Genetic and biochemical characterization of a short-chain alcohol dehydrogenase from the hyperthermophilic archaeon Pyrococcus furiosus
| Autor: | J, van der Oost, W G, Voorhorst, S W, Kengen, A C, Geerling, V, Wittenhorst, Y, Gueguen, W M, de Vos |
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| Rok vydání: | 2001 |
| Předmět: |
Sequence Homology
Amino Acid Blotting Western Molecular Sequence Data Alcohol Dehydrogenase Temperature Substrate Specificity Pyrococcus furiosus Alcohol Oxidoreductases Kinetics Fermentation Operon Escherichia coli Carbohydrate Metabolism Electrophoresis Polyacrylamide Gel Amino Acid Sequence Protein Structure Quaternary Dimerization Plasmids |
| Zdroj: | European journal of biochemistry. 268(10) |
| ISSN: | 0014-2956 |
| Popis: | The gene encoding a short-chain alcohol dehydrogenase, AdhA, has been identified in the hyperthermophilic archaeon Pyrococcus furiosus, as part of an operon that encodes two glycosyl hydrolases, the beta-glucosidase CelB and the endoglucanase LamA. The adhA gene was functionally expressed in Escherichia coli, and AdhA was subsequently purified to homogeneity. The quaternary structure of AdhA is a dimer of identical 26-kDa subunits. AdhA is an NADPH-dependent oxidoreductase that converts alcohols to the corresponding aldehydes/ketones and vice versa, with a rather broad substrate specificity. Maximal specific activities were observed with 2-pentanol (46 U x mg(-1)) and pyruvaldehyde (32 U x mg(-1)) in the oxidative and reductive reaction, respectively. AdhA has an optimal activity at 90 degrees C, at which temperature it has a half life of 22.5 h. The expression of the adhA gene in P. furiosus was demonstrated by activity measurements and immunoblot analysis of cell extracts. A role of this novel type of archaeal alcohol dehydrogenase in carbohydrate fermentation is discussed. |
| Databáze: | OpenAIRE |
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