Purification and properties of D-gluconate dehydratase fromAchromobacter

d-Gluconate dehydratase fromAchromobacter, grown ond-gluconate, was purified 100-fold by a procedure involving ammonium sulfate fractionation and preparative acrylamide gel electrophoresis. The purified enzyme appeared to be homogeneous by disc gel electrophoresis. It is an inducible enzyme with an optimal activity in the pH region 8.4–8.8, a Km value of 2.08 × 10^–2 m ford-gluconate and a molecular weight of 270,000 ± 25,000. Only C₅ and C₆ aldonic acids possessing al-threo configuration at C₂ and C₃ are dehydrated. The dehydration products ofd-gluconate,d-xylonate,d-galactonate,d-fuconate andl-arabonate were identified as 2-keto-3-deoxy compounds by specific colour reactions and thin layer chromatography. Onemm Mg^{+ +} is a powerful activator, 0.1 mm Mn^{+ +} activates poorly and EDTA inhibits. Glutatione, dithiothreitol and mercaptoethanol had no effect, althoughp-chloromercuribenzoate (0.01 mm) decreased enzyme activity.We wish to thank Mr D. Dewettinck for skilful technical assistance. The senior author (J.D.L.) is indebted to the Fonds voor Kollektief en Fundamenteel Onderzoek (Belgium) for research and personnel grants. J.K.-M. is indebted to the Belgian government for a travel and study grant.