Alessandra Griffo - RNA Origami Nanopores spanning lipid membranes

Artificial membrane nanopores have significance either in the field of synthetic biology and of single molecule detection. Recent advancements in DNA nanotechnology have yielded functional synthetic pores as noteworthy examples. Nevertheless, their full potential is constrained due to the requirement for chemical modification, which results in high production costs and incompatibility with cellular expression. In this study, we introduce an RNA origami nanopore that spans into lipid membranes through aptamer-target interactions, eliminating the need for chemical modification. To surmount the energy barrier for membrane insertion, we opted for aptamers that bind to lipid membranes, specifically employing a biotin aptamer to bind to biotinylated DOPE lipids. The RNA nanopore is synthesized through in vitro transcription from a DNA template, yielding a high quantity of nanopores. We validate its correct assembly and successful membrane insertion using respectively atomic force microscopy (AFM) and recordings of ionic currents. The latter demonstrates distinct insertion steps with a conductance of 0.8nS (in 150 mM NaCl) and voltage-dependent closure and gating patterns, resembling those observed in traditional protein pores.
Genetically encoded RNA origami pores have the potential to be expressed both in vitro and in cells, serving as valuable biophysical tools, components of synthetic cells, or single-molecule sensors.
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