The tRNA identity landscape for aminoacylation and beyond.

tRNAs are key partners in ribosome-dependent protein synthesis. This process is highly dependent on the fidelity of tRNA aminoacylation by aminoacyl-tRNA synthetases and relies primarily on sets of identities within tRNA molecules composed of determinants and antideterminants preventing mischarging by non-cognate synthetases. Such identity sets were discovered in the tRNAs of a few model organisms, and their properties were generalized as universal identity rules.

History of tRNA research in strasbourg.

The tRNA molecules, in addition to translating the genetic code into protein and defining the second genetic code via their aminoacylation by aminoacyl-tRNA synthetases, act in many other cellular functions and dysfunctions. This article, illustrated by personal souvenirs, covers the history of ~60 years tRNA research in Strasbourg. Typical examples point up how the work in Strasbourg was a two-way street, influenced by and at the same time influencing investigators outside of France.

Structure of Escherichia coli Arginyl-tRNA Synthetase in Complex with tRNA: Pivotal Role of the D-loop.

Aminoacyl-tRNA synthetases are essential components in protein biosynthesis. Arginyl-tRNA synthetase (ArgRS) belongs to the small group of aminoacyl-tRNA synthetases requiring cognate tRNA for amino acid activation. The crystal structure of Escherichia coli (Eco) ArgRS has been solved in complex with tRNA at 3.

What macromolecular crystallogenesis tells us - what is needed in the future.

Crystallogenesis is a longstanding topic that has transformed into a discipline that is mainly focused on the preparation of crystals for practising crystallo-graphers. Although the idiosyncratic features of proteins have to be taken into account, the crystallization of proteins is governed by the same physics as the crystallization of inorganic materials. At present, a diversified panel of crystallization methods adapted to proteins has been validated, and although only a few methods are in current practice, the success rate of crystallization has increased constantly, leading to the determination of ∼10 X-ray structures.

Aminoacyl-tRNA Synthetases in the Bacterial World.

Aminoacyl-tRNA synthetases (aaRSs) are modular enzymes globally conserved in the three kingdoms of life. All catalyze the same two-step reaction, i.e.

Interplay between Catalysts and Substrates for Activity of Class Ib Aminoacyl-tRNA Synthetases and Implications for Pharmacology.

Aminoacyl-tRNA synthetase:transfer RNA (aaRS:tRNA) systems became recently essential targets in molecular medicine, because perturbed recognition of cognate tRNAs by aaRSs and poor precision in tRNA aminoacylation do not guarantee accurate protein biosynthesis, thus leading to diseases. Sets of identity determinants situated at particular zones of tRNA are responsible for functional accuracy. Recent work in X-ray crystallography has revealed various snapshots of aaRS:ligand complexes which represent the stages required for aminoacylation.

tRNA biology in mitochondria.

Mitochondria are the powerhouses of eukaryotic cells. They are considered as semi-autonomous because they have retained genomes inherited from their prokaryotic ancestor and host fully functional gene expression machineries. These organelles have attracted considerable attention because they combine bacterial-like traits with novel features that evolved in the host cell.

A historical perspective on protein crystallization from 1840 to the present day.

Protein crystallization has been known since 1840 and can prove to be straightforward but, in most cases, it constitutes a real bottleneck. This stimulated the birth of the biocrystallogenesis field with both 'practical' and 'basic' science aims. In the early years of biochemistry, crystallization was a tool for the preparation of biological substances.

Aminoacyl-tRNA Synthetases in the Bacterial World.

Aminoacyl-tRNAsynthetases (aaRSs) are modular enzymesglobally conserved in the three kingdoms of life. All catalyze the same two-step reaction, i.e.

Predicting protein crystallizability and nucleation.

The outcome of protein crystallization attempts is often uncertain due to inherent features of the protein or to the crystallization process that are not fully under control of the experimentalist. The aim of this contribution is to propose user-friendly tools that can increase the success rate of a protein crytallization project. Different bioinformatic approaches to predict the crystallization feasibility (before any crystallization attempts are undertaken) are discussed and a novel approach to assess the nucleation process of a given protein is proposed.

Structure of transfer RNAs: similarity and variability.

Transfer RNAs (tRNAs) are ancient molecules whose origin goes back to the beginning of life on Earth. Key partners in the ribosome-translation machinery, tRNAs read genetic information on messenger RNA and deliver codon specified amino acids attached to their distal 3'-extremity for peptide bond synthesis on the ribosome. In addition to this universal function, tRNAs participate in a wealth of other biological processes and undergo intricate maturation events.

Biocrystallography: past, present, future.

The evolution of biocrystallography from the pioneers' time to the present era of global biology is presented in relation to the development of methodological and instrumental advances for molecular sample preparation and structure elucidation over the last 6 decades. The interdisciplinarity of the field that generated cross-fertilization between physics- and biology-focused themes is emphasized. In particular, strategies to circumvent the main bottlenecks of biocrystallography are discussed.

Mutation of the mitochondrial tyrosyl-tRNA synthetase gene, YARS2, causes myopathy, lactic acidosis, and sideroblastic anemia--MLASA syndrome.

Mitochondrial respiratory chain disorders are a heterogeneous group of disorders in which the underlying genetic defect is often unknown. We have identified a pathogenic mutation (c.156C>G [p.

Crystal growth of proteins, nucleic acids, and viruses in gels.

Medium-sized single crystals with perfect habits and no defect producing intense and well-resolved diffraction patterns are the dream of every protein crystallographer. Crystals of biological macromolecules possessing these characteristics can be prepared within a medium in which mass transport is restricted to diffusion. Chemical gels (like polysiloxane) and physical gels (such as agarose) provide such an environment and are therefore suitable for the crystallisation of biological macromolecules.

Diversity and similarity in the tRNA world: overall view and case study on malaria-related tRNAs.

Transfer RNAs (tRNAs) are ancient macromolecules that have evolved under various environmental pressures as adaptors in translation in all forms of life but also towards alternative structures and functions. The present knowledge on both "canonical" and "deviating" signature motifs retrieved from vertical and horizontal sequence comparisons is briefly reviewed. Novel characteristics, proper to tRNAs from a given translation system, are revealed by a case study on the nuclear and organellar tRNA sets from malaria-related organisms.

Microfluidic chips for the crystallization of biomacromolecules by counter-diffusion and on-chip crystal X-ray analysis.

Microfluidic devices were designed to perform on micromoles of biological macromolecules and viruses the search and the optimization of crystallization conditions by counter-diffusion, as well as the on-chip analysis of crystals by X-ray diffraction. Chips composed of microchannels were fabricated in poly-dimethylsiloxane (PDMS), poly-methyl-methacrylate (PMMA) and cyclo-olefin-copolymer (COC) by three distinct methods, namely replica casting, laser ablation and hot embossing. The geometry of the channels was chosen to ensure that crystallization occurs in a convection-free environment.

Peculiar inhibition of human mitochondrial aspartyl-tRNA synthetase by adenylate analogs.

Human mitochondrial aminoacyl-tRNA synthetases (mt-aaRSs), the enzymes which esterify tRNAs with the cognate specific amino acid, form mainly a different set of proteins than those involved in the cytosolic translation machinery. Many of the mt-aaRSs are of bacterial-type in regard of sequence and modular structural organization. However, the few enzymes investigated so far do have peculiar biochemical and enzymological properties such as decreased solubility, decreased specific activity and enlarged spectra of substrate tRNAs (of same specificity but from various organisms and kingdoms), as compared to bacterial aaRSs.

Toward a more complete view of tRNA biology.

Transfer RNAs are ancient molecules present in all domains of life. In addition to translating the genetic code into protein and defining the second genetic code together with aminoacyl-tRNA synthetases, tRNAs act in many other cellular functions. Robust phenomenological observations on the role of tRNAs in translation, together with massive sequence and crystallographic data, have led to a deeper physicochemical understanding of tRNA architecture, dynamics and identity.

Decreased aminoacylation in pathology-related mutants of mitochondrial tRNATyr is associated with structural perturbations in tRNA architecture.

A growing number of human pathologies are ascribed to mutations in mitochondrial tRNA genes. Here, we report biochemical investigations on three mt-tRNA(Tyr) molecules with point substitutions associated with diseases. The mutations occur in the atypical T- and D-loops at positions homologous to those involved in the tertiary interaction network of canonical tRNAs.

Crystal structure of human mitochondrial tyrosyl-tRNA synthetase reveals common and idiosyncratic features.

We report the structure of a strictly mitochondrial human synthetase, namely tyrosyl-tRNA synthetase (mt-TyrRS), in complex with an adenylate analog at 2.2 A resolution. The structure is that of an active enzyme deprived of the C-terminal S4-like domain and resembles eubacterial TyrRSs with a canonical tyrosine-binding pocket and adenylate-binding residues typical of class I synthetases.

Virus-encoded aminoacyl-tRNA synthetases: structural and functional characterization of mimivirus TyrRS and MetRS.

Aminoacyl-tRNA synthetases are pivotal in determining how the genetic code is translated in amino acids and in providing the substrate for protein synthesis. As such, they fulfill a key role in a process universally conserved in all cellular organisms from their most complex to their most reduced parasitic forms. In contrast, even complex viruses were not found to encode much translation machinery, with the exception of isolated components such as tRNAs.

Tyrosyl-tRNA synthetase: the first crystallization of a human mitochondrial aminoacyl-tRNA synthetase.

Human mitochondrial tyrosyl-tRNA synthetase and a truncated version with its C-terminal S4-like domain deleted were purified and crystallized. Only the truncated version, which is active in tyrosine activation and Escherichia coli tRNA(Tyr) charging, yielded crystals suitable for structure determination. These tetragonal crystals, belonging to space group P4(3)2(1)2, were obtained in the presence of PEG 4000 as a crystallizing agent and diffracted X-rays to 2.

Deinococcus glutaminyl-tRNA synthetase is a chimer between proteins from an ancient and the modern pathways of aminoacyl-tRNA formation.

Glutaminyl-tRNA synthetase from Deinococcus radiodurans possesses a C-terminal extension of 215 residues appending the anticodon-binding domain. This domain constitutes a paralog of the Yqey protein present in various organisms and part of it is present in the C-terminal end of the GatB subunit of GatCAB, a partner of the indirect pathway of Gln-tRNA(Gln) formation. To analyze the peculiarities of the structure-function relationship of this GlnRS related to the Yqey domain, a structure of the protein was solved from crystals diffracting at 2.