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A new method for evaluating the specificity of indirect readout in protein-DNA recognition.

ABSTRACT: Proteins recognize a specific DNA sequence not only through direct contact (direct readout) with base pairs but also through sequence-dependent conformation and/or flexibility of DNA (indirect readout). However, it is difficult to assess the contribution of indirect readout to the sequence specificity. What is needed is a straightforward method for quantifying its contributions to specificity. Using Bayesian statistics, we derived the probability of a particular sequence for a given DNA structure from the trajectories of molecular dynamics (MD) simulations of DNAs containing all possible tetramer sequences. Then, we quantified the specificity of indirect readout based on the information entropy associated with the probability. We tested this method with known structures of protein-DNA complexes. This method enabled us to correctly predict those regions where experiments suggested the involvement of indirect readout. The results also indicated new regions where the indirect readout mechanism makes major contributions to the recognition. The present method can be used to estimate the contribution of indirect readout without approximations to the distributions in the conformational ensembles of DNA, and would serve as a powerful tool to study the mechanism of protein-DNA recognition.

SUBMITTER: Yamasaki S

PROVIDER: S-EPMC3458528 | biostudies-literature | 2012 Sep

REPOSITORIES: biostudies-literature

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Project description:The catabolite activator protein (CAP) bends DNA in the CAP-DNA complex, typically introducing a sharp DNA kink, with a roll angle of approximately 40 degrees and a twist angle of approximately 20 degrees, between positions 6 and 7 of the DNA half-site, 5'-A1A2A3T4G5T6G7A8T9C10T11 -3' ("primary kink"). In previous work, we showed that CAP recognizes the nucleotide immediately 5' to the primary-kink site, T6, 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 T6G7 and the non-consensus pyrimidine-purine step C6G7 (roll angles of approximately 42 degrees, twist angles of approximately 16 degrees ), but is much less sharp with the non-consensus purine-purine steps A6G7 and G6G7 (roll angles of approximately 20 degrees, twist angles of approximately 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, approximately 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, approximately 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.

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A new method for evaluating the specificity of indirect readout in protein-DNA recognition.

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