Redox Properties of the Prosthetic Groups of Na+-Translocating NADH:Quinone Oxidoreductase. 2. Study of the Enzyme by Optical Spectroscopy

Redox titration of the electronic spectra of the prosthetic groups of the Na+-translocating NADH: quinone oxidoreductase (Na+-NQR) from Vibrio harveyi at different pH values showed five redox transitions corresponding to the four flavin cofactors of the enzyme and one additional transition reflecting oxidoreduction of the [2Fe-2S] cluster. The pH dependence of he measured midpoint redox potentials showed that the two-electron reduction of the FAD located in the NqrF subunit was Coupled with the uptake of only one H+. The one-electron reduction of neutral semiquinone of riboflavin and the formation of anion flavosemiquinone from the oxidized FMN bound to the NqrB subunit were not coupled to any proton uptake. The two sequential one-electron reductions of the FMN residue bound to the NqrC subunit showed pH-independent formation of anion radical in the first step and the formation of fully reduced flavin coupled to the uptake of one H+ in the second step. All four flavins stayed in the anionic form in the fully reduced enzyme. None of the six redox transitions in Na+-NQR showed dependence of its midpoint redox potential on the concentration of sodium ions. A model of the sequence of electron transfer steps In the enzyme is suggested.