The Structural Organization of Cationic Lipid-DNA Complexes

Abstract

The interaction of cationic liposomes with supercoiled plasmid DNA results in a major rearrangement of each component to form compact multilamellar structures comprised of alternating layers of two-dimensional arrays of DNA sandwiched between lipid bilayers. Fluorescence resonance energy transfer was used to estimate the distance of closest approach of DNA to the lipid bilayers in these complexes. The effect of several compositional variables on this distance, including the ratio of cationic lipid to DNA, and the charge density, intrinsic curvature, and fluidity of the lipid bilayer were examined. Additionally, the effect of ionic strength was studied. For complexes prepared at or above a 3:1 charge ratio (؉/؊), the observed distance of closest approach was found to be in agreement with the intercalation of DNA between lipid bilayers. As the charge ratio was decreased, a monotonic increase in the distance was observed with a maximum observed at 0.5:1. Correlations between differences in the proximity of DNA to the lipid bilayer and the hydrodynamic size of the complexes were also found. A model based on these observations and previous reports suggests the formation of discrete populations of complexes below a charge ratio of 0.5:1 and above 3:1. The structure of the negatively charged complexes is consistent with DNA extending from the surface of the particles, whereas those possessing excess positive charge were multilamellar aggregates with the DNA effectively condensed between lipid bilayers. Complexes between these two states consist of weighted fractions of these two species.

Voltammetric behaviour of oligonucleotide lipoplexes adsorbed onto glassy carbon electrodes

“…The results are in agreement with spectroscopic and enzymatic data, reported recently, in which the detection of partial denaturation in plasmidic DNA lipoplexes by increased hyperchromicity and nuclease susceptibility [20], and other methodologies was described [6][7][8][9][10]17]. They strongly suggest that the ODN composition can affect both the electrostatic and aggregation processes that occur during an ODN cationic liposome complexation event, and consequently can influence the efficiency of lipoplex transfection capacity.…”

“…It was reported that DOTAP cationic liposomes form more stable lipoplexes with relaxed DNA structures when plasmidic DNA has been used [20] and some similar features of the lipoplex generation driving force may be connected with the present work.…”

“…Recent literature reported that double strand DNA suffers conformational transitions [18][19][20][21] and denaturation can occur [22] when mixed with cationic liposomes to generate lipoplexes of low (+/)) charge ratio. By voltammetry, it is possible to distinguish between single strand and double helix DNA [14,15] since the peaks due to guanosine and adenosine nucleotide oxidation in DNA are comparably much higher and well defined for the single strand.…”

Analysis of Cationic-Lipid?Plasmid-DNA Complexes

“…DLS studies have identified CL-DNA particles in the size range 50-300 nm and confirmed their intrinsic heterogeneity (23)(24)(25). Their  potentials range from the highly negative (an excess of DNA) to the highly positive (an excess of CL).…”

“…In the case of lipoplexes, it has been possible to label the DNA by either intercalation of a dye between the bases or binding of a dye to the minor groove (donors). Then, a different dye can be placed within the bilayer of the CLs (acceptors); this makes it possible to estimate the mean distance between various sites within the complexes (23,42). Although they have only been explored to a limited extent, versions of this approach may turn out to be quite useful.…”