Genetic and biochemical characterization of a shortâchain alcohol dehydrogenase from the hyperthermophilic archaeon Pyrococcus furiosus
2001
Oost, John van der | Voorhorst, Wilfried G. B. | Kengen, Servé W. M. | Geerling, Ans C. M. | Wittenhorst, Vincent | Gueguen, Yannick | Vos, Willem M. de
The gene encoding a shortâchain alcohol dehydrogenase, AdhA, has been identified in the hyperthermophilic archaeon Pyrococcus furiosus, as part of an operon that encodes two glycosyl hydrolases, the βâglucosidase 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 26âkDa subunits. AdhA is an NADPHâdependent oxidoreductase that converts alcohols to the corresponding aldehydes/ketones and vice versa, with a rather broad substrate specificity. Maximal specific activities were observed with 2âpentanol (46âU·mg−1) and pyruvaldehyde (32âU·mg−1) in the oxidative and reductive reaction, respectively. AdhA has an optimal activity at 90â°C, at which temperature it has a half life of 22.5âh. The expression of the adhA gene in P.âfuriosus 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.
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