THERMODYNAMIC CHARACTERIZATION OF THE STRUCTURAL STABILITY OF THE COILED-COIL REGION OF THE BZIP TRANSCRIPTION FACTOR GCN4

The thermal stability of a 56 amino acid fragment of GCN4 has been stu died by high-sensitivity differential scanning calorimetry and circula r dichroism spectroscopy. This fragment contains the leucine zipper an d part of the basic region. The thermal unfolding of GCN4-56 is a reve rsible process and can be well represented by a reaction of the form N 2 <-> 2U, indicating that the unfolding of the leuzine zipper is a two -state process in which the helices are only stable when they are in t he coiled-coil conformation. As expected, the transition temperature i s concentration dependent. At pH 7.06 and a protein concentration of 5 X 10(-4) M the transition temperature is close to 70-degrees-C while at 5 X 10(-6) M it is close to 50-degrees-C. The enthalpy change for u nfolding is 31.5 kcal mol-1 at 70-degrees-C. Since the isolated helice s are unstable, interactions at the interface between the two helices play a key role in the stabilization of the native dimer. These intera ctions primarily involve the burial of apolar surface from the solvent (hydrophobic effect) and electrostatic interactions. Structural therm odynamic calculations have permitted a dissection of the magnitude of the various contributions to the total Gibbs free energy of stabilizat ion.