Ionic, Structural, and Temperature Effects on DNA Nanoparticles Formed by Natural and Synthetic Polyamines

Vijayanathan, Veena; Lyall, Jasleen; Shirahata, and Akira; Thomas, Thresia

doi:10.1021/bm0493234

Cited by 32 publications

(40 citation statements)

References 65 publications

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“…Similarly, Vijayanathan et al reported that spermine-like polyamines were stabilized in the major groove of DNA. 26) Our molecular dynamics studies also showed that a 600 Da branched PEI binds to 4 base pairs in the major groove of DNA. 32) This corresponds to l=8, which was the parameter used in excluded site model in this study.…”

Section: Discussionsupporting

confidence: 51%

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Low Molecular Weight Branched PEI Binding to Linear DNA

Utsuno

Kono

Tanaka

et al. 2016

CHEMICAL & PHARMACEUTICAL BULLETIN

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Polyethylenimine (PEI) is one of the most versatile non-viral vectors used in gene therapy, especially for delivering plasmid DNA to human cells. However, a good understanding of PEI binding to DNA, the fundamental basis for the functioning of PEI as a vector, has been missing in the literature. In this study, PEI (branched, 600 Da) binding to DNA was examined by isothermal titration calorimetry (ITC), quartz crystal microbalance (QCM) and a complementary set of analysis tools. We demonstrated that a separation between the binding heat and the condensation heat is needed and that the excluded site model should be used for PEI binding stage in the ITC analysis. The equilibrium constant for PEI binding to DNA was determined to be 2.5 10 5 M 1 from the ITC analysis, and as 2.3 10 5 M 1 from the QCM analysis. Additionally, we suggested that the 600 Da branched PEI binds to the major groove of DNA and the rearrangement of PEI on DNA may be difficult to occur because of the small dissociation rate. The binding analysis presented here can be employed to improve our understanding of the functioning of PEI and PEI-like non-viral vectors.Key words DNA condensation; polyethylenimine; isothermal titration calorimetry; quartz crystal microbalance; thermodynamics; kinetics The cationic polyelectrolyte polyethylenimine (PEI) is one of the most versatile polymers used for non-viral gene delivery.1-3) PEI binds to DNA via electrostatic interactions between the positively charged PEI amine groups and the negatively charged nucleic acid phosphate groups in the DNA backbone. PEI, by neutralizing the DNA molecule, facilitates its condensation into particles in the 'nano-meter' range.4) It is believed that PEI's strong transfection ability is due to its effective ability to condense the DNA molecule into a nanoparticle, 5-7) which facilitates its cellular uptake and subsequent intracellular trafficking. PEI is available in a broad range of molecular weights. Higher molecular weight PEIs have high transfection efficiency, but lead to excessive cytotoxicity. Low molecular weight PEIs have demonstrated low toxicity on cells, but they also display low transfection efficiency.5) Recently, highly efficient non-viral vectors have been developed by modifying low molecular weight (600 Da) PEI with a variety of functional groups. [8][9][10][11][12] To better understand PEI interactions with the DNA molecule, we previously determined the thermodynamic parameters of PEI binding to DNA using isothermal titration calorimetry (ITC). 13) In that study, two types of binding modes were found to describe the interactions between PEI and DNA. One type of binding involves PEI binding to the DNA groove, another likely binding mode involves external binding of PEI to the DNA phosphate backbone. Recently, Ketola et al. showed that the equilibrium constant of 25 kDa branched PEI to plasmid DNA (7164 bp) using time-resolved fluorescence spectroscopy was 7.3×10 ). 13)The observed difference may be due to the difference in the mass and architecture of the PEI u...

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Section: Discussionsupporting

confidence: 51%

“…The excitation and emission monochromators were set to 480 nm, and the scattered light was measured at 90° angle with respect to the incident beam. 26) PEI solution was added to 30 μM DNA in TAE buffer at room temperature (25±2°C) so that it became the same N/P ratio as the ITC study and the measurements were repeated.…”

Section: Itc Model Fittingmentioning

confidence: 99%

Low Molecular Weight Branched PEI Binding to Linear DNA

Utsuno

Kono

Tanaka

et al. 2016

CHEMICAL & PHARMACEUTICAL BULLETIN

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“…At the low salt concentrations used here, 10 mM Tris, pH 7.5, the binding of most tetra- and higher valent ions is close to stoichiometric (35). None-the-less, Thomas and coworkers (36, 45) at similar low salt conditions have found that considerably more tetra-lys 4+ was necessary for DNA condensation than spermine 4+ and that significantly more penta-lys 5+ was necessary than several +5 analogs of spermine. Brewer et al (46) reported that hexa-arg 6+ is much more effective than hexa-lys 6+ at condensing DNA.…”

Section: Discussionmentioning

confidence: 98%

A Comparison of DNA Compaction by Arginine and Lysine Peptides: A Physical Basis for Arginine Rich Protamines

2013

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Protamines are small, highly positively charged peptides used to package DNA to very high densities in sperm nuclei. Tight DNA packing is considered essential to minimize DNA damage by mutagens and reactive oxidizing species. A striking and general feature of protamines is the almost exclusive use of arginine over lysine for the positive charge to neutralize DNA. We have investigated whether this preference for arginine might arise from a difference in DNA condensation by arginine and lysine peptides. The forces underlying DNA compaction by arginine, lysine, and ornithine peptides are measured using the osmotic stress technique coupled with x-ray scattering. The equilibrium spacings between DNA helices condensed by lysine and ornithine peptides are significantly larger than the interhelical distances with comparable arginine peptides. The DNA surface-to-surface separation, for example, is some 50% larger with poly-lysine compared to poly-arginine. DNA packing by lysine rich peptides in sperm nuclei would allow much greater accessibility to small molecules that could damage DNA. The larger spacing with lysine peptides is due to both a weaker attraction and a stronger short ranged repulsion relative to the arginine peptides. A previously proposed model for poly-arginine and protamine binding to DNA provides a convenient framework for understanding the differences between the ability of lysine and arginine peptides to assemble DNA.

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“…The ability of polyamines and their analogues to promote conformational changes within DNA and its condensation to nanoparticles can facilitate the transport of oligonucleotides through the cell membrane and be helpful in developing simple carrier systems for gene delivery [ 17 , 21 , 22 , 23 ].…”

Section: Introductionmentioning

confidence: 99%

DNA Interaction Studies of Selected Polyamine Conjugates

Szumilak

Merecz

Strek

et al. 2016

IJMS

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The interaction of polyamine conjugates with DNA double helix has been studied. Binding properties were examined by ethidium bromide (EtBr) displacement and DNA unwinding/topoisomerase I/II (Topo I/II) activity assays, as well as dsDNA thermal stability studies and circular dichroism spectroscopy. Genotoxicity of the compounds was estimated by a comet assay. It has been shown that only compound 2a can interact with dsDNA via an intercalative binding mode as it displaced EtBr from the dsDNA-dye complex, with Kapp = 4.26 × 106 M−1; caused an increase in melting temperature; changed the circular dichroism spectrum of dsDNA; converted relaxed plasmid DNA into a supercoiled molecule in the presence of Topo I and reduced the amount of short oligonucleotide fragments in the comet tail. Furthermore, preliminary theoretical study has shown that interaction of the discussed compounds with dsDNA depends on molecule linker length and charge distribution over terminal aromatic chromophores.

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Ionic, Structural, and Temperature Effects on DNA Nanoparticles Formed by Natural and Synthetic Polyamines

Cited by 32 publications

References 65 publications

Low Molecular Weight Branched PEI Binding to Linear DNA

Low Molecular Weight Branched PEI Binding to Linear DNA

A Comparison of DNA Compaction by Arginine and Lysine Peptides: A Physical Basis for Arginine Rich Protamines

DNA Interaction Studies of Selected Polyamine Conjugates

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