A Fluorescence Study of the Structure and Accessibility of Plasmid DNA Condensed with Cationic Gene Delivery Vehicles

Abstract

The cationic lipids 1,2‐dioleoyl‐3‐trimethylammonium‐propane and dimethyldioctadecylammonium bromide, with or without the helper lipids 1,2‐dioleoyl‐sn‐glycero‐3‐phosphoethanolamine or cholesterol, and the cationic polymer polyethyleneimine, were compared for their ability to displace fluorescent dyes from DNA. Differences in displacement of the intercalating dyes ethidium bromide and ethidium homodimer correlate with their relative affinities with DNA, with the extent of ethidium homodimer displacement significantly less. Differences in ethidium homodimer and ethidium bromide displacement as a function of the ratio of polycation to DNA and the charge density of the polycation suggest a greater sensitivity of the former to topological changes in condensed DNA. Marked differences in the ability of these cationic delivery systems to displace the minor groove binding dyes 4′,6‐diamidino‐2‐phenylindole and Hoechst 33258 upon interaction with DNA are also apparent, with the majority of Hoechst 33258 remaining bound to DNA. Changes in the spectral properties of Hoechst 33258 were further used to characterize polycation‐induced changes in solvent accessibility of the DNA minor groove. Taken together, these studies demonstrate differences in the interaction of various cationic lipids and polyethyleneimine in terms of regional displacement of dyes, polycation‐induced structural changes in DNA, as well as polycation‐mediated changes in solvent accessibility of the minor groove. The relevance of these studies to current models of the structure and assembly of polycation/DNA complexes are discussed. © 2003 Wiley‐Liss, Inc. and the American Pharmaceutical Association J Pharm Sci 92:1272–1285, 2003