Thermodynamic parameters of specific and nonspecific protein-DNA binding

Proteins that bind preferentially to specific recognition sites on DNA also bind more weakly to nonspecific DNA. We have studied both specific and non -specific binding of the EcoRI and BamHI restriction endonucleases, and det ermined enthalpic and entropic contributions to binding free energy (DeltaG degrees (bind)) using both the van't Hoff method and isothermal titration calorimetry. Specific binding is characterized by a strongly negative Delta C degrees (p) and can be either enthalpy-driven or entropy-driven, dependin g on temperature. Nonspecific binding has DeltaC degrees (p) approximate to 0 and is enthalpy-driven. A strongly negative DeltaC degrees (p) is the "t hermodynamic signature" of site-specific binding, because it reflects the c haracteristics of a tight complementary recognition interface: the burial o f previously hydrated nonpolar surface and restriction of configurational-v ibrational freedoms of protein, DNA, and water molecules trapped at the pro tein-DNA interface. These factors are absent in nonspecific complexes. We p robed the contributions to DeltaC degrees (p) by varying the sequence conte xt surrounding the recognition site. As DeltaG degrees (bind) improves, Del taC degrees (p), DeltaH degrees and DeltaS degrees all become more negative , and there is a linear correlation between DeltaH degrees and DeltaS degre es (enthalpy-entropy compensation). Because these context variations do not change the protein-base or protein-phosphate contacts, the hydrophobic con tribution or the number of trapped water molecules at the interface, we con clude that a better sequence context improves the "goodness of fit" in the interface and and thus increases the magnitude of the negative configuratio nal-vibrational contribution to DeltaC degrees (p).