Purification and characterization of eugenol dehydrogenase from Pseudomonas fluorescens E118 |
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Authors: | Hirotaka Furukawa Marco Wieser Hiroshi Morita Tsuyoshi Sugio T. Nagasawa |
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Affiliation: | (1) Department of Biological Function & Genetic Resources Science, Okayama University, Tsushima Naka, Okayama, Japan 700-11, JP;(2) Department of Biomolecular Science, Faculty of Engineering, Gifu University, Gifu-Yanagido, Japan 501-11 E-mail: tonagasa@apchem.gifu-u.ac.jp Tel. +81-58-293-2647; Fax +81-58-293-2647, JP;(3) Chisso Corporation, Yokohama Research Center, Kanazawa-ku, Yokohama, Japan 236-8605, JP |
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Abstract: | Pseudomonas fluorescens E118 was isolated from soil as an effective eugenol-degrading organism by a screening using eugenol as enrichment substrate. The first enzyme involved in the degradation of eugenol in this organism, eugenol dehydrogenase, was purified after induction by eugenol, and the purity of the enzyme was shown by SDS-PAGE and gel-permeation HLPC. The enzyme is a heterodimer that consists of a 10-kDa cytochrome c and a 58-kDa subunit. The larger subunit presumably contains flavin, suggesting a flavocytochrome c structure and an electron transfer via flavin and cytochrome c during dehydrogenation. The activity of the purified enzyme depended on the addition of a final electron acceptor such as phenazine methosulfate, 2,6-dichlorophenol-indophenol, cytochrome c, or potassium ferricyanide. The enzyme catalyzed the dehydrogenation of three different 4-hydroxybenzylic structures including the conversion of eugenol to coniferyl alcohol, 4-alkylphenols to 1-(4-hydroxyphenyl)alcohols, and 4-hydroxybenzylalcohols to the corresponding aldehydes. The catalytic and structural similarity between this enzyme and a Penicillium vanillyl-alcohol oxidase and 4-alkylphenol methylhydroxylases from several Pseudomonas species is discussed. Received: 17 June 1998 / Accepted: 12 October 1998 |
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Keywords: | Dehydrogenase Eugenol Purification Flavocytochrome c Pseudomonas fluorescens E118 Substrate specificity 4-Alkylphenol 4-Hydroxybenzyl alcohol Electron acceptor |
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