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Cloning and Characterization of a 4-Hydroxyphenylacetate 3-Hydroxylase From the Thermophile Geobacillus sp. PA-9

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Abstract

A 4-hydroxyphenylacetic acid (4-HPA) hydroxylase-encoding gene, on a 2.7-kb genomic DNA fragment, was cloned from the thermophile Geobacillus sp. PA-9. The Geobacillus sp. PA-9 4-HPA hydroxylase gene, designated hpaH, encodes a protein of 494 amino acids with a predicted molecular mass of 56.269 Da. The deduced amino-acid sequence of the hpaH gene product displayed <30% amino-acid sequence identity with the larger monooxygenase components of the previously characterized two-component 4-HPA 3-hydroxylases from Escherichia coli W and Klebsiella pneumoniae M5a1. A second oxidoreductase component was not present on the 2.7-kb genomic DNA fragment. The deduced amino-acid sequence of a second C-terminal truncated open reading frame, designated hpaI, exhibited homology to extradiol oxygenases and displayed the highest amino-acid sequence identity (43%) with the 3,4-dihydroxyphenylacetate 2,3-dioxygenase of Arthrobacter globiformis, encoded by mndD. These results, along with catalytic activity observed in crude intracellular extracts prepared from Escherichia coli cells expressing hpaH, is in support of a role for hpaH in the 4-HPA degradative pathway of Geobacillus sp. PA-9.

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Acknowledgments

This work was supported through grants provided by the International Foundation for Science, the Council for Development of Social Science Research in Africa and by the Makerere University Staff Development Fund.

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Authors and Affiliations

  1. Department of Microbiology and Plant Pathology, University of Pretoria, 0002, Pretoria, South Africa

    J. F. Hawumba, V. S. Brözel & J. Theron

  2. Biochemistry Department, Makerere University, P. O. Box 7062, Kampala, Uganda

    J. F. Hawumba

  3. Department of Biology and Microbiology, South Dakota State University, Brookings, SD, 57007, USA

    V. S. Brözel

Authors
  1. J. F. Hawumba
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  2. V. S. Brözel
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  3. J. Theron
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Correspondence to J. Theron.

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Hawumba, J.F., Brözel, V.S. & Theron, J. Cloning and Characterization of a 4-Hydroxyphenylacetate 3-Hydroxylase From the Thermophile Geobacillus sp. PA-9. Curr Microbiol 55, 480–484 (2007). https://doi.org/10.1007/s00284-007-9016-5

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  • DOI: https://doi.org/10.1007/s00284-007-9016-5

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