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.
Highlights
Dehalospirillum multivorans is a strictly anaerobic, Gramnegative bacterium, which is able to grow with tetrachloroethene (PCE)1 as terminal electron acceptor for the oxidation of different electron donors [1, 2]
Purification of Tetrachloroethene Dehalogenase —The enzyme was purified from D. multivorans grown on pyruvate and fumarate in the presence of yeast extract
The specific activity of the purified PCE dehalogenase was determined with methyl viologen as electron donor to be about 2.6 microkatal/mg with a yield of nearly 64%
Summary
It was slightly oxygen-sensitive and was thermolabile above 50 °C. The dechlorination of PCE was stimulated by ammonium ions. Chlorinated methanes severely inhibited PCE dehalogenase activity. Dehalospirillum multivorans is a strictly anaerobic, Gramnegative bacterium, which is able to grow with tetrachloroethene (PCE) as terminal electron acceptor for the oxidation of different electron donors [1, 2]. The bacterium reduces tetrachloroethene via trichloroethene (TCE) to cis-1,2-dichlo-
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