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Orthric Rieske dioxygenases for degrading mixtures of 2,4-dinitrotoluene/naphthalene and 2-amino-4,6-dinitrotoluene/4-amino-2,6-dinitrotoluene

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Abstract

Pollutants are frequently found as mixtures yet it is difficult to engineer enzymes with broad substrate ranges on aromatics. Inspired by the archetypal nitroarene dioxygenase, which shares its electron transport with a salicylate monooxygenase, we have created an innovative and general approach to expand the substrate range of dioxygenase enzymes in a single cell. We have developed here a series of novel, hybrid dioxygenase enzymes that function with a single ferredoxin reductase and ferredoxin that are used to transport two electrons from nicotinamide adenine dinucleotide to the two independent terminal oxygenases. Each independent alpha-oxygenase may then be used simultaneously to create orthric enzymes that degrade mixtures of environmental pollutants. Specifically, we created a hybrid dioxygenase system consisting of naphthalene dioxygenase/dinitrotoluene dioxygenase to simultaneously degrade 2,4-dinitrotoluene and naphthalene (neither enzyme alone had significant activity on both compounds) and dinitrotoluene dioxygenase/nitrobenzene dioxygenase to simultaneously degrade the frequently encountered 2,4,6-trinitrotoluene reduction products 2-amino-4,6-dinitrotoluene and 4-amino-2,6-dinitrotoluene.

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Acknowledgements

This study was supported by the National Science Foundation (BES-0114126). We thank P. A. Williams for plasmid pWWF6, I. B. Lambert for E. coli JVQ2, J. C. Spain for 4M5NC, 3A4M5NC, and for plasmid pJS765, A. Fishman for her assistance with the HPLC analyses, and M. Thompson for his assistance with the GC-MS analyses.

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  1. Artie McFerrin Department of Chemical Engineering, Texas A and M University, College Station, TX, 77843-3122, USA

    Brendan G. Keenan & Thomas K. Wood

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  1. Brendan G. Keenan
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  2. Thomas K. Wood
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Correspondence to Thomas K. Wood.

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Keenan, B.G., Wood, T.K. Orthric Rieske dioxygenases for degrading mixtures of 2,4-dinitrotoluene/naphthalene and 2-amino-4,6-dinitrotoluene/4-amino-2,6-dinitrotoluene. Appl Microbiol Biotechnol 73, 827–838 (2006). https://doi.org/10.1007/s00253-006-0538-8

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  • DOI: https://doi.org/10.1007/s00253-006-0538-8

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