Dynamic Behavior of DNA Cages Anchored on Spherically Supported Lipid Bilayers

Abstract

We report the anchoring of 3D-DNA-cholesterol labeled cages on spherically supported lipid bilayer membranes (SSLBM) formed on silica beads, and their addressability through strand displacement reactions, controlled membrane orientation and templated dimerization. The bilayer-anchored cages can load three different DNA-fluorophores by hybridization to their "top" face (furthest from bilayer) and unload each of them selectively upon addition of a specific input displacement strand. We introduce a method to control strand displacement from their less accessible "bottom" face (closest to the bilayer), by adding cholesterol-substituted displacing strands that insert into the bilayer themselves in order to access the toehold region. The orientation of DNA cages within the bilayer is tunable by positioning multiple cholesterol anchoring units on the opposing two faces of the cage, thereby controlling their accessibility to proteins and enzymes. A population of two distinct DNA cages anchored to the SSLBMs exhibited significant membrane fluidity and have been directed into dimer assemblies on bilayer via input of a complementary linking strand. Displacement experiments performed on these anchored dimers indicate that removal of only one prism's anchoring cholesterol strand was not sufficient to release the dimers from the bilayer; however, removal of both cholesterol anchors from the dimerized prisms via two displacement strands cleanly released the dimers from the bilayer. This methodology allows for the anchoring of DNA cages on supported lipid bilayers, the control of their orientation and accessibility within the bilayer, and the programmable dimerization and selective removal of any of their components. The facile coupling of DNA to other functional materials makes this an attractive method for developing stimuli-responsive protein or nanoparticle arrays, drug releasing biomedical device surfaces and self-healing materials for light harvesting applications, using a highly modular, DNA-economic scaffold.

DNA Nanostructures on Membranes as Tools for Synthetic Biology

“…A recent approach further reported small membrane-anchored 3D-cages, which upon strand displacement could unload DNA-fluorophores, assemble/disassemble into dimers/monomers, or even detach from the membrane. These features allow programmable dynamic control of binding and signaling events attributed to living cells ( 49 ).…”

Spatial Presentation of Cholesterol Units on a DNA Cube as a Determinant of Membrane Protein-Mimicking Functions

“…[24][25] Another exciting focus is membrane functionalization to control membrane's properties and interactions as well as to use membranes as platform for DNA self-assembly. [26][27][28][29][30][31][32][33][34][35][36] As a recent example, cell clustering was programmed by higher-order assembly of DNA origami, imitating cell-adhesion molecules. 37 These studies raise an important fundamental question: What are the general design rules that specify the behavior of DNA nanostructures with lipid bilayers, and can a combination of these structural variations be used to change functions of membrane protein mimics?…”

A Microsphere-Supported Lipid Bilayer Platform for DNA Reactions on a Fluid Surface

“…Above the transition temperature of the base lipid (−2 °C for POPC, and <−2 °C for DOPC), bilayers formed on mesoporous silica microspheres maintain the ability to diffuse laterally. 35,36 Multiplexed µ SLB populations are created by incorporation of lipids with fluorescently-labeled headgroups (one with a yellow rhodamine dye (PELR) and one with a green fluorescein dye (PECF)) at varying percentages (Figure 1B). µ SLB populations were analyzed via multi-parameter flow cytometry and demonstrated a well resolved 4-plex (Figure 1C).…”