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Journal of Bacteriology, April 2002, p. 1988-1997, Vol. 184, No. 7
0021-9193/02/$04.00+0     DOI: 10.1128/JB.184.7.1988-1997.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Differential DNA Binding of Transcriptional Regulator PcaU from Acinetobacter sp. Strain ADP1

Roland Popp, Tobias Kohl,{dagger}, Patricia Patz, Gaby Trautwein, and Ulrike Gerischer*

Mikrobiologie und Biotechnologie, Universität Ulm, D-89069 Ulm, Germany

Received 28 August 2001/ Accepted 4 January 2002

Transcriptional regulator PcaU from Acinetobacter sp. strain ADP1 governs expression of genes for protocatechuate degradation (pca genes) as a repressor or an activator depending on the levels of the inducer protocatechuate and of its own gene. PcaU is a member of the IclR protein family. Here the DNA binding properties of the purified protein are described in terms of the location of the binding sites and the affinity to these sites. Native PcaU was purified after overexpression of the pcaU gene in Escherichia coli. It is a dimer in solution. The binding site in the pcaU-pcaI intergenic region is located between the two divergent promoters covering 45 bp, which includes three perfect 10-bp repetitions. A PcaU binding site downstream of pcaU is covered by PcaU across two palindromic sequence repetitions. The affinity of PcaU for the intergenic binding sites is 50-fold higher (dissociation constant [Kd], 0.16 nM) than the affinity for the site downstream of pcaU (Kd, 8 nM). The binding of PcaU was tested after modifications of the intergenic binding site. Removal of any external sequence repetition still allowed for specific binding of PcaU, but the affinity was significantly reduced, suggesting an important role for all three sequence repetitions in gene expression. The involvement of DNA bending in the regulatory process is suggested by the observed strong intrinsic curvature displayed by the pcaU-pcaI intergenic DNA.

* Corresponding author. Mailing address: Mikrobiologie und Biotechnologie, Universität Ulm, D-89069 Ulm, Germany. Phone: 49-731-502-2715. Fax: 49-731-502-2719. E-mail: ulrike.gerischer{at}biologie.uni-ulm.de.

{dagger} Present address: Max-Planck-Institut für Biophysikalische Chemie, D-37077 Göttingen, Germany.

Journal of Bacteriology, April 2002, p. 1988-1997, Vol. 184, No. 7
0021-9193/02/$04.00+0     DOI: 10.1128/JB.184.7.1988-1997.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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