Characterization of sequence-specific DNA binding by the transcription factor Oct-1

The DNA-binding domain of the Oct-1 transcription factor, POU, recognizes a defined DNA sequence known as the octamer element to regulate the expressi on of both general and cell-type-specific genes. The two-part DNA-binding d omain partially encircles the DNA to recognize the eight base pairs of the octamer element. We have characterized the binding of Oct-1/POU to an oct a mer element using isothermal titration calorimetry. As found for other cogn ate protein/DNA complexes, the formation of the Oct-1 POU/DNA complex is as sociated with a large negative heat capacity change, Delta C-p,C-obs. Howev er, the observed change is much greater than expected by empirical relation ships with buried surface area. Supported by data from proteolysis studies on the free and DNA-bound protein, we propose that the discrepancy in heat capacity arises principally from the partial folding of the Oct-1 POU prote in upon complex formation. Formation of the Oct-1 POU/DNA complex is strong ly dependent on ionic strength, and the detailed quantification of this rel ationship suggests that six charged contacts are made between the protein a nd the phosphate groups of the DNA. This agrees with observations from the crystal structure of an Oct-1 POU/DNA complex.