Characterization of a Lipophilic Plasmid DNA Condensate Formed with a Cationic Peptide Fatty Acid Conjugate

Abstract

In the presence of cationic ligands, DNA molecules can become aggregated into larger particles in a process known as condensation. DNA condensates are of interest as models for the dense packing found in naturally occurring structures such as phage heads and chromatin. They have found extensive application in DNA transfection and also provide convenient models with which to study DNA damage by the direct effect of ionizing radiation. Further, conjugates of cationic peptides with fatty acids may represent a class of attractive ligands for these areas because of their simple synthesis. When plasmid pUC18 is used as the DNA target and N-caproyl-penta-arginine amide (Cap-R(5)-NH(2)) is used as the ligand, the physical properties of the resulting mixtures were characterized using static and dynamic light scattering, sedimentation, dye exclusion, circular dichroism, nanoparticle tracking, and atomic force microscopy. Their chemical properties were assayed using solvent extraction and protection against hydroxyl radical attack and nuclease digestion. Titration of the plasmid with the Cap-R(5)-NH(2) ligand produced sharply defined changes in both chemical and physical properties, which was associated with the formation of condensed DNA particles in the 100-2000 nm size range. The caproyl group at the ligand's N-terminus produced a large increase in the partitioning of the resulting condensate from water into chloroform and in its binding to the neutral detergent Pluronic F-127. Both the physical and chemical data were all consistent with condensation of the plasmid by the ligand where the presence in the ligand of the caproyl group conferred an extensive lipophilic character upon the condensate.

Biodegradable Polymers for Gene Delivery

“…These polymers interact with DNA and provoke the formation of compact nano-sized polyplexes [12]. The charge neutralized compact polyplex core protects enclosed nucleic acids from nucleases, and hence, maintain their stability and integrity until the cellular uptake process occurs [39,93,94]. Although PEI is a non-biodegradable polymer, it is included herein because it represents a polymer showing high transfection efficiency and could be made biodegradable by appropriate modifications.…”

Supramolecular Langmuir monolayers and multilayered vesicles of self-assembling DNA–lipid surface structures and their further implications in polyelectrolyte-based cell transfections

“…4a). The addition of increasing concentration of DPPC vesicles drive these bands in the negative directions but their locations remain unchanged, suggesting that DNA condensation takes place in a similar manner as observed with plasmids interacting with cationic peptide ligand conjugates (Do et al 2011). To confirm the importance of the lipid charge for membrane interactions of T7 DNA and to study the potential changes of the lipid secondary structures upon binding we have performed CD experiments depicting lipid spectral signals (Fig.…”

“…5). This type of polynucleotide helix possesses approximately equal positive (275 nm) and negative (245 nm) components centered at around 260 nm (Zuidam et al 1999;Ranjbar and Gill 2009;Do et al 2011;Chang et al 2012). At 220 nm the CD signal is either positive or less negative.…”

Peptide nucleic acid-ionic self-complementary peptide conjugates: highly efficient DNA condensers with specific condensing mechanism

“…The ethidium bromide (EtBr) displacement assay was further used to assess the relative binding affinity of various condensation agents to DNA. [28][29][30][31] Upon mixing EtBr with DNA, the uorescence of EtBr enhances dramatically, as a result of its intercalation into DNA. The binding of condensation agents to DNA usually displaces the intercalated EtBr and causes a decrease in the uorescence emission intensity of EtBr at 610 nm.…”