Erratum: Structure of the 70S ribosome from human pathogen Staphylococcus aureus (Nucleic Acids Research (2017) DOI: 10.1093/nar/gkw933)

© The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.The authors wish to correct their Funding statement as follows: FUNDING. 'Centre National de la Recherche Scientifique' (CNRS) and the 'Agence Nationale de la Recherche' as part of the 'Investissements d'Avenir' program [LabEx: ANR-10-LABX-0036-NETRNA to P.R., Y.H.; ANR-15-CE11-0021-01 to G.Y.]; 'Fondation pour la Recherche Médicale en France' [FDT20140930867 to I.K; 'European Research Council advanced grant' [294312 to M.Y.]; the 'Russian Science Foundation' [Project No. 16-14-10014 to I.K., M.Y.]. Funding for open access charge: Centre National de la Recherche Scientifique (CNRS). In addition, Marat Yusupov is associated with both affiliations1 and 2. The authors apologise to the readers for this error

Structure of the 70S ribosome from human pathogen Staphylococcus aureus

© 2016 The Author(s).Comparative structural studies of ribosomes from various organisms keep offering exciting insights on how species-specific or environment-related structural features of ribosomes may impact translation specificity and its regulation. Although the importance of such features may be less obvious within more closely related organisms, their existence could account for vital yet species-specific mechanisms of translation regulation that would involve stalling, cell survival and antibiotic resistance. Here, we present the first full 70S ribosome structure from Staphylococcus aureus, a Gram-positive pathogenic bacterium, solved by cryo-electron microscopy. Comparative analysis with other known bacterial ribosomes pinpoints several unique features specific to S. aureus around a conserved core, at both the protein and the RNA levels. Our work provides the structural basis for the many studies aiming at understanding translation regulation in S. aureus and for designing drugs against this often multi-resistant pathogen

Preparation of Group I Introns for Biochemical Studies and Crystallization Assays by Native Affinity Purification

The study of functional RNAs of various sizes and structures requires efficient methods for their synthesis and purification. Here, 23 group I intron variants ranging in length from 246 to 341 nucleotides—some containing exons—were subjected to a native purification technique previously applied only to shorter RNAs (<160 nucleotides). For the RNAs containing both exons, we adjusted the original purification protocol to allow for purification of radiolabeled molecules. The resulting RNAs were used in folding assays on native gel electrophoresis and in self-splicing assays. The intron-only RNAs were subjected to the regular native purification scheme, assayed for folding and employed in crystallization screens. All RNAs that contained a 3′ overhang of one nucleotide were efficiently cleaved off from the support and were at least 90% pure after the non-denaturing purification. A representative subset of these RNAs was shown to be folded and self-splicing after purification. Additionally, crystals were grown for a 286 nucleotide long variant of the Clostridium botulinum intron. These results demonstrate the suitability of the native affinity purification method for the preparation of group I introns. We hope these findings will stimulate a broader application of this strategy to the preparation of other large RNA molecules

Understanding the Origins of Bacterial Resistance to Aminoglycosides through Molecular Dynamics Mutational Study of the Ribosomal A-Site

Paromomycin is an aminoglycosidic antibiotic that targets the RNA of the bacterial small ribosomal subunit. It binds in the A-site, which is one of the three tRNA binding sites, and affects translational fidelity by stabilizing two adenines (A1492 and A1493) in the flipped-out state. Experiments have shown that various mutations in the A-site result in bacterial resistance to aminoglycosides. In this study, we performed multiple molecular dynamics simulations of the mutated A-site RNA fragment in explicit solvent to analyze changes in the physicochemical features of the A-site that were introduced by substitutions of specific bases. The simulations were conducted for free RNA and in complex with paromomycin. We found that the specific mutations affect the shape and dynamics of the binding cleft as well as significantly alter its electrostatic properties. The most pronounced changes were observed in the U1406C∶U1495A mutant, where important hydrogen bonds between the RNA and paromomycin were disrupted. The present study aims to clarify the underlying physicochemical mechanisms of bacterial resistance to aminoglycosides due to target mutations

Ten Simple Rules for Effective Computational Research

<p>Ten Simple Rules for Effective Computational Research</p

A quick guide for building a successful bioinformatics community

“Scientific community” refers to a group of people collaborating together on scientific-research-related activities who also share common goals, interests, and values. Such communities play a key role in many bioinformatics activities. Communities may be linked to a specific location or institute, or involve people working at many different institutions and locations. Education and training is typically an important component of these communities, providing a valuable context in which to develop skills and expertise, while also strengthening links and relationships within the community. Scientific communities facilitate: (i) the exchange and development of ideas and expertise; (ii) career development; (iii) coordinated funding activities; (iv) interactions and engagement with professionals from other fields; and (v) other activities beneficial to individual participants, communities, and the scientific field as a whole. It is thus beneficial at many different levels to understand the general features of successful, high-impact bioinformatics communities; how individual participants can contribute to the success of these communities; and the role of education and training within these communities. We present here a quick guide to building and maintaining a successful, high-impact bioinformatics community, along with an overview of the general benefits of participating in such communities. This article grew out of contributions made by organizers, presenters, panelists, and other participants of the ISMB/ECCB 2013 workshop “The ‘How To Guide’ for Establishing a Successful Bioinformatics Network” at the 21st Annual International Conference on Intelligent Systems for Molecular Biology (ISMB) and the 12th European Conference on Computational Biology (ECCB)

Labeled EF-Tus for rapid kinetic studies of pretranslocation complex formation

The universally conserved translation elongation factor EF-Tu delivers aminoacyl(aa)-tRNA in the form of an aa-tRNA·EF-Tu·GTP ternary complex (TC) to the ribosome where it binds to the cognate mRNA codon within the ribosomal A-site, leading to formation of a pretranslocation (PRE) complex. Here we describe preparation of QSY9 and Cy5 derivatives of the variant E348C-EF-Tu that are functional in translation elongation. Together with fluorophore derivatives of aa-tRNA and of ribosomal protein L11, located within the GTPase associated center (GAC), these labeled EF-Tus allow development of two new FRET assays that permit the dynamics of distance changes between EF-Tu and both L11 (Tu-L11 assay) and aa-tRNA (Tu-tRNA assay) to be determined during the decoding process. We use these assays to examine: (i) the relative rates of EF-Tu movement away from the GAC and from aa-tRNA during decoding, (ii) the effects of the misreading-inducing antibiotics streptomycin and paromomycin on tRNA selection at the A-site, and (iii) how strengthening the binding of aa-tRNA to EF-Tu affects the rate of EF-Tu movement away from L11 on the ribosome. These FRET assays have the potential to be adapted for high throughput screening of ribosomal antibiotics

Ten Simple Rules for a Successful Cross-Disciplinary Collaboration

Cross-disciplinary collaborations have become an increasingly important part of science. They are seen as key if we are to find solutions to pressing, global-scale societal challenges, including green technologies, sustainable food production, and drug development. Regulators and policy- makers have realized the power of such collaborations, for example, in the 80 billion Euro "Horizon 2020" EU Framework Programme for Research and Innovation. This programme puts special emphasis on “breaking down barriers to create a genuine single market for knowledge, research and innovation