In vitro selection of integration host factor binding sites.
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.
D001426:Bacterial Proteins / Q000378:metabolism*; D010582:Bacteriophage lambda / Q000235:genetics; D001482:Base Composition; D001483:Base Sequence; D001667:Binding, Competitive; D003001:Cloning, Molecular; D016384:Consensus Sequence / Q000235:genetics; D004247:DNA / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018983:DNA Footprinting; D004268:DNA-Binding Proteins / Q000378:metabolism; D015723:Gene Library; D020128:Inhibitory Concentration 50; D034982:Integration Host Factors; D008024:Ligands; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D011995:Recombination, Genetic / Q000235:genetics; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics*; D017422:Sequence Analysis, DNA