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At least 793 records · Page 44Linked to original sources
PubMed1979

Attenuation in the Escherichia coli tryptophan operon: role of RNA secondary structure involving the tryptophan codon region.

The secondary structure of the terminated trp leader transcript from Escherichia coli was analyzed by RNase T1 partial digestion. Base-paired regions were recovered by nondenaturing gel electrophoresis and identified by denaturing gel electrophoresis and fingerprinting. The tandem tryptophan codons in the leader peptide coding region were found to be base paired with a more distal region of the transcript. This and other secondary structures that the trp leader RNA can form help explain the physiological response of the operon as well as the behavior of regulatory mutants.

Codon↗
PubMed1999

Estimating the contributions of selection and self-organization in RNA secondary structure.

In addition to characteristic structural properties imposed by evolutionary modification, evolved, single-stranded RNAs also display characteristic structural properties imposed by intrinsic physical constraints on RNA polymer folding. The balance of intrinsic and functionally selected characters in the folded conformation of evolved secondary structures was determined by comparing the predicted secondary structures of evolved and unevolved (random) RNA sequences. Though evolved conformations are significantly more ordered than conformations of random-sequence RNA, this analysis demonstrates that the majority of conformational order within evolved structures results not from evolutionary optimization but from constraints imposed by rules intrinsic to RNA polymer folding.

Algorithms↗
PubMed1997

Structure-based discovery of ligands targeted to the RNA double helix.

Ligands capable of specific recognition of RNA structures are of interest in terms of the principles of molecular recognition as well as potential chemotherapeutic applications. We have approached the problem of identifying small molecules with binding specificity for the RNA double helix through application of the DOCK program [Kuntz, I. D., Meng, E. C., and Shoichet, B. K. (1994) Acc. Chem. Res. 27, 117-123], a structure-based method for drug discovery. A series of lead compounds was generated through a database search for ligands with shape complementarity to the RNA deep major groove. Compounds were then evaluated with regard to their fit into the minor groove of B DNA. Those compounds predicted to have an optimal fit to the RNA groove and strong discrimination against DNA were examined experimentally. Of the 11 compounds tested, 3, all aminoglycosides, exhibited pronounced stabilization of RNA duplexes against thermal denaturation with only marginal effects on DNA duplexes. One compound, lividomycin, was examined further, and shown to facilitate the ethanol-induced B to A transition in calf thymus DNA. Fluorine NMR solvent isotope shift measurements on RNA duplexes containing 5-fluorouracil provided evidence that lividomycin binds in the RNA major groove. Taken together, these results indicate that lividomycin recognizes the general features of the A conformation of nucleic acids through deep groove binding, confirming the predictions of our DOCK analysis. This approach may be of general utility for identifying ligands possessing specificity for additional RNA structures as well as other nucleic acid structural motifs.

Aminoglycosides↗
PubMed2003

A cyclopentane conformational restraint for a peptide nucleic acid: design, asymmetric synthesis, and improved binding affinity to DNA and RNA.

[structure: see text] A strategy to restrict the highly flexible backbone conformation of a peptide nucleic acid (PNA) by incorporation of a cyclopentane ring is proposed. An asymmetric synthesis of cyclopentane-modified PNA is reported, and its binding properties were determined. The cyclopentane ring leads to a significant improvement in the binding properties of the resulting PNA to DNA and RNA.

Alkylation↗
PubMed1969

Studies on a nucleoprotein prepared from rat liver polysomes by digestion with T1 ribonuclease.

1. Treatment of rat liver polysomes in a buffer containing 2.5mm-magnesium chloride with T(1) ribonuclease at a concentration of 330units/ml. of reaction medium at 37 degrees for 2hr. leads to the production of an insoluble nucleoprotein. 2. On the bases of analysis for protein and RNA and of u.v.-absorption spectra the nucleoprotein appears to have lost approx. 60% of the structural RNA originally present in the ribosome. Degradation of (3)H-labelled polysomes (structural RNA labelled with orotic acid) with T(1) ribonuclease leads to nucleoprotein preparations retaining approx. 30% of the radioactivity originally present in the polysomes. By means of sucrose-density-gradient centrifugation it is shown that the nucleoprotein preparations are free of single 73s ribosomes and ribosomal subunits. No evidence for the presence of 28s and 18s structural RNA was obtained on examination of extracted nucleoprotein-particle RNA by means of sucrose-density-gradient centrifugation. 3. Digestion of washed polysomes carrying (14)C-labelled nascent peptide chains with T(1) ribonuclease gives a nucleoprotein particle that retains approx. 70% of the original labelled chains. Treatment of labelled nucleoprotein particles with 1mm-puromycin in the absence of transfer factors releases 20% of the labelled chains. Addition of GTP (0.48mumole) increases this release to 37%. 4. Treatment of nucleoprotein particles carrying (14)C-labelled peptide chains with either EDTA (50mm) or ammonium chloride (0.5m) brings about a small release of labelled material (approx. 15%). 5. Disruption of nucleoprotein particles carrying (14)C-labelled peptide chains with either sodium dodecyl sulphate or 2m-lithium chloride, followed by addition of transfer RNA as marker and chromatography on Sephadex G-200, show in both cases that considerable amounts of labelled peptide material move well ahead of the added transfer RNA marker. Further, if nucleoprotein particles carrying labelled peptide chains are treated with 0.3m-potassium hydroxide at 20 degrees for 24 hr., neutralized to pH7.6, and then chromatographed on Sephadex G-200, the labelled peptide material moves much closer to the added transfer RNA marker. These results suggest that a proportion of the nascent (14)C-labelled peptides on the nucleoprotein are attached to transfer RNA or large fragments of transfer RNA. 6. [(3)H]Polyuridylic acid binds to nucleoprotein particles in 1mm-magnesium chloride. The rate of binding is rapid when measured at 20 degrees .

Ammonium Chloride↗
PubMed2004

Application of NMR and EPR methods to the study of RNA.

The application of techniques based on magnetic resonance, specifically electron paramagnetic resonance (EPR) and nuclear magnetic resonance (NMR), has provided a wealth of new information on RNA structures, as well as insights into the dynamics and function of these important biomolecules. NMR spectroscopy is very successful for determining the solution structures of small RNA domains, aptamers and ribozymes, and exploring their intramolecular dynamics and interactions with ligands. EPR-based methods have been used to map local dynamic and structural features of RNA, to explore different modes of RNA-ligand interaction, to obtain long-range structural restraints and to probe metal-ion-binding sites.

Binding Sites↗
PubMedDate not supplied

Pseudoknots in RNA secondary structures: representation, enumeration, and prevalence.

A number of non-coding RNA are known to contain functionally important or conserved pseudoknots. However, pseudoknotted structures are more complex than orthodox, and most methods for analyzing secondary structures do not handle them. I present here a way to decompose and represent general secondary structures which extends the tree representation of the stem-loop structure, and use this to analyze the frequency of pseudoknots in known and in random secondary structures. This comparison shows that, though a number of pseudoknots exist, they are still relatively rare and mostly of the simpler kinds. In contrast, random secondary structures tend to be heavily knotted, and the number of available structures increases dramatically when allowing pseudoknots. Therefore, methods for structure prediction and non-coding RNA identification that allow pseudoknots are likely to be much less powerful than those that do not, unless they penalize pseudoknots appropriately.

Algorithms↗
PubMed2005

HIV-1 can escape from RNA interference by evolving an alternative structure in its RNA genome.

HIV-1 replication can be efficiently inhibited by intracellular expression of an siRNA targeting the viral RNA. However, HIV-1 escape variants emerged after prolonged culturing. These RNAi-resistant viruses contain nucleotide substitutions or deletions in or near the targeted sequence. We observed an inverse correlation between the level of resistance and the stability of the siRNA/target-RNA duplex. However, two escape variants showed a higher level of resistance than expected based on the duplex stability. We demonstrate that these mutations induce alternative folding of the RNA such that the target sequence is occluded from binding to the siRNA, resulting in reduced RNAi efficiency. HIV-1 can thus escape from RNAi-mediated inhibition not only through nucleotide substitutions or deletions in the siRNA target sequence, but also through mutations that alter the local RNA secondary structure. The results highlight the enormous genetic flexibility of HIV-1 and provide detailed molecular insight into the sequence specificity of RNAi and the impact of target RNA secondary structure.

Anti-HIV Agents↗
PubMed1991

Recognition of the folding consensus in RNA secondary structures by the topological-filtering method.

Functionally homologous RNA sequences can substantially diverge in their primary sequences but it can be reasonably assumed that they are related in their higher-degree structures. The problem to find such structures and simultaneously satisfy as far as possible the free-energy-minimization criterion, is considered here in two aspects. Firstly a quantitative measure of the folding consensus among secondary structures is defined, translating each structure into a linear representation and using the correlation theorem to compare them. Secondly an algorithm for the parallel search for secondary structures according to the free-energy-minimization criterion, but with a filtering action on the basis of the folding consensus measure is presented. The method is tested on groups of RNA sequences different in origin and in functions, for which proposals of homologous secondary structures based on experimental data exist. A comparison of the results with a blank consisting of a search on the basis of the free energy minimization alone is always performed. In these tests the method shows its ability in obtaining, from different sequences, secondary structures characterized by a high-folding consensus measure also when lower free energy but not homologous structures are possible. Two applications are also shown. The first demonstrates the transfer of experimental data available for one sequence, to a functionally related and therefore homologous one. The second application is the possibility of using a topological probe in the search for precise structural motifs.

Algorithms↗
PubMed2004

Prediction of locally stable RNA secondary structures for genome-wide surveys.

MOTIVATION: Recently novel classes of functional RNAs, most prominently the miRNAs have been discovered, strongly suggesting that further types of functional RNAs are still hidden in the recently completed genomic DNA sequences. Only few techniques are known, however, to survey genomes for such RNA genes. When sufficiently similar sequences are not available for comparative approaches the only known remedy is to search directly for structural features. RESULTS: We present here efficient algorithms for computing locally stable RNA structures at genome-wide scales. Both the minimum energy structure and the complete matrix of base pairing probabilities can be computed in theta(N x L2) time and theta(N + L2) memory in terms of the length N of the genome and the size L of the largest secondary structure motifs of interest. In practice, the 100 Mb of the complete genome of Caenorhabditis elegans can be folded within about half a day on a modern PC with a search depth of L = 100. This is sufficient example for a survey for miRNAs. AVAILABILITY: The software described in this contribution will be available for download at http://www.tbi.univie.ac.at/~ivo/RNA/ as part of the Vienna RNA Package.

Algorithms↗
PubMed2006

RNAmute: RNA secondary structure mutation analysis tool.

BACKGROUND: RNAMute is an interactive Java application that calculates the secondary structure of all single point mutations, given an RNA sequence, and organizes them into categories according to their similarity with respect to the wild type predicted structure. The secondary structure predictions are performed using the Vienna RNA package. Several alternatives are used for the categorization of single point mutations: Vienna's RNAdistance based on dot-bracket representation, as well as tree edit distance and second eigenvalue of the Laplacian matrix based on Shapiro's coarse grain tree graph representation. RESULTS: Selecting a category in each one of the processed tables lists all single point mutations belonging to that category. Selecting a mutation displays a graphical drawing of the single point mutation and the wild type, and includes basic information such as associated energies, representations and distances. RNAMute can be used successfully with very little previous experience and without choosing any parameter value alongside the initial RNA sequence. The package runs under LINUX operating system. CONCLUSION: RNAMute is a user friendly tool that can be used to predict single point mutations leading to conformational rearrangements in the secondary structure of RNAs. In several cases of substantial interest, notably in virology, a point mutation may lead to a loss of important functionality such as the RNA virus replication and translation initiation because of a conformational rearrangement in the secondary structure.

Animals↗
PubMed2004

ILM: a web server for predicting RNA secondary structures with pseudoknots.

The ILM web server provides a web interface to two algorithms, iterated loop matching and maximum weighted matching, for efficiently predicting RNA secondary structures with pseudoknots. The algorithms can utilize either thermodynamic or comparative information or both, and thus can work on both aligned and individual sequences. Predicted secondary structures are presented in several formats compatible with a variety of existing visualization tools. The service can be accessed at http://cic.cs.wustl.edu/RNA/.

Algorithms↗
PubMed1978

The evolution of 5S RNA secondary structures.

We have applied the Pipas-McMahon algorithm based on free energy calculations to the search for a 5S RNA base-pair structure common to all known sequences. We find that a 'Y' shaped model is consistently among the structures having the lowest free energy using 5S RNA sequences from either eukaryotic or prokaryotic sources. Compaison of this 'Y' structure with models which have recently been proposed show these models to be remarkably similar, and the minor differences are explicable based on the technique used to obtain the model. That prokaryotic and eukaryotic 5S RNA can adopt a similar secondary structure is strong support for its resistance to change during evolution.

Animals↗
PubMed2025

CAR-SPLASH identifies nascent pre-mRNA structures implicated in kinetic coupling and alternative splicing.

Pre-mRNA splicing is kinetically coupled to transcription as shown by the widespread effects of transcription speed on alternative splicing (AS) outcomes. The molecular basis for such kinetic coupling is incompletely understood, but one potential mechanism is through elongation rate-dependent alternative folding pathways of the nascent pre-messenger RNA (pre-mRNA). To search for RNA structures in nascent pre-mRNA, we modified Sequencing of Psoralen Crosslinked, Ligated And Selected Hybrids (SPLASH) [J. G. Ashley Aw et al., Mol. Cell 62, 603-617 (2016)] for use with Chromatin Associated RNA. We applied this method called Chromatin Associated RNA (CAR)-SPLASH to cells expressing wild-type and slow mutant RNA polymerase II and identified >3,000 intramolecular RNA duplexes of which >400 are proximal to splice sites. Antisense oligonucleotide (ASO) disruption of several such duplexes that sequester splice sites has a major impact on AS outcomes, even though the ASOs do not directly disrupt splice sites. ASO disruption of these regulatory elements that we designate "RNA kinetic switches" modified AS of NISCH Exon 18, GAK Exon 7, and MEGF8 Exon 14 in a way that depends on the rate of transcription elongation. We propose that these switches mediate kinetic coupling via the effects of transcription speed on folding of nascent RNA structures that modulate AS and that many nascent RNA structures can thereby serve as targets for splice-modifying ASOs.

RNA Precursors↗
PubMed1990

Stable RNA secondary structure in a retroviral vector insert terminates reverse transcriptase elongation in vitro but not in cultured cells.

We wished to test whether an RNA signal that causes termination of elongation by reverse transcriptase in vitro would affect retroviral vector function. A synthetic oligonucleotide containing a sequence capable of forming a very stable RNA secondary structure was subcloned into the retrovirus vector N2. The integration of this sequence into N2 causes termination of elongation by reverse transcriptase in vitro at the precise positions previously reported in a different sequence context. However, no premature termination of DNA synthesis was observed in the unintegrated DNA of vector transduced cells. Likewise, there was no deleterious effect of the sequence insert on vector titer. These results indicate that termination signals defined in in vitro systems cannot be used as predictors of in vivo function and suggest that viral proteins in addition to reverse transcriptase play an important role in transcript initiation and elongation.

Animals↗
PubMed2001

A novel family of RNA tetraloop structure forms the recognition site for Saccharomyces cerevisiae RNase III.

RNases III are a family of double-stranded RNA (dsRNA) endoribonucleases involved in the processing and decay of a large number of cellular RNAs as well as in RNA interference. The dsRNA substrates of Saccharomyces cerevisiae RNase III (Rnt1p) are capped by tetraloops with the consensus sequence AGNN, which act as the primary docking site for the RNase. We have solved the solution structures of two RNA hairpins capped by AGNN tetraloops, AGAA and AGUU, using NMR spectroscopy. Both tetraloops have the same overall structure, in which the backbone turn occurs on the 3' side of the syn G residue in the loop, with the first A and G in a 5' stack and the last two residues in a 3' stack. A non-bridging phosphate oxygen and the universal G which are essential for Rnt1p binding are strongly exposed. The compared biochemical and structural analysis of various tetraloop sequences defines a novel family of RNA tetraloop fold with the consensus (U/A)GNN and implicates this conserved structure as the primary determinant for specific recognition of Rnt1p substrates.

Base Sequence↗
PubMed2003

Compensatory evolution of a precursor messenger RNA secondary structure in the Drosophila melanogaster Adh gene.

Evidence for the evolutionary maintenance of a hairpin structure possibly involved in intron processing had been found in intron 1 of the alcohol dehydrogenase gene (Adh) in diverse Drosophila species. In this study, the putative hairpin structure was evaluated systematically in Drosophila melanogaster by elimination of either side of the stem using site-directed mutagenesis. The effects of these mutations and the compensatory double mutant on intron splicing efficiency and ADH protein production were assayed in Drosophila melanogaster Schneider L2 cells and germ-line transformed adult flies. Mutations that disrupt the putative hairpin structure right upstream of the intron branch point were found to cause a significant reduction in both splicing efficiency and ADH protein production. In contrast, the compensatory double mutant that restores the putative hairpin structure was indistinguishable from the WT in both splicing efficiency and ADH level. It was also observed by mutational analysis that a more stable secondary structure (with a longer stem) in this intron decreases both splicing efficiency and ADH protein production. Implications for RNA secondary structure and intron evolution are discussed.

Alcohol Dehydrogenase↗
PubMed2000

Escherichia coli RNA polymerase core and holoenzyme structures.

Multisubunit RNA polymerase is an essential enzyme for regulated gene expression. Here we report two Escherichia coli RNA polymerase structures: an 11.0 A structure of the core RNA polymerase and a 9.5 A structure of the sigma(70) holoenzyme. Both structures were obtained by cryo-electron microscopy and angular reconstitution. Core RNA polymerase exists in an open conformation. Extensive conformational changes occur between the core and the holoenzyme forms of the RNA polymerase, which are largely associated with movements in ss'. All common RNA polymerase subunits (alpha(2), ss, ss') could be localized in both structures, thus suggesting the position of sigma(70) in the holoenzyme.

Bacterial Proteins↗