DNA release dynamics from reducible polyplexes by atomic force microscopy.

Controlled intracellular disassembly of polyelectrolyte complexes of polycations and DNA (polyplexes) is a crucial step for the success of nonviral gene delivery. Motivated by our previous observation of different gene delivery performances among multiblock reducible copolypeptide vectors ( Manickam, D. S. ; Oupicky, D. Bioconjugate Chem. 2006, 17, 1395- 1403 ), atomic force microscopy is used to visualize plasmid DNA in various decondensed states from reducible polypeptide polyplexes under simulated physiological reducing conditions. DNA decondensation is triggered by reductive degradation of disulfide-containing cationic polypeptides. Striking differences in DNA release dynamics between polyplexes based on polypeptides of histidine-rich peptide (HRP, CKHHHKHHHKC) and nuclear localization signal (NLS, CGAGPKKKRKVC) peptide are presented. The HRP and NLS polyplexes are similar to each other in their initial morphology with a majority of them containing only one DNA plasmid. Upon reductive degradation by dithiothreitol, DNA is released from NLS abruptly regardless of the initial polyplex morphology, while DNA release from HRP polyplexes displays a gradual decondensation that is dependent on the size of polyplexes. The release rate is higher for larger HRP polyplexes. The smaller HRP polyplexes become unstable when they are in contact with expanding chains nearby. The results reveal potentially rich DNA release dynamics that can be controlled by subtle variation in multivalent counterion binding to DNA as well as the cellular matrix.