Compaction of DNA in an anionic micelle environment followed by assembly into phosphatidylcholine liposomes

A difficult problem concerning the interaction of DNA with amphiphiles of o pposite charge above their critical micelle concentration is the propensity for aggregation of the condensed DNA complexes. In this study, this proble m was addressed by attenuating amphiphile charge density within a cholate m icelle environment. The amphiphile consisted of a cationic peptide, acetyl- CWKKKPKK-amide, conjugated to dilaurylphosphatidylethanolamine. In the pres ence of cholate, multiple equivalents of cationic charge were required to b ring about the completion of DNA condensation. At the end point of condensa tion, stable, soluble DNA-micelle complexes were formed, which by dynamic l ight scattering exhibited apparent hydrodynamic diameters between 30 and 60 nm, Aggregation, as measured by static light scattering at 90 degrees and by turbidity, was not observed until further additions of peptide-lipid con jugate were made beyond the end point of DNA condensation. Liposome complex es containing the non-aggregated, compacted DNA were formed by adding diole oylphosphatidylcholine followed by removing the cholate by dialysis, The re sulting complexes were distributed within a narrow density range, the DNA w as quantitatively assembled into the liposomes, and liposomes without DNA w ere not detected. Small particles were formed with a mean hydrodynamic diam eter of 77 nm, The liposomal DNA showed complete retention of its supercoil ed form and no detectable sensitivity to DNase (25 U/10 mu g DNA, 1.5 h, 37 degrees C). The use of an anionic, dialyzable amphiphile to attenuate char ge interactions between DNA and cationic amphiphiles is a useful technology for the quantitative assembly of compacted DNA into conventional liposomes , with complete protection against nuclease activity.