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Journal of Bacteriology, May 1999, p. 3246-3255, Vol. 181, No. 10
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

In Vitro Selection of Integration Host Factor Binding Sites

Steven D. Goodman,1,* Nerissa J. Velten,1 Qian Gao,1 Scott Robinson,2 and Anca M. Segall2

Department of Basic Sciences, University of Southern California School of Dentistry, Los Angeles, California,1 and Department of Biology, San Diego State University, San Diego, California 92182-46142

Received 24 September 1998/Accepted 9 March 1999

Integration host factor (IHF) is a bacterial protein that binds and severely bends a specific DNA target. IHF binding sites are approximately 30 to 35 bp long and are apparently divided into two domains. While the 3' domain is conserved, the 5' domain is degenerate but is typically AT rich. As a result of physical constraints that IHF must impose on DNA in order to bind, it is believed that this 5' domain must possess structural characteristics conducive for both binding and bending with little regard for specific contacts between the protein and the DNA. We have examined the sequence requirements of the 5' binding domain of the IHF binding target. Using a SELEX procedure, we randomized and selected variants of a natural IHF site. We then analyzed these variants to determine how the 5' binding domain affects the structure, affinity, and function of an IHF-DNA complex in a native system. Despite finding individual sequences that varied over 100-fold in affinity for IHF, we found no apparent correlation between affinity and function.

* Corresponding author. Mailing address: Department of Basic Sciences, University of Southern California School of Dentistry, 925 West 34th St., Los Angeles, CA 90089. Phone: (213) 740-3867. Fax: (213) 740-7560. E-mail: sgoodman{at}hsc.usc.edu.

Journal of Bacteriology, May 1999, p. 3246-3255, Vol. 181, No. 10
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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