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Journal of Bacteriology, August 1998, p. 3940-3945, Vol. 180, No. 15
0021-9193/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Identification of an Intragenic Ribosome Binding Site That Affects Expression of the uncB Gene of the Escherichia coli Proton-Translocating ATPase (unc) Operon

Sharlene R. Matten,1,dagger Thomas D. Schneider,2 Steven Ringquist,3,Dagger and William S. A. Brusilow1,*

Department of Biochemistry and Molecular Biology, Wayne State University School of Medicine, Detroit, Michigan 482011; Laboratory of Computational and Experimental Biology, National Cancer Institute, Frederick, Maryland 21702-12012; and Department of Molecular, Cellular, and Developmental Biology, University of Colorado, Boulder, Colorado 803093

Received 7 August 1997/Accepted 1 June 1998

The uncB gene codes for the a subunit of the Fo proton channel sector of the Escherichia coli F1 Fo ATPase. Control of expression of uncB appears to be exerted at some step after translational initiation. Sequence analysis by the perceptron matrices (G. D. Stormo, T. D. Schneider, L. Gold, and A. Ehrenfeucht, Nucleic Acids Res. 10:2997-3011, 1982) identified a potential ribosome binding site within the uncB reading frame preceding a five-codon reading frame which is shifted one base relative to the uncB reading frame. Elimination of this binding site by mutagenesis resulted in a four- to fivefold increase in expression of an uncB'-'lacZ fusion gene containing most of uncB. Primer extension inhibition (toeprint) analysis to measure ribosome binding demonstrated that ribosomes could form an initiation complex at this alternative start site. Two fusions of lacZ to the alternative reading frame demonstrated that this site is recognized by ribosomes in vivo. The results suggest that expression of uncB is reduced by translational frameshifting and/or a translational false start at this site within the uncB reading frame.

* Corresponding author. Mailing address: Department of Biochemistry and Molecular Biology, Wayne State University School of Medicine, Scott Hall, 540 E. Canfield Ave., Detroit, MI 48201. Phone: (313) 577-6659. Fax: (313) 577-2765. E-mail: wbrusilow{at}med.wayne.edu.

dagger Present address: U.S. Environmental Protection Agency, Washington, DC 20460.

Dagger Present address: Sidney Kimmel Cancer Center, San Diego, CA 92121.

Journal of Bacteriology, August 1998, p. 3940-3945, Vol. 180, No. 15
0021-9193/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.


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