This study investigates the genetic architecture of a transcription factor's specificity for DNA, using a new method based on ordinal logistic regression to analyze 20-state combinatorial deep mutational scanning (DMS) data. The researchers found that the genetic architecture of DNA recognition is characterized by a dense set of main and pairwise effects involving almost every possible amino acid state in the protein-DNA interface, but higher-order epistasis plays a minor role. Pairwise interactions significantly expand the set of functional sequences and are the primary determinants of specificity for different DNA elements. These interactions also increase the number of opportunities for single-residue mutations to switch specificity between different DNA targets. By bringing variants with different functions close together in sequence space, pairwise epistasis facilitates the evolution of new functions rather than constraining it. The study provides compelling evidence that changes in function are largely due to pairwise interactions, and that epistasis potentiates rather than constrains evolutionary paths.This study investigates the genetic architecture of a transcription factor's specificity for DNA, using a new method based on ordinal logistic regression to analyze 20-state combinatorial deep mutational scanning (DMS) data. The researchers found that the genetic architecture of DNA recognition is characterized by a dense set of main and pairwise effects involving almost every possible amino acid state in the protein-DNA interface, but higher-order epistasis plays a minor role. Pairwise interactions significantly expand the set of functional sequences and are the primary determinants of specificity for different DNA elements. These interactions also increase the number of opportunities for single-residue mutations to switch specificity between different DNA targets. By bringing variants with different functions close together in sequence space, pairwise epistasis facilitates the evolution of new functions rather than constraining it. The study provides compelling evidence that changes in function are largely due to pairwise interactions, and that epistasis potentiates rather than constrains evolutionary paths.