The interaction of
DNA with a novel cationic
phospholipid transfection
reagent, 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine (
EDOPC), was investigated by monitoring thermal effects, particle size, vesicle
rupture, and
lipid mixing. By isothermal titration calorimetry, the heat of interaction between large unilamellar
EDOPC vesicles and plasmid
DNA was endothermic at both physiological and low ionic strength, although the heat absorbed was slightly larger at the higher ionic strength. The energetic driving force for
DNA-
EDOPC association is thus an increase in entropy, presumably due to release of counterions and water. The estimated minimum entropy gain per released counterion was 1.4 cal/mole- degrees K (about 0.7 kT), consistent with previous theoretical predictions. All experimental approaches revealed significant differences in the
DNA-
lipid particle, depending upon whether complexes were formed by the addition of
DNA to
lipid or vice versa. When
EDOPC vesicles were titrated with
DNA at physiological ionic strength, particle size increased, vesicles ruptured, and
membrane lipids became mixed as the amount of
DNA was added up to a 1.6:1 (+:-) charge ratio. This charge ratio also corresponded to the calorimetric end point. In contrast, when
lipid was added to
DNA, vesicles remained separate and intact until a charge ratio of 1:1 (+:-) was exceeded. Under such conditions, the calorimetric end point was 3:1 (+:-). Thus it is clear that fundamental differences in
DNA-cationic
lipid complexes exist, depending upon their mode of formation. A model is proposed to explain the major differences between these two situations. Significant effects of ionic strength were observed; these are rationalized in terms of the model. The implications of the analysis are that considerable control can be exerted over the structure of the complex by exploiting vectorial preparation methods and manipulating ionic strength.