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J Biol Chem, Vol. 275, Issue 16, 12214-12222, April 21, 2000
From the Department of Molecular Microbiology, Centro de
Investigaciones Biológicas, Consejo Superior de Investigaciones
Científicas, 28006 Madrid, Spain
The expression of the divergently transcribed
paaZ and paaABCDEFGHIJK catabolic operons,
which are responsible for phenylacetic acid (PA) degradation in
Escherichia coli, is driven by the Pz and
Pa promoters, respectively. To study the transcriptional
regulation of the inducible paa catabolic genes, genetic
and biochemical approaches were used. Gel retardation assays showing
that the PaaX regulator binds specifically to the Pa and
Pz promoters were complemented with in vivo
experiments that indicated a PaaX-mediated repression effect on the
expression of Pa-lacZ and Pz-lacZ reporter fusions. The
region within the Pa and Pz promoters that is
protected by the PaaX repressor in DNase I footprinting assays contains a conserved 15-base pair imperfect palindromic sequence motif that was
shown, through mutational analysis, to be indispensable for PaaX
binding and repression. PA-coenzyme A (PA-CoA), but not PA,
specifically inhibited binding of PaaX to the target sequences, thus
confirming the first intermediate of the pathway as the true inducer
and PaaX as the only bacterial regulatory protein described so far that
responds to an aryl-CoA compound. Superimposed in the specific
PaaX-mediated regulation is transcriptional activation by the cAMP
receptor protein and the integration host factor protein. These global
regulators may adjust the transcriptional output from Pa
and Pz promoters to the overall growth status of the cell.
Transcriptional Regulation of the Divergent paa
Catabolic Operons for Phenylacetic Acid Degradation in
Escherichia coli*
,
*
This work was supported in part by Grants AMB97-063-C02-02
from the Comisión Interministerial de Ciencia y
Tecnología and 06M/029/96 from the Comunidad Autónoma de
Madrid, Spain.The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
Recipient of a predoctoral fellowship from the Plan Nacional de
Formación de Personal Investigador-MEC. Present address: Dept. of
Microbiology, the University of Iowa, Iowa City, IA 52242.
§
To whom correspondence should be addressed: Dept. of Molecular
Microbiology, Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, Velázquez, 144, 28006 Madrid, Spain. Tel.: 34-91-5611800; Fax: 34-91-5627518; E-mail: ediaz@cib.csic.es.
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