Composing RNA Nanostructures from a Syntax of RNA Structural Modules.

Natural stable RNAs fold and assemble into complex three-dimensional architectures by relying on the hierarchical formation of intricate, recurrent networks of noncovalent tertiary interactions. These sequence-dependent networks specify RNA structural modules enabling orientational and topological control of helical struts to form larger self-folding domains. Borrowing concepts from linguistics, we defined an extended structural syntax of RNA modules for programming RNA strands to assemble into complex, responsive nanostructures under both thermodynamic and kinetic control.

Stability of Cucumber Necrosis Virus at the Quasi-6-Fold Axis Affects Zoospore Transmission.

(CNV) is a member of the genus and has a monopartite positive-sense RNA genome. CNV is transmitted in nature via zoospores of the fungus As with other members of the genus, the CNV capsid swells when exposed to alkaline pH and EDTA. We previously demonstrated that a P73G mutation blocks the virus from zoospore transmission while not significantly affecting replication in plants (K.

Visualization and quality assessment of the contrast transfer function estimation.

The contrast transfer function (CTF) describes an undesirable distortion of image data from a transmission electron microscope. Many users of full-featured processing packages are often new to electron microscopy and are unfamiliar with the CTF concept. Here we present a common graphical output to clearly demonstrate the CTF fit quality independent of estimation software.

Engineered mutations change the structure and stability of a virus-like particle.

The single-coat protein (CP) of bacteriophage Qβ self-assembles into T = 3 icosahedral virus-like particles (VLPs), of interest for a wide range of applications. These VLPs are very stable, but identification of the specific molecular determinants of this stability is lacking. To investigate these determinants along with manipulations that confer more capabilities to our VLP material, we manipulated the CP primary structure to test the importance of various putative stabilizing interactions.

Software tools for molecular microscopy: an open-text Wikibook.

We provide a brief description of a Wikibook open-content textbook that was set up with a goal of providing a comprehensive and continually updated list of all of the software packages of interest to the cryo-EM community. While the content of the Wikibook will change over time, here we provide a snapshot of the current state of software tools available, and encourage the members of this community to view the pages, add content, correct errors, and make any other contributions that might be useful.

Automation in single-particle electron microscopy connecting the pieces.

Throughout the history of single-particle electron microscopy (EM), automated technologies have seen varying degrees of emphasis and development, usually depending upon the contemporary demands of the field. We are currently faced with increasingly sophisticated devices for specimen preparation, vast increases in the size of collected data sets, comprehensive algorithms for image processing, sophisticated tools for quality assessment, and an influx of interested scientists from outside the field who might lack the skills of experienced microscopists. This situation places automated techniques in high demand.

In vitro assembly of cubic RNA-based scaffolds designed in silico.

The organization of biological materials into versatile three-dimensional assemblies could be used to build multifunctional therapeutic scaffolds for use in nanomedicine. Here, we report a strategy to design three-dimensional nanoscale scaffolds that can be self-assembled from RNA with precise control over their shape, size and composition. These cubic nanoscaffolds are only approximately 13 nm in diameter and are composed of short oligonucleotides, making them amenable to chemical synthesis, point modifications and further functionalization.

A polyhedron made of tRNAs.

Supramolecular assembly is a powerful strategy used by nature to build nanoscale architectures with predefined sizes and shapes. With synthetic systems, however, numerous challenges remain to be solved before precise control over the synthesis, folding and assembly of rationally designed three-dimensional nano-objects made of RNA can be achieved. Here, using the transfer RNA molecule as a structural building block, we report the design, efficient synthesis and structural characterization of stable, modular three-dimensional particles adopting the polyhedral geometry of a non-uniform square antiprism.

3V: cavity, channel and cleft volume calculator and extractor.

As larger macromolecular structures become available, there is a growing need to understand their 'internal' volumes--such as deep clefts, channels and cavities--as these often play critical roles in their function. The 3V web server can automatically extract and comprehensively analyze all the internal volumes from input RNA and protein structures. It rapidly finds internal volumes by taking the difference between two rolling-probe solvent-excluded surfaces, one with as large as possible a probe radius and the other with a solvent radius (typically 1.

High-resolution cryo-electron microscopy structures of murine norovirus 1 and rabbit hemorrhagic disease virus reveal marked flexibility in the receptor binding domains.

Our previous structural studies on intact, infectious murine norovirus 1 (MNV-1) virions demonstrated that the receptor binding protruding (P) domains are lifted off the inner shell of the virus. Here, the three-dimensional (3D) reconstructions of recombinant rabbit hemorrhagic disease virus (rRHDV) virus-like particles (VLPs) and intact MNV-1 were determined to approximately 8-A resolution. rRHDV also has a raised P domain, and therefore, this conformation is independent of infectivity and genus.

A toolbox for ab initio 3-D reconstructions in single-particle electron microscopy.

Structure determination of a novel macromolecular complex via single-particle electron microscopy depends upon overcoming the challenge of establishing a reliable 3-D reconstruction using only 2-D images. There are a variety of strategies that deal with this issue, but not all of them are readily accessible and straightforward to use. We have developed a "toolbox" of ab initio reconstruction techniques that provide several options for calculating 3-D volumes in an easily managed and tightly controlled work-flow that adheres to standard conventions and formats.

A test-bed for optimizing high-resolution single particle reconstructions.

It is becoming routine for cryoEM single particle reconstructions to result in 3D electron density maps with resolutions of approximately 10A, but maps with resolutions of 5A or better are still celebrated events. The electron microscope has a resolving power to better than 2A, and thus should not be a limiting factor; instead the practical limitations in resolution most likely arise from a combination of specimen preparation methods, data collection parameters, and data analysis procedures. With the aid of a highly automated system for acquiring images, coupled to a relational database to keep track of all processing parameters, we have taken a systematic approach to optimizing parameters affecting the resolution of single particle reconstructions.