Temperature- and Composition-Dependent DNA Condensation by Thermosensitive Block Copolymers
journal contributionposted on 2017-11-15, 18:11 authored by Satyagopal Sahoo, Sharmita Bera, Saikat Maiti, Dibakar Dhara
Successful intracellular delivery of genes requires an efficient carrier, as genes by themselves cannot diffuse across cell membranes. Because of the toxicity and immunogenicity of viral vectors, nonviral vectors are gaining tremendous interest in research. In this work, we have investigated the temperature-dependent DNA condensation efficiency of various compositions of a thermosensitive block copolymer viz., poly(N-isopropylacrylamide)-b-poly(2-(diethylamino)ethyl methacrylate) (PNIPA-b-PDMAEMA). Three different copolymer compositions of varying molecular weights were successfully synthesized via the RAFT polymerization technique. Steady-state fluorescence and circular dichroism (CD) spectroscopies, dynamic light scattering (DLS) and zeta potential measurements, agarose gel electrophoresis, and atomic force microscopy techniques were utilized to study the interaction of the copolymers with DNA at temperatures above and below the critical aggregation temperature (CAT). All these experiments revealed that, above the CAT, there was formation of highly stable and tight polymer–DNA complexes (polyplexes). The size of polyplexes was dependent on the temperature up to a certain charge ratio, as determined by the DLS results. The results obtained from temperature-dependent fluorescence spectroscopy, CD, and gel electrophoresis indicated that the DNA molecules were shielded more from aqueous exposure above the CAT because of the formation of relatively more compact complexes. The polyplexes also exhibited changes in the particle morphology below and above the CAT, with particles generated above CAT being more spherical in morphology. These results suggested at the possibility of modulating the complex formation by temperature modification. The present biophysical studies would provide new physical insight into the design of novel gene carriers.
Steady-state fluorescenceforce microscopy techniquesnonviral vectorsComposition-Dependent DNA Condensationintracellular deliverygel electrophoresiscopolymer compositionsparticle morphologyDNA moleculestemperature-dependent fluorescence spectroscopynovel gene carriersagarose gel electrophoresisformationPDMAEMAPNIPARAFT polymerization techniqueThermosensitive Block CopolymersDLS resultspolyplexeaggregation temperaturetemperature-dependent DNA condensation efficiencycharge ratiocomplexcell membranestemperature modification