Interaction of the lambda site-specific recombination protein Xis with attachment site DNA.

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RESUMO

Nuclease protection experiments show that Xis protein of bacteriophage lambda specifically binds attachment (att) site DNA. The region of Xis binding, present in both the phage att site and the right prophage att site, extends from position -102 to position -62 in the P arm. The sequence of this region, the positions of purines protected by Xis against methylation, and the binding of Xis to a resected att site indicate the presence of two binding sites. The postulated recognition elements, contained in 13-base-pair direct repeats separated by 7 base pairs, are situated on the same face of the DNA helix. Protection experiments performed with DNase I suggest that the DNA wraps around (or along the surface of) the bound Xis protein. The Xis binding data presented here establishes that Xis, like the other two proteins involved in lambda site-specific recombination, interacts specifically with att DNA. This rules out that class of models in which the profound effects of Xis on the directionality of site-specific recombination are mediated solely through protein-protein interactions or modification of another protein. In addition, nuclease protection experiments with pairwise combinations of the proteins show that Xis and integration host factor (IHF), or Xis and Int, can bind simultaneously to either the phage or right prophage att sites, and the DNA sequences protected are the sum of those protected with each protein alone. It is therefore unlikely that the effect of Xis on the direction of recombination is exerted by directly blocking the binding of Int or IHF to one or more of their respective binding sites.

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