus 26 kDa) and oligomeric (dimeric versus pentameric) structure but exhibit similar chlorite degradation activity. The E°’ values for free and cyanide-bound proteins were determined to be -119 mV and -397 mV for NwCld as well as -113 mV and -404 mV for NdCld, respectively (pH 7.0, 25 °C). Variable-temperature spectroelectrochemical experiments revealed that the oxidized state of both proteins is enthalpically stabilized. Molecular dynamics simulations suggest that changes in the protein structure are negligible, whereas solvent reorganization is mainly responsible for the increase of entropy during the redox reaction. Obtained data are discussed with respect to the known structures of the two Clds and the proposed reaction mechanism.