Identification of the Prosthetic Group and Further Characterization of a Novel Enzyme, Polyethylene Glycol Dehydrogenase

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Abstract
The purified polyethylene glycol (PEG) dehydrogenase from cells of a synergistic mixed culture of Flavobacterium and Pseudomonas spp. showed a similar absorption spectrum to those of other quinoproteins reported so far. The prosthetic group of the PEG dehydrogenase after extraction with cold methanol and purification by DEAE-Sephadex A-25 column chromatography and Sephadex G-25 gel filtration showed the same elution profiles as those of authentic pyrroloquinoline quinone (PQQ). Absorption and fluorescence spectra of the purified prosthetic group and its prosthetic group capability for glucose dehydrogenase indicated that it was identical with authentic PQQ. The enzyme was induced during bacterial cell growth on a medium containing PEG 6000 as a sole source of Ca. The purified enzyme oxidized primary alcohols of C2-C16 and the corresponding aldehydes of C4-C7. The enzyme also reacted with nonionic surfactants containing PEG residues. The enzyme reduced 2,6-dichlorophenolindophenol (DCIP) and the Km value for DCIP was calculated to be 1.4 .times. 10-4 M. The DCIP reductase activity was inhibited by carbonyl reagents like semicarbazide, hydrazine, hydroxylamine and 1,4-benzoquinone. 1,4-Benzoquinone inhibited the DCIP reductase activity competitively as to DCIP.