NF-kappa B p65 (RelA) homodimer uses distinct mechanisms to recognize DNA targets

Background: The NF-kappa B family of dimeric transcription factors regulate s the expression of several genes by binding to a variety of related DNA se quences. One of these dimers, p65(RelA), regulates a subclass of these targ ets. We have shown previously that p65 binds to the 5'-GGAA T TTTC-3' seque nce asymmetrically. In that complex one subunit base specifically interacts with the preferred 5' half site and the other subunit binds non-specifical ly to the 3' half site. Results: Here we describe the crystal structures of two new p65-DNA complex es. One complex contains a pseudosymmetric 5'-GGAA T TTCC-3' DNA sequence t aken from the enhancer of the gene encoding interleukin 8 (IL-8) and the ot her contains the asymmetric 5'-GGAA T TCCC-3' target DNA taken from the enh ancer of the gene encoding type VH collagen. As expected, the global positi oning of the dimer on both DNA targets is roughly symmetric, however, the h ydrogen-bonding patterns at the protein-DNA interfaces differ significantly . One of the p65 monomers in complex with the asymmetric DNA binds to an ex tra base pair located immediately upstream of the 5'-GGAA-3' half site. We also show that p65 binds to these targets with almost equal affinity and th at different residues have variable roles in binding different kappa B targ ets. Conclusions: Taken together, these structures reveal that p65 exhibits the unique capability to specifically bind DNA targets of variable lengths from four to ten base pairs. Also, the small protein segment Arg41-Ser42-Ala43 is at least partially responsible for flexibility in DNA-binding modes.