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Purification and characterization of a dehalogenase from Pseudomonas stutzeri DEH130 isolated from the marine sponge Hymeniacidon perlevis

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Abstract

2-haloacid dehalogenases are enzymes that are capable of degrading 2-haloacid compounds. These enzymes are produced by bacteria, but so far they have only been purified and characterized from terrestrial bacteria. The present study describes the purification and characterization of 2-haloacid dehalogenase from the marine bacterium Pseudomonas stutzeri DEH130. P. Stutzeri DEH130 contained two kinds of 2-haloacid dehalogenase (designated as Dehalogenase I and Dehalogenase II) as detected in the crude cell extract after ammonium sulfate fractionation. Both enzymes appeared to exhibit stereo-specificity with respect to substrate. Dehalogenase I was a 109.9-kDa enzyme that preferentially utilized D-2-chloropropropionate and had optimum activity at pH 7.5. Dehalogenase II, which preferentially utilized L-2-chloropropionate, was further purified by ion-exchange chromatography and gel filtration. Purified Dehalogenase II appeared to be a dimeric enzyme with a subunit of 26.0-kDa. It had maximum activity at pH 10.0 and a temperature of 40 °C. Its activity was not inhibited by DTT and EDTA, but strongly inhibited by Cu2+, Zn2+, and Co2+. The K m and V max for L-2-chloropropionate were 0.3 mM and 23.8 μmol/min/mg, respectively. Its substrate specificity was limited to short chain mono-substituted 2-halocarboxylic acids, with no activity detected toward fluoropropionate and monoiodoacetate. This is the first report on the purification and characterization of 2-haloacid dehalogenase from a marine bacterium.

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Acknowledgments

This work was financially supported by National Basic Research Program of China (973 program, Grant No. 2009CB724700), the Hundred Talent Program of the Chinese Academy of Sciences (A1097), the Knowledge Innovation Program of the Chinese Academy of Sciences (Grant No.KSCX2-YW-G-073).

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Authors and Affiliations

  1. Marine Bioproducts Engineering Group, Dalian Institution of chemical physics, Chinese Academy of Sciences, Zhongshan Road 457, Dalian, 116023, People’s Republic of China

    Jinyou Zhang, Xupeng Cao, Yanjuan Xin & Song Xue

  2. Flinders Centre for Marine Bioproducts Development (FCMB2), Medical Biotechnology, School of Medicine, Flinders University, Adelaide, SA, 5042, Australia

    Wei Zhang

  3. University of Chinese Academy of Sciences, Beijing, 100049, China

    Jinyou Zhang

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  1. Jinyou Zhang
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  2. Xupeng Cao
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  3. Yanjuan Xin
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  4. Song Xue
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  5. Wei Zhang
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Correspondence to Song Xue.

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Zhang, J., Cao, X., Xin, Y. et al. Purification and characterization of a dehalogenase from Pseudomonas stutzeri DEH130 isolated from the marine sponge Hymeniacidon perlevis . World J Microbiol Biotechnol 29, 1791–1799 (2013). https://doi.org/10.1007/s11274-013-1340-2

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