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 PQQH(2) to generate semiquinone radicals. No such increase in the EPR signal was observed in UQ8-free mGDH under the same conditions. Moreover, a UQ(2) 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 QI site acts as a single electron mediator in the intramolecular electron transfer in mGDH.