Indirect readout of DNA sequence at the primary-kink site in the CAP-DNA complex: Recognition of pyrimidine-purine and purine-purine steps

The catabolite activator protein (CAP) bends DNA in the CAP-DNA complex, typically introducing a sharp DNA kink, with a roll angle of similar to 40 degrees and a twist angle of similar to 20 degrees, between positions 6 and 7 of the DNA half-site, 5'-A(1)A(2)A(3)T(4)G(5)T(6)G(7)A(8)T(9)C(10)T(11)-3' ("primary kink").In previous wor, we showed that CAP recognizes the nucleotide immediatly 5' to the primary-kink site, T-6, through an "indirect-readout" mechanism involving sequence effects on energetics of primary-kink formation. Here, to understand further this example of indirect readout, we have determined crystal structures of CAP-DNA complexes containing each possible nucleotide at position 6. The structures show that CAP can introduce a DNA kink at the primary-kink site with any nucleotide at position 6. The DNA kink is sharp with the consensus pyrimidine-purine step T(6)G(7) and the non-consensus pyrimidine-purine step C(6)G(7) (roll angles of similar to 42 degrees, twist angles of similar to 16 degrees), but is much less sharp with the non-consensun purine-purine steps A(6)G(7) and G(6)G(7) (roll angles of similar to 20 degrees, twist angles of similar to 17 degrees). We infer that CAP discriminates between consensus and non-consensus pyrimidine-purine steps at positions 6-7 solely based on differences in the energetics of DNA deformation, but that CAP discriminates between the consensus pyrimidine-purine step and non-consensus purine-purine steps at positions 6-7 both based on differences in the energetics of DNA deformation and based on qualitative differences in DNA deformation. The structures further show that CAP can achieve a similar, similar to 46 degrees per DNA half-site, overall DNA bend through a sharp DNA kink, a less sharp DNA kink, or a smooth DNA bend. Analysis of these and other crystal structures of CAP-DNA complexes indicates that there is a large, similar to 28 degrees per DNA half-site, out-of-plane component of CAP-induced DNA bending in structures not constrained by end-to-end DNA lattice interactions and that lattice contacts involving CAP tend to involve residues in or near biologically functional surfaces. (c) 2005 Elsevier Ltd. All rights reserved.