The complex between phage 434 repressor DNA-binding domain and operator site O_R3: structural differences between consensus and non-consensus half-sites

Background: The repressor of phage 434 binds to a set of operator sites as a homodimer. Its relative affinities for these sites determine the switch from lysogenic to lytic growth. The six 434 operator sites (O_R1, O_R2, O_R3, O_L1, O_L2 and O_L3) have a particularly simple organization; all are 14 base pairs long, with a conserved 5′-ACAA sequence symmetrically placed at either end, and a variable central six base pairs. O_R3 is unique among naturally-occurring 434 operator sites in that it contains a non-consensus base pair, G·C, at the fourth position of the otherwise invariant 5′-ACAA sequence. Comparisons among structures of the 434 repressor DNA-binding domain, R1-69, bound to various operator sites, allow us to analyze differential specificity in regulatory complexes of this kind. Results: We have determined the structure at 2.5 Å resolution of a complex of R1-69 with DNA containing the O_R3 site and compared it with previously studied complexes of R1-69 bound to O_R1 and O_R2. There are surprisingly extensive structural differences between the consensus and non-consensus half-sites of O_R3 with respect to their interactions with R1-69, including a shift in the DNA backbone and a small rotation of the entire R1-69 monomer. Conclusions: Recognition of the base pair difference that is critical for the 434 regulatory switch involves a number of amino acid residues, not just the one or two side chains in direct contact with the G·C base pair. Moreover, the repressor imposes a somewhat altered DNA conformation on the non-consensus half-site.