화학공학소재연구정보센터
Langmuir, Vol.19, No.22, 9387-9394, 2003
Thermodynamic analysis of polycation-DNA interaction applying titration microcalorimetry
Nonviral gene delivery systems based on DNA complexes with polycations have recently been extensively studied. An important consideration in the design of such therapeutic systems is the understanding of forces governing the DNA-polycation interaction and the complex stabilization. In this work, the interaction of a cationic polymer from the poly(amidoamine) family (poly(bis-acryloylpiperazine-2-methyl-piperazine), p(BAP-2MP)) with DNA at different salt concentrations and pHs and in different buffers was studied using isothermal titration microcalorimetry as a principal tool. The binding isotherms were analyzed using a proposed complexation-condensation model, to calculate the observed binding constant, K-obs, enthalpies of the complex formation and condensation process, DeltaH(com)(o) and DeltaH(con)(o) and complexation site and condensation site exclusion sizes, n(com), and n(con), respectively. K-obs, DeltaH(com)(o), and DeltaH(con)(o) decreased with increasing salt concentration, indicating involvement of electrostatic interactions. However, noon, was rather independent of salt concentration and nearly equal to the average number of ionized charges at the polycation, indicating that the ratio of nucleotide to p(BAP-2MP) monomer units is close to 1. Dissection of the free energy of interaction showed that electrostatic contributions dominate the p(BAP-2MP)-DNA interaction. The use of buffers with different protonation enthalpies demonstrated that the p(BAP-2MP) interaction with DNA is coupled with a proton extraction from the buffer resulting in an enthalpy change and enabling the polymer ionization and interaction at a stoichiometric ratio. Obtained intrinsic binding parameters (in a buffer with DeltaH(ion) = 0) reveal that p(BAP-2MP) interaction with DNA is nonspecific with a moderate binding constant and that it contains the enthalpic contribution (probably due to the formation of hydrogen bonding). The values for the condensation process, n(con) and DeltaH(con)(o), indicate that there is no further ionization of the p(BAP-2MP) polymer to interact with DNA and that the stoichiometric ratio is preserved.