Non-complementary mismatched base pairs locally distort DNA structure, leading to increased DNA-binding by transcription factor proteins

Mismatched base-pairs represent a form of DNA damage frequently formed in living cells. Mismatches alter the local DNA structure, which can affect interactions with DNA-binding proteins, including regulatory transcription-factors (TFs).

Recent studies found that interactions between TFs and damaged DNA may play an important role in mutagenesis. However, DNA damages significantly distort the DNA, and the structural impact of damage-induced and other distorted DNA shapes on protein-DNA recognition has not been well characterized.

We present Saturation Mismatch Binding Assay (SaMBA), a new technique to characterize the effects of mismatches on TF-DNA binding in high throughput. SaMBA generates DNA duplexes containing all possible single-base mismatches and quantitatively assesses the effects of the mismatches on TF-DNA interactions.

We applied SaMBA to measure binding of 21 TFs to thousands of mismatched sequences and mapped the impact of mismatches on these TFs. Remarkably, for all TFs examined, the introduction of mismatches at certain positions resulted in significantly increased binding, with some mismatches creating high-affinity binding sites in nonspecific DNA and some converting known binding sites into “super-sites” stronger than any canonical Watson-Crick site.

Structural analyses revealed that these mismatches are oftentimes distorting the naked DNA such that its structure becomes similar to that of bound DNA sites, thus explaining the increased binding measured in our assay. Our results reveal that the energy cost of deforming the DNA structure is a major determinant of protein-DNA recognition, and reveal mechanisms by which mismatches can recruit TFs and thus modulate replication and repair activities in the cell.