Skip to main content
PMC home page
Biophysical Journal logoLink to Biophysical Journal
. 2002 Jul;83(1):556–565. doi: 10.1016/S0006-3495(02)75191-6

Thermodynamics of cationic lipid-DNA complex formation as studied by isothermal titration calorimetry.

Edwin Pozharski 1, Robert C MacDonald 1
PMCID: PMC1302169  PMID: 12080142

Abstract

The detailed analysis of the cationic lipid-DNA complex formation by means of isothermal titration calorimetry is presented. Most experiments were done using 1,2-dioleyl-sn-glycero-3-ethylphosphocholine (EDOPC), but basic titrations were also done using DOTAP, DOTAP:DOPC, and DOTAP:DOPE mixtures. Complex formation was endothermic with less than 1 kcal absorbed per mole of lipid or DNA charge. This enthalpy change was attributed to DNA-DNA mutual repulsion within the lamellar complex. The exception was DOTAP:DOPE-containing lipoplex for which the enthalpy of formation was exothermic, presumably because of DOPE amine group protonation. Experimental conditions, namely, direction and titration increment as well as concentration of titrant, which dictate the structure of resulting lipoplex (whether lamellar complex or DNA-coated vesicle), were found to affect the apparent thermodynamics of complex formation. The structure, in turn, influences the biological properties of the lipoplex. If the titration of lipid into DNA was carried out in large increments, the DeltaH was larger than when the injection increments were smaller, a finding that is consistent with increased vesicle disruption under large increments and which is expected theoretically. Cationic lipid-DNA binding was weak in high ionic strength solutions, however, the effective binding constant is within micromolar range because of macromolecular nature of the interaction.

Full Text

The Full Text of this article is available as a PDF (226.1 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. BARTLETT G. R. Phosphorus assay in column chromatography. J Biol Chem. 1959 Mar;234(3):466–468. [PubMed] [Google Scholar]
  2. Banerjee R., Das P. K., Chaudhuri A. Interfacial indazolization: novel chemical evidence for remarkably high exo-surface pH of cationic liposomes used in gene transfection. Biochim Biophys Acta. 1998 Sep 2;1373(2):299–308. doi: 10.1016/s0005-2736(98)00113-8. [DOI] [PubMed] [Google Scholar]
  3. Boukhnikachvili T., Aguerre-Chariol O., Airiau M., Lesieur S., Ollivon M., Vacus J. Structure of in-serum transfecting DNA-cationic lipid complexes. FEBS Lett. 1997 Jun 9;409(2):188–194. doi: 10.1016/s0014-5793(97)00505-x. [DOI] [PubMed] [Google Scholar]
  4. Cowley A. C., Fuller N. L., Rand R. P., Parsegian V. A. Measurement of repulsive forces between charged phospholipid bilayers. Biochemistry. 1978 Jul 25;17(15):3163–3168. doi: 10.1021/bi00608a034. [DOI] [PubMed] [Google Scholar]
  5. Dan N. Multilamellar structures of DNA complexes with cationic liposomes. Biophys J. 1997 Oct;73(4):1842–1846. doi: 10.1016/S0006-3495(97)78214-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Dan N. The structure of DNA complexes with cationic liposomes-cylindrical or flat bilayers? Biochim Biophys Acta. 1998 Feb 2;1369(1):34–38. doi: 10.1016/s0005-2736(97)00171-5. [DOI] [PubMed] [Google Scholar]
  7. Felgner P. L., Gadek T. R., Holm M., Roman R., Chan H. W., Wenz M., Northrop J. P., Ringold G. M., Danielsen M. Lipofection: a highly efficient, lipid-mediated DNA-transfection procedure. Proc Natl Acad Sci U S A. 1987 Nov;84(21):7413–7417. doi: 10.1073/pnas.84.21.7413. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Gustafsson J., Arvidson G., Karlsson G., Almgren M. Complexes between cationic liposomes and DNA visualized by cryo-TEM. Biochim Biophys Acta. 1995 May 4;1235(2):305–312. doi: 10.1016/0005-2736(95)80018-b. [DOI] [PubMed] [Google Scholar]
  9. Harries D., May S., Gelbart W. M., Ben-Shaul A. Structure, stability, and thermodynamics of lamellar DNA-lipid complexes. Biophys J. 1998 Jul;75(1):159–173. doi: 10.1016/S0006-3495(98)77503-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Hirsch-Lerner D., Barenholz Y. Hydration of lipoplexes commonly used in gene delivery: follow-up by laurdan fluorescence changes and quantification by differential scanning calorimetry. Biochim Biophys Acta. 1999 Nov 9;1461(1):47–57. doi: 10.1016/s0005-2736(99)00145-5. [DOI] [PubMed] [Google Scholar]
  11. Huebner S., Battersby B. J., Grimm R., Cevc G. Lipid-DNA complex formation: reorganization and rupture of lipid vesicles in the presence of DNA as observed by cryoelectron microscopy. Biophys J. 1999 Jun;76(6):3158–3166. doi: 10.1016/S0006-3495(99)77467-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Kennedy M. T., Pozharski E. V., Rakhmanova V. A., MacDonald R. C. Factors governing the assembly of cationic phospholipid-DNA complexes. Biophys J. 2000 Mar;78(3):1620–1633. doi: 10.1016/S0006-3495(00)76714-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Koltover I., Salditt T., Rädler J. O., Safinya C. R. An inverted hexagonal phase of cationic liposome-DNA complexes related to DNA release and delivery. Science. 1998 Jul 3;281(5373):78–81. doi: 10.1126/science.281.5373.78. [DOI] [PubMed] [Google Scholar]
  14. Koltover I., Salditt T., Safinya C. R. Phase diagram, stability, and overcharging of lamellar cationic lipid-DNA self-assembled complexes. Biophys J. 1999 Aug;77(2):915–924. doi: 10.1016/S0006-3495(99)76942-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Kreiss P., Cameron B., Rangara R., Mailhe P., Aguerre-Charriol O., Airiau M., Scherman D., Crouzet J., Pitard B. Plasmid DNA size does not affect the physicochemical properties of lipoplexes but modulates gene transfer efficiency. Nucleic Acids Res. 1999 Oct 1;27(19):3792–3798. doi: 10.1093/nar/27.19.3792. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Leventis R., Silvius J. R. Interactions of mammalian cells with lipid dispersions containing novel metabolizable cationic amphiphiles. Biochim Biophys Acta. 1990 Mar 30;1023(1):124–132. doi: 10.1016/0005-2736(90)90017-i. [DOI] [PubMed] [Google Scholar]
  17. Lin A. J., Slack N. L., Ahmad A., Koltover I., George C. X., Samuel C. E., Safinya C. R. Structure and structure-function studies of lipid/plasmid DNA complexes. J Drug Target. 2000;8(1):13–27. doi: 10.3109/10611860009009206. [DOI] [PubMed] [Google Scholar]
  18. Lobo B. A., Davis A., Koe G., Smith J. G., Middaugh C. R. Isothermal titration calorimetric analysis of the interaction between cationic lipids and plasmid DNA. Arch Biochem Biophys. 2001 Feb 1;386(1):95–105. doi: 10.1006/abbi.2000.2196. [DOI] [PubMed] [Google Scholar]
  19. Loosley-Millman M. E., Rand R. P., Parsegian V. A. Effects of monovalent ion binding and screening on measured electrostatic forces between charged phospholipid bilayers. Biophys J. 1982 Dec;40(3):221–232. doi: 10.1016/S0006-3495(82)84477-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. MacDonald R. C., Ashley G. W., Shida M. M., Rakhmanova V. A., Tarahovsky Y. S., Pantazatos D. P., Kennedy M. T., Pozharski E. V., Baker K. A., Jones R. D. Physical and biological properties of cationic triesters of phosphatidylcholine. Biophys J. 1999 Nov;77(5):2612–2629. doi: 10.1016/S0006-3495(99)77095-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. MacDonald R. C., MacDonald R. I., Menco B. P., Takeshita K., Subbarao N. K., Hu L. R. Small-volume extrusion apparatus for preparation of large, unilamellar vesicles. Biochim Biophys Acta. 1991 Jan 30;1061(2):297–303. doi: 10.1016/0005-2736(91)90295-j. [DOI] [PubMed] [Google Scholar]
  22. Marra J. Direct measurement of the interaction between phosphatidylglycerol bilayers in aqueous electrolyte solutions. Biophys J. 1986 Nov;50(5):815–825. doi: 10.1016/S0006-3495(86)83522-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Matulis D., Rouzina I., Bloomfield V. A. Thermodynamics of DNA binding and condensation: isothermal titration calorimetry and electrostatic mechanism. J Mol Biol. 2000 Mar 3;296(4):1053–1063. doi: 10.1006/jmbi.1999.3470. [DOI] [PubMed] [Google Scholar]
  24. May S., Ben-Shaul A. DNA-lipid complexes: stability of honeycomb-like and spaghetti-like structures. Biophys J. 1997 Nov;73(5):2427–2440. doi: 10.1016/S0006-3495(97)78271-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. May S., Harries D., Ben-Shaul A. The phase behavior of cationic lipid-DNA complexes. Biophys J. 2000 Apr;78(4):1681–1697. doi: 10.1016/S0006-3495(00)76720-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. McDaniel R. V., McIntosh T. J. Neutron and X-ray diffraction structural analysis of phosphatidylinositol bilayers. Biochim Biophys Acta. 1989 Aug 7;983(2):241–246. doi: 10.1016/0005-2736(89)90239-3. [DOI] [PubMed] [Google Scholar]
  27. McIntosh T. J., Magid A. D., Simon S. A. Interactions between charged, uncharged, and zwitterionic bilayers containing phosphatidylglycerol. Biophys J. 1990 Jun;57(6):1187–1197. doi: 10.1016/S0006-3495(90)82638-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Meidan V. M., Cohen J. S., Amariglio N., Hirsch-Lerner D., Barenholz Y. Interaction of oligonucleotides with cationic lipids: the relationship between electrostatics, hydration and state of aggregation. Biochim Biophys Acta. 2000 Apr 5;1464(2):251–261. doi: 10.1016/s0005-2736(00)00151-6. [DOI] [PubMed] [Google Scholar]
  29. Mitrakos P., Macdonald P. M. Nucleotide chain length and the morphology of complexes with cationic amphiphiles: (31)P-NMR observations. Biochim Biophys Acta. 2000 Feb 15;1463(2):355–373. doi: 10.1016/s0005-2736(99)00232-1. [DOI] [PubMed] [Google Scholar]
  30. Oberle V., Bakowsky U., Zuhorn I. S., Hoekstra D. Lipoplex formation under equilibrium conditions reveals a three-step mechanism. Biophys J. 2000 Sep;79(3):1447–1454. doi: 10.1016/S0006-3495(00)76396-X. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Pector V., Backmann J., Maes D., Vandenbranden M., Ruysschaert J. M. Biophysical and structural properties of DNA.diC(14)-amidine complexes. Influence of the DNA/lipid ratio. J Biol Chem. 2000 Sep 22;275(38):29533–29538. doi: 10.1074/jbc.M909996199. [DOI] [PubMed] [Google Scholar]
  32. Rau D. C., Lee B., Parsegian V. A. Measurement of the repulsive force between polyelectrolyte molecules in ionic solution: hydration forces between parallel DNA double helices. Proc Natl Acad Sci U S A. 1984 May;81(9):2621–2625. doi: 10.1073/pnas.81.9.2621. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Ross P. C., Hui S. W. Lipoplex size is a major determinant of in vitro lipofection efficiency. Gene Ther. 1999 Apr;6(4):651–659. doi: 10.1038/sj.gt.3300863. [DOI] [PubMed] [Google Scholar]
  34. Rädler J. O., Koltover I., Salditt T., Safinya C. R. Structure of DNA-cationic liposome complexes: DNA intercalation in multilamellar membranes in distinct interhelical packing regimes. Science. 1997 Feb 7;275(5301):810–814. doi: 10.1126/science.275.5301.810. [DOI] [PubMed] [Google Scholar]
  35. Strey H. H., Podgornik R., Rau D. C., Parsegian V. A. DNA--DNA interactions. Curr Opin Struct Biol. 1998 Jun;8(3):309–313. doi: 10.1016/s0959-440x(98)80063-8. [DOI] [PubMed] [Google Scholar]
  36. Templeton N. S., Lasic D. D., Frederik P. M., Strey H. H., Roberts D. D., Pavlakis G. N. Improved DNA: liposome complexes for increased systemic delivery and gene expression. Nat Biotechnol. 1997 Jul;15(7):647–652. doi: 10.1038/nbt0797-647. [DOI] [PubMed] [Google Scholar]
  37. Wiseman T., Williston S., Brandts J. F., Lin L. N. Rapid measurement of binding constants and heats of binding using a new titration calorimeter. Anal Biochem. 1989 May 15;179(1):131–137. doi: 10.1016/0003-2697(89)90213-3. [DOI] [PubMed] [Google Scholar]
  38. Xu Y., Hui S. W., Frederik P., Szoka F. C., Jr Physicochemical characterization and purification of cationic lipoplexes. Biophys J. 1999 Jul;77(1):341–353. doi: 10.1016/S0006-3495(99)76894-3. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Yang J. P., Huang L. Overcoming the inhibitory effect of serum on lipofection by increasing the charge ratio of cationic liposome to DNA. Gene Ther. 1997 Sep;4(9):950–960. doi: 10.1038/sj.gt.3300485. [DOI] [PubMed] [Google Scholar]
  40. Yang J. P., Huang L. Time-dependent maturation of cationic liposome-DNA complex for serum resistance. Gene Ther. 1998 Mar;5(3):380–387. doi: 10.1038/sj.gt.3300596. [DOI] [PubMed] [Google Scholar]
  41. Zuidam N. J., Barenholz Y. Electrostatic and structural properties of complexes involving plasmid DNA and cationic lipids commonly used for gene delivery. Biochim Biophys Acta. 1998 Jan 5;1368(1):115–128. doi: 10.1016/s0005-2736(97)00187-9. [DOI] [PubMed] [Google Scholar]
  42. Zuidam N. J., Barenholz Y. Electrostatic parameters of cationic liposomes commonly used for gene delivery as determined by 4-heptadecyl-7-hydroxycoumarin. Biochim Biophys Acta. 1997 Oct 23;1329(2):211–222. doi: 10.1016/s0005-2736(97)00110-7. [DOI] [PubMed] [Google Scholar]
  43. Zuidam N. J., Hirsch-Lerner D., Margulies S., Barenholz Y. Lamellarity of cationic liposomes and mode of preparation of lipoplexes affect transfection efficiency. Biochim Biophys Acta. 1999 Jul 15;1419(2):207–220. doi: 10.1016/s0005-2736(99)00069-3. [DOI] [PubMed] [Google Scholar]

Articles from Biophysical Journal are provided here courtesy of The Biophysical Society

RESOURCES