Abstract
3-Nitotoluene dioxygenase (3-NTDO) is the first enzyme in the degradation pathway of 3-nitrotoluene (3-NT) by Diaphorobacter sp. strain DS2. The complete gene sequences of 3-NTDO were PCR amplified from genomic DNA of Diaphorobacter sp., cloned, sequenced and expressed. The 3-NTDO gene revealed a multi component structure having a reductase, a ferredoxin and two oxygenase subunits. Clones expressing the different subunits were constructed in pET21a expression vector system and overexpressed in E. coli BL21(DE3) host. Each subunit was individually purified separately to homogeneity. The active recombinant enzyme was reconstituted in vitro by mixing all three purified subunits. The reconstituted recombinant enzyme could catalyse biotransformations on a variety of organic aromatics.
Keywords:
3-Nitotoluene dioxygenase (3-NTDO); Biodegradation; Diaphorobacter; Nitrotoluene (NT); Rieske dioxygenase.
Copyright © 2014 Elsevier Inc. All rights reserved.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Bacterial Proteins / genetics
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Bacterial Proteins / metabolism*
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Comamonadaceae / enzymology*
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Comamonadaceae / genetics
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Comamonadaceae / metabolism
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Dinitrobenzenes / chemistry
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Dinitrobenzenes / metabolism
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Dioxygenases / genetics
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Dioxygenases / metabolism*
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Electrophoresis, Polyacrylamide Gel
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Escherichia coli / genetics
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Ferredoxins / genetics
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Ferredoxins / metabolism
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Gene Expression Regulation, Bacterial
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Gene Expression Regulation, Enzymologic
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Hydrocarbons, Aromatic / chemistry
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Hydrocarbons, Aromatic / metabolism
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Molecular Structure
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Oxidoreductases / genetics
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Oxidoreductases / metabolism
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Oxygenases / genetics
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Oxygenases / metabolism
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Protein Subunits / genetics
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Protein Subunits / metabolism
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Recombinant Proteins / isolation & purification
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Recombinant Proteins / metabolism
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Substrate Specificity
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Toluene / analogs & derivatives*
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Toluene / chemistry
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Toluene / metabolism
Substances
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Bacterial Proteins
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Dinitrobenzenes
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Ferredoxins
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Hydrocarbons, Aromatic
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Protein Subunits
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Recombinant Proteins
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3-nitrotoluene
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Toluene
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2,4-dinitrotoluene
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Oxidoreductases
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Oxygenases
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Dioxygenases