Isolation and Characterization of Membrane-bound L-Sorbose Dehydrogenase from Gluconobacter melanogenus UV10

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Membrane-bound L-sorbose dehydrogenase was purified from the membrane fraction of Gluconobacter melanogenus UV10. The reaction with 2, 6-dichlorophenolindophenol or phenazine methosulfate as acceptor was formulated as; L-sorbose + acceptor →-sorbosone + reduced acceptor.
The enzyme was purified by DEAE-Cellulose, DEAE-Sepharose, and Sephadex G-200 column chromatographies after differential solubilization with Triton X-100 and Tween 80, and appeared as a single band in a SDS-PAGE gel at the molecular weight of about 58, 000. It was most active at pH 7.0 and 48°C, and rather stable at around 7.0 in a buffer containing 200 mM L-sorbose, below 30°C. It showed high substrate specificity for L-sorbose, and its activity was stimulated by Fe³⁺ and Co²⁺, and inhibited by Cu²⁺, quinine, mono-iodoacetate, and sodium fluoroacetate.