Fluorescence and Förster resonance energy transfer investigations on DNA oligonucleotide and PAMAM dendrimer packing interactions in dendriplexes.
Considering the importance of short oligonucleotide packing in dendriplex-mediated gene delivery, a direct insight into the 14-mer oligonucleotide and dendrimer interactions using fluorescence and FRET techniques is the focus of this study. Fluorometric titrations of various fluorophore-tagged oligonucleotides with the first three PAMAM dendrimer generations showed a decrease in the fluorescence intensity with two break points, namely Z and Z, for each titration. The first break point for each dendrimer was identical to the neutralization point observed by basic biophysical studies for the corresponding dendrimer generations. Additionally, FRET studies on dual tagged oligonucleotide (DFT) molecules revealed a third break point at the charge ratio (Z) where there was the highest fluorescence energy transfer from the donor to the acceptor fluorophores. Altogether, dendriplex formation was considered to take place via three steps with an increase in the dendrimer concentration, where initially there was monomeric complexation at the neutralization point (Z) followed by loosely held molecular aggregation of the dendrimer (Z). In the final step, dendrimer molecular aggregates were held tightly together for the closest possible packing of the oligonucleotide molecules onto their surface. The effective molecular packing is identified by the highest FRET intensity for the dendrimer of generation 2 at a charge ratio of 0.34 (Z±).
Department of Applied Chemistry and Polymer Technology, Delhi Technological University, Bawana Road, Delhi-110 042, India. deenan.santhiya@dce.ac.in.
Hema Kumari
HK
Alajangi
Deenan
D
Santhiya
D004247:DNA / Q000737:chemistry*; D050091:Dendrimers / Q000737:chemistry*; D005453:Fluorescence; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D005456:Fluorescent Dyes / Q000737:chemistry*; D009841:Oligonucleotides / Q000737:chemistry*
D050091:Dendrimers; D005456:Fluorescent Dyes; D009841:Oligonucleotides; C104700:PAMAM Starburst; D004247:DNA