Publications

2014

• Structural and functional studies of Bud23-Trm112 reveal 18S rRNA N7-G1575 methylation occurs on late 40S precursor ribosomes
  
  • Létoquart Juliette
  • Huvelle E.
  • Wacheul L.
  • Bourgeois Gabrielle
  • Zorbas C.
  • Graille Marc
  • Heurgué-Hamard V.
  • Lafontaine D.
  Proceedings of the National Academy of Sciences of the United States of America, National Academy of Sciences, 2014, 111 (51), pp.E5518-E5526. Ribosomes are essential cellular nanomachines responsible for all protein synthesis in vivo. Efficient and faithful ribosome biogenesis requires a plethora of assembly factors whose precise role and timing of action remains to be established. Here we determined the crystal structure of Bud23–Trm112, which is required for efficient pre-rRNA processing steps leading to 18S rRNA synthesis and methylation of 18S rRNA at position G1575. For the first time, to our knowledge, we identified where on Bud23–Trm112 the contacts with precursor ribosomes occur. We further report that the essential helicase Dhr1 interacts directly with Bud23–Trm112, proposing a concerted action of these proteins in ribosome assembly. Finally, we reveal that the methyltransferase activity of Bud23–Trm112 and its requirement for pre-rRNA processing are disconnected in time. (10.1073/pnas.1413089111)
  DOI : 10.1073/pnas.1413089111
• Structure and Thermodynamics of Mg:Phosphate Interactions in Water: A Simulation Study
  
  • Kumar Manjeet
  • Simonson Thomas
  • Ohanessian Gilles
  • Clavaguéra Carine
  ChemPhysChem, Wiley-VCH Verlag, 2014, pp.201402685. The association of Mg2+ and H2PO4− in water can give insights into Mg:phosphate interactions in general, which are very widespread, but for which experimental data is surprisingly sparse. It is studied through molecular dynamics simulations (>100 ns) by using the polarizable AMOEBA force field, and the association free energy is computed for the first time. Explicit consideration of outer-sphere and two types of inner-sphere association provides considerable insight into the dynamics and thermodynamics of ion pairing. After careful assessment of the computational approximations, the agreement with experimental values indicates that the methodology can be extended to other inorganic and biological Mg:phosphate interactions in solution. (10.1002/cphc.201402685)
  DOI : 10.1002/cphc.201402685
• Solvation and stabilization of palladium nanoparticles in phosphonium-based ionic liquids: a combined infrared spectroscopic and density functional theory study.
  
  • Zvereva Elena E
  • Grimme Stefan
  • Katsyuba Sergey A
  • Ermolaev Vadim V
  • Arkhipova Daria A
  • Yan Ning
  • Miluykov Vasili A
  • Sinyashin Oleg G
  • Aleksandrov Alexey
  Physical Chemistry Chemical Physics, Royal Society of Chemistry, 2014, 16 (38), pp.20672-80. Analysis of infrared spectra of palladium nanoparticles (NPs) immersed in the tri-tert-butyl-R-phosphonium-based ionic liquids (ILs) demonstrates that both cations and anions of the ILs interact with the NPs. According to quantum-chemical simulations of these interactions, the binding energy of anions to the Pd6 cluster, taken as a minimal-size model of the NPs, increases from ∼6 to ∼27 kcal mol(-1) in the order [PF6](-)≈ [BF4](-) < [Tf2N](-) < [OTf](-) < [Br](-)≪ [TFA](-). In contrast, the binding energy for all types of the [Bu(t)3PR](+) cations slightly varies at about ∼22 kcal mol(-1) only moderately depending on the choice of the R moiety (n-pentyl, 2-hydroxyethyl, 2-methoxyethyl, 2-ethoxy-2-oxoethyl). As a result, the energies of interaction between a Pd6 cluster and various ion pairs, formed by the abovementioned counter-ions, follow the order found for the anions and vary from ∼24 to ∼47 kcal mol(-1). These values are smaller than the energy of addition of a Pd atom to a Pdn cluster (∼58 kcal mol(-1)), which suggests kinetic stabilization of the NPs in phosphonium-based ILs rather than thermodynamic stabilization. The results are qualitatively similar to the trends found earlier for interactions between palladium clusters and components of imidazolium-based ILs, in spite of much larger contributions of the London dispersion forces to the binding of the [Bu(t)3PR](+) cations to the cluster (up to 80%) relative to the case of 1-R-3-methylimidazolium cations (up to 40%). (10.1039/c4cp02547d)
  DOI : 10.1039/c4cp02547d
• Unravelling the mechanism of non-ribosomal peptide synthesis by cyclodipeptide synthases.
  
  • Moutiez Mireille
  • Schmitt Emmanuelle
  • Seguin Jérôme
  • Thai Robert
  • Favry Emmanuel
  • Belin Pascal
  • Mechulam Yves
  • Gondry Muriel
  Nature Communications, Nature Publishing Group, 2014, 5 (October), pp.5141. Cyclodipeptide synthases form cyclodipeptides from two aminoacyl transfer RNAs. They use a ping-pong mechanism that begins with transfer of the aminoacyl moiety of the first aminoacyl tRNA onto a conserved serine, yielding an aminoacyl enzyme. Combining X-ray crystallography, site-directed mutagenesis and affinity labelling of the cyclodipeptide synthase AlbC, we demonstrate that the covalent intermediate reacts with the aminoacyl moiety of the second aminoacyl tRNA, forming a dipeptidyl enzyme, and identify the aminoacyl-binding sites of the aminoacyl tRNAs. (10.1038/ncomms6141)
  DOI : 10.1038/ncomms6141
• Structure-function analysis of the Trm9-Trm112 complex, a methyltransferase involved in the mcm5U34 tRNA modification and in response to genotoxic stresses.
  
  • Graille Marc
  , 2014.
• Etudes structurales et fonctionnelles de complexes entre Trm112 et différentes méthyltransférases impliquées dans la traduction
  
  • Létoquart Juliette
  , 2014. La traduction représente un processus central au sein de la cellule, elle assure le transfert de l’information génétique de l’ARNm vers les protéines. De nombreux acteurs y sont impliqués directement ou indirectement et parmi eux, chez les eucaryotes, la petite protéine Trm112. Celle-ci participe à la modification de plusieurs acteurs directs en interagissant et en activant quatre MTases. Le facteur de terminaison eRF1 est méthylé par le complexe Mtq2-Trm112, l’ARNr 18S par Bud23-Trm112 et certains ARNt par les complexes Trm9-Trm112 et Trm11-Trm112. Au cours de ce travail, les structures cristallographiques de Trm9-Trm112 et de Bud23-Trm112 de levure ont été résolues. L’étude comparative structurale de ces complexes et de la structure connue de Mtq2-Trm112, a permis de mettre en évidence que dans un même organisme, les séquences des trois protéines ont évolué de manière à conserver l’interaction avec Trm112. Même si les quatre partenaires présentent moins de 20% d’identité de séquence, les résidus clés pour l’interaction avec la petite protéine activatrice sont conservés ou partagent des caractéristiques identiques. En plus de l’analyse structurale, le complexe Trm9-Trm112 a fait l’objet d’une étude fonctionnelle chez S. cerevisiae ce qui a permis de cartographier le site actif de l’enzyme et de proposer un modèle de mécanisme d’action. Enfin, les premières études in vivo réalisées chez Haloferax volcanii suggèrent que cette plateforme serait également présente chez certains organismes procaryotes.
• Pairwise Decomposition of an MMGBSA Energy Function for Computational Protein Design
  
  • Gaillard Thomas
  • Simonson Thomas
  Journal of Computational Chemistry, Wiley, 2014, 35 (18), pp.1371-1387. Computational protein design (CPD) aims at predicting new proteins or modifying existing ones. The computational challenge is huge as it requires exploring an enormous sequence and conformation space. The difficulty can be reduced by considering a fixed backbone and a discrete set of sidechain conformations. Another common strategy consists in precalculating a pairwise energy matrix, from which the energy of any sequence/conformation can be quickly obtained. In this work, we examine the pairwise decomposition of protein MMGBSA energy functions from a general theoretical perspective, and an implementation proposed earlier for CPD. It includes a Generalized Born term, whose many-body character is overcome using an effective dielectric environment, and a Surface Area term, for which we present an improved pairwise decomposition. A detailed evaluation of the error introduced by the decomposition on the different energy components is performed. We show that the error remains reasonable, compared to other uncertainties. © 2014 Wiley Periodicals, Inc. (10.1002/jcc.23637)
  DOI : 10.1002/jcc.23637
• The C-Terminal Domain from S. cerevisiae Pat1 Displays Two Conserved Regions Involved in Decapping Factor Recruitment
  
  • Fourati Zaineb
  • Kolesnikova Olga A
  • Back Régis
  • Keller Jenny
  • Charenton Clément
  • Taverniti Valério
  • Gaudon Plesse Claudine
  • Lazar Noureddine
  • Durand Dominique
  • van Tilbeurgh Herman
  • Séraphin Bertrand
  • Graille Marc
  PLoS ONE, Public Library of Science, 2014, 9 (5), pp.e96828. Eukaryotic mRNA decay is a highly regulated process allowing cells to rapidly modulate protein production in response to internal and environmental cues. Mature translatable eukaryotic mRNAs are protected from fast and uncontrolled degradation in the cytoplasm by two cis-acting stability determinants: a methylguanosine (m7G) cap and a poly(A) tail at their 5′ and 3′ extremities, respectively. The hydrolysis of the m7G cap structure, known as decapping, is performed by the complex composed of the Dcp2 catalytic subunit and its partner Dcp1. The Dcp1-Dcp2 decapping complex has a low intrinsic activity and requires accessory factors to be fully active. Among these factors, Pat1 is considered to be a central scaffolding protein involved in Dcp2 activation but also in inhibition of translation initiation. Here, we present the structural and functional study of the C-terminal domain from S. cerevisiae Pat1 protein. We have identified two conserved and functionally important regions located at both extremities of the domain. The first region is involved in binding to Lsm1-7 complex. The second patch is specific for fungal proteins and is responsible for Pat1 interaction with Edc3. These observations support the plasticity of the protein interaction network involved in mRNA decay and show that evolution has extended the C-terminal alpha-helical domain from fungal Pat1 proteins to generate a new binding platform for protein partners. (10.1371/journal.pone.0096828)
  DOI : 10.1371/journal.pone.0096828
• An Overview of Electrostatic Free Energy Computations for Solutions and Proteins
  
  • Lin Yen-Lin
  • Aleksandrov Alexey
  • Simonson Thomas
  • Roux Benoit
  Journal of Chemical Theory and Computation, American Chemical Society, 2014, 10 (7), pp.2690-2709. Free energy simulations for electrostatic and charging processes in complex molecular systems encounter specific difficulties owing to the long-range, 1/r Coulomb interaction. To calculate the solvation free energy of a simple ion, it is essential to take into account the polarization of nearby solvent but also the electrostatic potential drop across the liquid-gas boundary, however distant. The latter does not exist in a simulation model based on periodic boundary conditions because there is no physical boundary to the system. An important consequence is that the reference value of the electrostatic potential is not an ion in a vacuum. Also, in an infinite system, the electrostatic potential felt by a perturbing charge is conditionally convergent and dependent on the choice of computational conventions. Furthermore, with Ewald lattice summation and tinfoil conducting boundary conditions, the charges experience a spurious shift in the potential that depends on the details of the simulation system such as the volume fraction occupied by the solvent. All these issues can be handled with established computational protocols, as reviewed here and illustrated for several small ions and three solvated proteins. (10.1021/ct500195p)
  DOI : 10.1021/ct500195p
• A Highly Conserved Region Essential for NMD in the Upf2 N-Terminal Domain
  
  • Fourati Zaineb
  • Roy Bijoyita
  • Millan Claudia
  • Coureux Pierre-Damien
  • Kervestin Stéphanie
  • van Tilbeurgh Herman
  • He Feng
  • Usón Isabel
  • Jacobson Allan
  • Graille Marc
  Journal of Molecular Biology, Elsevier, 2014, 426 (22), pp.3689 - 3702. Upf1, Upf2, and Upf3 are the principal regulators of nonsense-mediated mRNA decay (NMD), a cytoplasmic surveillance pathway that accelerates the degradation of mRNAs undergoing premature translation termination. These three proteins interact with each other, the ribosome, the translation termination machinery, and multiple mRNA decay factors, but the precise mechanism allowing the selective detection and degradation of nonsense-containing transcripts remains elusive. Here, we have determined the crystal structure of the N-terminal mIF4G domain from Saccharomyces cerevisiae Upf2 and identified a highly conserved region in this domain that is essential for NMD and independent of Upf2's binding sites for Upf1 and Upf3. Mutations within this conserved region not only inactivate NMD but also disrupt Upf2 binding to specific proteins, including Dbp6, a DEAD-box helicase. Although current models indicate that Upf2 functions principally as an activator of Upf1 and a bridge between Upf1 and Upf3, our data suggest that it may also serve as a platform for the association of additional factors that play roles in premature translation termination and NMD. (10.1016/j.jmb.2014.09.015)
  DOI : 10.1016/j.jmb.2014.09.015
• Iron is essential for living!
  
  • Lasocki Sigismond
  • Gaillard Thomas
  • Rineau Emmanuel
  Critical Care, BioMed Central, 2014, 18 (678), pp.Non spécifié. <p>Iron as an element is a double-edged sword, essential for living but also potentially toxic through the generation of oxidative stress. The recent study by Chen and colleagues in Critical Care reminds us of this elegantly. In a mouse model of acute lung injury, they showed that silencing hepcidin (the master regulator of iron metabolism) locally in airway epithelial cells aggravates lung injury by increasing the release of iron from alveolar macrophages, which in turn enhances pulmonary bacterial growth and reduces the macrophages’ killing properties. This work underscores that hepcidin acts not only systematically (as a hormone) but also locally for iron metabolism regulation. This opens areas of research for sepsis treatment but also for iron deficiency or anaemia treatment, since the local and systemic iron regulation appear to be independent.</p> (10.1186/s13054-014-0678-7)
  DOI : 10.1186/s13054-014-0678-7
• Cytoplasmic mRNA surveillance pathways
  
  • Fourati Zaineb
  • Graille Marc
  , 2014, pp.195-216. During mRNA synthesis and maturation, the introduction of errors can strongly influence the expression of certain genes and/or the activity of the proteins for which they encode. To minimise these defects, eukaryotic cells have evolved several cytoplasmic and translation-dependent quality control pathways aimed at detecting and degrading mRNAs that would lead to the production of aberrant proteins. The nonsense-mediated mRNA decay pathway (NMD) clears cells from mRNAs harbouring premature in-frame stop codons. Two other pathways (NSD for nonstop decay and NGD for No-Go decay) degrade mRNAs on which ribosomes have stalled during elongation. In this chapter, we describe the current knowledge on the biological roles and molecular mechanisms of these surveillance pathways, which were mainly unravelled using baker’s yeast as model system. (10.1007/978-3-319-05687-6_8)
  DOI : 10.1007/978-3-319-05687-6_8
• The Mechanism of Citryl-Coenzyme A Formation Catalyzed by Citrate Synthase
  
  • Aleksandrov Alexey
  • Zvereva Elena
  • Field Martin J
  Journal of Physical Chemistry B, American Chemical Society, 2014, 118 (17), pp.4505-4513. The enzyme citrate synthase is used by all living cells to catalyze the first step of the citric acid cycle. In this work, we have investigated the enolization and condensation steps catalyzed by citrate synthase, using ab initio (B3LYP/def2-TZVP and MP2/aug-cc-pVDZ) quantum chemical/molecular mechanical hybrid potentials in conjunction with reaction-path-location algorithms and molecular dynamics free energy simulations. The results of the latter indicate that the catalytic His238 residue is in its neutral form, and also argue strongly for the presence of a water molecule in the enzyme's catalytic center. Such a water is observed in some, but not all, of the experimentally resolved structures of the protein. The mechanism itself starts with an enolization that proceeds via an enolate intermediate rather than the enol form, which is much more unstable. This is in agreement with the results of other workers. For the condensation step, we investigated two mechanisms in which there is a direct nucleophilic attack of the enolate intermediate on the oxaloacetate carbonyl carbon, and found the one in which there is no proton transfer from the neighboring arginine to be preferred. Although this residue, Arg329, is not implicated directly in the reaction, it helps to stabilize the negative citryl-CoA formed during the condensation step. (10.1021/jp412346g)
  DOI : 10.1021/jp412346g