Occurrence of a Bound Ubiquinone and Its Function in Escherichia coli Membrane-bound Quinoprotein Glucose Dehydrogenase*

The membrane-bound pyrroloquinoline quinone (PQQ)-containing quinoprotein glucose dehydrogenase (mGDH) in Escherichia coli functions by catalyzing glucose oxidation in the periplasm and by transferring electrons directly to ubiquinone (UQ) in the respiratory chain. To clarify the intramolecular electron transfer of mGDH, quantitation and identification of UQ were performed, indicating that purified mGDH contains a tightly bound UQ₈ in its molecule. A significant increase in the EPR signal was observed following glucose addition in mGDH reconstituted with PQQ and Mg²⁺, suggesting that bound UQ₈ accepts a single electron from PQQH₂ to generate semiquinone radicals. No such increase in the EPR signal was observed in UQ₈-free mGDH under the same conditions. Moreover, a UQ₂ reductase assay with a UQ-related inhibitor (C49) revealed different inhibition kinetics between the wild-type mGDH and UQ₈-free mGDH. From these findings, we propose that the native mGDH bears two ubiquinone-binding sites, one (Q_I) for bound UQ₈ in its molecule and the other (Q_II) for UQ₈ in the ubiquinone pool, and that the bound UQ₈ in the Q_I site acts as a single electron mediator in the intramolecular electron transfer in mGDH.