Biology subjects

Geary, C.

Publications and source records attributed to Geary, C..

4 recordsLinked to original sources

bioRxiv2026

Programmable Edge-to-Edge Assembly of RNA Nanostructures

Constructing complex three-dimensional RNA nanostructures requires precise molecular connectors for controlled self-assembly. Existing RNA-RNA connectors, such as kissing loops, are restricted to coaxial, end-to-end joining, limiting the range of accessible geometries. Here, we introduce the alpha kissing loop (alphaKL), a compact, sequence-programmable RNA connector that enables edge-to-edge helix association. The alphaKL combines a four-nucleotide kissing loop with minor- and major-groove triplex interactions that pre-organize an -shaped conformation compatible with co-transcriptional folding. Embedded into RNA origami tiles, alphaKLs drive multivalent assembly into filaments and lattices, visualized at nanoscale resolution by atomic force microscopy. All-atom molecular dynamics simulations reveal that triplex occupancy at each sequence position controls the preferred inter-helical angle. Cooperative backbone contacts progressively rigidify the multi-alphaKL interface beyond what individual motifs achieve alone. By linking helices along their edges rather than their ends, the alphaKL expands the structural design space of programmable RNA nanostructures, unlocking previously inaccessible architectures and applications.

biophysics↗

bioRxiv2026

Fluorinated RNA origami enables serum-stable nanodevices for sensing and targeting

Chemically modified RNAs with increased stability and reduced immunogenicity have transformed RNA therapeutics. Rational RNA design methods, including RNA origami, seek to further extend RNA medicine and biotechnology by encoding advanced functions such as signalling, targeting, and controlled release within the RNA polymer. However, current design methods lack the ability to integrate chemical modification or predict how it shapes the structure of large RNA assemblies inhibiting its use in RNA therapeutics. Here we demonstrate that 2-fluoro pyrimidine RNA (FY-RNA) origami structures can be co-transcriptionally folded to generate serum-stable nanodevices. Cryogenic electron microscopy reveals that FY-RNA can alter folding pathways and perturb tertiary motifs, while molecular dynamics simulations show how 2-fluoro modification affects hydrogen bonding, sugar pucker, and helix-helix interactions. Despite these structural perturbations, fluorogenic aptamers embedded within RNA origami retain partial activity and enable logic-based molecular sensing in human serum. Finally, we use an FY-RNA scaffold to determine the structure of an FY-RNA anti-Spike aptamer bound to the Spike protein at 3.4 [A] resolution, uncovering fluorine-specific structural motifs and protein interactions. Together, our results establish design principles for nuclease-resistant RNA architectures and position FY-RNA as a versatile polymer for constructing medical nanodevices and environmental sensors. More broadly, this work provides a framework for systematically exploring the folding landscape of chemically modified RNAs, expanding the chemical and functional diversity accessible to nucleic acid nanotechnology and RNA medicine.

biochemistry↗

bioRxiv2023

An RNA origami robot that traps and releases a fluorescent aptamer

RNA nanotechnology aims at using RNA as a programmable material to create self-assembling nanodevices for application in medicine and synthetic biology. RNA devices have been developed by adopting mechanisms such as allosteric binding and toehold-mediated strand displacement. There are, however, no examples of RNA "robotic" devices that sense, compute, and actuate through mechanical reconfiguration as has been demonstrated in DNA nanotechnology. Here we use the RNA origami method to prototype an RNA robotic device, named the "Traptamer", that senses two RNA key strands, acts as a Boolean AND gate, and activates the fluorescent aptamer iSpinach through release from a mechanical trap. The Traptamer depends on binding of two different RNA key strands to achieve full activation and can be reversed by addition of two complementary RNA anti-key strands. Cryo-EM of the closed Traptamer structure at 5.45 [A] resolution reveals a hinge-like mechanical distortion of the iSpinach motif. Our RNA robot prototype opens the door to build more sophisticated RNA machines that use sensing, computing, and acting modules to precisely control RNA functionalities.

bioengineering↗

bioRxiv2023

Tertiary structure of single-instant RNA molecule reveals folding landscape

The folding of RNA and protein molecules during their synthesis is a crucial self-assembly process that nature employs to convert genetic information into the complex molecular machinery that supports life. Misfolding events are the cause of several diseases, and the folding pathway of central biomolecules, such as the ribosome, is strictly regulated by programmed maturation processes and folding chaperones. However, the dynamic folding processes are challenging to study because current structure determination methods heavily rely on averaging, and existing computational methods do not efficiently simulate non-equilibrium dynamics. Here we utilize individual-particle cryo-electron tomography (IPET) to investigate the folding landscape of a rationally designed RNA origami 6-helix bundle that undergoes slow maturation from a "young" to "mature" conformation. By optimizing the IPET imaging and electron dose conditions, we obtain 3D reconstructions of 120 individual particles at resolutions ranging from 23-35 [A], enabling us first-time to observe individual RNA helices and tertiary structures without averaging. Statistical analysis of 120 tertiary structures confirms the two main conformations and suggests a possible folding pathway driven by helix-helix compaction. Studies of the full conformational landscape reveal both trapped states, misfolded states, intermediate states, and fully compacted states. The study provides novel insight into RNA folding pathways and paves the way for future studies of the energy landscape of molecular machines and self-assembly processes.

biochemistry↗