title: Purification and characterization of tetrachloroethene reductive dehalogenase from Dehalospirillum multivorans.
abstract: Tetrachloroethene reductive dehalogenase from the tetrachloroethene-utilizing anaerobe, Dehalospirillum multivorans, was purified approximately 100-fold to apparent homogeneity. The purified dehalogenase catalyzed the reductive dechlorination of tetrachloroethene (PCE) to trichloroethene and of trichloroethene to cis-1,2-dichloroethene with reduced methyl viologen as the electron donor at a specific activity of 2.6 microkatal/mg. The apparent Km values for tetrachloroethene and trichloroethene were 0.20 and 0.24 mM, respectively. The apparent molecular mass of the native enzyme was determined by gel filtration to be 58 kDa. Sodium dodecyl sulfate-gel electrophoresis revealed a single protein band with a molecular mass of 57 kDa. One mol of dehalogenase contained 1.0 mol of corrinoid, 9.8 mol of iron, and 8.0 mol of acid-labile sulfur. The pH optimum was about 8.0. The enzyme had a temperature optimum of 42 degrees C. It was slightly oxygen-sensitive and was thermolabile above 50 degrees C. The dechlorination of PCE was stimulated by ammonium ions. Chlorinated methanes severely inhibited PCE dehalogenase activity.
authors.affiliation: Department of Microbiology, University of Stuttgart, Allmandring 31, D-70550 Stuttgart, Federal Republic of Germany.
authors.forename: A
authors.initials: A
authors.lastname: Neumann
authors.forename: G
authors.initials: G
authors.lastname: Wohlfarth
authors.forename: G
authors.initials: G
authors.lastname: Diekert
mesh_terms: D002384:Catalysis; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D006089:Gram-Negative Anaerobic Bacteria / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D010088:Oxidoreductases / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D013379:Substrate Specificity; D013750:Tetrachloroethylene / Q000378:metabolism
chemical_list: D010088:Oxidoreductases; C098849:tetrachloroethene reductase; D013750:Tetrachloroethylene