Strategies for constructing and operating DNA origami linear actuators

Small Wiley 17:20 (2021) 2007704

Authors:

Erik Benson, Rafael Carrascosa Marzo, Jonathan Bath, Andrew Turberfield

Abstract:

Linear actuators are ubiquitous components at all scales of engineering. DNA nanotechnology offers a unique opportunity for bottom-up assembly at the molecular scale, providing nanoscale precision with multiple methods for constructing and operating devices. In this paper, DNA origami linear actuators with up to 200 nm travel, based on a rail threading a topologically locked slider, are demonstrated. Two strategies, one- and two-pot assembly, are demonstrated whereby the two components are folded from one or two DNA scaffold strands, respectively. In order to control the position of the slider on the rail, the rail and the inside of the slider are decorated with single-stranded oligonucleotides with distinct sequences. Two positioning strategies, based on diffusion and capture of signaling strands, are used to link the slider reversibly to determined positions on the rail with high yield and precision. These machine components provide a basis for applications in molecular machinery and nanoscale manufacture including programmed chemical synthesis.

DNA origami signposts for identifying proteins on cell membranes by electron cryotomography

Cell Cell Press 184:4 (2021) 1110-1121.e16

Authors:

Emma Silvester, Benjamin Vollmer, Vojtěch Pražák, Daven Vasishtan, Emily A Machala, Catheryne Whittle, Susan Black, Jonathan Bath, Andrew J Turberfield, Kay Grünewald, Lindsay A Baker

Abstract:

Electron cryotomography (cryoET), an electron cryomicroscopy (cryoEM) modality, has changed our understanding of biological function by revealing the native molecular details of membranes, viruses, and cells. However, identification of individual molecules within tomograms from cryoET is challenging because of sample crowding and low signal-to-noise ratios. Here, we present a tagging strategy for cryoET that precisely identifies individual protein complexes in tomograms without relying on metal clusters. Our method makes use of DNA origami to produce “molecular signposts” that target molecules of interest, here via fluorescent fusion proteins, providing a platform generally applicable to biological surfaces. We demonstrate the specificity of signpost origami tags (SPOTs) in vitro as well as their suitability for cryoET of membrane vesicles, enveloped viruses, and the exterior of intact mammalian cells.

Reconfigurable T‐junction DNA origami

Angewandte Chemie International Edition Wiley 59:37 (2020) 15942-15946

Authors:

Katherine Young, Behnam Najafi, William Sant, Sonia Contera, Ard Louis, Jonathan Doye, Andrew Turberfield, Jonathan Bath

Abstract:

DNA self‐assembly allows the construction of nanometre‐scale structures and devices. Structures with thousands of unique components are routinely assembled in good yield. Experimental progress has been rapid, based largely on empirical design rules. Here we demonstrate a DNA origami technique designed as a model system with which to explore the mechanism of assembly. The origami fold is controlled through single‐stranded loops embedded in a double‐stranded DNA template and is programmed by a set of double‐stranded linkers that specify pairwise interactions between loop sequences. Assembly is via T‐junctions formed by hybridization of single‐stranded overhangs on the linkers with the loops. The sequence of loops on the template and the set of interaction rules embodied in the linkers can be reconfigured with ease. We show that a set of just two interaction rules can be used to assemble simple T‐junction origami motifs and that assembly can be performed at room temperature.

Reconfigurable T‐junction DNA origami

Angewandte Chemie Wiley (2020) ange.202006281

Authors:

Katherine Young, Behnam Najafi, William Sant, Sonia Contera, Ard Louis, Jonathan Doye, Andrew Turberfield, Jonathan Bath

Design, optimization and analysis of large DNA and RNA nanostructures through interactive visualization, editing and molecular simulation

Nucleic Acids Research Oxford University Press (OUP) (2020)

Authors:

Erik Poppleton, Joakim Bohlin, Michael Matthies, Shuchi Sharma, Fei Zhang, Petr Šulc

Abstract:

<jats:title>Abstract</jats:title> <jats:p>This work seeks to remedy two deficiencies in the current nucleic acid nanotechnology software environment: the lack of both a fast and user-friendly visualization tool and a standard for structural analyses of simulated systems. We introduce here oxView, a web browser-based visualizer that can load structures with over 1 million nucleotides, create videos from simulation trajectories, and allow users to perform basic edits to DNA and RNA designs. We additionally introduce open-source software tools for extracting common structural parameters to characterize large DNA/RNA nanostructures simulated using the coarse-grained modeling tool, oxDNA, which has grown in popularity in recent years and is frequently used to prototype new nucleic acid nanostructural designs, model biophysics of DNA/RNA processes, and rationalize experimental results. The newly introduced software tools facilitate the computational characterization of DNA/RNA designs by providing multiple analysis scripts, including mean structures and structure flexibility characterization, hydrogen bond fraying, and interduplex angles. The output of these tools can be loaded into oxView, allowing users to interact with the simulated structure in a 3D graphical environment and modify the structures to achieve the required properties. We demonstrate these newly developed tools by applying them to design and analysis of a range of DNA/RNA nanostructures.</jats:p>