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- Title
A bacterial chloroform reductive dehalogenase: purification and biochemical characterization.
- Authors
Jugder, Bat‐Erdene; Bohl, Susanne; Lebhar, Helene; Healey, Robert D.; Manefield, Mike; Marquis, Christopher P.; Lee, Matthew
- Abstract
We report herein the purification of a chloroform ( CF)-reducing enzyme, TmrA, from the membrane fraction of a strict anaerobe Dehalobacter sp. strain UNSWDHB to apparent homogeneity with an approximate 23-fold increase in relative purity compared to crude lysate. The membrane fraction obtained by ultracentrifugation was solubilized in Triton X-100 in the presence of glycerol, followed by purification by anion exchange chromatography. The molecular mass of the purified TmrA was determined to be 44.5 kDa by SDS- PAGE and MALDI-TOF/TOF. The purified dehalogenase reductively dechlorinated CF to dichloromethane in vitro with reduced methyl viologen as the electron donor at a specific activity of (1.27 ± 0.04) × 103 units mg protein−1. The optimum temperature and pH for the activity were 45°C and 7.2, respectively. The UV-visible spectrometric analysis indicated the presence of a corrinoid and two [4Fe-4S] clusters, predicted from the amino acid sequence. This is the first report of the production, purification and biochemical characterization of a CF reductive dehalogenase.
- Subjects
CHLOROFORM; DEHALOGENASES; HOMOGENEITY; ULTRACENTRIFUGATION; MOLECULAR weights
- Publication
Microbial Biotechnology, 2017, Vol 10, Issue 6, p1640
- ISSN
1751-7907
- Publication type
Article
- DOI
10.1111/1751-7915.12745