CALORIMETRIC STUDIES OF THE INTERACTIONS OF CYTOCHROME-C WITH DIOLEOYLPHOSPHATIDYLGLYCEROL EXTRUDED VESICLES - IONIC-STRENGTH EFFECTS

Cytochrome c has been studied as an example of a peripheral membrane p rotein which interacts with the lipids as well as the proteins of the inner mitochondrial membrane. In order to elucidate the thermodynamic properties of these interactions, isothermal titration calorimetry and differential scanning calorimetry (DSC) were used to study the bindin g of cytochrome c to negatively charged dioleoylphosphatidylglycerol ( DOPG) extruded vesicles as a function of ionic strength. The binding c onstant and enthalpy of association decrease with increasing ionic str ength, with no binding detected above 0.5 M NaCl. The enthalpy of the binding of cytochrome c to DOPG-extruded vesicles was 15 kcal/mol, and the binding constant was 6.10(6) M(-1) at the lowest ionic strengths. The minimum size of the lipid cluster to which the protein bound was found to be approx. 9 lipid molecules in the titration calorimetry mea surements and as low as 5 lipid molecules in the DSC measurements. The stability of the bound cytochrome c was found to be reduced; the ther mal denaturation temperature was lowered from 83 to 50 degrees when bo und to DOPG. The results of this study support previous suggestions th at cytochrome c may undergo a conformational change when it binds to c harged lipids such as DOPG. The results also support the suggestion th at the protein penetrates partially into the lipid bilayer.