Abstract
DNA nanopores offer a unique nano-scale foothold at the membrane interface that can help advance the life sciences as biophysical research tools or gate-keepers for drug delivery. Biological applications require sufficient physiological stability and membrane activity for viable biological action. In this report, we determine essential parameters for efficient nanopore folding and membrane binding in biocompatible cell media. The parameters are identified for an archetypal DNA nanopore composed of six interwoven strands carrying cholesterol lipid anchors. Using gel electrophoresis and fluorescence spectroscopy, the nanostructures are found to assemble efficiently in cell media, such as LB and DMEM, and remain structurally stable at physiological temperatures. Furthermore, the pores’ oligomerization state is monitored using fluorescence spectroscopy and confocal microscopy. The pores remain predominately water-soluble over 24 h in all buffer systems, and were able to bind to lipid vesicles after 24 h to confirm membrane activity. However, the addition of fetal bovine serum to DMEM causes a significant reduction in nanopore activity. Serum proteins complex rapidly to the pore, most likely via ionic interactions, to reduce the effective nanopore concentration in solution. Our findings outline crucial conditions for maintaining lipidated DNA nanodevices, structurally and functionally intact in cell media, and pave the way for biological studies in the future.
Highlights
DNA nanotechnology excels at the bottom-up fabrication of engineered nanostructures
Identifying Nanopore Melting Temperatures in Biological Media The thermal stability of the pores was established using DNA nanostructures labeled with a fluorophore pair for Förster resonance energy transfer (FRET)
The DNA nanopores binding towards giant unilamellar vesicles (GUVs) was identified using Confocal laser scanning microscopy (CLSM) (Figure 5)
Summary
DNA nanotechnology excels at the bottom-up fabrication of engineered nanostructures. DNA duplexes can be manipulated into user-defined shapes by exploiting the base-pairing rules for duplex formation [1,2,3]. DNA nanostructures can help control processes within cells, or at the membrane interface to advance biological understanding [17,18,19,20]. This progress includes the generation of novel diagnostic tools [21,22,23], the enhancement of existing drugs [24], and devices with novel therapeutic action [25]. With the knowledge gained using our approach, new pore formulations and folding protocols can be established which should help provide insights for future applications across the life sciences
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
