Membrane-bound D-glucose dehydrogenase from Pseudomonas sp.: Solubilization, purification and characterization.

A membrane-bound n-glucose dehydrogenase [E. C. 1.1.99. a] was solubilized from the membrane of Pseudomonas sp. and purified to a nearly homogeneous state. The solubilized enzyme was monomeric in the presence of 1% Triton X-100 but aggregated after removing the detergent. The enzyme was a single peptide having a molecular weight of about 90, 000. The enzyme reacted with various artificial electron acceptors such as phenazine methosulfate, 2, 6-dichlorophenolindophenol, Wurster's blue, coenzyme Q1 and ferricyanide. The enzyme had a dual optimum pH depending on the electron acceptor. Reductase activities of the enzyme for 2, 6-dichlorophenolindophenol, ferricyanide and coenzyme Q1, were found in more acidic pH region, whereas its activities for phenazine methosulfate and Wurster's blue were observed in more alkaline region. p-Benzoquinone inhibited phenazine metho-sulfate reductase activity non-competitively but it inhibited 2, 6-dichlorophenolindophenol reductase activity competitively against the acceptor. The enzyme possessed fairly broad substrate specificity, and the reaction product was a gluconolactone.