Thermodynamic Contribution to the Regulation of Electron Transfer in the Na⁺-Pumping NADH:Quinone Oxidoreductase from <i>Vibrio cholerae</i>

The Na⁺-pumping NADH:quinone oxidoreductase (Na⁺-NQR) is a fundamental enzyme of the oxidative phosphorylation metabolism and ionic homeostasis in several pathogenic and marine bacteria. To understand the mechanism that couples electron transfer with sodium translocation in Na⁺-NQR, the ion dependence of the redox potential of the individual cofactors was studied using a spectroelectrochemical approach. The redox potential of one of the FMN cofactors increased 90 mV in the presence of Na⁺ or Li⁺, compared to the redox potentials measured in the presence of other cations that are not transported by the enzyme, such as K⁺, Rb⁺, and NH₄⁺. This shift in redox potential of one FMN confirms the crucial role of the FMN anionic radicals in the Na⁺ pumping mechanism and demonstrates that the control of the electron transfer rate has both kinetic (via conformational changes) and thermodynamic components