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J. Bacteriol. doi:10.1128/JB.01379-07
Copyright (c) 2007, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Transcriptome analysis of a phenol producing Pseudomonas putida S12 construct: Genetic and physiological basis for improved production

TNO Quality of Life, Department of Bioconversion. Julianalaan 67, 2628 BC, Delft, The Netherlands; Kluyver Centre for Genomics of Industrial Fermentation. PO box 5057, 2600 GB, Delft, The Netherlands; Delft University of Technology, Department of Biotechnology. Julianalaan 67, 2628 BC, Delft, The Netherlands

^* To whom correspondence should be addressed. Email: nick.wierckx{at}tno.nl.

	Abstract

The unknown genetic basis for improved phenol production by a recombinant Pseudomonas putida S12 derivative bearing the tpl (tyrosine-phenol lyase) gene was investigated via comparative transcriptomics, nucleotide sequence analysis and targeted gene disruption. We show upregulation of tyrosine biosynthetic genes, and the possibly decreased biosynthesis of tryptophan caused by a mutation in the trpE gene, as the genetic basis for the enhanced phenol production. In addition, several genes in degradation routes connected to the tyrosine biosynthetic pathway were upregulated. This may either be a side effect that negatively affects phenol production or it may point to intracellular accumulation of tyrosine or its intermediates. A number of genes identified by the transcriptome analysis were selected for targeted disruption in P. putida S12TPL3. Physiological and biochemical examination of P. putida S12TPL3 and these mutants led to the conclusion that the metabolic flux toward tyrosine in P. putida S12TPL3 was improved to such an extent that the heterologous TPL enzyme has become the rate-limiting step in phenol biosynthesis.