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Journal of Bacteriology, March 1999, p. 1719-1727, Vol. 181, No. 6
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Identification and Characterization of a DeoR-Specific Operator Sequence Essential for Induction of dra-nupC-pdp Operon Expression in Bacillus subtilis

Xianmin Zeng* and Hans H. Saxild

Department of Microbiology, Technical University of Denmark, DK-2800 Lyngby, Denmark

Received 8 October 1998/Accepted 9 January 1999

The deoR gene located just upstream the dra-nupC-pdp operon of Bacillus subtilis encodes the DeoR repressor protein that negatively regulates the expression of the operon at the level of transcription. The control region upstream of the operon was mapped by the use of transcriptional lacZ fusions. It was shown that all of the cis-acting elements, which were necessary for full DeoR regulation of the operon, were included in a 141-bp sequence just upstream of dra. The increased copy number of this control region resulted in titration of the DeoR molecules of the cell. By using mutagenic PCR and site-directed mutagenesis techniques, a palindromic sequence located from position -60 to position -43 relative to the transcription start point was identified as a part of the operator site for the binding of DeoR. Furthermore, it was shown that a direct repeat of five nucleotides, which was identical to the 3' half of the palindrome and was located between the -10 and -35 regions of the dra promoter, might function as a half binding site involved in cooperative binding of DeoR to the regulatory region. Binding of DeoR protein to the operator DNA was confirmed by a gel electrophoresis mobility shift assay. Moreover, deoxyribose-5-phosphate was shown to be a likely candidate for the true inducer of the dra-nupC-pdp expression.

* Corresponding author. Mailing address: Department of Microbiology, Technical University of Denmark, Building 301, DK-2800 Lyngby, Denmark. Phone: 45 45 25 22 81. Fax: 45 45 88 26 60. E-mail: imxz{at}pop.dtu.dk.

Journal of Bacteriology, March 1999, p. 1719-1727, Vol. 181, No. 6
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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