Publications

2007

• Asmparts: assembly of biological model parts.
  
  • Rodrigo G.
  • Carrera J.
  • Jaramillo A.
  Systems and Synthetic Biology, Springer Verlag (Germany), 2007, 1 (4), pp.167-70. We propose a new computational tool to produce models of biological systems by assembling models from biological parts. Our software not only takes advantage of modularity, but it also enforces standardisation in part characterisation by considering a model of each part. We have used model parts in SBML to design transcriptional networks. Our software is open source, it works in linux and windows platforms, and it could be used to automatically produce models in a server. Our tool not only facilitates model design, but it will also help to promote the establishment of a registry of model parts. (10.1007/s11693-008-9013-4)
  DOI : 10.1007/s11693-008-9013-4
• Computational design of digital and memory biological devices.
  
  • Rodrigo G.
  • Jaramillo A.
  Systems and Synthetic Biology, Springer Verlag (Germany), 2007, 1 (4), pp.183-95. The use of combinatorial optimization techniques with computational design allows the development of automated methods to design biological systems. Automatic design integrates design principles in an unsupervised algorithm to sample a larger region of the biological network space, at the topology and parameter levels. The design of novel synthetic transcriptional networks with targeted behaviors will be key to understand the design principles underlying biological networks. In this work, we evolve transcriptional networks towards a targeted dynamics, by using a library of promoters and coding sequences, to design a complex biological memory device. The designed sequential transcription network implements a JK-Latch, which is fully predictable and richer than other memory devices. Furthermore, we present designs of transcriptional devices behaving as logic gates, and we show how to create digital behavior from analog promoters. Our procedure allows us to propose a scenario for the evolution of multi-functional genetic networks. In addition, we discuss the decomposability of regulatory networks in terms of genetic modules to develop a given cellular function. Summary. We show how to use an automated procedure to design logic and sequential transcription circuits. This methodology will allow advancing the rational design of biological devices to more complex systems, and we propose the first design of a biological JK-latch memory device. (10.1007/s11693-008-9017-0)
  DOI : 10.1007/s11693-008-9017-0
• Structure of an archaeal heterotrimeric initiation factor 2 reveals a nucleotide state between the GTP and the GDP states.
  
  • Yatime Laure
  • Mechulam Yves
  • Blanquet Sylvain
  • Schmitt Emmanuelle
  Proceedings of the National Academy of Sciences of the United States of America, National Academy of Sciences, 2007, 104 (47), pp.18445-50. Initiation of translation in eukaryotes and in archaea involves eukaryotic/archaeal initiation factor (e/aIF)1 and the heterotrimeric initiation factor e/aIF2. In its GTP-bound form, e/aIF2 provides the initiation complex with Met-tRNA(i)(Met). After recognition of the start codon by initiator tRNA, e/aIF1 leaves the complex. Finally, e/aIF2, now in a GDP-bound form, loses affinity for Met-tRNA(i)(Met) and dissociates from the ribosome. Here, we report a 3D structure of an aIF2 heterotrimer from the archeon Sulfolobus solfataricus obtained in the presence of GDP. Our report highlights how the two-switch regions involved in formation of the tRNA-binding site on subunit gamma exchange conformational information with alpha and beta. The zinc-binding domain of beta lies close to the guanine nucleotide and directly contacts the switch 1 region. As a result, switch 1 adopts a not yet described conformation. Moreover, unexpectedly for a GDP-bound state, switch 2 has the "ON" conformation. The stability of these conformations is accounted for by a ligand, most probably a phosphate ion, bound near the nucleotide binding site. The structure suggests that this GDP-inorganic phosphate (Pi) bound state of aIF2 may be proficient for tRNA binding. Recently, it has been proposed that dissociation of eIF2 from the initiation complex is closely coupled to that of Pi from eIF2gamma upon start codon recognition. The nucleotide state of aIF2 shown here is indicative of a similar mechanism in archaea. Finally, we consider the possibility that release of Pi takes place after e/aIF2gamma has been informed of e/aIF1 dissociation by e/aIF2beta. (10.1073/pnas.0706784104)
  DOI : 10.1073/pnas.0706784104
• Wear mechanism of WC-Co cutting tool from turning experiments and high speed tribological tests
  
  • Kagnaya Tchadja
  • Boher Christine
  • Lambert Laurence
  • Lazard Myriam
  • Cutard Thierry
  , 2007, pp.8 p..
• Ammonium scanning in an enzyme active site. The chiral specificity of aspartyl-tRNA synthetase.
  
  • Thompson Damien
  • Lazennec Christine
  • Plateau Pierre
  • Simonson Thomas
  Journal of Biological Chemistry, American Society for Biochemistry and Molecular Biology, 2007, 282 (42), pp.30856-68. D-amino acids are largely excluded from protein synthesis, yet they are of great interest in biotechnology. Aspartyl-tRNA synthetase (AspRS) can misacylate tRNA(Asp) with D-aspartate instead of its usual substrate, L-Asp. We investigate how the preference for L-Asp arises, using molecular dynamics simulations. Asp presents a special problem, having pseudosymmetry broken only by its ammonium group, and AspRS must protect not only against D-Asp, but against an "inverted" orientation where the two substrate carboxylates are swapped. We compare L-Asp and D-Asp, in both orientations, and succinate, where the ammonium group is removed and the ligand has an additional negative charge. All possible ammonium positions on the ligand are thus scanned, providing information on electrostatic interactions. As controls, we simulate a Q199E mutation, obtaining a reduction in binding free energy in agreement with experiment, and we simulate TyrRS, which can misacylate tRNA(Tyr) with D-Tyr. For both TyrRS and AspRS, we obtain a moderate binding free energy difference DeltaDeltaG between the L- and D-amino acids, in agreement with their known ability to misacylate their tRNAs. In contrast, we predict that AspRS is strongly protected against inverted L-Asp binding. For succinate, kinetic measurements reveal a DeltaDeltaG of over 5 kcal/mol, favoring L-Asp. The simulations show how chiral discriminations arises from the structures, with two AspRS conformations acting in different ways and proton uptake by nearby histidines playing a role. A complex network of charges protects AspRS against most binding errors, making the engineering of its specificity a difficult challenge. (10.1074/jbc.M704788200)
  DOI : 10.1074/jbc.M704788200
• GEK1, une protéine d'Arabidopsis thaliana impliquée dans la tolérance à l'éthanol, est une D-aminoacyl-ARNt désacylase
  
  • Wydau Sandra
  • Ferri-Fioni Maria-Laura
  • Blanquet Sylvain
  • Plateau Pierre
  , 2007, pp.p. 121.
• Nanobiosensors based on silicon nanowires
  
  • Eude L.
  • Pribat D.
  • Charlot S.
  • Gué Anne Marie
  • Mugherli Laurent
  • Fromant Michel
  • Plateau Pierre
  • Blanquet Sylvain
  • Lehoucq G.
  • Bondavalli P.
  • Legagneux P.
  , 2007.
• Genetdes: automatic design of transcriptional networks.
  
  • Rodrigo G.
  • Carrera J.
  • Jaramillo A.
  Bioinformatics, Oxford University Press (OUP), 2007, 23 (14), pp.1857-8. MOTIVATION: The rational design of biological networks with prescribed functions is limited to gene circuits of a few genes. Larger networks involve complex interactions with many parameters and the use of automated computational tools can be very valuable. We propose a new tool to design transcriptional networks with targeted behavior that could be used to better understand the design principles of genetic circuits. RESULTS: We have implemented a Simulated Annealing optimization algorithm that explores throughout the space of transcription networks to obtain a specific behavior. The software outputs a transcriptional network with all the corresponding kinetic parameters in SBML format. We provide examples of transcriptional circuits with logical and oscillatory behaviors. Our tool can also be applied to design networks with multiple external input and output genes. AVAILABILITY: The software, a tutorial manual, parameter sets and examples are freely available at http://synth-bio.yi.org/genetdes.html. (10.1093/bioinformatics/btm237)
  DOI : 10.1093/bioinformatics/btm237
• Protonation patterns in tetracycline:tet repressor recognition: simulations and experiments.
  
  • Aleksandrov Alexey
  • Proft Juliane
  • Hinrichs Winfried
  • Simonson Thomas
  ChemBioChem, Wiley-VCH Verlag, 2007, 8 (6), pp.675-85. Resistance to the antibiotic tetracycline (Tc) is regulated by its binding as a Tc:Mg2+ complex to the Tet Repressor protein (TetR). Tc:TetR recognition is a complex problem, with the protein and ligand each having several possible conformations and protonation states, which are difficult to elucidate by experiment alone. We used a combination of free-energy simulations and crystallographic analysis to investigate the electrostatic interactions between protein and ligand and the possible role of induced fit in Tc binding. Tc in solution was described quantum mechanically, while Tc:TetR interactions were described by a recent, high-quality molecular-mechanics model. The orientations of the amide and imidazole groups were determined experimentally by a careful analysis of Debye-Waller factors in alternate crystallographic models. The agreement with experiment for these orientations suggested that the simulations and their more detailed, thermodynamic predictions were reliable. We found that the ligand prefers an extended, zwitterionic state both in solution and in complexation with the protein. Tc is thus preorganized for binding, while the protein combines lock-and-key behavior for regions close to the ligand's amide, enolate, and ammonium groups, with an induced fit for regions close to the Mg2+ ion. These insights and the modeling techniques employed should be of interest for engineering improved TetR ligands and improved TetR proteins for gene regulation, as well as for drug design. (10.1002/cbic.200600535)
  DOI : 10.1002/cbic.200600535
• Extracellular production of hydrogen selenide accounts for thiol-assisted toxicity of selenite against Saccharomyces cerevisiae.
  
  • Tarze Agathe
  • Dauplais Marc
  • Grigoras Ioana
  • Lazard Myriam
  • Ha-Duong Nguyet-Thanh
  • Barbier Frédérique
  • Blanquet Sylvain
  • Plateau Pierre
  Journal of Biological Chemistry, American Society for Biochemistry and Molecular Biology, 2007, 282 (12), pp.8759-67. Administration of selenium in humans has anticarcinogenic effects. However, the boundary between cancer-protecting and toxic levels of selenium is extremely narrow. The mechanisms of selenium toxicity need to be fully understood. In Saccharomyces cerevisiae, selenite in the millimolar range is well tolerated by cells. Here we show that the lethal dose of selenite is reduced to the micromolar range by the presence of thiols in the growth medium. Glutathione and selenite spontaneously react to produce several selenium-containing compounds (selenodiglutathione, glutathioselenol, hydrogen selenide, and elemental selenium) as well as reactive oxygen species. We studied which compounds in the reaction pathway between glutathione and sodium selenite are responsible for this toxicity. Involvement of selenodiglutathione, elemental selenium, or reactive oxygen species could be ruled out. In contrast, extracellular formation of hydrogen selenide can fully explain the exacerbation of selenite toxicity by thiols. Indeed, direct production of hydrogen selenide with D-cysteine desulfhydrase induces high mortality. Selenium uptake by S. cerevisiae is considerably enhanced in the presence of external thiols, most likely through internalization of hydrogen selenide. Finally, we discuss the possibility that selenium exerts its toxicity through consumption of intracellular reduced glutathione, thus leading to severe oxidative stress. (10.1074/jbc.M610078200)
  DOI : 10.1074/jbc.M610078200
• Recognizing protein-protein interfaces with empirical potentials and reduced amino acid alphabets.
  
  • Launay Guillaume
  • Mendez Raul
  • Wodak Shoshana J
  • Simonson Thomas
  BMC Bioinformatics, BioMed Central, 2007, 8, pp.270. BACKGROUND: In structural genomics, an important goal is the detection and classification of protein-protein interactions, given the structures of the interacting partners. We have developed empirical energy functions to identify native structures of protein-protein complexes among sets of decoy structures. To understand the role of amino acid diversity, we parameterized a series of functions, using a hierarchy of amino acid alphabets of increasing complexity, with 2, 3, 4, 6, and 20 amino acid groups. Compared to previous work, we used the simplest possible functional form, with residue-residue interactions and a stepwise distance-dependence. We used increased computational resources, however, constructing 290,000 decoys for 219 protein-protein complexes, with a realistic docking protocol where the protein partners are flexible and interact through a molecular mechanics energy function. The energy parameters were optimized to correctly assign as many native complexes as possible. To resolve the multiple minimum problem in parameter space, over 64000 starting parameter guesses were tried for each energy function. The optimized functions were tested by cross validation on subsets of our native and decoy structures, by blind tests on series of native and decoy structures available on the Web, and on models for 13 complexes submitted to the CAPRI structure prediction experiment. RESULTS: Performance is similar to several other statistical potentials of the same complexity. For example, the CAPRI target structure is correctly ranked ahead of 90% of its decoys in 6 cases out of 13. The hierarchy of amino acid alphabets leads to a coherent hierarchy of energy functions, with qualitatively similar parameters for similar amino acid types at all levels. Most remarkably, the performance with six amino acid classes is equivalent to that of the most detailed, 20-class energy function. CONCLUSION: This suggests that six carefully chosen amino acid classes are sufficient to encode specificity in protein-protein interactions, and provide a starting point to develop more complicated energy functions. (10.1186/1471-2105-8-270)
  DOI : 10.1186/1471-2105-8-270
• Computational sidechain placement and protein mutagenesis with implicit solvent models
  
  • Lopes Anne
  • Alexandrov Alexey
  • Bathelt Christine
  • Archontis Georgios
  • Simonson Thomas
  Proteins - Structure, Function and Bioinformatics, Wiley, 2007, 67 (4), pp.853-867. Structure prediction and computational protein design should benefit from accurate solvent models. We have applied implicit solvent models to two problems that are central to this area. First, we performed sidechain placement for 29 proteins, using a solvent model that combines a screened Coulomb term with an Accessible Surface Area term (CASA model). With optimized parameters, the prediction quality is comparable with earlier work that omitted electrostatics and solvation altogether. Second, we computed the stability changes associated with point mutations involving ionized sidechains. For over 1000 mutations, including many fully or partly buried positions, we compared CASA and two generalized Born models (GB) with a more accurate model, which solves the Poisson equation of continuum electrostatics numerically. CASA predicts the correct sign and order of magnitude of the stability change for 81% of the mutations, compared to 97% with the best GB. We also considered 140 mutations for which experimental data are available. Comparing to experiment requires additional assumptions about the unfolded protein structure, protein relaxation in response to the mutations, and contributions from the hydrophobic effect. With a simple, commonly‐used unfolded state model, the mean unsigned error is 2.1 kcal/mol with both CASA and the best GB. Overall, the electrostatic model is not important for sidechain placement; CASA and GB are equivalent for surface mutations, while GB is far superior for fully or partly buried positions. Thus, for problems like protein design that involve all these aspects, the most recent GB models represent an important step forward. Along with the recent discovery of efficient, pairwise implementations of GB, this will open new possibilities for the computational engineering of proteins. Proteins 2007. © 2007 Wiley‐Liss, Inc. (10.1002/prot.21379)
  DOI : 10.1002/prot.21379
• Vanillin cell sensor
  
  • Rodrigo G.
  • Montagud A.
  • Aparici A.
  • Aroca M.C.
  • Baguena M.
  • Carrera J.
  • Edo C.
  • Fernandez-De-Cordoba P.
  • Ferrando A.
  • Fuertes G.
  • Gimenez D.
  • Mata C.
  • Medrano J.V.
  • Navarrete C.
  • Navarro E.
  • Salgado J.
  • Tortosa Pablo
  • Urchueguia J.
  • Jaramillo A.
  IET Synthetic Biology, 2007, 1 (40940), pp.74. Our project for iGEM 2006 consisted of designing a cellular vanillin biosensor. We used an EnvZ-E. coli strain as a chassis, and constructed two different devices: a sensor and an actuator, assembled using OmpR-P as a standardised mediator. The sensor device contained a computationally designed vanillin receptor and a synthetic two-component signal transduction protein (Trz). The receptor protein was based on a ribose-binding protein as scaffold. The Trz was built by fusion of the periplasmic and transmembrane domains of a Trg protein with an EnvZ kinase domain. When the receptor complex binds Trg, an allosteric motion is propagated to the cytoplasmic EnvZ kinase domain, resulting in autophosphorylation and subsequent phosphate transfer to the OmpR transcription factor, which finally induces transcription of the ompC promoter. As actuator, we used a synthetic transcriptional circuit, which implements an OmpR-P band detector having GFP and RFP as an output. We designed this circuit using a synthetic promoter working as an AND gate, which is synergistically activated by cI and CRP. Our constructed Trg-EnvZ fusion and AND promoter will be very useful to future synthetic biology projects. © 2007 The Institution of Engineering and Technology. (10.1049/iet-stb:20060003)
  DOI : 10.1049/iet-stb:20060003
• Evolutionary mechanisms of circadian clocks
  
  • Rodrigo G.
  • Carrera J.
  • Jaramillo A.
  Central European Journal of Biology, Springer Verlag, 2007, 2 (2), pp.233. An intriguing question in biology is to know how circadian molecular networks could have evolved their particular topologies to adjust to a daily period. We analyze the mechanism of the evolution of such networks by using a computational design strategy that allows for the generation of synthetic networks with a targeted 24 hours oscillation. We have performed a systematic analysis of all possible two-gene network topologies based on a core activator-repressor frequently found in circadian mechanisms. We have considered transcriptional and post-translational regulations to implement this core. We have applied our analysis to both, eukaryotic and prokaryotic circadian machinery. Finally, we conjecture a possible mechanism for the evolution of circadian clocks. © Versita Warsaw and Springer-Verlag Berlin Heidelberg 2007. (10.2478/s11535-007-0016-z)
  DOI : 10.2478/s11535-007-0016-z
• Applications of Free Energy Calculations to Chemistry and Biology
  
  • Chipot Christophe
  • Mark Alan E.
  • Pande Vijay S.
  • Simonson Thomas
  , 2007, 86, pp.463-501. (10.1007/978-3-540-38448-9_13)
  DOI : 10.1007/978-3-540-38448-9_13
• Neutral evolution of protein-protein interactions: a computational study using simple models.
  
  • Noirel Josselin
  • Simonson Thomas
  BMC Structural Biology, BioMed Central, 2007, 7, pp.79-93. BACKGROUND: Protein-protein interactions are central to cellular organization, and must have appeared at an early stage of evolution. To understand better their role, we consider a simple model of protein evolution and determine the effect of an explicit selection for Protein-protein interactions. RESULTS: In the model, viable sequences all have the same fitness, following the neutral evolution theory. A very simple, two-dimensional lattice representation of the protein structures is used, and the model only considers two kinds of amino acids: hydrophobic and polar. With these approximations, exact calculations are performed. The results do not depend too strongly on these assumptions, since a model using a 3D, off-lattice representation of the proteins gives results in qualitative agreement with the 2D one. With both models, the evolutionary dynamics lead to a steady state population that is enriched in sequences that dimerize with a high affinity, well beyond the minimal level needed to survive. Correspondingly, sequences close to the viability threshold are less abundant in the steady state, being subject to a larger proportion of lethal mutations. The set of viable sequences has a "funnel" shape, consistent with earlier studies: sequences that are highly populated in the steady state are "close" to each other (with proximity being measured by the number of amino acids that differ). CONCLUSION: This bias in the the steady state sequences should lead to an increased resistance of the population to environmental change and an increased ability to evolve. (10.1186/1472-6807-7-79)
  DOI : 10.1186/1472-6807-7-79
• Protection-based assays to measure aminoacyl-tRNA binding to translation initiation factors.
  
  • Mechulam Yves
  • Guillon Laurent
  • Yatime Laure
  • Blanquet Sylvain
  • Schmitt Emmanuelle
  Methods in Enzymology, Elsevier, 2007, 430, pp.265-81. To decipher the mechanisms of translation initiation, the stability of the complexes between tRNA and initiation factors has to be evaluated in a routine manner. A convenient method to measure the parameters of binding of an aminoacyl-tRNA to an initiation factor results from the property that, when specifically complexed to a protein, the aminoacyl-tRNA often resists spontaneous deacylation. This chapter describes the preparation of suitable aminoacyl-tRNA ligands and their use in evaluating the stability of their complexes with various initiation factors, such as e/aIF2 and e/aIF5B. The advantages and the limitations of the method are discussed. (10.1016/S0076-6879(07)30011-6)
  DOI : 10.1016/S0076-6879(07)30011-6
• GEK1, a gene product of Arabidopsis thaliana involved in ethanol tolerance, is a D-aminoacyl-tRNA deacylase.
  
  • Wydau Sandra
  • Ferri-Fioni Maria-Laura
  • Blanquet Sylvain
  • Plateau Pierre
  Nucleic Acids Research, Oxford University Press, 2007, 35 (3), pp.930-8. GEK1, an Arabidopsis thaliana gene product, was recently identified through its involvement in ethanol tolerance. Later, this protein was shown to display 26% strict identity with archaeal d-Tyr-tRNA(Tyr) deacylases. To determine whether it actually possessed deacylase activity, the product of the GEK1 open reading frame was expressed in Escherichia coli from a multi-copy plasmid. Purified GEK1 protein contains two zinc ions and proves to be a broad-specific, markedly active d-aminoacyl-tRNA deacylase in vitro. Moreover, GEK1 expression is capable of functionally compensating in E. coli for the absence of endogeneous d-Tyr- tRNA(Tyr) deacylase. Possible connections between exposure of plants to ethanol/acetaldehyde and misaminoacylation of tRNA by d-amino acids are considered. (10.1093/nar/gkl1145)
  DOI : 10.1093/nar/gkl1145
• Free Energy Calculations: Approximate Methods for Biological Macromolecules
  
  • Simonson Thomas
  , 2007, 86, pp.423–461. (10.1007/978-3-540-38448-9_12)
  DOI : 10.1007/978-3-540-38448-9_12