The folding space of protein β2-microglobulin is modulated by a single disulfide bond

Protein beta-2-microglobulin (β2m) is classically considered the causative agent of dialysis related amyloidosis (DRA), a conformational disorder that affects patients undergoing long-term hemodialysis. Together with the wild type form, the ΔN6 structural variant, and the D76N mutant, have been extensively used as model systems of β2m aggregation. In all of them, the native structure is stabilized by a disulfide bridge between the sulphur atoms of the cysteine residues 25 (at B strand) and 80 (at F strand), which has been considered fundamental in β2m fibrillogenesis. Here, we use extensive Discrete Molecular Dynamics simulations of a full atomistic structure-based model to explore the role of this disulfide bridge as a modulator of the folding space of β2m. In particular, by considering different models for the disulfide bridge, we explore the thermodynamics of the folding transition, and the formation of intermediate states that may have the potential to trigger the aggregation cascade. Our results show that the dissulfide bridge affects folding transition and folding thermodynamics of the considered model systems, although to different extents. In particular, when the interaction between the sulphur atoms is stabilized relative to the other intramolecular interactions, or even locked (i.e. permanently established), the WT form populates an intermediate state featuring a well preserved core, and two unstructured termini, which was previously detected only for the D76N mutant. The formation of this intermediate state may have important implications in our understanding of β2m fibrillogenesis.

Simulation of Folding Kinetics for Aligned RNAs

Studying the folding kinetics of an RNA can provide insight into its function and is thus a valuable method for RNA analyses. Computational approaches to the simulation of folding kinetics suffer from the exponentially large folding space that needs to be evaluated. Here, we present a new approach that combines structure abstraction with evolutionary conservation to restrict the analysis to common parts of folding spaces of related RNAs. The resulting algorithm can recapitulate the folding kinetics known for single RNAs and is able to analyse even long RNAs in reasonable time. Our program RNAliHiKinetics is the first algorithm for the simulation of consensus folding kinetics and addresses a long-standing problem in a new and unique way.

RNAxplorer: harnessing the power of guiding potentials to sample RNA landscapes

Motivation Predicting the folding dynamics of RNAs is a computationally difficult problem, first and foremost due to the combinatorial explosion of alternative structures in the folding space. Abstractions are therefore needed to simplify downstream analyses, and thus make them computationally tractable. This can be achieved by various structure sampling algorithms. However, current sampling methods are still time consuming and frequently fail to represent key elements of the folding space. Method We introduce RNAxplorer, a novel adaptive sampling method to efficiently explore the structure space of RNAs. RNAxplorer uses dynamic programming to perform an efficient Boltzmann sampling in the presence of guiding potentials, which are accumulated into pseudo-energy terms and reflect similarity to already well-sampled structures. This way, we effectively steer sampling toward underrepresented or unexplored regions of the structure space. Results We developed and applied different measures to benchmark our sampling methods against its competitors. Most of the measures show that RNAxplorer produces more diverse structure samples, yields rare conformations that may be inaccessible to other sampling methods and is better at finding the most relevant kinetic traps in the landscape. Thus, it produces a more representative coarse graining of the landscape, which is well suited to subsequently compute better approximations of RNA folding kinetics. Availabilityand implementation https://github.com/ViennaRNA/RNAxplorer/. Supplementary information Supplementary data are available at Bioinformatics online.

RNAxplorer: Harnessing the Power of Guiding Potentials to Sample RNA Landscapes

MotivationPredicting the folding dynamics of RNAs is a computationally difficult problem, first and foremost due to the combinatorial explosion of alternative structures in the folding space. Abstractions are therefore needed to simplify downstream analyses, and thus make them computationally tractable. This can be achieved by various structure sampling algorithms. However, current sampling methods are still time consuming and frequently fail to represent key elements of the folding space.MethodWe introduceRNAxplorer, a novel adaptive sampling method to efficiently explore the structure space of RNAs.RNAxploreruses dynamic programming to perform an efficient Boltzmann sampling in the presence of guiding potentials, which are accumulated into pseudo-energy terms and reflect similarity to already well-sampled structures. This way, we effectively steer sampling towards underrepresented or unexplored regions of the structure space.ResultsWe developed and applied different measures to benchmark our sampling methods against its competitors. Most of the measures show thatRNAxplorerproduces more diverse structure samples, yields rare conformations that may be inaccessible to other sampling methods and is better at finding the most relevant kinetic traps in the landscape. Thus, it produces a more representative coarse graining of the landscape, which is well suited to subsequently compute better approximations of RNA folding kinetics.Availabilityhttps://github.com/ViennaRNA/RNAxplorer/[email protected],[email protected]

Structure and function of virion RNA polymerase of crAss-like phage

CrAss-like phages are a recently described family-level group of viruses that includes the most abundant virus in the human gut1,2. Genomes of all crAss-like phages encode a large virion-packaged protein2,3 that contains a DFDxD sequence motif, which forms the catalytic site in cellular multisubunit RNA polymerases (RNAPs)4. Using Cellulophaga baltica crAss-like phage phi14:2 as a model system, we show that this protein is a novel DNA-dependent RNAP that is translocated into the host cell along with the phage DNA and transcribes early phage genes. We determined the crystal structure of this 2,180-residue enzyme in a self-inhibited, likely pre-virion-packaged state. This conformation is attained with the help of a Cleft-blocking domain that interacts with the active site motif and occupies the RNA-DNA hybrid binding grove. Structurally, phi14:2 RNAP is most similar to eukaryotic RNAPs involved in RNA interference5,6, although most of phi14:2 RNAP structure (nearly 1,600 residues) maps to a new region of protein folding space. Considering the structural similarity, we propose that eukaryal RNA interference polymerases take their origin in a phage, which parallels the emergence of the mitochondrial transcription apparatus7.