• ~,'ey words." DNA-lipid interaction; Transfection; 1 ipid polymorphism; Membrane ultrastructure Materials and methods Preparation of complexCalf thymus DNA was purchased from Sigma, additionally purified using phenol and chloroform (A26olA2so value being greater than 1.9) and mildly ultrasonicated until the formation of rather homogeneous native fragments with a chain length of about 300-500 bp. The DNA fragments were mixed with a 1 mg/ml suspension of sonicated small unilamellar vesicles of egg PC or DPPC from Avanti in 0.5 mM HEPES solution (pH 7.5) and then a 100 mM stock solution of CaC12 was slowly added with rapid stirring to yield a final Ca 2+ concentration of 20 mM. The mixture was lyophilized and treated with chloroform to disintegrate initially existing membrane bilayer structures and to favour the formation of the inverted phase in organic solvent [19]. After the removal of chloroform under vacuum, the DNA-Ca2+-lipid complex was rehydrated with the initial volume of water. MicrocalorimetryMeasurements were performed using a DASM-4 (Russia) differential scanning microcalorimeter at a heating of rate of 0.25 K/rain. The

DNA-DPPC complexes can be prepared by means of a single step procedure of mixing DNA solution and aqueous lipid dispersion in the presence of calcium ions. Interaction between DPPC and DNA brings about a biphasic shape of melting curves corresponding to the free lipid and the strongly bound one. The amount of the strongly bound lipid is 5 molecules per nucleotide which is close to the size of the first lipid monolayer around DNA molecule.z 1999 Federation of European Biochemical Societies.

The structure of aggregates formed due to DNA interaction with dioleoylphosphatidylcholine (DOPC) vesicles in presence of Ca(2+) and Mg(2+) cations was investigated using synchrotron small-angle X-ray diffraction. For DOPC/DNA=1:1 mol/base and in the range of concentration of the cation(2+) 0-76.5 mM, the diffractograms show the coexistence of two lamellar phases: L(x) phase with repeat distance d(Lx) approximately 8.26-7.39 nm identified as a phase where the DNA strands are intercalated in water layers between adjacent lipid bilayers, and L(DOPC) phase with repeat distance d(DOPC) approximately 6.45-5.65 nm identified as a phase of partially dehydrated DOPC bilayers without any divalent cations and DNA strands. The coexistence of these phases was investigated as a function of DOPC/DNA molar ratio, length of DNA fragments and temperature. If the amount of lipid increases, the fraction of partially dehydrated L(DOPC) phase is limited, depends on the portion of DNA in the sample and also on the length of DNA fragments. Thermal behaviour of DOPC+DNA+Ca(2+) aggregates was investigated in the range 20-80 degrees C. The transversal thermal expansivities of both phases were evaluated.

No abstract

Scanning microcalorimetry has been used to study the high pressure effect on the main transition from the ripple gel P'(beta) phase to the liquid crystal (L(alpha)) phase in DPPC (dipalmitoylphosphatidylcholine). It has been demonstrated that an increase of the pressure by 200 MPa shifts the transition to higher temperatures by 36.4 degrees. The pressure increase does not affect the cooperativity of transition but reduces noticeably its enthalpy. The changes of the molar partial volume, isothermal compressibility as well as volume thermal expansibility during transition in DPPC suspension have been estimated. It has been shown that monovalent ions (Na(+), Cl(-)) in solution slightly affect the main thermodynamic parameters of the transition. Calcium ions significantly decrease distinction in compressibility and thermal expansibility between liquid-crystal and ripple gel phases of lipid suspension, which in its turn reflects less difference in their volume fluctuations.