Anatomy of specific interactions between lambda repressor and operator DNA

Recognition of specific DNA sequences by proteins is essential for regulation of gene expression. To fully understand the recognition mechanism, it is necessary to understand not only the structure of the specific protein-DNA interactions but also the energetics. We therefore performed a computer analysis in which a phage DNA-binding protein, lambda repressor, was used to examine the changes in binding free energy (DeltaDeltaG) and its energy components caused by single base mutations. We then determined which of the calculated energy components best correlated with the experimental data. The experimental DeltaDeltaG values were well reproduced by the calculations. Component analysis revealed that the electrostatic and hydrogen bond energies were most strongly correlated with the experimental data. Among the 51 single base-substitution mutants examined, positive DeltaDeltaG values, corresponding to weakened binding, were caused by the loss of favorable electrostatic interactions and hydrogen bonds, the introduction of steric collisions and electrostatic repulsion, the loss of favorable interactions with a thymine methyl group, and the increase of unfavorable hydration energy from isolated DNA. This analysis also showed distinct patterns of recognition at A-T and G-C positions, as different combinations of energy components were involved in DeltaDeltaG caused by the two substitution types. We have thus been able to identify the energy components that most strongly correlate with sequence-dependent DeltaDeltaG and determine their contribution to the specificity of DNA sequence recognition by the lambda repressor. Application of this method to other systems should provide additional insight into the molecular mechanism of protein-DNA recognition.