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Directed evolution of aniline dioxygenase for enhanced bioremediation of aromatic amines

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Abstract

The objective of this study was to enhance the activity of aniline dioxygenase (AtdA), a multi-component Rieske non-heme iron dioxygenase enzyme isolated from Acinetobacter sp. strain YAA, so as to create an enhanced biocatalyst for the bioremediation of aromatic amines. Previously, the mutation V205A was found to widen the substrate specificity of AtdA to accept 2-isopropylaniline (2IPA) for which the wild-type enzyme has no activity (Ang EL, Obbard JP, Zhao HM, FEBS J, 274:928–939, 2007). Using mutant V205A as the parent and applying one round of saturation mutagenesis followed by a round of random mutagenesis, the activity of the final mutant, 3-R21, was increased by 8.9-, 98.0-, and 2.0-fold for aniline, 2,4-dimethylaniline (24DMA), and 2-isopropylaniline (2IPA), respectively, over the mutant V205A. In particular, the activity of the mutant 3-R21 for 24DMA, which is a carcinogenic aromatic amine pollutant, was increased by 3.5-fold over the wild-type AtdA, while the AN activity was restored to the wild-type level, thus yielding a mutant aniline dioxygenase with enhanced activity and capable of hydroxylating a wider range of aromatic amines than the wild type.

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Authors and Affiliations

  1. Department of Chemical Engineering and Biomolecular Engineering, University of Illinois at Urbana-Champaign, 600 South Mathews Avenue, Urbana, IL, 61801, USA

    Ee Lui Ang & Huimin Zhao

  2. Center for Biophysics and Computational Biology, University of Illinois at Urbana-Champaign, 600 South Mathews Avenue, Urbana, IL, 61801, USA

    Huimin Zhao

  3. Department of Chemistry, University of Illinois at Urbana-Champaign, 600 South Mathews Avenue, Urbana, IL, 61801, USA

    Huimin Zhao

  4. Department of Chemical and Biomolecular Engineering, National University of Singapore, 21 Lower Kent Ridge Road, Singapore, 119077, Singapore

    Ee Lui Ang

  5. Division of Environmental Science and Engineering, National University of Singapore, 21 Lower Kent Ridge Road, Singapore, 119077, Singapore

    Jeffrey P. Obbard

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  1. Ee Lui Ang
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  2. Jeffrey P. Obbard
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  3. Huimin Zhao
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Correspondence to Huimin Zhao.

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Ang, E.L., Obbard, J.P. & Zhao, H. Directed evolution of aniline dioxygenase for enhanced bioremediation of aromatic amines. Appl Microbiol Biotechnol 81, 1063–1070 (2009). https://doi.org/10.1007/s00253-008-1710-0

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  • DOI: https://doi.org/10.1007/s00253-008-1710-0

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