Multi-functional DNA nanostructures that puncture and remodel lipid membranes into hybrid materials

Abstract

Synthetically replicating key biological processes requires the ability to puncture lipid bilayer membranes and to remodel their shape. Recently developed artificial DNA nanopores are one possible synthetic route due to their ease of fabrication. However, an unresolved fundamental question is how DNA nanopores bind to and dynamically interact with lipid bilayers. Here we use single-molecule fluorescence microscopy to establish that DNA nanopores carrying cholesterol anchors insert via a two-step mechanism into membranes. Nanopores are furthermore shown to locally cluster and remodel membranes into nanoscale protrusions. Most strikingly, the DNA pores can function as cytoskeletal components by stabilizing autonomously formed lipid nanotubes. The combination of membrane puncturing and remodeling activity can be attributed to the DNA pores’ tunable transition between two orientations to either span or co-align with the lipid bilayer. This insight is expected to catalyze the development of future functional nanodevices relevant in synthetic biology and nanobiotechnology.

Cholesterol Anchors Enable Efficient Binding and Intracellular Uptake of DNA Nanostructures

“…However, due to the chemical and structural complexity of biological cell membranes, one cannot deduce that NB-3C also punctures cells. To measure nanobundle-cell interactions, Alexaflour-647 was incorporated into the nanostructure at the DNA bundle exterior via the 5' terminus of a non-cholesterol modified DNA strands, as described in ref 54 . Alexafluor has a high fluorescence stability and can, like other fluorophores, partition into the membrane.…”

“…This nanobunlde can also span synthetic lipid bilayers as opposed to NB-1C that just tethers to membranes. 53,54 However, due to the chemical and structural complexity of biological cell membranes, one cannot deduce that NB-3C did punctured also these bilayers. Future research may explore uptake and trafficking pathways, as well as endosomal fate of the DNA nanobundles in relation to their behavior in different media conditions.…”

Graphene Templated DNA Arrays and Biotin-Streptavidin Sensitive Bio-Transistors Patterned by Dynamic Self-Assembly of Polymeric Films Confined within a Roll-on-Plate Geometry

“…Over the past decades, there has been considerable progress in the development of multifunctional electronic devices capable of detecting chemical and biological substances and measuring nanoscale phenomena [ 1 ]. Recent advances in integrated circuit technology and controlled nanofabrication with nanomaterials have enabled implementations of microelectronic sensors with a variety of novel sensing mechanisms in conjunction with their collective properties [ 2 ], which can provide viable prototype solutions in applications ranging from environmental monitoring to biomedical diagnostic sensors [ 3 ].…”

Structural and Functional Stability of DNA Nanopores in Biological Media

“…DNA-based nanopores are the most recent class of membrane channels which can potentially offer a unique degree of control at the membrane interface [40,41,42,43,44]. Naturally occurring nanopores are usually composed of proteins or peptides to help regulate ion transport across cell membranes [45].…”

“…In contrast, utilizing DNA as a construction material can help overcome this issue. To date, DNA nanotechnology has produced nanopores with highly customizable properties, including channel diameter, length, functionalized groups within the lumen, and ligand-controlled pore opening [40,41,42,43,44]. For future biological applications, including pore-mediated drug delivery, nanopore stability and solubility within biological media must be maintained.…”