Formation of Supramolecular Structures between DNA and Starburst Dendrimers Studied by EPR, CD, UV, and Melting Profiles

Abstract

Poly(amidoamine) starburst dendrimers (SBDs) form stable supramolecular structures (complexes) with DNA and confer protection against degradation by nucleases. Such properties make SBDs excellent candidates for application in gene delivery strategies. Formation of supramolecular structures by two generations of SBDs, namely 2SBD and 6SBD, with calf thymus DNA was examined at varying r = [SBD](surface groups)/[DNA](base pairs) through various physicochemical techniques including electronic absorption spectroscopy (UV), circular dichroism (CD), DNA thermal denaturation studies (melting profiles), and electron paramagnetic resonance (EPR) spectroscopy. EPR spectroscopy of nitroxide-labeled dendrimers provides information on the interactions between SBDs and DNA, whereas the other techniques mainly monitor the structural variations of DNA following formation of SBD−DNA complexes. At the lowest r values (r < 1), both dendrimers slightly affect DNA conformation within the general framework of B-type structure; minor stabilization effects of the double helix were detected through analysis of the melting profiles. When r increases, extensive precipitation of the SBD/DNA adducts takes place due to charge neutralization effects. A persistent opalescence of the solution prevents recording of CD spectra up to r = 100 for 2SBD and r = 200 for 6SBD. However, at these r values, EPR analysis indicates that a saturation of the interacting sites on DNA occurs which allows calculation of the formation constant of the SBD−DNA adducts. By further increasing the [SBD]/[DNA] ratio, DNA resolubilizes. At the high r values, both SBDs form stable soluble supramolecular structures with DNA due to a “salting in” effect. EPR, CD, and UV results allow us to propose a model for the formation of different supramolecular structures in the various r ranges.