Reversible Binding and Trafficking of DNA Origami Between Lipid Membrane Phases

Cellular membranes are inherently heterogeneous in their lipid composition and can exhibit varying degrees of stiffness and phase separation. Many proteins that reversibly traffic between membrane compartments and/or domains are modified with lipid anchors that enable them to bind and localize to desired membrane regions. Inspired by the lipidation patterns of membrane trafficking elements and using DNA origami as bioengineering material, this work aims to develop biomimetic lipid‐anchored DNA nanostructures that selectively bind to and reversibly shuttle between membranes of different rigidities. Here, origami structures are functionalized with different types and numbers of lipophilic groups, namely cholesterol, tocopherol, oleyl, palmitoyl, and stearoyl, which mimic important physicochemical properties of naturally occurring lipid anchors. Their membrane interaction and phase selectivity toward homogeneous and liquid‐disordered/liquid‐ordered phase‐separated giant unilamellar vesicles are quantitatively characterized by fluorescence confocal microscopy. Key findings show that the lipid‐anchored DNA nanostructures selectively bind to varying membrane phases, depending on the type of anchor and membrane composition/rigidity. Finally, by using strand displacement to control the dimerization of DNA nanostructures carrying distinct lipid anchors, a reversible shuttle between different membrane domains and vesicles is accomplished, recapitulating key principles of membrane trafficking in vitro and opening new perspectives for more precise membrane‐active nanocarriers.