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Applied and Environmental Microbiology, March 2004, p. 1749-1757, Vol. 70, No. 3
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.3.1749-1757.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Elucidation of the Antibacterial Mechanism of the Curvularia Haloperoxidase System by DNA Microarray Profiling

Eva H. Hansen,^1,2 Mark A. Schembri,^3* Per Klemm,³ Thomas Schäfer,¹ Søren Molin,³ and Lone Gram²

Novozymes A/S, DK-2880 Bagsværd,¹ Department of Seafood Research, Danish Institute for Fisheries Research,² Centre for Biomedical Microbiology, BioCentrum-DTU, Technical University of Denmark, DK-2800 Kgs. Lyngby, Denmark³

Received 3 October 2003/ Accepted 26 November 2003

A novel antimicrobial enzyme system, the Curvularia haloperoxidase system, was examined with the aim of elucidating its mechanism of antibacterial action. Escherichia coli strain MG1655 was stressed with sublethal concentrations of the enzyme system, causing a temporary arrest of growth. The expression of genes altered upon exposure to the Curvularia haloperoxidase system was analyzed by using DNA microarrays. Only a limited number of genes were involved in the response to the Curvularia haloperoxidase system. Among the induced genes were the ibpA and ibpB genes encoding small heat shock proteins, a gene cluster of six genes (b0301-b0306) of unknown function, and finally, cpxP, a member of the Cpx pathway. Knockout mutants were constructed with deletions in b0301-b0306, cpxP, and cpxARP, respectively. Only the mutant lacking cpxARP was significantly more sensitive to the enzyme system than was the wild type. Our results demonstrate that DNA microarray technology cannot be used as the only technique to investigate the mechanisms of action of new antimicrobial compounds. However, by combining DNA microarray analysis with the subsequent creation of knockout mutants, we were able to pinpoint one of the specific responses of E. coli—namely, the Cpx pathway, which is important for managing the stress response from the Curvularia haloperoxidase system.

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* Corresponding author. Mailing address: Microbial Adhesion Group, Centre for Biomedical Microbiology, BioCentrum-DTU, Bldg. 301, Technical University of Denmark, DK-2800 Kgs. Lyngby, Denmark. Phone: (45) 45252519. Fax: (45) 45932809. E-mail: msc{at}biocentrum.dtu.dk.

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Applied and Environmental Microbiology, March 2004, p. 1749-1757, Vol. 70, No. 3
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.3.1749-1757.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

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