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Sample records for helicase mechanism based

  1. A mechanical mechanism for translocation of ring-shaped helicases on DNA and its demonstration in a macroscopic simulation system

    Chou, Y. C.

    2018-04-01

    The asymmetry in the two-layered ring structure of helicases and the random thermal fluctuations of the helicase and DNA molecules are considered as the bases for the generation of the force required for translocation of the ring-shaped helicase on DNA. The helicase comprises a channel at its center with two unequal ends, through which strands of DNA can pass. The random collisions between the portion of the DNA strand in the central channel and the wall of the channel generate an impulsive force toward the small end. This impulsive force is the starting point for the helicase to translocate along the DNA with the small end in front. Such a physical mechanism may serve as a complementary for the chemomechanical mechanism of the translocation of helicase on DNA. When the helicase arrives at the junction of ssDNA and dsDNA (a fork), the collision between the helicase and the closest base pair may produce a sufficient impulsive force to break the weak hydrogen bond of the base pair. Thus, the helicase may advance and repeat the process of unwinding the dsDNA strand. This mechanism was tested in a macroscopic simulation system where the helicase was simulated using a truncated-cone structure and DNA was simulated with bead chains. Many features of translocation and unwinding such as translocation on ssDNA and dsDNA, unwinding of dsDNA, rewinding, strand switching, and Holliday junction resolution were reproduced.

  2. Interplay of cis- and trans-regulatory mechanisms in the spliceosomal RNA helicase Brr2.

    Absmeier, Eva; Becke, Christian; Wollenhaupt, Jan; Santos, Karine F; Wahl, Markus C

    2017-01-02

    RNA helicase Brr2 is implicated in multiple phases of pre-mRNA splicing and thus requires tight regulation. Brr2 can be auto-inhibited via a large N-terminal region folding back onto its helicase core and auto-activated by a catalytically inactive C-terminal helicase cassette. Furthermore, it can be regulated in trans by the Jab1 domain of the Prp8 protein, which can inhibit Brr2 by intermittently inserting a C-terminal tail in the enzyme's RNA-binding tunnel or activate the helicase after removal of this tail. Presently it is unclear, whether these regulatory mechanisms functionally interact and to which extent they are evolutionarily conserved. Here, we report crystal structures of Saccharomyces cerevisiae and Chaetomium thermophilum Brr2-Jab1 complexes, demonstrating that Jab1-based inhibition of Brr2 presumably takes effect in all eukaryotes but is implemented via organism-specific molecular contacts. Moreover, the structures show that Brr2 auto-inhibition can act in concert with Jab1-mediated inhibition, and suggest that the N-terminal region influences how the Jab1 C-terminal tail interacts at the RNA-binding tunnel. Systematic RNA binding and unwinding studies revealed that the N-terminal region and the Jab1 C-terminal tail specifically interfere with accommodation of double-stranded and single-stranded regions of an RNA substrate, respectively, mutually reinforcing each other. Additionally, such analyses show that regulation based on the N-terminal region requires the presence of the inactive C-terminal helicase cassette. Together, our results outline an intricate system of regulatory mechanisms, which control Brr2 activities during snRNP assembly and splicing.

  3. A Brownian motor mechanism of translocation and strand separation by hepatitis C virus helicase.

    Levin, Mikhail K; Gurjar, Madhura; Patel, Smita S

    2005-05-01

    Helicases translocate along their nucleic acid substrates using the energy of ATP hydrolysis and by changing conformations of their nucleic acid-binding sites. Our goal is to characterize the conformational changes of hepatitis C virus (HCV) helicase at different stages of ATPase cycle and to determine how they lead to translocation. We have reported that ATP binding reduces HCV helicase affinity for nucleic acid. Now we identify the stage of the ATPase cycle responsible for translocation and unwinding. We show that a rapid directional movement occurs upon helicase binding to DNA in the absence of ATP, resulting in opening of several base pairs. We propose that HCV helicase translocates as a Brownian motor with a simple two-stroke cycle. The directional movement step is fueled by single-stranded DNA binding energy while ATP binding allows for a brief period of random movement that prepares the helicase for the next cycle.

  4. DNA-conjugated gold nanoparticles based colorimetric assay to assess helicase activity: a novel route to screen potential helicase inhibitors

    Deka, Jashmini; Mojumdar, Aditya; Parisse, Pietro; Onesti, Silvia; Casalis, Loredana

    2017-03-01

    Helicase are essential enzymes which are widespread in all life-forms. Due to their central role in nucleic acid metabolism, they are emerging as important targets for anti-viral, antibacterial and anti-cancer drugs. The development of easy, cheap, fast and robust biochemical assays to measure helicase activity, overcoming the limitations of the current methods, is a pre-requisite for the discovery of helicase inhibitors through high-throughput screenings. We have developed a method which exploits the optical properties of DNA-conjugated gold nanoparticles (AuNP) and meets the required criteria. The method was tested with the catalytic domain of the human RecQ4 helicase and compared with a conventional FRET-based assay. The AuNP-based assay produced similar results but is simpler, more robust and cheaper than FRET. Therefore, our nanotechnology-based platform shows the potential to provide a useful alternative to the existing conventional methods for following helicase activity and to screen small-molecule libraries as potential helicase inhibitors.

  5. Mechanism of Archaeal MCM Helicase Recruitment to DNA Replication Origins

    Samson, Rachel Y.; Abeyrathne, Priyanka D.; Bell, Stephen D.

    2015-01-01

    Summary Cellular DNA replication origins direct the recruitment of replicative helicases via the action of initiator proteins belonging to the AAA+ superfamily of ATPases. Archaea have a simplified subset of the eukaryotic DNA replication machinery proteins and possess initiators that appear ancestral to both eukaryotic Orc1 and Cdc6. We have reconstituted origin-dependent recruitment of the homohexameric archaeal MCM in vitro with purified recombinant proteins. Using this system, we reveal that archaeal Orc1-1 fulfills both Orc1 and Cdc6 functions by binding to a replication origin and directly recruiting MCM helicase. We identify the interaction interface between these proteins and reveal how ATP binding by Orc1-1 modulates recruitment of MCM. Additionally, we provide evidence that an open-ring form of the archaeal MCM homohexamer is loaded at origins. PMID:26725007

  6. Targeting Dengue Virus NS-3 Helicase by Ligand based Pharmacophore Modeling and Structure based Virtual Screening

    Halim, Sobia A.; Khan, Shanza; Khan, Ajmal; Wadood, Abdul; Mabood, Fazal; Hussain, Javid; Al-Harrasi, Ahmed

    2017-10-01

    Dengue fever is an emerging public health concern, with several million viral infections occur annually, for which no effective therapy currently exist. Non-structural protein 3 (NS-3) Helicase encoded by the dengue virus (DENV) is considered as a potential drug target to design new and effective drugs against dengue. Helicase is involved in unwinding of dengue RNA. This study was conducted to design new NS-3 Helicase inhibitor by in silico ligand- and structure based approaches. Initially ligand-based pharmacophore model was generated that was used to screen a set of 1201474 compounds collected from ZINC Database. The compounds matched with the pharmacophore model were docked into the active site of NS-3 helicase. Based on docking scores and binding interactions, twenty five compounds are suggested to be potential inhibitors of NS3 Helicase. The pharmacokinetic properties of these hits were predicted. The selected hits revealed acceptable ADMET properties. This study identified potential inhibitors of NS-3 Helicase in silico, and can be helpful in the treatment of Dengue.

  7. Single molecule measurements of DNA helicase activity with magnetic tweezers and t-test based step-finding analysis

    Seol, Yeonee; Strub, Marie-Paule; Neuman, Keir C.

    2016-01-01

    Magnetic tweezers is a versatile and easy to implement single-molecule technique that has become increasingly prevalent in the study of nucleic acid based molecular motors. Here, we provide a description of the magnetic tweezers instrument and guidelines for measuring and analyzing DNA helicase activity. Along with experimental methods, we describe a robust method of single-molecule trajectory analysis based on the Student’s t-test that accommodates continuous transitions in addition to the discrete transitions assumed in most widely employed analysis routines. To illustrate the single-molecule unwinding assay and the analysis routine, we provide DNA unwinding measurements of Escherichia coli RecQ helicase under a variety of conditions (Na+, ATP, temperature, and DNA substrate geometry). These examples reveal that DNA unwinding measurements under various conditions can aid in elucidating the unwinding mechanism of DNA helicase but also emphasize that environmental effects on DNA helicase activity must be considered in relation to in vivo activity and mechanism. PMID:27131595

  8. Structure-Based Mutational Analysis of the Hepatitis C Virus NS3 Helicase

    Tai, Chun-Ling; Pan, Wen-Ching; Liaw, Shwu-Huey; Yang, Ueng-Cheng; Hwang, Lih-Hwa; Chen, Ding-Shinn

    2001-01-01

    The carboxyl terminus of the hepatitis C virus (HCV) nonstructural protein 3 (NS3) possesses ATP-dependent RNA helicase activity. Based on the conserved sequence motifs and the crystal structures of the helicase domain, 17 mutants of the HCV NS3 helicase were generated. The ATP hydrolysis, RNA binding, and RNA unwinding activities of the mutant proteins were examined in vitro to determine the functional role of the mutated residues. The data revealed that Lys-210 in the Walker A motif and Asp-290, Glu-291, and His-293 in the Walker B motif were crucial to ATPase activity and that Thr-322 and Thr-324 in motif III and Arg-461 in motif VI significantly influenced ATPase activity. When the pairing between His-293 and Gln-460, referred to as gatekeepers, was replaced with the Asp-293/His-460 pair, which makes the NS3 helicase more like the DEAD helicase subgroup, ATPase activity was not restored. It thus indicated that the whole microenvironment surrounding the gatekeepers, rather than the residues per se, was important to the enzymatic activities. Arg-461 and Trp-501 are important residues for RNA binding, while Val-432 may only play a coadjutant role. The data demonstrated that RNA helicase activity was possibly abolished by the loss of ATPase activity or by reduced RNA binding activity. Nevertheless, a low threshold level of ATPase activity was found sufficient for helicase activity. Results in this study provide a valuable reference for efforts under way to develop anti-HCV therapeutic drugs targeting NS3. PMID:11483774

  9. Structural mechanisms of human RecQ helicases WRN and BLM

    Ken eKitano

    2014-10-01

    Full Text Available The RecQ family DNA helicases WRN (Werner syndrome protein and BLM (Bloom syndrome protein play a key role in protecting the genome against deleterious changes. In humans, mutations in these proteins lead to rare genetic diseases associated with cancer predisposition and accelerated aging. WRN and BLM are distinguished from other helicases by possessing signature tandem domains toward the C terminus, referred to as the RecQ C-terminal (RQC and helicase-and-ribonuclease D-C-terminal (HRDC domains. Although the precise function of the HRDC domain remains unclear, the previous crystal structure of a WRN RQC-DNA complex visualized a central role for the RQC domain in recognizing, binding and unwinding DNA at branch points. In particular, a prominent hairpin structure (the β-wing within the RQC winged-helix motif acts as a scalpel to induce the unpairing of a Watson-Crick base pair at the DNA duplex terminus. A similar RQC-DNA interaction was also observed in the recent crystal structure of a BLM-DNA complex. I review the latest structures of WRN and BLM, and then provide a docking simulation of BLM with a Holliday junction. The model offers an explanation for the efficient branch migration activity of the RecQ family toward recombination and repair intermediates.

  10. Substrate-assisted mechanism of RNP disruption by the spliceosomal Brr2 RNA helicase

    Theuser, Matthias; Höbartner, Claudia; Wahl, Markus C.; Santos, Karine F.

    2016-01-01

    The Brr2 RNA helicase disrupts the U4/U6 di-small nuclear RNA–protein complex (di-snRNP) during spliceosome activation via ATP-driven translocation on the U4 snRNA strand. However, it is unclear how bound proteins influence U4/U6 unwinding, which regions of the U4/U6 duplex the helicase actively unwinds, and whether U4/U6 components are released as individual molecules or as subcomplexes. Here, we set up a recombinant Brr2-mediated U4/U6 di-snRNP disruption system, showing that sequential addition of the U4/U6 proteins small nuclear ribonucleoprotein-associated protein 1 (Snu13), pre-mRNA processing factor 31 (Prp31), and Prp3 to U4/U6 di-snRNA leads to a stepwise decrease of Brr2-mediated U4/U6 unwinding, but that unwinding is largely restored by a Brr2 cofactor, the C-terminal Jab1/MPN domain of the Prp8 protein. Brr2-mediated U4/U6 unwinding was strongly inhibited by mutations in U4/U6 di-snRNAs that diminish the ability of U6 snRNA to adopt an alternative conformation but leave the number and kind of U4/U6 base pairs unchanged. Irrespective of the presence of the cofactor, the helicase segregated a Prp3-Prp31-Snu13-U4/U6 RNP into an intact Prp31-Snu13-U4 snRNA particle, free Prp3, and free U6 snRNA. Together, these observations suggest that Brr2 translocates only a limited distance on the U4 snRNA strand and does not actively release RNA-bound proteins. Unwinding is then completed by the partially displaced U6 snRNA adopting an alternative conformation, which leads to dismantling of the Prp3-binding site on U4/U6 di-snRNA but leaves the Prp31- and Snu13-binding sites on U4 snRNA unaffected. In this fashion, Brr2 can activate the spliceosome by stripping U6 snRNA of all precatalytic binding partners, while minimizing logistic requirements for U4/U6 di-snRNP reassembly after splicing. PMID:27354531

  11. Velocity and processivity of helicase unwinding of double-stranded nucleic acids

    Betterton, M D; Juelicher, F

    2005-01-01

    Helicases are molecular motors which unwind double-stranded nucleic acids (dsNA) in cells. Many helicases move with directional bias on single-stranded (ss) nucleic acids, and couple their directional translocation to strand separation. A model of the coupling between translocation and unwinding uses an interaction potential to represent passive and active helicase mechanisms. A passive helicase must wait for thermal fluctuations to open dsNA base pairs before it can advance and inhibit NA closing. An active helicase directly destabilizes dsNA base pairs, accelerating the opening rate. Here we extend this model to include helicase unbinding from the nucleic-acid strand. The helicase processivity depends on the form of the interaction potential. A passive helicase has a mean attachment time which does not change between ss translocation and ds unwinding, while an active helicase in general shows a decrease in attachment time during unwinding relative to ss translocation. In addition, we describe how helicase unwinding velocity and processivity vary if the base-pair binding free energy is changed

  12. AAA-ATPase FIDGETIN-LIKE 1 and Helicase FANCM Antagonize Meiotic Crossovers by Distinct Mechanisms.

    Chloe Girard

    2015-07-01

    Full Text Available Meiotic crossovers (COs generate genetic diversity and are critical for the correct completion of meiosis in most species. Their occurrence is tightly constrained but the mechanisms underlying this limitation remain poorly understood. Here we identified the conserved AAA-ATPase FIDGETIN-LIKE-1 (FIGL1 as a negative regulator of meiotic CO formation. We show that Arabidopsis FIGL1 limits CO formation genome-wide, that FIGL1 controls dynamics of the two conserved recombinases DMC1 and RAD51 and that FIGL1 hinders the interaction between homologous chromosomes, suggesting that FIGL1 counteracts DMC1/RAD51-mediated inter-homologue strand invasion to limit CO formation. Further, depleting both FIGL1 and the previously identified anti-CO helicase FANCM synergistically increases crossover frequency. Additionally, we showed that the effect of mutating FANCM on recombination is much lower in F1 hybrids contrasting from the phenotype of inbred lines, while figl1 mutation equally increases crossovers in both contexts. This shows that the modes of action of FIGL1 and FANCM are differently affected by genomic contexts. We propose that FIGL1 and FANCM represent two successive barriers to CO formation, one limiting strand invasion, the other disassembling D-loops to promote SDSA, which when both lifted, leads to a large increase of crossovers, without impairing meiotic progression.

  13. Structural view of the helicase reveals that Zika virus uses a conserved mechanism for unwinding RNA.

    Li, Lei; Wang, Jin; Jia, Zhihui; Shaw, Neil

    2018-04-01

    Recent studies suggest a link between infection by Zika virus (ZIKV) and the development of neurological complications. The lack of ZIKV-specific therapeutics has alarmed healthcare professionals worldwide. Here, crystal structures of apo and AMPPNP- and Mn 2+ -bound forms of the essential helicase of ZIKV refined to 1.78 and 1.3 Å resolution, respectively, are reported. The structures reveal a conserved trimodular topology of the helicase. ATP and Mn 2+ are tethered between two RecA-like domains by conserved hydrogen-bonding interactions. The binding of ligands induces the movement of backbone Cα and side-chain atoms. Numerous solvent molecules are observed in the vicinity of the AMPPNP, suggesting a role in catalysis. These high-resolution structures could be useful for the design of inhibitors targeting the helicase of ZIKV for the treatment of infections caused by ZIKV.

  14. Dna2 nuclease-helicase structure, mechanism and regulation by Rpa.

    Zhou, Chun; Pourmal, Sergei; Pavletich, Nikola P

    2015-11-02

    The Dna2 nuclease-helicase maintains genomic integrity by processing DNA double-strand breaks, Okazaki fragments and stalled replication forks. Dna2 requires ssDNA ends, and is dependent on the ssDNA-binding protein Rpa, which controls cleavage polarity. Here we present the 2.3 Å structure of intact mouse Dna2 bound to a 15-nucleotide ssDNA. The nuclease active site is embedded in a long, narrow tunnel through which the DNA has to thread. The helicase domain is required for DNA binding but not threading. We also present the structure of a flexibly-tethered Dna2-Rpa interaction that recruits Dna2 to Rpa-coated DNA. We establish that a second Dna2-Rpa interaction is mutually exclusive with Rpa-DNA interactions and mediates the displacement of Rpa from ssDNA. This interaction occurs at the nuclease tunnel entrance and the 5' end of the Rpa-DNA complex. Hence, it only displaces Rpa from the 5' but not 3' end, explaining how Rpa regulates cleavage polarity.

  15. Assessment of Dengue virus helicase and methyltransferase as targets for fragment-based drug discovery.

    Coutard, Bruno; Decroly, Etienne; Li, Changqing; Sharff, Andrew; Lescar, Julien; Bricogne, Gérard; Barral, Karine

    2014-06-01

    Seasonal and pandemic flaviviruses continue to be leading global health concerns. With the view to help drug discovery against Dengue virus (DENV), a fragment-based experimental approach was applied to identify small molecule ligands targeting two main components of the flavivirus replication complex: the NS3 helicase (Hel) and the NS5 mRNA methyltransferase (MTase) domains. A library of 500 drug-like fragments was first screened by thermal-shift assay (TSA) leading to the identification of 36 and 32 fragment hits binding Hel and MTase from DENV, respectively. In a second stage, we set up a fragment-based X-ray crystallographic screening (FBS-X) in order to provide both validated fragment hits and structural binding information. No fragment hit was confirmed for DENV Hel. In contrast, a total of seven fragments were identified as DENV MTase binders and structures of MTase-fragment hit complexes were solved at resolution at least 2.0Å or better. All fragment hits identified contain either a five- or six-membered aromatic ring or both, and three novel binding sites were located on the MTase. To further characterize the fragment hits identified by TSA and FBS-X, we performed enzymatic assays to assess their inhibition effect on the N7- and 2'-O-MTase enzymatic activities: five of these fragment hits inhibit at least one of the two activities with IC50 ranging from 180μM to 9mM. This work validates the FBS-X strategy for identifying new anti-flaviviral hits targeting MTase, while Hel might not be an amenable target for fragment-based drug discovery (FBDD). This approach proved to be a fast and efficient screening method for FBDD target validation and discovery of starting hits for the development of higher affinity molecules that bind to novel allosteric sites. Copyright © 2014 Elsevier B.V. All rights reserved.

  16. RecQL5 promotes genome stabilization through two parallel mechanisms--interacting with RNA polymerase II and acting as a helicase.

    Islam, M Nurul; Fox, David; Guo, Rong; Enomoto, Takemi; Wang, Weidong

    2010-05-01

    The RecQL5 helicase is essential for maintaining genome stability and reducing cancer risk. To elucidate its mechanism of action, we purified a RecQL5-associated complex and identified its major component as RNA polymerase II (Pol II). Bioinformatics and structural modeling-guided mutagenesis revealed two conserved regions in RecQL5 as KIX and SRI domains, already known in transcriptional regulators for Pol II. The RecQL5-KIX domain binds both initiation (Pol IIa) and elongation (Pol IIo) forms of the polymerase, whereas the RecQL5-SRI domain interacts only with the elongation form. Fully functional RecQL5 requires both helicase activity and associations with the initiation polymerase, because mutants lacking either activity are partially defective in the suppression of sister chromatid exchange and resistance to camptothecin-induced DNA damage, and mutants lacking both activities are completely defective. We propose that RecQL5 promotes genome stabilization through two parallel mechanisms: by participation in homologous recombination-dependent DNA repair as a RecQ helicase and by regulating the initiation of Pol II to reduce transcription-associated replication impairment and recombination.

  17. The RNA-mediated, asymmetric ring regulatory mechanism of the transcription termination Rho helicase decrypted by time-resolved nucleotide analog interference probing (trNAIP).

    Soares, Emilie; Schwartz, Annie; Nollmann, Marcello; Margeat, Emmanuel; Boudvillain, Marc

    2014-08-01

    Rho is a ring-shaped, ATP-dependent RNA helicase/translocase that dissociates transcriptional complexes in bacteria. How RNA recognition is coupled to ATP hydrolysis and translocation in Rho is unclear. Here, we develop and use a new combinatorial approach, called time-resolved Nucleotide Analog Interference Probing (trNAIP), to unmask RNA molecular determinants of catalytic Rho function. We identify a regulatory step in the translocation cycle involving recruitment of the 2'-hydroxyl group of the incoming 3'-RNA nucleotide by a Rho subunit. We propose that this step arises from the intrinsic weakness of one of the subunit interfaces caused by asymmetric, split-ring arrangement of primary RNA tethers around the Rho hexamer. Translocation is at highest stake every seventh nucleotide when the weak interface engages the incoming 3'-RNA nucleotide or breaks, depending on RNA threading constraints in the Rho pore. This substrate-governed, 'test to run' iterative mechanism offers a new perspective on how a ring-translocase may function or be regulated. It also illustrates the interest and versatility of the new trNAIP methodology to unveil the molecular mechanisms of complex RNA-based systems. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

  18. Structural basis of Zika virus helicase in recognizing its substrates

    Hongliang Tian

    2016-07-01

    Full Text Available Abstract The recent explosive outbreak of Zika virus (ZIKV infection has been reported in South and Central America and the Caribbean. Neonatal microcephaly associated with ZIKV infection has already caused a public health emergency of international concern. No specific vaccines or drugs are currently available to treat ZIKV infection. The ZIKV helicase, which plays a pivotal role in viral RNA replication, is an attractive target for therapy. We determined the crystal structures of ZIKV helicase-ATP-Mn2+ and ZIKV helicase-RNA. This is the first structure of any flavivirus helicase bound to ATP. Comparisons with related flavivirus helicases have shown that although the critical P-loop in the active site has variable conformations among different species, it adopts an identical mode to recognize ATP/Mn2+. The structure of ZIKV helicase-RNA has revealed that upon RNA binding, rotations of the motor domains can cause significant conformational changes. Strikingly, although ZIKV and dengue virus (DENV apo-helicases share conserved residues for RNA binding, their different manners of motor domain rotations result in distinct individual modes for RNA recognition. It suggests that flavivirus helicases could have evolved a conserved engine to convert chemical energy from nucleoside triphosphate to mechanical energy for RNA unwinding, but different motor domain rotations result in variable RNA recognition modes to adapt to individual viral replication.

  19. General Methods for Analysis of Sequential “n-step” Kinetic Mechanisms: Application to Single Turnover Kinetics of Helicase-Catalyzed DNA Unwinding

    Lucius, Aaron L.; Maluf, Nasib K.; Fischer, Christopher J.; Lohman, Timothy M.

    2003-01-01

    Helicase-catalyzed DNA unwinding is often studied using “all or none” assays that detect only the final product of fully unwound DNA. Even using these assays, quantitative analysis of DNA unwinding time courses for DNA duplexes of different lengths, L, using “n-step” sequential mechanisms, can reveal information about the number of intermediates in the unwinding reaction and the “kinetic step size”, m, defined as the average number of basepairs unwound between two successive rate limiting steps in the unwinding cycle. Simultaneous nonlinear least-squares analysis using “n-step” sequential mechanisms has previously been limited by an inability to float the number of “unwinding steps”, n, and m, in the fitting algorithm. Here we discuss the behavior of single turnover DNA unwinding time courses and describe novel methods for nonlinear least-squares analysis that overcome these problems. Analytic expressions for the time courses, fss(t), when obtainable, can be written using gamma and incomplete gamma functions. When analytic expressions are not obtainable, the numerical solution of the inverse Laplace transform can be used to obtain fss(t). Both methods allow n and m to be continuous fitting parameters. These approaches are generally applicable to enzymes that translocate along a lattice or require repetition of a series of steps before product formation. PMID:14507688

  20. General methods for analysis of sequential "n-step" kinetic mechanisms: application to single turnover kinetics of helicase-catalyzed DNA unwinding.

    Lucius, Aaron L; Maluf, Nasib K; Fischer, Christopher J; Lohman, Timothy M

    2003-10-01

    Helicase-catalyzed DNA unwinding is often studied using "all or none" assays that detect only the final product of fully unwound DNA. Even using these assays, quantitative analysis of DNA unwinding time courses for DNA duplexes of different lengths, L, using "n-step" sequential mechanisms, can reveal information about the number of intermediates in the unwinding reaction and the "kinetic step size", m, defined as the average number of basepairs unwound between two successive rate limiting steps in the unwinding cycle. Simultaneous nonlinear least-squares analysis using "n-step" sequential mechanisms has previously been limited by an inability to float the number of "unwinding steps", n, and m, in the fitting algorithm. Here we discuss the behavior of single turnover DNA unwinding time courses and describe novel methods for nonlinear least-squares analysis that overcome these problems. Analytic expressions for the time courses, f(ss)(t), when obtainable, can be written using gamma and incomplete gamma functions. When analytic expressions are not obtainable, the numerical solution of the inverse Laplace transform can be used to obtain f(ss)(t). Both methods allow n and m to be continuous fitting parameters. These approaches are generally applicable to enzymes that translocate along a lattice or require repetition of a series of steps before product formation.

  1. RecQ Helicases

    Larsen, Nicolai Balle; Hickson, Ian D

    2013-01-01

    The RecQ family of DNA helicases is highly conserved throughout -evolution, and is important for the maintenance of genome stability. In humans, five RecQ family members have been identified: BLM, WRN, RECQ4, RECQ1 and RECQ5. Defects in three of these give rise to Bloom's syndrome (BLM), Werner...

  2. Helicase-dependent isothermal amplification: a novel tool in the development of molecular-based analytical systems for rapid pathogen detection.

    Barreda-García, Susana; Miranda-Castro, Rebeca; de-Los-Santos-Álvarez, Noemí; Miranda-Ordieres, Arturo J; Lobo-Castañón, María Jesús

    2018-01-01

    Highly sensitive testing of nucleic acids is essential to improve the detection of pathogens, which pose a major threat for public health worldwide. Currently available molecular assays, mainly based on PCR, have a limited utility in point-of-need control or resource-limited settings. Consequently, there is a strong interest in developing cost-effective, robust, and portable platforms for early detection of these harmful microorganisms. Since its description in 2004, isothermal helicase-dependent amplification (HDA) has been successfully applied in the development of novel molecular-based technologies for rapid, sensitive, and selective detection of viruses and bacteria. In this review, we highlight relevant analytical systems using this simple nucleic acid amplification methodology that takes place at a constant temperature and that is readily compatible with microfluidic technologies. Different strategies for monitoring HDA amplification products are described. In addition, we present technological advances for integrating sample preparation, HDA amplification, and detection. Future perspectives and challenges toward point-of-need use not only for clinical diagnosis but also in food safety testing and environmental monitoring are also discussed. Graphical Abstract Expanding the analytical toolbox for the detection of DNA sequences specific of pathogens with isothermal helicase dependent amplification (HDA).

  3. Overcoming natural replication barriers: differential helicase requirements.

    Anand, Ranjith P; Shah, Kartik A; Niu, Hengyao; Sung, Patrick; Mirkin, Sergei M; Freudenreich, Catherine H

    2012-02-01

    DNA sequences that form secondary structures or bind protein complexes are known barriers to replication and potential inducers of genome instability. In order to determine which helicases facilitate DNA replication across these barriers, we analyzed fork progression through them in wild-type and mutant yeast cells, using 2-dimensional gel-electrophoretic analysis of the replication intermediates. We show that the Srs2 protein facilitates replication of hairpin-forming CGG/CCG repeats and prevents chromosome fragility at the repeat, whereas it does not affect replication of G-quadruplex forming sequences or a protein-bound repeat. Srs2 helicase activity is required for hairpin unwinding and fork progression. Also, the PCNA binding domain of Srs2 is required for its in vivo role of replication through hairpins. In contrast, the absence of Sgs1 or Pif1 helicases did not inhibit replication through structural barriers, though Pif1 did facilitate replication of a telomeric protein barrier. Interestingly, replication through a protein barrier but not a DNA structure barrier was modulated by nucleotide pool levels, illuminating a different mechanism by which cells can regulate fork progression through protein-mediated stall sites. Our analyses reveal fundamental differences in the replication of DNA structural versus protein barriers, with Srs2 helicase activity exclusively required for fork progression through hairpin structures.

  4. Inhibition of RNA Helicases of ssRNA+ Virus Belonging to Flaviviridae, Coronaviridae and Picornaviridae Families

    Irene Briguglio

    2011-01-01

    Full Text Available Many viral pathogens encode the motor proteins named RNA helicases which display various functions in genome replication. General strategies to design specific and selective drugs targeting helicase for the treatment of viral infections could act via one or more of the following mechanisms: inhibition of the NTPase activity, by interferences with ATP binding and therefore by limiting the energy required for the unwinding and translocation, or by allosteric mechanism and therefore by stabilizing the conformation of the enzyme in low helicase activity state; inhibition of nucleic acids binding to the helicase; inhibition of coupling of ATP hydrolysis to unwinding; inhibition of unwinding by sterically blocking helicase translocation. Recently, by in vitro screening studies, it has been reported that several benzotriazole, imidazole, imidazodiazepine, phenothiazine, quinoline, anthracycline, triphenylmethane, tropolone, pyrrole, acridone, small peptide, and Bananin derivatives are endowed with helicase inhibition of pathogen viruses belonging to Flaviviridae, Coronaviridae, and Picornaviridae families.

  5. GINS complex protein Sld5 recruits SIK1 to activate MCM helicase during DNA replication.

    Joshi, Kiranmai; Shah, Varun Jayeshkumar; Maddika, Subbareddy

    2016-12-01

    In eukaryotes, proper loading and activation of MCM helicase at chromosomal origins plays a central role in DNA replication. Activation of MCM helicase requires its association with CDC45-GINS complex, but the mechanism of how this complex activates MCM helicase is poorly understood. Here we identified SIK1 (salt-inducible kinase 1), an AMPK related protein kinase, as a molecular link that connects GINS complex with MCM helicase activity. We demonstrated that Sld5 a component of GINS complex interacts with SIK1 and recruits it to the sites of DNA replication at the onset of S phase. Depletion of SIK1 leads to defective DNA replication. Further, we showed that SIK1 phosphorylates MCM2 at five conserved residues at its N-terminus, which is essential for the activation of MCM helicase. Collectively, our results suggest SIK1 as a novel integral component of CMG replicative helicase during eukaryotic DNA replication. Copyright © 2016 Elsevier Inc. All rights reserved.

  6. Preliminary crystallographic characterization of an RNA helicase from Kunjin virus

    Mastrangelo, Eloise; Bollati, Michela; Milani, Mario; Brisbarre, Nadège; Lamballerie, Xavier de; Coutard, Bruno; Canard, Bruno; Khromykh, Alexander; Bolognesi, Martino

    2006-01-01

    The C-terminal 440 amino acids of the NS3 protein from Kunjin virus (Flaviviridae) code for a helicase. The protein has been overexpressed and crystallized. Characterization of the isolated monoclinic crystal form and diffraction data (at 3.0 Å resolution) are presented, together with a preliminary molecular-replacement solution. Kunjin virus is a member of the Flavivirus genus and is an Australian variant of West Nile virus. The C-terminal domain of the Kunjin virus NS3 protein displays helicase activity. The protein is thought to separate daughter and template RNA strands, assisting the initiation of replication by unwinding RNA secondary structure in the 3′ nontranslated region. Expression, purification and preliminary crystallographic characterization of the NS3 helicase domain are reported. It is shown that Kunjin virus helicase may adopt a dimeric assembly in absence of nucleic acids, oligomerization being a means to provide the helicases with multiple nucleic acid-binding capability, facilitating translocation along the RNA strands. Kunjin virus NS3 helicase domain is an attractive model for studying the molecular mechanisms of flavivirus replication, while simultaneously providing a new basis for the rational development of anti-flaviviral compounds

  7. Structure based modification of Bluetongue virus helicase protein VP6 to produce a viable VP6-truncated BTV

    Matsuo, Eiko [Microbiology and Immunology, Division of Animal Science, Department of Bioresource Science, Graduate School of Agricultural Science, Kobe University, 1-1, Rokkodai, Nada-ku, Kobe-City 657-8501 (Japan); Faculty of Infectious and Tropical Diseases, London School of Hygiene and Tropical Medicine, Keppel Street, London WC1E 7HT (United Kingdom); Leon, Esther; Matthews, Steve J. [Division of Molecular Biosciences, Centre for Structural Biology, Imperial College London, South Kensington, London SW7 2AZ (United Kingdom); Roy, Polly, E-mail: polly.roy@lshtm.ac.uk [Faculty of Infectious and Tropical Diseases, London School of Hygiene and Tropical Medicine, Keppel Street, London WC1E 7HT (United Kingdom)

    2014-09-05

    Highlights: • NMR analysis on BTV VP6 reveals two large loop regions. • The loss of a loop (aa 34–130) does not affect the overall fold of the protein. • A region of VP6 (aa 34–92) is not required for BTV replication. • A region of VP6 (aa 93–130) plays an essential role in the virus replication. - Abstract: Bluetongue virus core protein VP6 is an ATP hydrolysis dependent RNA helicase. However, despite much study, the precise role of VP6 within the viral capsid and its structure remain unclear. To investigate the requirement of VP6 in BTV replication, we initiated a structural and biological study. Multinuclear nuclear magnetic resonance spectra were assigned on his-tagged full-length VP6 (329 amino acid residues) as well as several truncated VP6 variants. The analysis revealed a large structured domain with two large loop regions that exhibit significant conformational exchange. One of the loops (amino acid position 34–130) could be removed without affecting the overall fold of the protein. Moreover, using a BTV reverse genetics system, it was possible to demonstrate that the VP6-truncated BTV was viable in BHK cells in the absence of any helper VP6 protein, suggesting that a large portion of this loop region is not absolutely required for BTV replication.

  8. RecQ helicases and cellular responses to DNA damage

    Wu, Leonard; Hickson, Ian D.

    2002-01-01

    The faithful replication of the genome is essential for the survival of all organisms. It is not surprising therefore that numerous mechanisms have evolved to ensure that duplication of the genome occurs with only minimal risk of mutation induction. One mechanism of genome destabilization is replication fork demise, which can occur when a translocating fork meets a lesion or adduct in the template. Indeed, the collapse of replication forks has been suggested to occur in every replicative cell cycle making this a potentially significant problem for all proliferating cells. The RecQ helicases, which are essential for the maintenance of genome stability, are thought to function during DNA replication. In particular, RecQ helicase mutants display replication defects and have phenotypes consistent with an inability to efficiently reinitiate replication following replication fork demise. Here, we review some current models for how replication fork repair might be effected, and discuss potential roles for RecQ helicases in this process

  9. Mitochondrial helicases and mitochondrial genome maintenance

    de Souza-Pinto, Nadja C; Aamann, Maria Diget; Kulikowicz, Tomasz

    2010-01-01

    Helicases are essential enzymes that utilize the energy of nucleotide hydrolysis to drive unwinding of nucleic acid duplexes. Helicases play roles in all aspects of DNA metabolism including DNA repair, DNA replication and transcription. The subcellular locations and functions of several helicases...

  10. Identification of Hydroxyanthraquinones as Novel Inhibitors of Hepatitis C Virus NS3 Helicase

    Furuta, Atsushi; Tsubuki, Masayoshi; Endoh, Miduki; Miyamoto, Tatsuki; Tanaka, Junichi; Abdus Salam, Kazi; Akimitsu, Nobuyoshi; Tani, Hidenori; Yamashita, Atsuya; Moriishi, Kohji; Nakakoshi, Masamichi; Sekiguchi, Yuji; Tsuneda, Satoshi; Noda, Naohiro

    2015-01-01

    Hepatitis C virus (HCV) is an important etiological agent of severe liver diseases, including cirrhosis and hepatocellular carcinoma. The HCV genome encodes nonstructural protein 3 (NS3) helicase, which is a potential anti-HCV drug target because its enzymatic activity is essential for viral replication. Some anthracyclines are known to be NS3 helicase inhibitors and have a hydroxyanthraquinone moiety in their structures; mitoxantrone, a hydroxyanthraquinone analogue, is also known to inhibit NS3 helicase. Therefore, we hypothesized that the hydroxyanthraquinone moiety alone could also inhibit NS3 helicase. Here, we performed a structure–activity relationship study on a series of hydroxyanthraquinones by using a fluorescence-based helicase assay. Hydroxyanthraquinones inhibited NS3 helicase with IC50 values in the micromolar range. The inhibitory activity varied depending on the number and position of the phenolic hydroxyl groups, and among different hydroxyanthraquinones examined, 1,4,5,8-tetrahydroxyanthraquinone strongly inhibited NS3 helicase with an IC50 value of 6 µM. Furthermore, hypericin and sennidin A, which both have two hydroxyanthraquinone-like moieties, were found to exert even stronger inhibition with IC50 values of 3 and 0.8 µM, respectively. These results indicate that the hydroxyanthraquinone moiety can inhibit NS3 helicase and suggest that several key chemical structures are important for the inhibition. PMID:26262613

  11. Identification of Hydroxyanthraquinones as Novel Inhibitors of Hepatitis C Virus NS3 Helicase

    Atsushi Furuta

    2015-08-01

    Full Text Available Hepatitis C virus (HCV is an important etiological agent of severe liver diseases, including cirrhosis and hepatocellular carcinoma. The HCV genome encodes nonstructural protein 3 (NS3 helicase, which is a potential anti-HCV drug target because its enzymatic activity is essential for viral replication. Some anthracyclines are known to be NS3 helicase inhibitors and have a hydroxyanthraquinone moiety in their structures; mitoxantrone, a hydroxyanthraquinone analogue, is also known to inhibit NS3 helicase. Therefore, we hypothesized that the hydroxyanthraquinone moiety alone could also inhibit NS3 helicase. Here, we performed a structure–activity relationship study on a series of hydroxyanthraquinones by using a fluorescence-based helicase assay. Hydroxyanthraquinones inhibited NS3 helicase with IC50 values in the micromolar range. The inhibitory activity varied depending on the number and position of the phenolic hydroxyl groups, and among different hydroxyanthraquinones examined, 1,4,5,8-tetrahydroxyanthraquinone strongly inhibited NS3 helicase with an IC50 value of 6 µM. Furthermore, hypericin and sennidin A, which both have two hydroxyanthraquinone-like moieties, were found to exert even stronger inhibition with IC50 values of 3 and 0.8 µM, respectively. These results indicate that the hydroxyanthraquinone moiety can inhibit NS3 helicase and suggest that several key chemical structures are important for the inhibition.

  12. Ebselen Inhibits Hepatitis C Virus NS3 Helicase Binding to Nucleic Acid and Prevents Viral Replication

    Mukherjee, Sourav; Weiner, Warren S.; Schroeder, Chad E.; Simpson, Denise S.; Hanson, Alicia M.; Sweeney, Noreena L.; Marvin, Rachel K.; Ndjomou, Jean; Kolli, Rajesh; Isailovic, Dragan; Schoenen, Frank J.; Frick, David N.

    2014-01-01

    The hepatitis C virus (HCV) nonstructural protein 3 (NS3) is both a protease, which cleaves viral and host proteins, and a helicase that separates nucleic acid strands, using ATP hydrolysis to fuel the reaction. Many antiviral drugs, and compounds in clinical trials, target the NS3 protease, but few helicase inhibitors that function as antivirals have been reported. This study focuses on the analysis of the mechanism by which ebselen (2-phenyl-1,2-benzisoselenazol-3-one), a compound previousl...

  13. Genome-Wide Analysis of the RNA Helicase Gene Family in Gossypium raimondii

    Jie Chen

    2014-03-01

    Full Text Available The RNA helicases, which help to unwind stable RNA duplexes, and have important roles in RNA metabolism, belong to a class of motor proteins that play important roles in plant development and responses to stress. Although this family of genes has been the subject of systematic investigation in Arabidopsis, rice, and tomato, it has not yet been characterized in cotton. In this study, we identified 161 putative RNA helicase genes in the genome of the diploid cotton species Gossypium raimondii. We classified these genes into three subfamilies, based on the presence of either a DEAD-box (51 genes, DEAH-box (52 genes, or DExD/H-box (58 genes in their coding regions. Chromosome location analysis showed that the genes that encode RNA helicases are distributed across all 13 chromosomes of G. raimondii. Syntenic analysis revealed that 62 of the 161 G. raimondii helicase genes (38.5% are within the identified syntenic blocks. Sixty-six (40.99% helicase genes from G. raimondii have one or several putative orthologs in tomato. Additionally, GrDEADs have more conserved gene structures and more simple domains than GrDEAHs and GrDExD/Hs. Transcriptome sequencing data demonstrated that many of these helicases, especially GrDEADs, are highly expressed at the fiber initiation stage and in mature leaves. To our knowledge, this is the first report of a genome-wide analysis of the RNA helicase gene family in cotton.

  14. Robust translocation along a molecular monorail: the NS3 helicase from hepatitis C virus traverses unusually large disruptions in its track.

    Beran, Rudolf K F; Bruno, Michael M; Bowers, Heath A; Jankowsky, Eckhard; Pyle, Anna Marie

    2006-05-12

    The NS3 helicase is essential for replication of the hepatitis C virus. This multifunctional Superfamily 2 helicase protein unwinds nucleic acid duplexes in a stepwise, ATP-dependent manner. Although kinetic features of its mechanism are beginning to emerge, little is known about the physical determinants for NS3 translocation along a strand of nucleic acid. For example, it is not known whether NS3 can traverse covalent or physical discontinuities on the tracking strand. Here we provide evidence that NS3 translocates with a mechanism that is different from its well-studied relative, the Vaccinia helicase NPH-II. Like NPH-II, NS3 translocates along the loading strand (the strand bearing the 3'-overhang) and it fails to unwind substrates that contain nicks, or covalent discontinuities in the loading strand. However, unlike NPH-II, NS3 readily unwinds RNA duplexes that contain long stretches of polyglycol, which are moieties that bear no resemblance to nucleic acid. Whether located on the tracking strand, the top strand, or both, long polyglycol regions fail to disrupt the function of NS3. This suggests that NS3 does not require the continuous formation of specific contacts with the ribose-phosphate backbone as it translocates along an RNA duplex, which is an observation consistent with the large NS3 kinetic step size (18 base-pairs). Rather, once NS3 loads onto a substrate, the helicase can translocate along the loading strand of an RNA duplex like a monorail train following a track. Bumps in the track do not significantly disturb NS3 unwinding, but a break in the track de-rails the helicase.

  15. High-throughput screening assay of hepatitis C virus helicase inhibitors using fluorescence-quenching phenomenon

    Tani, Hidenori; Akimitsu, Nobuyoshi; Fujita, Osamu; Matsuda, Yasuyoshi; Miyata, Ryo; Tsuneda, Satoshi; Igarashi, Masayuki; Sekiguchi, Yuji; Noda, Naohiro

    2009-01-01

    We have developed a novel high-throughput screening assay of hepatitis C virus (HCV) nonstructural protein 3 (NS3) helicase inhibitors using the fluorescence-quenching phenomenon via photoinduced electron transfer between fluorescent dyes and guanine bases. We prepared double-stranded DNA (dsDNA) with a 5'-fluorescent-dye (BODIPY FL)-labeled strand hybridized with a complementary strand, the 3'-end of which has guanine bases. When dsDNA is unwound by helicase, the dye emits fluorescence owing to its release from the guanine bases. Our results demonstrate that this assay is suitable for quantitative assay of HCV NS3 helicase activity and useful for high-throughput screening for inhibitors. Furthermore, we applied this assay to the screening for NS3 helicase inhibitors from cell extracts of microorganisms, and found several cell extracts containing potential inhibitors.

  16. Authentic interdomain communication in an RNA helicase reconstituted by expressed protein ligation of two helicase domains.

    Karow, Anne R; Theissen, Bettina; Klostermeier, Dagmar

    2007-01-01

    RNA helicases mediate structural rearrangements of RNA or RNA-protein complexes at the expense of ATP hydrolysis. Members of the DEAD box helicase family consist of two flexibly connected helicase domains. They share nine conserved sequence motifs that are involved in nucleotide binding and hydrolysis, RNA binding, and helicase activity. Most of these motifs line the cleft between the two helicase domains, and extensive communication between them is required for RNA unwinding. The two helicase domains of the Bacillus subtilis RNA helicase YxiN were produced separately as intein fusions, and a functional RNA helicase was generated by expressed protein ligation. The ligated helicase binds adenine nucleotides with very similar affinities to the wild-type protein. Importantly, its intrinsically low ATPase activity is stimulated by RNA, and the Michaelis-Menten parameters are similar to those of the wild-type. Finally, ligated YxiN unwinds a minimal RNA substrate to an extent comparable to that of the wild-type helicase, confirming authentic interdomain communication.

  17. Ebselen inhibits hepatitis C virus NS3 helicase binding to nucleic acid and prevents viral replication.

    Mukherjee, Sourav; Weiner, Warren S; Schroeder, Chad E; Simpson, Denise S; Hanson, Alicia M; Sweeney, Noreena L; Marvin, Rachel K; Ndjomou, Jean; Kolli, Rajesh; Isailovic, Dragan; Schoenen, Frank J; Frick, David N

    2014-10-17

    The hepatitis C virus (HCV) nonstructural protein 3 (NS3) is both a protease, which cleaves viral and host proteins, and a helicase that separates nucleic acid strands, using ATP hydrolysis to fuel the reaction. Many antiviral drugs, and compounds in clinical trials, target the NS3 protease, but few helicase inhibitors that function as antivirals have been reported. This study focuses on the analysis of the mechanism by which ebselen (2-phenyl-1,2-benzisoselenazol-3-one), a compound previously shown to be a HCV antiviral agent, inhibits the NS3 helicase. Ebselen inhibited the abilities of NS3 to unwind nucleic acids, to bind nucleic acids, and to hydrolyze ATP, and about 1 μM ebselen was sufficient to inhibit each of these activities by 50%. However, ebselen had no effect on the activity of the NS3 protease, even at 100 times higher ebselen concentrations. At concentrations below 10 μM, the ability of ebselen to inhibit HCV helicase was reversible, but prolonged incubation of HCV helicase with higher ebselen concentrations led to irreversible inhibition and the formation of covalent adducts between ebselen and all 14 cysteines present in HCV helicase. Ebselen analogues with sulfur replacing the selenium were just as potent HCV helicase inhibitors as ebselen, but the length of the linker between the phenyl and benzisoselenazol rings was critical. Modifications of the phenyl ring also affected compound potency over 30-fold, and ebselen was a far more potent helicase inhibitor than other, structurally unrelated, thiol-modifying agents. Ebselen analogues were also more effective antiviral agents, and they were less toxic to hepatocytes than ebselen. Although the above structure-activity relationship studies suggest that ebselen targets a specific site on NS3, we were unable to confirm binding to either the NS3 ATP binding site or nucleic acid binding cleft by examining the effects of ebselen on NS3 proteins lacking key cysteines.

  18. Close encounters for the first time: Helicase interactions with DNA damage.

    Khan, Irfan; Sommers, Joshua A; Brosh, Robert M

    2015-09-01

    DNA helicases are molecular motors that harness the energy of nucleoside triphosphate hydrolysis to unwinding structured DNA molecules that must be resolved during cellular replication, DNA repair, recombination, and transcription. In vivo, DNA helicases are expected to encounter a wide spectrum of covalent DNA modifications to the sugar phosphate backbone or the nitrogenous bases; these modifications can be induced by endogenous biochemical processes or exposure to environmental agents. The frequency of lesion abundance can vary depending on the lesion type. Certain adducts such as oxidative base modifications can be quite numerous, and their effects can be helix-distorting or subtle perturbations to DNA structure. Helicase encounters with specific DNA lesions and more novel forms of DNA damage will be discussed. We will also review the battery of assays that have been used to characterize helicase-catalyzed unwinding of damaged DNA substrates. Characterization of the effects of specific DNA adducts on unwinding by various DNA repair and replication helicases has proven to be insightful for understanding mechanistic and biological aspects of helicase function in cellular DNA metabolism. Published by Elsevier B.V.

  19. ARCPHdb: A comprehensive protein database for SF1 and SF2 helicase from archaea.

    Moukhtar, Mirna; Chaar, Wafi; Abdel-Razzak, Ziad; Khalil, Mohamad; Taha, Samir; Chamieh, Hala

    2017-01-01

    Superfamily 1 and Superfamily 2 helicases, two of the largest helicase protein families, play vital roles in many biological processes including replication, transcription and translation. Study of helicase proteins in the model microorganisms of archaea have largely contributed to the understanding of their function, architecture and assembly. Based on a large phylogenomics approach, we have identified and classified all SF1 and SF2 protein families in ninety five sequenced archaea genomes. Here we developed an online webserver linked to a specialized protein database named ARCPHdb to provide access for SF1 and SF2 helicase families from archaea. ARCPHdb was implemented using MySQL relational database. Web interfaces were developed using Netbeans. Data were stored according to UniProt accession numbers, NCBI Ref Seq ID, PDB IDs and Entrez Databases. A user-friendly interactive web interface has been developed to browse, search and download archaeal helicase protein sequences, their available 3D structure models, and related documentation available in the literature provided by ARCPHdb. The database provides direct links to matching external databases. The ARCPHdb is the first online database to compile all protein information on SF1 and SF2 helicase from archaea in one platform. This database provides essential resource information for all researchers interested in the field. Copyright © 2016 Elsevier Ltd. All rights reserved.

  20. Knowledge base mechanisms

    Suwa, M; Furukawa, K; Makinouchi, A; Mizoguchi, T; Mizoguchi, F; Yamasaki, H

    1982-01-01

    One of the principal goals of the Fifth Generation Computer System Project for the coming decade is to develop a methodology for building knowledge information processing systems which will provide people with intelligent agents. The key notion of the fifth generation computer system is knowledge used for problem solving. In this paper the authors describe the plan of Randd on knowledge base mechanisms. A knowledge representation system is to be designed to support knowledge acquisition for the knowledge information processing systems. The system will include a knowledge representation language, a knowledge base editor and a debugger. It is also expected to perform as a kind of meta-inference system. In order to develop the large scale knowledge base systems, a knowledge base mechanism based on the relational model is to be studied in the earlier stage of the project. Distributed problem solving is also one of the main issues of the project. 19 references.

  1. RNA helicase HEL-1 promotes longevity by specifically activating DAF-16/FOXO transcription factor signaling in Caenorhabditis elegans

    Seo, Mihwa; Seo, Keunhee; Hwang, Wooseon; Koo, Hee Jung; Hahm, Jeong-Hoon; Yang, Jae-Seong; Han, Seong Kyu; Hwang, Daehee; Kim, Sanguk; Jang, Sung Key; Lee, Yoontae; Nam, Hong Gil; Lee, Seung-Jae V.

    2015-01-01

    The homeostatic maintenance of the genomic DNA is crucial for regulating aging processes. However, the role of RNA homeostasis in aging processes remains unknown. RNA helicases are a large family of enzymes that regulate the biogenesis and homeostasis of RNA. However, the functional significance of RNA helicases in aging has not been explored. Here, we report that a large fraction of RNA helicases regulate the lifespan of Caenorhabditis elegans. In particular, we show that a DEAD-box RNA helicase, helicase 1 (HEL-1), promotes longevity by specifically activating the DAF-16/forkhead box O (FOXO) transcription factor signaling pathway. We find that HEL-1 is required for the longevity conferred by reduced insulin/insulin-like growth factor 1 (IGF-1) signaling (IIS) and is sufficient for extending lifespan. We further show that the expression of HEL-1 in the intestine and neurons contributes to longevity. HEL-1 enhances the induction of a large fraction of DAF-16 target genes. Thus, the RNA helicase HEL-1 appears to promote longevity in response to decreased IIS as a transcription coregulator of DAF-16. Because HEL-1 and IIS are evolutionarily well conserved, a similar mechanism for longevity regulation via an RNA helicase-dependent regulation of FOXO signaling may operate in mammals, including humans. PMID:26195740

  2. Helicase and Polymerase Move Together Close to the Fork Junction and Copy DNA in One-Nucleotide Steps

    Manjula Pandey

    2014-03-01

    Full Text Available By simultaneously measuring DNA synthesis and dNTP hydrolysis, we show that T7 DNA polymerase and T7 gp4 helicase move in sync during leading-strand synthesis, taking one-nucleotide steps and hydrolyzing one dNTP per base-pair unwound/copied. The cooperative catalysis enables the helicase and polymerase to move at a uniformly fast rate without guanine:cytosine (GC dependency or idling with futile NTP hydrolysis. We show that the helicase and polymerase are located close to the replication fork junction. This architecture enables the polymerase to use its strand-displacement synthesis to increase the unwinding rate, whereas the helicase aids this process by translocating along single-stranded DNA and trapping the unwound bases. Thus, in contrast to the helicase-only unwinding model, our results suggest a model in which the helicase and polymerase are moving in one-nucleotide steps, DNA synthesis drives fork unwinding, and a role of the helicase is to trap the unwound bases and prevent DNA reannealing.

  3. Physical and functional interactions of Caenorhabditis elegans WRN-1 helicase with RPA-1.

    Hyun, Moonjung; Park, Sojin; Kim, Eunsun; Kim, Do-Hyung; Lee, Se-Jin; Koo, Hyeon-Sook; Seo, Yeon-Soo; Ahn, Byungchan

    2012-02-21

    The Caenorhabditis elegans Werner syndrome protein, WRN-1, a member of the RecQ helicase family, has a 3'-5' DNA helicase activity. Worms with defective wrn-1 exhibit premature aging phenotypes and an increased level of genome instability. In response to DNA damage, WRN-1 participates in the initial stages of checkpoint activation in concert with C. elegans replication protein A (RPA-1). WRN-1 helicase is stimulated by RPA-1 on long DNA duplex substrates. However, the mechanism by which RPA-1 stimulates DNA unwinding and the function of the WRN-1-RPA-1 interaction are not clearly understood. We have found that WRN-1 physically interacts with two RPA-1 subunits, CeRPA73 and CeRPA32; however, full-length WRN-1 helicase activity is stimulated by only the CeRPA73 subunit, while the WRN-1(162-1056) fragment that harbors the helicase activity requires both the CeRPA73 and CeRPA32 subunits for the stimulation. We also found that the CeRPA73(1-464) fragment can stimulate WRN-1 helicase activity and that residues 335-464 of CeRPA73 are important for physical interaction with WRN-1. Because CeRPA73 and the CeRPA73(1-464) fragment are able to bind single-stranded DNA (ssDNA), the stimulation of WRN-1 helicase by RPA-1 is most likely due to the ssDNA binding activity of CeRPA73 and the direct interaction of WRN-1 and CeRPA73.

  4. Purification and crystallization of Kokobera virus helicase

    De Colibus, Luigi; Speroni, Silvia; Coutard, Bruno; Forrester, Naomi L.; Gould, Ernest; Canard, Bruno; Mattevi, Andrea

    2007-01-01

    Kokobera virus is a mosquito-borne flavivirus belonging, like West Nile virus, to the Japanese encephalitis virus serocomplex. Crystals of the Kokobera virus helicase domain were obtained by the hanging-drop vapour-diffusion method and exhibit a diffraction limit of 2.3 Å. Kokobera virus is a mosquito-borne flavivirus belonging, like West Nile virus, to the Japanese encephalitis virus serocomplex. The flavivirus genus is characterized by a positive-sense single-stranded RNA genome. The unique open reading frame of the viral RNA is transcribed and translated as a single polyprotein which is post-translationally cleaved to yield three structural and seven nonstructural proteins, one of which is the NS3 gene that encodes a C-terminal helicase domain consisting of 431 amino acids. Helicase inhibitors are potential antiviral drugs as the helicase is essential to viral replication. Crystals of the Kokobera virus helicase domain were obtained by the hanging-drop vapour-diffusion method. The crystals belong to space group P3 1 21 (or P3 2 21), with unit-cell parameters a = 88.6, c = 138.6 Å, and exhibit a diffraction limit of 2.3 Å

  5. Helicase-dependent amplification of nucleic acids.

    Cao, Yun; Kim, Hyun-Jin; Li, Ying; Kong, Huimin; Lemieux, Bertrand

    2013-10-11

    Helicase-dependent amplification (HDA) is a novel method for the isothermal in vitro amplification of nucleic acids. The HDA reaction selectively amplifies a target sequence by extension of two oligonucleotide primers. Unlike the polymerase chain reaction (PCR), HDA uses a helicase enzyme to separate the deoxyribonucleic acid (DNA) strands, rather than heat denaturation. This allows DNA amplification without the need for thermal cycling. The helicase used in HDA is a helicase super family II protein obtained from a thermophilic organism, Thermoanaerobacter tengcongensis (TteUvrD). This thermostable helicase is capable of unwinding blunt-end nucleic acid substrates at elevated temperatures (60° to 65°C). The HDA reaction can also be coupled with reverse transcription for ribonucleic acid (RNA) amplification. The products of this reaction can be detected during the reaction using fluorescent probes when incubations are conducted in a fluorimeter. Alternatively, products can be detected after amplification using a disposable amplicon containment device that contains an embedded lateral flow strip. Copyright © 2013 John Wiley & Sons, Inc.

  6. Purification and crystallization of Kokobera virus helicase

    De Colibus, Luigi; Speroni, Silvia [Department of Genetics and Microbiology, University of Pavia, Via Ferrata 1, 27100 Pavia (Italy); Coutard, Bruno [Architecture et Fonction des Macromolécules Biologiques, UMR 6098 CNRS et Université Aix-Marseille I et II, ESIL, Campus de Luminy, 13288 Marseille CEDEX 09 (France); Forrester, Naomi L.; Gould, Ernest [Centre for Ecology and Hydrology (formerly Institute of Virology), Mansfield Road, Oxford OX1 3SR (United Kingdom); Canard, Bruno [Architecture et Fonction des Macromolécules Biologiques, UMR 6098 CNRS et Université Aix-Marseille I et II, ESIL, Campus de Luminy, 13288 Marseille CEDEX 09 (France); Mattevi, Andrea, E-mail: mattevi@ipvgen.unipv.it [Department of Genetics and Microbiology, University of Pavia, Via Ferrata 1, 27100 Pavia (Italy)

    2007-03-01

    Kokobera virus is a mosquito-borne flavivirus belonging, like West Nile virus, to the Japanese encephalitis virus serocomplex. Crystals of the Kokobera virus helicase domain were obtained by the hanging-drop vapour-diffusion method and exhibit a diffraction limit of 2.3 Å. Kokobera virus is a mosquito-borne flavivirus belonging, like West Nile virus, to the Japanese encephalitis virus serocomplex. The flavivirus genus is characterized by a positive-sense single-stranded RNA genome. The unique open reading frame of the viral RNA is transcribed and translated as a single polyprotein which is post-translationally cleaved to yield three structural and seven nonstructural proteins, one of which is the NS3 gene that encodes a C-terminal helicase domain consisting of 431 amino acids. Helicase inhibitors are potential antiviral drugs as the helicase is essential to viral replication. Crystals of the Kokobera virus helicase domain were obtained by the hanging-drop vapour-diffusion method. The crystals belong to space group P3{sub 1}21 (or P3{sub 2}21), with unit-cell parameters a = 88.6, c = 138.6 Å, and exhibit a diffraction limit of 2.3 Å.

  7. RNA helicase A is not required for RISC activity.

    Liang, Xue-Hai; Crooke, Stanley T

    2013-10-01

    It has been shown that siRNAs can compete with each other or with endogenous miRNAs for RISC components. This competition may complicate the interpretations of phenotypes observed through siRNA-mediated knockdown of genes, especially those genes implicated in the RISC pathway. In this study, we re-examined the function of RNA helicase A (RHA), which has been previously proposed to function in RISC loading based on siRNA-mediated knockdown studies. Here we show that reduced RISC activity or loading of siRNAs was observed only in cells depleted of RHA using siRNA, but not using RNaseH-dependent antisense oligonucleotides (ASOs), suggesting that the impaired RISC function stems from the competition between pre-existing and newly transfected siRNAs, but not from reduction of the RHA protein. This view is further supported by the findings that cells depleted of a control protein, NCL1, using siRNA, but not ASO, exhibited similar defects on the loading and activity of a subsequently transfected siRNA. Transfection of RHA or NCL1 siRNAs, but not ASOs, reduced the levels of endogenous miRNAs, suggesting a competition mechanism. As a positive control, we showed that reduction of MOV10 by either siRNA or ASO decreased siRNA activity, confirming its role in RISC function. Together, our results indicate that RHA is not required for RISC activity or loading, and suggest that proper controls are required when using siRNAs to functionalize genes to avoid competition effects. © 2013. Published by Elsevier B.V. All rights reserved.

  8. Chl1 DNA helicase regulates Scc2 deposition specifically during DNA-replication in Saccharomyces cerevisiae.

    Soumya Rudra

    Full Text Available The conserved family of cohesin proteins that mediate sister chromatid cohesion requires Scc2, Scc4 for chromatin-association and Eco1/Ctf7 for conversion to a tethering competent state. A popular model, based on the notion that cohesins form huge ring-like structures, is that Scc2, Scc4 function is essential only during G1 such that sister chromatid cohesion results simply from DNA replisome passage through pre-loaded cohesin rings. In such a scenario, cohesin deposition during G1 is temporally uncoupled from Eco1-dependent establishment reactions that occur during S-phase. Chl1 DNA helicase (homolog of human ChlR1/DDX11 and BACH1/BRIP1/FANCJ helicases implicated in Fanconi anemia, breast and ovarian cancer and Warsaw Breakage Syndrome plays a critical role in sister chromatid cohesion, however, the mechanism through which Chl1 promotes cohesion remains poorly understood. Here, we report that Chl1 promotes Scc2 loading unto DNA such that both Scc2 and cohesin enrichment to chromatin are defective in chl1 mutant cells. The results further show that both Chl1 expression and chromatin-recruitment are tightly regulated through the cell cycle, peaking during S-phase. Importantly, kinetic ChIP studies reveals that Chl1 is required for Scc2 chromatin-association specifically during S-phase, but not during G1. Despite normal chromatin enrichment of both Scc2 and cohesin during G1, chl1 mutant cells exhibit severe chromosome segregation and cohesion defects--revealing that G1-loaded cohesins is insufficient to promote cohesion. Based on these findings, we propose a new model wherein S-phase cohesin loading occurs during DNA replication and in concert with both cohesion establishment and chromatin assembly reactions--challenging the notion that DNA replication fork navigates through or around pre-loaded cohesin rings.

  9. Comparative structural analysis of human DEAD-box RNA helicases.

    Patrick Schütz

    2010-09-01

    Full Text Available DEAD-box RNA helicases play various, often critical, roles in all processes where RNAs are involved. Members of this family of proteins are linked to human disease, including cancer and viral infections. DEAD-box proteins contain two conserved domains that both contribute to RNA and ATP binding. Despite recent advances the molecular details of how these enzymes convert chemical energy into RNA remodeling is unknown. We present crystal structures of the isolated DEAD-domains of human DDX2A/eIF4A1, DDX2B/eIF4A2, DDX5, DDX10/DBP4, DDX18/myc-regulated DEAD-box protein, DDX20, DDX47, DDX52/ROK1, and DDX53/CAGE, and of the helicase domains of DDX25 and DDX41. Together with prior knowledge this enables a family-wide comparative structural analysis. We propose a general mechanism for opening of the RNA binding site. This analysis also provides insights into the diversity of DExD/H- proteins, with implications for understanding the functions of individual family members.

  10. Comparative structural analysis of human DEAD-box RNA helicases.

    Schütz, Patrick; Karlberg, Tobias; van den Berg, Susanne; Collins, Ruairi; Lehtiö, Lari; Högbom, Martin; Holmberg-Schiavone, Lovisa; Tempel, Wolfram; Park, Hee-Won; Hammarström, Martin; Moche, Martin; Thorsell, Ann-Gerd; Schüler, Herwig

    2010-09-30

    DEAD-box RNA helicases play various, often critical, roles in all processes where RNAs are involved. Members of this family of proteins are linked to human disease, including cancer and viral infections. DEAD-box proteins contain two conserved domains that both contribute to RNA and ATP binding. Despite recent advances the molecular details of how these enzymes convert chemical energy into RNA remodeling is unknown. We present crystal structures of the isolated DEAD-domains of human DDX2A/eIF4A1, DDX2B/eIF4A2, DDX5, DDX10/DBP4, DDX18/myc-regulated DEAD-box protein, DDX20, DDX47, DDX52/ROK1, and DDX53/CAGE, and of the helicase domains of DDX25 and DDX41. Together with prior knowledge this enables a family-wide comparative structural analysis. We propose a general mechanism for opening of the RNA binding site. This analysis also provides insights into the diversity of DExD/H- proteins, with implications for understanding the functions of individual family members.

  11. MOV10 RNA helicase is a potent inhibitor of retrotransposition in cells.

    John L Goodier

    Full Text Available MOV10 protein, a putative RNA helicase and component of the RNA-induced silencing complex (RISC, inhibits retrovirus replication. We show that MOV10 also severely restricts human LINE1 (L1, Alu, and SVA retrotransposons. MOV10 associates with the L1 ribonucleoprotein particle, along with other RNA helicases including DDX5, DHX9, DDX17, DDX21, and DDX39A. However, unlike MOV10, these other helicases do not strongly inhibit retrotransposition, an activity dependent upon intact helicase domains. MOV10 association with retrotransposons is further supported by its colocalization with L1 ORF1 protein in stress granules, by cytoplasmic structures associated with RNA silencing, and by the ability of MOV10 to reduce endogenous and ectopic L1 expression. The majority of the human genome is repetitive DNA, most of which is the detritus of millions of years of accumulated retrotransposition. Retrotransposons remain active mutagens, and their insertion can disrupt gene function. Therefore, the host has evolved defense mechanisms to protect against retrotransposition, an arsenal we are only beginning to understand. With homologs in other vertebrates, insects, and plants, MOV10 may represent an ancient and innate form of immunity against both infective viruses and endogenous retroelements.

  12. DNA unwinding by ring-shaped T4 helicase gp41 is hindered by tension on the occluded strand.

    Ribeck, Noah; Saleh, Omar A

    2013-01-01

    The replicative helicase for bacteriophage T4 is gp41, which is a ring-shaped hexameric motor protein that achieves unwinding of dsDNA by translocating along one strand of ssDNA while forcing the opposite strand to the outside of the ring. While much study has been dedicated to the mechanism of binding and translocation along the ssDNA strand encircled by ring-shaped helicases, relatively little is known about the nature of the interaction with the opposite, 'occluded' strand. Here, we investigate the interplay between the bacteriophage T4 helicase gp41 and the ss/dsDNA fork by measuring, at the single-molecule level, DNA unwinding events on stretched DNA tethers in multiple geometries. We find that gp41 activity is significantly dependent on the geometry and tension of the occluded strand, suggesting an interaction between gp41 and the occluded strand that stimulates the helicase. However, the geometry dependence of gp41 activity is the opposite of that found previously for the E. coli hexameric helicase DnaB. Namely, tension applied between the occluded strand and dsDNA stem inhibits unwinding activity by gp41, while tension pulling apart the two ssDNA tails does not hinder its activity. This implies a distinct variation in helicase-occluded strand interactions among superfamily IV helicases, and we propose a speculative model for this interaction that is consistent with both the data presented here on gp41 and the data that had been previously reported for DnaB.

  13. [RTEL1 (regulator of telomere elongation helicase 1), a DNA helicase essential for genome stability].

    Le Guen, Tangui; Jullien, Laurent; Schertzer, Mike; Lefebvre, Axelle; Kermasson, Laetitia; de Villartay, Jean-Pierre; Londoño-Vallejo, Arturo; Revy, Patrick

    2013-12-01

    RTEL1 (regulator of telomere length helicase 1) is a DNA helicase that has been identified more than 10 years ago. Many works since, mainly in the nematode Caenorhabditis elegans and the mouse, have highlighted its role in chromosomal stability, maintenance of telomere length, and DNA repair. Recently, four laboratories have characterized RTEL1 mutations in patients with dyskeratosis congenita (DC) and Hoyeraal-Hreidarsson (HH) syndrome, a rare and severe variant of DC. We here summarize the current knowledge on RTEL1 and discuss the possible other functions that RTEL1 could play. © 2013 médecine/sciences – Inserm.

  14. Enzymatic activities and DNA substrate specificity of Mycobacterium tuberculosis DNA helicase XPB.

    Balasingham, Seetha V; Zegeye, Ephrem Debebe; Homberset, Håvard; Rossi, Marie L; Laerdahl, Jon K; Bohr, Vilhelm A; Tønjum, Tone

    2012-01-01

    XPB, also known as ERCC3 and RAD25, is a 3' → 5' DNA repair helicase belonging to the superfamily 2 of helicases. XPB is an essential core subunit of the eukaryotic basal transcription factor complex TFIIH. It has two well-established functions: in the context of damaged DNA, XPB facilitates nucleotide excision repair by unwinding double stranded DNA (dsDNA) surrounding a DNA lesion; while in the context of actively transcribing genes, XPB facilitates initiation of RNA polymerase II transcription at gene promoters. Human and other eukaryotic XPB homologs are relatively well characterized compared to conserved homologs found in mycobacteria and archaea. However, more insight into the function of bacterial helicases is central to understanding the mechanism of DNA metabolism and pathogenesis in general. Here, we characterized Mycobacterium tuberculosis XPB (Mtb XPB), a 3'→5' DNA helicase with DNA-dependent ATPase activity. Mtb XPB efficiently catalyzed DNA unwinding in the presence of significant excess of enzyme. The unwinding activity was fueled by ATP or dATP in the presence of Mg(2+)/Mn(2+). Consistent with the 3'→5' polarity of this bacterial XPB helicase, the enzyme required a DNA substrate with a 3' overhang of 15 nucleotides or more. Although Mtb XPB efficiently unwound DNA model substrates with a 3' DNA tail, it was not active on substrates containing a 3' RNA tail. We also found that Mtb XPB efficiently catalyzed ATP-independent annealing of complementary DNA strands. These observations significantly enhance our understanding of the biological roles of Mtb XPB.

  15. Enzymatic activities and DNA substrate specificity of Mycobacterium tuberculosis DNA helicase XPB.

    Seetha V Balasingham

    Full Text Available XPB, also known as ERCC3 and RAD25, is a 3' → 5' DNA repair helicase belonging to the superfamily 2 of helicases. XPB is an essential core subunit of the eukaryotic basal transcription factor complex TFIIH. It has two well-established functions: in the context of damaged DNA, XPB facilitates nucleotide excision repair by unwinding double stranded DNA (dsDNA surrounding a DNA lesion; while in the context of actively transcribing genes, XPB facilitates initiation of RNA polymerase II transcription at gene promoters. Human and other eukaryotic XPB homologs are relatively well characterized compared to conserved homologs found in mycobacteria and archaea. However, more insight into the function of bacterial helicases is central to understanding the mechanism of DNA metabolism and pathogenesis in general. Here, we characterized Mycobacterium tuberculosis XPB (Mtb XPB, a 3'→5' DNA helicase with DNA-dependent ATPase activity. Mtb XPB efficiently catalyzed DNA unwinding in the presence of significant excess of enzyme. The unwinding activity was fueled by ATP or dATP in the presence of Mg(2+/Mn(2+. Consistent with the 3'→5' polarity of this bacterial XPB helicase, the enzyme required a DNA substrate with a 3' overhang of 15 nucleotides or more. Although Mtb XPB efficiently unwound DNA model substrates with a 3' DNA tail, it was not active on substrates containing a 3' RNA tail. We also found that Mtb XPB efficiently catalyzed ATP-independent annealing of complementary DNA strands. These observations significantly enhance our understanding of the biological roles of Mtb XPB.

  16. Crystal structure of the FeS cluster-containing nucleotide excision repair helicase XPD.

    Stefanie C Wolski

    2008-06-01

    Full Text Available DNA damage recognition by the nucleotide excision repair pathway requires an initial step identifying helical distortions in the DNA and a proofreading step verifying the presence of a lesion. This proofreading step is accomplished in eukaryotes by the TFIIH complex. The critical damage recognition component of TFIIH is the XPD protein, a DNA helicase that unwinds DNA and identifies the damage. Here, we describe the crystal structure of an archaeal XPD protein with high sequence identity to the human XPD protein that reveals how the structural helicase framework is combined with additional elements for strand separation and DNA scanning. Two RecA-like helicase domains are complemented by a 4Fe4S cluster domain, which has been implicated in damage recognition, and an alpha-helical domain. The first helicase domain together with the helical and 4Fe4S-cluster-containing domains form a central hole with a diameter sufficient in size to allow passage of a single stranded DNA. Based on our results, we suggest a model of how DNA is bound to the XPD protein, and can rationalize several of the mutations in the human XPD gene that lead to one of three severe diseases, xeroderma pigmentosum, Cockayne syndrome, and trichothiodystrophy.

  17. A holistic evolutionary and structural study of flaviviridae provides insights into the function and inhibition of HCV helicase

    Dimitrios Vlachakis

    2013-05-01

    Full Text Available Viral RNA helicases are involved in duplex unwinding during the RNA replication of the virus. It is suggested that these helicases represent very promising antiviral targets. Viruses of the flaviviridae family are the causative agents of many common and devastating diseases, including hepatitis, yellow fever and dengue fever. As there is currently no available anti-Flaviviridae therapy, there is urgent need for the development of efficient anti-viral pharmaceutical strategies. Herein, we report the complete phylogenetic analysis across flaviviridae alongside a more in-depth evolutionary study that revealed a series of conserved and invariant amino acids that are predicted to be key to the function of the helicase. Structural molecular modelling analysis revealed the strategic significance of these residues based on their relative positioning on the 3D structures of the helicase enzymes, which may be used as pharmacological targets. We previously reported a novel series of highly potent HCV helicase inhibitors, and we now re-assess their antiviral potential using the 3D structural model of the invariant helicase residues. It was found that the most active compound of the series, compound C4, exhibited an IC50 in the submicromolar range, whereas its stereoisomer (compound C12 was completely inactive. Useful insights were obtained from molecular modelling and conformational search studies via molecular dynamics simulations. C12 tends to bend and lock in an almost “U” shape conformation, failing to establish vital interactions with the active site of HCV. On the contrary, C4 spends most of its conformational time in a straight, more rigid formation that allows it to successfully block the passage of the oligonucleotide in the ssRNA channel of the HCV helicase. This study paves the way and provides the necessary framework for the in-depth analysis required to enable the future design of new and potent anti-viral agents.

  18. Human Enterovirus Nonstructural Protein 2CATPase Functions as Both an RNA Helicase and ATP-Independent RNA Chaperone

    Xia, Hongjie; Wang, Peipei; Wang, Guang-Chuan; Yang, Jie; Sun, Xianlin; Wu, Wenzhe; Qiu, Yang; Shu, Ting; Zhao, Xiaolu; Yin, Lei; Qin, Cheng-Feng; Hu, Yuanyang; Zhou, Xi

    2015-01-01

    RNA helicases and chaperones are the two major classes of RNA remodeling proteins, which function to remodel RNA structures and/or RNA-protein interactions, and are required for all aspects of RNA metabolism. Although some virus-encoded RNA helicases/chaperones have been predicted or identified, their RNA remodeling activities in vitro and functions in the viral life cycle remain largely elusive. Enteroviruses are a large group of positive-stranded RNA viruses in the Picornaviridae family, which includes numerous important human pathogens. Herein, we report that the nonstructural protein 2CATPase of enterovirus 71 (EV71), which is the major causative pathogen of hand-foot-and-mouth disease and has been regarded as the most important neurotropic enterovirus after poliovirus eradication, functions not only as an RNA helicase that 3′-to-5′ unwinds RNA helices in an adenosine triphosphate (ATP)-dependent manner, but also as an RNA chaperone that destabilizes helices bidirectionally and facilitates strand annealing and complex RNA structure formation independently of ATP. We also determined that the helicase activity is based on the EV71 2CATPase middle domain, whereas the C-terminus is indispensable for its RNA chaperoning activity. By promoting RNA template recycling, 2CATPase facilitated EV71 RNA synthesis in vitro; when 2CATPase helicase activity was impaired, EV71 RNA replication and virion production were mostly abolished in cells, indicating that 2CATPase-mediated RNA remodeling plays a critical role in the enteroviral life cycle. Furthermore, the RNA helicase and chaperoning activities of 2CATPase are also conserved in coxsackie A virus 16 (CAV16), another important enterovirus. Altogether, our findings are the first to demonstrate the RNA helicase and chaperoning activities associated with enterovirus 2CATPase, and our study provides both in vitro and cellular evidence for their potential roles during viral RNA replication. These findings increase our

  19. Crystal structure of Middle East respiratory syndrome coronavirus helicase.

    Wei Hao

    2017-06-01

    Full Text Available Middle East respiratory syndrome coronavirus (MERS-CoV remains a threat to public health worldwide; however, effective vaccine or drug against CoVs remains unavailable. CoV helicase is one of the three evolutionary most conserved proteins in nidoviruses, thus making it an important target for drug development. We report here the first structure of full-length coronavirus helicase, MERS-CoV nsp13. MERS-CoV helicase has multiple domains, including an N-terminal Cys/His rich domain (CH with three zinc atoms, a beta-barrel domain and a C-terminal SF1 helicase core with two RecA-like subdomains. Our structural analyses show that while the domain organization of nsp13 is conserved throughout nidoviruses, the individual domains of nsp13 are closely related to the equivalent eukaryotic domains of Upf1 helicases. The most distinctive feature differentiating CoV helicases from eukaryotic Upf1 helicases is the interaction between CH domain and helicase core.

  20. Nucleolin inhibits G4 oligonucleotide unwinding by Werner helicase.

    Fred E Indig

    Full Text Available The Werner protein (WRNp, a member of the RecQ helicase family, is strongly associated with the nucleolus, as is nucleolin (NCL, an important nucleolar constituent protein. Both WRNp and NCL respond to the effects of DNA damaging agents. Therefore, we have investigated if these nuclear proteins interact and if this interaction has a possible functional significance in DNA damage repair.Here we report that WRNp interacts with the RNA-binding protein, NCL, based on immunoprecipitation, immunofluorescent co-localization in live and fixed cells, and direct binding of purified WRNp to nucleolin. We also map the binding region to the C-terminal domains of both proteins. Furthermore, treatment of U2OS cells with 15 µM of the Topoisomerase I inhibitor, camptothecin, causes the dissociation of the nucleolin-Werner complex in the nucleolus, followed by partial re-association in the nucleoplasm. Other DNA damaging agents, such as hydroxyurea, Mitomycin C, and aphidicolin do not have these effects. Nucleolin or its C-terminal fragment affected the helicase, but not the exonuclease activity of WRNp, by inhibiting WRN unwinding of G4 tetraplex DNA structures, as seen in activity assays and electrophoretic mobility shift assays (EMSA.These data suggest that nucleolin may regulate G4 DNA unwinding by WRNp, possibly in response to certain DNA damaging agents. We postulate that the NCL-WRNp complex may contain an inactive form of WRNp, which is released from the nucleolus upon DNA damage. Then, when required, WRNp is released from inhibition and can participate in the DNA repair processes.

  1. Structural basis for the function of DEAH helicases

    He, Yangzi; Andersen, Gregers Rom; Nielsen, Klaus Hvid

    2010-01-01

    DEAH helicases participate in pre‐messenger RNA splicing and ribosome biogenesis. The structure of yeast Prp43p‐ADP reveals the homology of DEAH helicases to DNA helicases and the presence of an oligonucleotide‐binding motif. A β‐hairpin from the second RecA domain is wedged between two carboxy......‐terminal domains and blocks access to the occluded RNA binding site formed by the RecA domains and a C‐terminal domain. ATP binding and hydrolysis are likely to induce conformational changes in the hairpin that are important for RNA unwinding or ribonucleoprotein remodelling. The structure of Prp43p provides...

  2. Mutational analysis of an archaeal minichromosome maintenance protein exterior hairpin reveals critical residues for helicase activity and DNA binding

    Brewster Aaron S

    2010-08-01

    Full Text Available Abstract Background The mini-chromosome maintenance protein (MCM complex is an essential replicative helicase for DNA replication in Archaea and Eukaryotes. While the eukaryotic complex consists of six homologous proteins (MCM2-7, the archaeon Sulfolobus solfataricus has only one MCM protein (ssoMCM, six subunits of which form a homohexamer. We have recently reported a 4.35Å crystal structure of the near full-length ssoMCM. The structure reveals a total of four β-hairpins per subunit, three of which are located within the main channel or side channels of the ssoMCM hexamer model generated based on the symmetry of the N-terminal Methanothermobacter thermautotrophicus (mtMCM structure. The fourth β-hairpin, however, is located on the exterior of the hexamer, near the exit of the putative side channels and next to the ATP binding pocket. Results In order to better understand this hairpin's role in DNA binding and helicase activity, we performed a detailed mutational and biochemical analysis of nine residues on this exterior β-hairpin (EXT-hp. We examined the activities of the mutants related to their helicase function, including hexamerization, ATPase, DNA binding and helicase activities. The assays showed that some of the residues on this EXT-hp play a role for DNA binding as well as for helicase activity. Conclusions These results implicate several current theories regarding helicase activity by this critical hexameric enzyme. As the data suggest that EXT-hp is involved in DNA binding, the results reported here imply that the EXT-hp located near the exterior exit of the side channels may play a role in contacting DNA substrate in a manner that affects DNA unwinding.

  3. Acute inactivation of the replicative helicase in human cells triggers MCM8-9-dependent DNA synthesis

    Natsume, Toyoaki; Nishimura, Kohei; Minocherhomji, Sheroy

    2017-01-01

    stemming from replisome dissociation during DNA replication perturbation, we used a degron-based system for inducible proteolysis of a subunit of the replicative helicase. We show that MCM2-depleted cells activate a DNA damage response pathway and generate replication-associated DNA double-strand breaks...

  4. Novel benzoxazole inhibitor of dengue virus replication that targets the NS3 helicase.

    Byrd, Chelsea M; Grosenbach, Douglas W; Berhanu, Aklile; Dai, Dongcheng; Jones, Kevin F; Cardwell, Kara B; Schneider, Christine; Yang, Guang; Tyavanagimatt, Shanthakumar; Harver, Chris; Wineinger, Kristin A; Page, Jessica; Stavale, Eric; Stone, Melialani A; Fuller, Kathleen P; Lovejoy, Candace; Leeds, Janet M; Hruby, Dennis E; Jordan, Robert

    2013-04-01

    Dengue virus (DENV) is the predominant mosquito-borne viral pathogen that infects humans with an estimated 50 to 100 million infections per year worldwide. Over the past 50 years, the incidence of dengue disease has increased dramatically and the virus is now endemic in more than 100 countries. Moreover, multiple serotypes of DENV are now found in the same geographic region, increasing the likelihood of more severe forms of disease. Despite extensive research, there are still no approved vaccines or therapeutics commercially available to treat DENV infection. Here we report the results of a high-throughput screen of a chemical compound library using a whole-virus assay that identified a novel small-molecule inhibitor of DENV, ST-610, that potently and selectively inhibits all four serotypes of DENV replication in vitro. Sequence analysis of drug-resistant virus isolates has identified a single point mutation, A263T, in the NS3 helicase domain that confers resistance to this compound. ST-610 inhibits DENV NS3 helicase RNA unwinding activity in a molecular-beacon-based helicase assay but does not inhibit nucleoside triphosphatase activity based on a malachite green ATPase assay. ST-610 is nonmutagenic, is well tolerated (nontoxic) in mice, and has shown efficacy in a sublethal murine model of DENV infection with the ability to significantly reduce viremia and viral load compared to vehicle controls.

  5. Evolution of the DEAD box helicase family in chicken: chickens have no DHX9 ortholog.

    Sato, Haruko; Oshiumi, Hiroyuki; Takaki, Hiromi; Hikono, Hirokazu; Seya, Tsukasa

    2015-10-01

    Viral RNA represents a pattern molecule that can be recognized by RNA sensors in innate immunity. Humans and mice possess cytoplasmic DNA/RNA sensors for detecting viral replication. There are a number of DEAD (Asp-Glu-Ala-Asp; DExD/H) box-type helicases in mammals, among which retinoic acid-inducible gene 1 (RIG-I) and melanoma differentiation-associated protein 5 (MDA50) are indispensable for RNA sensing; however, they are functionally supported by a number of sensors that directly bind viral RNA or replicative RNA intermediates to convey signals to RIG-I and MDA5. Some DEAD box helicase members recognize DNA irrespective of the origin. These sensors transmit IFN-inducing signals through adaptors, including mitochondrial antiviral signaling. Viral double-stranded RNAs are reportedly sensed by the helicases DDX1, DDX21, DHX36, DHX9, DDX3, DDX41, LGP2 and DDX60, in addition to RIG-I and MDA5, and induce type I IFNs, thereby blocking viral replication. Humans and mice have all nucleic acid sensors listed here. In the RNA sensing system in chicken, it was found in the present study that most DEAD box helicases are conserved; however, DHX9 is genetically deficient in addition to reported RIG-I. Based on the current genome databases, similar DHX9 deficiency was observed in ducks and several other bird species. Because chicken, but not duck, was found to be deficient in RIG-I, the RNA-sensing system of chicken lacks RIG-I and DHX9 and is thus more fragile than that of duck or mammal. DHX9 may generally compensate for the function of RIG-I and deficiency of DHX9 possibly participates in exacerbations of viral infection such as influenza in chickens. © 2015 The Societies and Wiley Publishing Asia Pty Ltd.

  6. Role of the hydrophilic channels of simian virus 40 T-antigen helicase in DNA replication.

    Wang, Weiping; Manna, David; Simmons, Daniel T

    2007-05-01

    The simian virus 40 (SV40) hexameric helicase consists of a central channel and six hydrophilic channels located between adjacent large tier domains within each hexamer. To study the function of the hydrophilic channels in SV40 DNA replication, a series of single-point substitutions were introduced at sites not directly involved in protein-protein contacts. The mutants were characterized biochemically in various ways. All mutants oligomerized normally in the absence of DNA. Interestingly, 8 of the 10 mutants failed to unwind an origin-containing DNA fragment and nine of them were totally unable to support SV40 DNA replication in vitro. The mutants fell into four classes based on their biochemical properties. Class A mutants bound DNA normally and had normal ATPase and helicase activities but failed to unwind origin DNA and support SV40 DNA replication. Class B mutants were compromised in single-stranded DNA and origin DNA binding at low protein concentrations. They were defective in helicase activity and unwinding of the origin and in supporting DNA replication. Class C and D mutants possessed higher-than-normal single-stranded DNA binding activity at low protein concentrations. The class C mutants failed to separate origin DNA and support DNA replication. The class D mutants unwound origin DNA normally but were compromised in their ability to support DNA replication. Taken together, these results suggest that the hydrophilic channels have an active role in the unwinding of SV40 DNA from the origin and the placement of the resulting single strands within the helicase.

  7. Dissection of the functional domains of an archaeal holliday junction helicase

    Hong, Ye; Chu, Mingzhu; Li, Yansheng

    2012-01-01

    Helicases and nucleases form complexes that play very important roles in DNA repair pathways some of which interact with each other at Holliday junctions. In this study, we present in vitro and in vivo analysis of Hjm and its interaction with Hjc in Sulfolobus. In vitro studies employed Hjm from...... conformation change of the enzyme. Furthermore, StoHjm is able to prevent the formation of Hjc/HJ high complex, suggesting a regulation mechanism of Hjm to the activity of Hjc. We show that Hjm is essential for cell viability using recently developed genetic system and mutant propagation assay, suggesting...

  8. Viral hijacking of a replicative helicase loader and its implications for helicase loading control and phage replication

    Hood, Iris V.; Berger, James M.

    2016-05-31

    Replisome assembly requires the loading of replicative hexameric helicases onto origins by AAA+ ATPases. How loader activity is appropriately controlled remains unclear. Here, we use structural and biochemical analyses to establish how an antimicrobial phage protein interferes with the function of theStaphylococcus aureusreplicative helicase loader, DnaI. The viral protein binds to the loader’s AAA+ ATPase domain, allowing binding of the host replicative helicase but impeding loader self-assembly and ATPase activity. Close inspection of the complex highlights an unexpected locus for the binding of an interdomain linker element in DnaI/DnaC-family proteins. We find that the inhibitor protein is genetically coupled to a phage-encoded homolog of the bacterial helicase loader, which we show binds to the host helicase but not to the inhibitor itself. These findings establish a new approach by which viruses can hijack host replication processes and explain how loader activity is internally regulated to prevent aberrant auto-association.

  9. Norm-based mechanism design

    Bulling, Nils; Dastani, Mehdi

    2016-01-01

    The increasing presence of autonomous (software) systems in open environments in general, and the complex interactions taking place among them in particular, require flexible control and coordination mechanisms to guarantee desirable overall system level properties without limiting the autonomy of

  10. MCM Paradox: Abundance of Eukaryotic Replicative Helicases and Genomic Integrity.

    Das, Mitali; Singh, Sunita; Pradhan, Satyajit; Narayan, Gopeshwar

    2014-01-01

    As a crucial component of DNA replication licensing system, minichromosome maintenance (MCM) 2-7 complex acts as the eukaryotic DNA replicative helicase. The six related MCM proteins form a heterohexamer and bind with ORC, CDC6, and Cdt1 to form the prereplication complex. Although the MCMs are well known as replicative helicases, their overabundance and distribution patterns on chromatin present a paradox called the "MCM paradox." Several approaches had been taken to solve the MCM paradox and describe the purpose of excess MCMs distributed beyond the replication origins. Alternative functions of these MCMs rather than a helicase had also been proposed. This review focuses on several models and concepts generated to solve the MCM paradox coinciding with their helicase function and provides insight into the concept that excess MCMs are meant for licensing dormant origins as a backup during replication stress. Finally, we extend our view towards the effect of alteration of MCM level. Though an excess MCM constituent is needed for normal cells to withstand stress, there must be a delineation of the threshold level in normal and malignant cells. This review also outlooks the future prospects to better understand the MCM biology.

  11. New roles of the human Suv3 helicase in genome maintenance

    Venø, Susanne Trillingsgaard

    During her PhD studies, Susanne Trillingsgaard Venø carried out research into the role of the human Suv3 protein in stabilising the human genome – DNA. Suv3 is a helicase that separates the two strands of the DNA’s double helix. Throughout our lives, the DNA in our cells is constantly exposed...... maintenance. Based on these new research results, the Suv3 protein could be a valuable model for genome stability as an important factor in our understanding of why we get old....

  12. Three-dimensional structure of N-terminal domain of DnaB helicase and helicase-primase interactions in Helicobacter pylori.

    Tara Kashav

    2009-10-01

    Full Text Available Replication initiation is a crucial step in genome duplication and homohexameric DnaB helicase plays a central role in the replication initiation process by unwinding the duplex DNA and interacting with several other proteins during the process of replication. N-terminal domain of DnaB is critical for helicase activity and for DnaG primase interactions. We present here the crystal structure of the N-terminal domain (NTD of H. pylori DnaB (HpDnaB helicase at 2.2 A resolution and compare the structural differences among helicases and correlate with the functional differences. The structural details of NTD suggest that the linker region between NTD and C-terminal helicase domain plays a vital role in accurate assembly of NTD dimers. The sequence analysis of the linker regions from several helicases reveals that they should form four helix bundles. We also report the characterization of H. pylori DnaG primase and study the helicase-primase interactions, where HpDnaG primase stimulates DNA unwinding activity of HpDnaB suggesting presence of helicase-primase cohort at the replication fork. The protein-protein interaction study of C-terminal domain of primase and different deletion constructs of helicase suggests that linker is essential for proper conformation of NTD to interact strongly with HpDnaG. The surface charge distribution on the primase binding surface of NTDs of various helicases suggests that DnaB-DnaG interaction and stability of the complex is most probably charge dependent. Structure of the linker and helicase-primase interactions indicate that HpDnaB differs greatly from E.coli DnaB despite both belong to gram negative bacteria.

  13. The nuclear import of RNA helicase A is mediated by importin-α3

    Aratani, Satoko; Oishi, Takayuki; Fujita, Hidetoshi; Nakazawa, Minako; Fujii, Ryouji; Imamoto, Naoko; Yoneda, Yoshihiro; Fukamizu, Akiyoshi; Nakajima, Toshihiro

    2006-01-01

    RNA helicase A (RHA), an ATPase/helicase, regulates the gene expression at various steps including transcriptional activation and RNA processing. RHA is known to shuttle between the nucleus and cytoplasm. We identified the nuclear localization signal (NLS) of RHA and analyzed the nuclear import mechanisms. The NLS of RHA (RHA-NLS) consisting of 19 amino acid residues is highly conserved through species and does not have the consensus classical NLS. In vitro nuclear import assays revealed that the nuclear import of RHA was Ran-dependent and mediated with the classical importin-α/β-dependent pathway. The binding assay indicated that the basic residues in RHA-NLS were used for interaction with importin-α. Furthermore, the nuclear import of RHA-NLS was supported by importin-α1 and preferentially importin-α3. Our results indicate that the nuclear import of RHA is mediated by the importin-α3/importin-β-dependent pathway and suggest that the specificity for importin may regulate the functions of cargo proteins

  14. The adnAB Locus, Encoding a Putative Helicase-Nuclease Activity, Is Essential in Streptomyces

    Zhang, Lingli; Nguyen, Hoang Chuong; Chipot, Ludovic; Piotrowski, Emilie; Bertrand, Claire

    2014-01-01

    Homologous recombination is a crucial mechanism that repairs a wide range of DNA lesions, including the most deleterious ones, double-strand breaks (DSBs). This multistep process is initiated by the resection of the broken DNA ends by a multisubunit helicase-nuclease complex exemplified by Escherichia coli RecBCD, Bacillus subtilis AddAB, and newly discovered Mycobacterium tuberculosis AdnAB. Here we show that in Streptomyces, neither recBCD nor addAB homologues could be detected. The only putative helicase-nuclease-encoding genes identified were homologous to M. tuberculosis adnAB genes. These genes are conserved as a single copy in all sequenced genomes of Streptomyces. The disruption of adnAB in Streptomyces ambofaciens and Streptomyces coelicolor could not be achieved unless an ectopic copy was provided, indicating that adnAB is essential for growth. Both adnA and adnB genes were shown to be inducible in response to DNA damage (mitomycin C) and to be independently transcribed. Introduction of S. ambofaciens adnAB genes in an E. coli recB mutant restored viability and resistance to UV light, suggesting that Streptomyces AdnAB could be a functional homologue of RecBCD and be involved in DNA damage resistance. PMID:24837284

  15. Mcm10 regulates DNA replication elongation by stimulating the CMG replicative helicase.

    Lõoke, Marko; Maloney, Michael F; Bell, Stephen P

    2017-02-01

    Activation of the Mcm2-7 replicative DNA helicase is the committed step in eukaryotic DNA replication initiation. Although Mcm2-7 activation requires binding of the helicase-activating proteins Cdc45 and GINS (forming the CMG complex), an additional protein, Mcm10, drives initial origin DNA unwinding by an unknown mechanism. We show that Mcm10 binds a conserved motif located between the oligonucleotide/oligosaccharide fold (OB-fold) and A subdomain of Mcm2. Although buried in the interface between these domains in Mcm2-7 structures, mutations predicted to separate the domains and expose this motif restore growth to conditional-lethal MCM10 mutant cells. We found that, in addition to stimulating initial DNA unwinding, Mcm10 stabilizes Cdc45 and GINS association with Mcm2-7 and stimulates replication elongation in vivo and in vitro. Furthermore, we identified a lethal allele of MCM10 that stimulates initial DNA unwinding but is defective in replication elongation and CMG binding. Our findings expand the roles of Mcm10 during DNA replication and suggest a new model for Mcm10 function as an activator of the CMG complex throughout DNA replication. © 2017 Lõoke et al.; Published by Cold Spring Harbor Laboratory Press.

  16. G-quadruplexes Significantly Stimulate Pif1 Helicase-catalyzed Duplex DNA Unwinding*

    Duan, Xiao-Lei; Liu, Na-Nv; Yang, Yan-Tao; Li, Hai-Hong; Li, Ming; Dou, Shuo-Xing; Xi, Xu-Guang

    2015-01-01

    The evolutionarily conserved G-quadruplexes (G4s) are faithfully inherited and serve a variety of cellular functions such as telomere maintenance, gene regulation, DNA replication initiation, and epigenetic regulation. Different from the Watson-Crick base-pairing found in duplex DNA, G4s are formed via Hoogsteen base pairing and are very stable and compact DNA structures. Failure of untangling them in the cell impedes DNA-based transactions and leads to genome instability. Cells have evolved highly specific helicases to resolve G4 structures. We used a recombinant nuclear form of Saccharomyces cerevisiae Pif1 to characterize Pif1-mediated DNA unwinding with a substrate mimicking an ongoing lagging strand synthesis stalled by G4s, which resembles a replication origin and a G4-structured flap in Okazaki fragment maturation. We find that the presence of G4 may greatly stimulate the Pif1 helicase to unwind duplex DNA. Further studies reveal that this stimulation results from G4-enhanced Pif1 dimerization, which is required for duplex DNA unwinding. This finding provides new insights into the properties and functions of G4s. We discuss the observed activation phenomenon in relation to the possible regulatory role of G4s in the rapid rescue of the stalled lagging strand synthesis by helping the replicator recognize and activate the replication origin as well as by quickly removing the G4-structured flap during Okazaki fragment maturation. PMID:25627683

  17. Saccharomyces cerevisiae Hrq1 requires a long 3′-tailed DNA substrate for helicase activity

    Kwon, Sung-Hun; Choi, Do-Hee; Lee, Rina; Bae, Sung-Ho

    2012-01-01

    Highlights: ► Hrq1 has intrinsic 3′–5′ helicase and DNA strand annealing activities. ► Hrq1 requires a long 3′-tail for efficient DNA unwinding. ► Helicase activity of Hrq1 is stimulated by a fork structure. ► Hrq1 is a moderately processive helicase. -- Abstract: RecQ helicases are well conserved proteins from bacteria to human and function in various DNA metabolism for maintenance of genome stability. Five RecQ helicases are found in humans, whereas only one RecQ helicase has been described in lower eukaryotes. However, recent studies predicted the presence of a second RecQ helicase, Hrq1, in fungal genomes and verified it as a functional gene in fission yeast. Here we show that 3′–5′ helicase activity is intrinsically associated with Hrq1 of Saccharomyces cerevisiae. We also determined several biochemical properties of Hrq1 helicase distinguishable from those of other RecQ helicase members. Hrq1 is able to unwind relatively long duplex DNA up to 120-bp and is significantly stimulated by a preexisting fork structure. Further, the most striking feature of Hrq1 is its absolute requirement for a long 3′-tail (⩾70-nt) for efficient unwinding of duplex DNA. We also found that Hrq1 has potent DNA strand annealing activity. Our results indicate that Hrq1 has vigorous helicase activity that deserves further characterization to expand our understanding of RecQ helicases.

  18. The MCM Helicase Motor of the Eukaryotic Replisome.

    Abid Ali, Ferdos; Costa, Alessandro

    2016-05-08

    The MCM motor of the CMG helicase powers ahead of the eukaryotic replication machinery to unwind DNA, in a process that requires ATP hydrolysis. The reconstitution of DNA replication in vitro has established the succession of events that lead to replication origin activation by the MCM and recent studies have started to elucidate the structural basis of duplex DNA unwinding. Despite the exciting progress, how the MCM translocates on DNA remains a matter of debate. Copyright © 2016. Published by Elsevier Ltd.

  19. Saccharomyces cerevisiae Hrq1 requires a long 3'-tailed DNA substrate for helicase activity.

    Kwon, Sung-Hun; Choi, Do-Hee; Lee, Rina; Bae, Sung-Ho

    2012-10-26

    RecQ helicases are well conserved proteins from bacteria to human and function in various DNA metabolism for maintenance of genome stability. Five RecQ helicases are found in humans, whereas only one RecQ helicase has been described in lower eukaryotes. However, recent studies predicted the presence of a second RecQ helicase, Hrq1, in fungal genomes and verified it as a functional gene in fission yeast. Here we show that 3'-5' helicase activity is intrinsically associated with Hrq1 of Saccharomyces cerevisiae. We also determined several biochemical properties of Hrq1 helicase distinguishable from those of other RecQ helicase members. Hrq1 is able to unwind relatively long duplex DNA up to 120-bp and is significantly stimulated by a preexisting fork structure. Further, the most striking feature of Hrq1 is its absolute requirement for a long 3'-tail (⩾70-nt) for efficient unwinding of duplex DNA. We also found that Hrq1 has potent DNA strand annealing activity. Our results indicate that Hrq1 has vigorous helicase activity that deserves further characterization to expand our understanding of RecQ helicases. Copyright © 2012 Elsevier Inc. All rights reserved.

  20. Bloom syndrome helicase in meiosis: Pro-crossover functions of an anti-crossover protein.

    Hatkevich, Talia; Sekelsky, Jeff

    2017-09-01

    The functions of the Bloom syndrome helicase (BLM) and its orthologs are well characterized in mitotic DNA damage repair, but their roles within the context of meiotic recombination are less clear. In meiotic recombination, multiple repair pathways are used to repair meiotic DSBs, and current studies suggest that BLM may regulate the use of these pathways. Based on literature from Saccharomyces cerevisiae, Arabidopsis thaliana, Mus musculus, Drosophila melanogaster, and Caenorhabditis elegans, we present a unified model for a critical meiotic role of BLM and its orthologs. In this model, BLM and its orthologs utilize helicase activity to regulate the use of various pathways in meiotic recombination by continuously disassembling recombination intermediates. This unwinding activity provides the meiotic program with a steady pool of early recombination substrates, increasing the probability for a DSB to be processed by the appropriate pathway. As a result of BLM activity, crossovers are properly placed throughout the genome, promoting proper chromosomal disjunction at the end of meiosis. This unified model can be used to further refine the complex role of BLM and its orthologs in meiotic recombination. © 2017 WILEY Periodicals, Inc.

  1. Real-time electrochemical monitoring of isothermal helicase-dependent amplification of nucleic acids.

    Kivlehan, Francine; Mavré, François; Talini, Luc; Limoges, Benoît; Marchal, Damien

    2011-09-21

    We described an electrochemical method to monitor in real-time the isothermal helicase-dependent amplification of nucleic acids. The principle of detection is simple and well-adapted to the development of portable, easy-to-use and inexpensive nucleic acids detection technologies. It consists of monitoring a decrease in the electrochemical current response of a reporter DNA intercalating redox probe during the isothermal DNA amplification. The method offers the possibility to quantitatively analyze target nucleic acids in less than one hour at a single constant temperature, and to perform at the end of the isothermal amplification a DNA melt curve analysis for differentiating between specific and non-specific amplifications. To illustrate the potentialities of this approach for the development of a simple, robust and low-cost instrument with high throughput capability, the method was validated with an electrochemical system capable of monitoring up to 48 real-time isothermal HDA reactions simultaneously in a disposable microplate consisting of 48-electrochemical microwells. Results obtained with this approach are comparable to that obtained with a well-established but more sophisticated and expensive fluorescence-based method. This makes for a promising alternative detection method not only for real-time isothermal helicase-dependent amplification of nucleic acid, but also for other isothermal DNA amplification strategies.

  2. In TFIIH, XPD helicase is exclusively devoted to DNA repair.

    Jochen Kuper

    2014-09-01

    Full Text Available The eukaryotic XPD helicase is an essential subunit of TFIIH involved in both transcription and nucleotide excision repair (NER. Mutations in human XPD are associated with several inherited diseases such as xeroderma pigmentosum, Cockayne syndrome, and trichothiodystrophy. We performed a comparative analysis of XPD from Homo sapiens and Chaetomium thermophilum (a closely related thermostable fungal orthologue to decipher the different molecular prerequisites necessary for either transcription or DNA repair. In vitro and in vivo assays demonstrate that mutations in the 4Fe4S cluster domain of XPD abrogate the NER function of TFIIH and do not affect its transcriptional activity. We show that the p44-dependent activation of XPD is promoted by the stimulation of its ATPase activity. Furthermore, we clearly demonstrate that XPD requires DNA binding, ATPase, and helicase activity to function in NER. In contrast, these enzymatic properties are dispensable for transcription initiation. XPD helicase is thus exclusively devoted to NER and merely acts as a structural scaffold to maintain TFIIH integrity during transcription.

  3. Genome-wide identification of SF1 and SF2 helicases from archaea.

    Chamieh, Hala; Ibrahim, Hiba; Kozah, Juliana

    2016-01-15

    Archaea microorganisms have long been used as model organisms for the study of protein molecular machines. Archaeal proteins are particularly appealing to study since archaea, even though prokaryotic, possess eukaryotic-like cellular processes. Super Family I (SF1) and Super Family II (SF2) helicase families have been studied in many model organisms, little is known about their presence and distribution in archaea. We performed an exhaustive search of homologs of SF1 and SF2 helicase proteins in 95 complete archaeal genomes. In the present study, we identified the complete sets of SF1 and SF2 helicases in archaea. Comparative analysis between archaea, human and the bacteria E. coli SF1 and SF2 helicases, resulted in the identification of seven helicase families conserved among representatives of the domains of life. This analysis suggests that these helicase families are highly conserved throughout evolution. We highlight the conserved motifs of each family and characteristic domains of the detected families. Distribution of SF1/SF2 families show that Ski2-like, Lhr, Sfth and Rad3-like helicases are ubiquitous among archaeal genomes while the other families are specific to certain archaeal groups. We also report the presence of a novel SF2 helicase specific to archaea domain named Archaea Specific Helicase (ASH). Phylogenetic analysis indicated that ASH has evolved in Euryarchaeota and is evolutionary related to the Ski2-like family with specific characteristic domains. Our study provides the first exhaustive analysis of SF1 and SF2 helicases from archaea. It expands the variety of SF1 and SF2 archaeal helicases known to exist to date and provides a starting point for new biochemical and genetic studies needed to validate their biological functions. Copyright © 2015 Elsevier B.V. All rights reserved.

  4. Requirement for the E1 Helicase C-Terminal Domain in Papillomavirus DNA Replication In Vivo.

    Bergvall, Monika; Gagnon, David; Titolo, Steve; Lehoux, Michaël; D'Abramo, Claudia M; Melendy, Thomas; Archambault, Jacques

    2016-01-06

    The papillomavirus (PV) E1 helicase contains a conserved C-terminal domain (CTD), located next to its ATP-binding site, whose function in vivo is still poorly understood. The CTD is comprised of an alpha helix followed by an acidic region (AR) and a C-terminal extension termed the C-tail. Recent biochemical studies on bovine papillomavirus 1 (BPV1) E1 showed that the AR and C-tail regulate the oligomerization of the protein into a double hexamer at the origin. In this study, we assessed the importance of the CTD of human papillomavirus 11 (HPV11) E1 in vivo, using a cell-based DNA replication assay. Our results indicate that combined deletion of the AR and C-tail drastically reduces DNA replication, by 85%, and that further truncation into the alpha-helical region compromises the structural integrity of the E1 helicase domain and its interaction with E2. Surprisingly, removal of the C-tail alone or mutation of highly conserved residues within the domain still allows significant levels of DNA replication (55%). This is in contrast to the absolute requirement for the C-tail reported for BPV1 E1 in vitro and confirmed here in vivo. Characterization of chimeric proteins in which the AR and C-tail from HPV11 E1 were replaced by those of BPV1 indicated that while the function of the AR is transferable, that of the C-tail is not. Collectively, these findings define the contribution of the three CTD subdomains to the DNA replication activity of E1 in vivo and suggest that the function of the C-tail has evolved in a PV type-specific manner. While much is known about hexameric DNA helicases from superfamily 3, the papillomavirus E1 helicase contains a unique C-terminal domain (CTD) adjacent to its ATP-binding site. We show here that this CTD is important for the DNA replication activity of HPV11 E1 in vivo and that it can be divided into three functional subdomains that roughly correspond to the three conserved regions of the CTD: an alpha helix, needed for the structural

  5. Requirement for the E1 Helicase C-Terminal Domain in Papillomavirus DNA Replication In Vivo

    Bergvall, Monika; Gagnon, David; Titolo, Steve; Lehoux, Michaël; D'Abramo, Claudia M.

    2016-01-01

    ABSTRACT The papillomavirus (PV) E1 helicase contains a conserved C-terminal domain (CTD), located next to its ATP-binding site, whose function in vivo is still poorly understood. The CTD is comprised of an alpha helix followed by an acidic region (AR) and a C-terminal extension termed the C-tail. Recent biochemical studies on bovine papillomavirus 1 (BPV1) E1 showed that the AR and C-tail regulate the oligomerization of the protein into a double hexamer at the origin. In this study, we assessed the importance of the CTD of human papillomavirus 11 (HPV11) E1 in vivo, using a cell-based DNA replication assay. Our results indicate that combined deletion of the AR and C-tail drastically reduces DNA replication, by 85%, and that further truncation into the alpha-helical region compromises the structural integrity of the E1 helicase domain and its interaction with E2. Surprisingly, removal of the C-tail alone or mutation of highly conserved residues within the domain still allows significant levels of DNA replication (55%). This is in contrast to the absolute requirement for the C-tail reported for BPV1 E1 in vitro and confirmed here in vivo. Characterization of chimeric proteins in which the AR and C-tail from HPV11 E1 were replaced by those of BPV1 indicated that while the function of the AR is transferable, that of the C-tail is not. Collectively, these findings define the contribution of the three CTD subdomains to the DNA replication activity of E1 in vivo and suggest that the function of the C-tail has evolved in a PV type-specific manner. IMPORTANCE While much is known about hexameric DNA helicases from superfamily 3, the papillomavirus E1 helicase contains a unique C-terminal domain (CTD) adjacent to its ATP-binding site. We show here that this CTD is important for the DNA replication activity of HPV11 E1 in vivo and that it can be divided into three functional subdomains that roughly correspond to the three conserved regions of the CTD: an alpha helix, needed

  6. A role for the fission yeast Rqh1 helicase in chromosome segregation

    Win, Thein Z; Mankouri, Hocine W; Hickson, Ian D

    2005-01-01

    Schizosaccharomyces pombe Rqh1 protein is a member of the RecQ DNA helicase family. Members of this protein family are mutated in several human genome instability syndromes, including Bloom, Werner and Rothmund-Thomson syndromes. RecQ helicases participate in recombination repair of stalled...

  7. Emerging Importance of Helicases in Plant Stress Tolerance: Characterization of Oryza sativa Repair Helicase XPB2 Promoter and Its Functional Validation in Tobacco under Multiple Stresses

    Raikwar, Shailendra; Srivastava, Vineet K.; Gill, Sarvajeet S.; Tuteja, Renu; Tuteja, Narendra

    2015-01-01

    Genetic material always remains at the risk of spontaneous or induced damage which challenges the normal functioning of DNA molecule, thus, DNA repair is vital to protect the organisms against genetic damage. Helicases, the unique molecular motors, are emerged as prospective molecules to engineer stress tolerance in plants and are involved in nucleic acid metabolism including DNA repair. The repair helicase, XPB is an evolutionary conserved protein present in different organisms, including pl...

  8. Emerging importance of helicases in plant stress tolerance: characterization of Oryza sativa repair helicase XPB2 promoter and its functional validation in tobacco under multiple stresses

    Shailendra eRaikwar; Vineet Kumar Shrivastava; Sarvajeet Singh Gill; Renu eTuteja; Narendra eTuteja; Narendra eTuteja

    2015-01-01

    Genetic material always remains at the risk of spontaneous or induced damage which challenges the normal functioning of DNA molecule, thus, DNA repair is vital to protect the organisms against genetic damage. DNA hHelicases, the unique molecular motors, are emerged as potentialprospective molecules to engineer stress tolerance in plants and are involved in a variety of DNA nucleic acid metabolismc processes including DNA repair. The DNA repair helicase, OsXPB2 is an evolutionary conserved pr...

  9. Uncoupling of Protease trans-Cleavage and Helicase Activities in Pestivirus NS3.

    Zheng, Fengwei; Lu, Guoliang; Li, Ling; Gong, Peng; Pan, Zishu

    2017-11-01

    The nonstructural protein NS3 from the Flaviviridae family is a multifunctional protein that contains an N-terminal protease and a C-terminal helicase, playing essential roles in viral polyprotein processing and genome replication. Here we report a full-length crystal structure of the classical swine fever virus (CSFV) NS3 in complex with its NS4A protease cofactor segment (PCS) at a 2.35-Å resolution. The structure reveals a previously unidentified ∼2,200-Å 2 intramolecular protease-helicase interface comprising three clusters of interactions, representing a "closed" global conformation related to the NS3-NS4A cis -cleavage event. Although this conformation is incompatible with protease trans -cleavage, it appears to be functionally important and beneficial to the helicase activity, as the mutations designed to perturb this conformation impaired both the helicase activities in vitro and virus production in vivo Our work reveals important features of protease-helicase coordination in pestivirus NS3 and provides a key basis for how different conformational states may explicitly contribute to certain functions of this natural protease-helicase fusion protein. IMPORTANCE Many RNA viruses encode helicases to aid their RNA genome replication and transcription by unwinding structured RNA. Being naturally fused to a protease participating in viral polyprotein processing, the NS3 helicases encoded by the Flaviviridae family viruses are unique. Therefore, how these two enzyme modules coordinate in a single polypeptide is of particular interest. Here we report a previously unidentified conformation of pestivirus NS3 in complex with its NS4A protease cofactor segment (PCS). This conformational state is related to the protease cis -cleavage event and is optimal for the function of helicase. This work provides an important basis to understand how different enzymatic activities of NS3 may be achieved by the coordination between the protease and helicase through different

  10. SU-8 Based Piezoresistive Mechanical Sensor

    Thaysen, Jacob; Yalcinkaya, Arda Deniz; Vestergaard, R.K.

    2002-01-01

    We present the first SU-8 based piezoresistive mechanical sensor. Conventionally, silicon has been used as a piezoresistive material due to its high gauge factor and thereby high sensitivity to strain changes in a sensor. By using the fact that SU-8 is much softer than silicon and that a gold...

  11. The Drosophila Helicase MLE Targets Hairpin Structures in Genomic Transcripts.

    Simona Cugusi

    2016-01-01

    Full Text Available RNA hairpins are a common type of secondary structures that play a role in every aspect of RNA biochemistry including RNA editing, mRNA stability, localization and translation of transcripts, and in the activation of the RNA interference (RNAi and microRNA (miRNA pathways. Participation in these functions often requires restructuring the RNA molecules by the association of single-strand (ss RNA-binding proteins or by the action of helicases. The Drosophila MLE helicase has long been identified as a member of the MSL complex responsible for dosage compensation. The complex includes one of two long non-coding RNAs and MLE was shown to remodel the roX RNA hairpin structures in order to initiate assembly of the complex. Here we report that this function of MLE may apply to the hairpins present in the primary RNA transcripts that generate the small molecules responsible for RNA interference. Using stocks from the Transgenic RNAi Project and the Vienna Drosophila Research Center, we show that MLE specifically targets hairpin RNAs at their site of transcription. The association of MLE at these sites is independent of sequence and chromosome location. We use two functional assays to test the biological relevance of this association and determine that MLE participates in the RNAi pathway.

  12. Molecular Dynamics of the ZIKA Virus NS3 Helicase

    Raubenolt, Bryan; Rick, Steven; The Rick Group Team

    The recent outbreaks of the ZIKA virus (ZIKV) and its connection to microcephaly in newborns has raised its awareness as a global threat and many scientific research efforts are currently underway in attempt to create a vaccine. Molecular Dynamics is a powerful method of investigating the physical behavior of protein complexes. ZIKV is comprised of 3 structural and 7 nonstructural proteins. The NS3 helicase protein appears to play a significant role in the replication complex and its inhibition could be a crucial source of antiviral drug design. This research primarily focuses on studying the structural dynamics, over the course of few hundred nanoseconds, of NS3 helicase in the free state, as well as in complex form with human ssRNA, ATP, and an analogue of GTP. RMSD and RMSF plots of each simulation will provide details on the forces involved in the overall stability of the active and inactive states. Furthermore, free energy calculations on a per residue level will reveal the most interactive residues between states and ultimately the primary driving force behind these interactions. Together these analyses will provide highly relevant information on the binding surface chemistry and thus serve as the basis for potential drug design.

  13. Molecular determinants of nucleolar translocation of RNA helicase A

    Liu Zhe; Kenworthy, Rachael; Green, Christopher; Tang, Hengli

    2007-01-01

    RNA helicase A (RHA) is a member of the DEAH-box family of DNA/RNA helicases involved in multiple cellular processes and the life cycles of many viruses. The subcellular localization of RHA is dynamic despite its steady-state concentration in the nucleoplasm. We have previously shown that it shuttles rapidly between the nucleus and the cytoplasm by virtue of a bidirectional nuclear transport domain (NTD) located in its carboxyl terminus. Here, we investigate the molecular determinants for its translocation within the nucleus and, more specifically, its redistribution from the nucleoplasm to nucleolus or the perinucleolar region. We found that low temperature treatment, transcription inhibition or replication of hepatitis C virus caused the intranuclear redistribution of the protein, suggesting that RHA shuttles between the nucleolus and nucleoplasm and becomes trapped in the nucleolus or the perinucleolar region upon blockade of transport to the nucleoplasm. Both the NTD and ATPase activity were essential for RHA's transport to the nucleolus or perinucleolar region. One of the double-stranded RNA binding domains (dsRBD II) was also required for this nucleolar translocation (NoT) phenotype. RNA interference studies revealed that RHA is essential for survival of cultured hepatoma cells and the ATPase activity appears to be important for this critical role

  14. The RNA helicase Rm62 cooperates with SU(VAR3-9 to re-silence active transcription in Drosophila melanogaster.

    Joern Boeke

    Full Text Available Gene expression is highly dynamic and many genes show a wide range in expression over several orders of magnitude. This regulation is often mediated by sequence specific transcription factors. In addition, the tight packaging of DNA into chromatin can provide an additional layer of control resulting in a dynamic range of gene expression covering several orders of magnitude. During transcriptional activation, chromatin barriers have to be eliminated to allow an efficient progression of the RNA polymerase. This repressive chromatin structure has to be re-established quickly after it has been activated in order to tightly regulate gene activity. We show that the DExD/H box containing RNA helicase Rm62 is targeted to a site of rapid induction of transcription where it is responsible for an increased degree of methylation at H3K9 at the heat shock locus after removal of the heat shock stimulus. The RNA helicase interacts with the well-characterized histone methyltransferase SU(VAR3-9 via its N-terminus, which provides a potential mechanism for the targeting of H3K9 methylation to highly regulated genes. The recruitment of SU(VAR3-9 through interaction with a RNA helicase to a site of active transcription might be a general mechanism that allows an efficient silencing of highly regulated genes thereby enabling a cell to fine tune its gene activity over a wide range.

  15. PBDE: Structure-Activity Studies for the Inhibition of Hepatitis C Virus NS3 Helicase

    Kazi Abdus Salam

    2014-04-01

    Full Text Available The helicase portion of the hepatitis C virus nonstructural protein 3 (NS3 is considered one of the most validated targets for developing direct acting antiviral agents. We isolated polybrominated diphenyl ether (PBDE 1 from a marine sponge as an NS3 helicase inhibitor. In this study, we evaluated the inhibitory effects of PBDE (1 on the essential activities of NS3 protein such as RNA helicase, ATPase, and RNA binding activities. The structure-activity relationship analysis of PBDE (1 against the HCV ATPase revealed that the biphenyl ring, bromine, and phenolic hydroxyl group on the benzene backbone might be a basic scaffold for the inhibitory potency.

  16. ATPase activity measurement of DNA replicative helicase from Bacillus stearothermophilus by malachite green method.

    Yang, Mu; Wang, Ganggang

    2016-09-15

    The DnaB helicase from Bacillus stearothermophilus (DnaBBst) was a model protein for studying the bacterial DNA replication. In this work, a non-radioactive method for measuring ATPase activity of DnaBBst helicase was described. The working parameters and conditions were optimized. Furthermore, this method was applied to investigate effects of DnaG primase, ssDNA and helicase loader protein (DnaI) on ATPase activity of DnaBBst. Our results showed this method was sensitive and efficient. Moreover, it is suitable for the investigation of functional interaction between DnaB and related factors. Copyright © 2016 Elsevier Inc. All rights reserved.

  17. Mechanics model for actin-based motility.

    Lin, Yuan

    2009-02-01

    We present here a mechanics model for the force generation by actin polymerization. The possible adhesions between the actin filaments and the load surface, as well as the nucleation and capping of filament tips, are included in this model on top of the well-known elastic Brownian ratchet formulation. A closed form solution is provided from which the force-velocity relationship, summarizing the mechanics of polymerization, can be drawn. Model predictions on the velocity of moving beads driven by actin polymerization are consistent with experiment observations. This model also seems capable of explaining the enhanced actin-based motility of Listeria monocytogenes and beads by the presence of Vasodilator-stimulated phosphoprotein, as observed in recent experiments.

  18. RTEL1: functions of a disease-associated helicase.

    Vannier, Jean-Baptiste; Sarek, Grzegorz; Boulton, Simon J

    2014-07-01

    DNA secondary structures that arise during DNA replication, repair, and recombination (3R) must be processed correctly to prevent genetic instability. Regulator of telomere length 1 (RTEL1) is an essential DNA helicase that disassembles a variety of DNA secondary structures to facilitate 3R processes and to maintain telomere integrity. The past few years have witnessed the emergence of RTEL1 variants that confer increased susceptibility to high-grade glioma, astrocytomas, and glioblastomas. Mutations in RTEL1 have also been implicated in Hoyeraal-Hreidarsson syndrome, a severe form of the bone-marrow failure and cancer predisposition disorder, dyskeratosis congenita. We review these recent findings and highlight its crucial link between DNA secondary-structure metabolism and human disease. Copyright © 2014 Elsevier Ltd. All rights reserved.

  19. Heteroduplex DNA position defines the roles of the Sgs1, Srs2, and Mph1 helicases in promoting distinct recombination outcomes.

    Katrina Mitchel

    Full Text Available The contributions of the Sgs1, Mph1, and Srs2 DNA helicases during mitotic double-strand break (DSB repair in yeast were investigated using a gap-repair assay. A diverged chromosomal substrate was used as a repair template for the gapped plasmid, allowing mismatch-containing heteroduplex DNA (hDNA formed during recombination to be monitored. Overall DSB repair efficiencies and the proportions of crossovers (COs versus noncrossovers (NCOs were determined in wild-type and helicase-defective strains, allowing the efficiency of CO and NCO production in each background to be calculated. In addition, the products of individual NCO events were sequenced to determine the location of hDNA. Because hDNA position is expected to differ depending on whether a NCO is produced by synthesis-dependent-strand-annealing (SDSA or through a Holliday junction (HJ-containing intermediate, its position allows the underlying molecular mechanism to be inferred. Results demonstrate that each helicase reduces the proportion of CO recombinants, but that each does so in a fundamentally different way. Mph1 does not affect the overall efficiency of gap repair, and its loss alters the CO-NCO by promoting SDSA at the expense of HJ-containing intermediates. By contrast, Sgs1 and Srs2 are each required for efficient gap repair, strongly promoting NCO formation and having little effect on CO efficiency. hDNA analyses suggest that all three helicases promote SDSA, and that Sgs1 and Srs2 additionally dismantle HJ-containing intermediates. The hDNA data are consistent with the proposed role of Sgs1 in the dissolution of double HJs, and we propose that Srs2 dismantles nicked HJs.

  20. Crystal structures of the methyltransferase and helicase from the ZIKA 1947 MR766 Uganda strain

    Bukrejewska, Malgorzata; Derewenda, Urszula; Radwanska, Malwina; Engel, Daniel A.; Derewenda, Zygmunt S.

    2017-08-15

    Two nonstructural proteins encoded byZika virusstrain MR766 RNA, a methyltransferase and a helicase, were crystallized and their structures were solved and refined at 2.10 and 2.01 Å resolution, respectively. The NS5 methyltransferase contains a boundS-adenosyl-L-methionine (SAM) co-substrate. The NS3 helicase is in the apo form. Comparison with published crystal structures of the helicase in the apo, nucleotide-bound and single-stranded RNA (ssRNA)-bound states suggests that binding of ssRNA to the helicase may occur through conformational selection rather than induced fit.

  1. Dissociation from DNA of Type III Restriction–Modification enzymes during helicase-dependent motion and following endonuclease activity

    Tóth, Júlia; van Aelst, Kara; Salmons, Hannah; Szczelkun, Mark D.

    2012-01-01

    DNA cleavage by the Type III Restriction–Modification (RM) enzymes requires the binding of a pair of RM enzymes at two distant, inversely orientated recognition sequences followed by helicase-catalysed ATP hydrolysis and long-range communication. Here we addressed the dissociation from DNA of these enzymes at two stages: during long-range communication and following DNA cleavage. First, we demonstrated that a communicating species can be trapped in a DNA domain without a recognition site, with a non-specific DNA association lifetime of ∼200 s. If free DNA ends were present the lifetime became too short to measure, confirming that ends accelerate dissociation. Secondly, we observed that Type III RM enzymes can dissociate upon DNA cleavage and go on to cleave further DNA molecules (they can ‘turnover’, albeit inefficiently). The relationship between the observed cleavage rate and enzyme concentration indicated independent binding of each site and a requirement for simultaneous interaction of at least two enzymes per DNA to achieve cleavage. In light of various mechanisms for helicase-driven motion on DNA, we suggest these results are most consistent with a thermally driven random 1D search model (i.e. ‘DNA sliding’). PMID:22523084

  2. Structural insights into RISC assembly facilitated by dsRNA-binding domains of human RNA helicase A (DHX9).

    Fu, Qinqin; Yuan, Y Adam

    2013-03-01

    Intensive research interest has focused on small RNA-processing machinery and the RNA-induced silencing complex (RISC), key cellular machines in RNAi pathways. However, the structural mechanism regarding RISC assembly, the primary step linking small RNA processing and RNA-mediated gene silencing, is largely unknown. Human RNA helicase A (DHX9) was reported to function as an RISC-loading factor, and such function is mediated mainly by its dsRNA-binding domains (dsRBDs). Here, we report the crystal structures of human RNA helicase A (RHA) dsRBD1 and dsRBD2 domains in complex with dsRNAs, respectively. Structural analysis not only reveals higher siRNA duplex-binding affinity displayed by dsRBD1, but also identifies a crystallographic dsRBD1 pair of physiological significance in cooperatively recognizing dsRNAs. Structural observations are further validated by isothermal titration calorimetric (ITC) assay. Moreover, co-immunoprecipitation (co-IP) assay coupled with mutagenesis demonstrated that both dsRBDs are required for RISC association, and such association is mediated by dsRNA. Hence, our structural and functional efforts have revealed a potential working model for siRNA recognition by RHA tandem dsRBDs, and together they provide direct structural insights into RISC assembly facilitated by RHA.

  3. Teletactile System Based on Mechanical Properties Estimation

    Mauro M. Sette

    2011-01-01

    Full Text Available Tactile feedback is a major missing feature in minimally invasive procedures; it is an essential means of diagnosis and orientation during surgical procedures. Previous works have presented a remote palpation feedback system based on the coupling between a pressure sensor and a general haptic interface. Here a new approach is presented based on the direct estimation of the tissue mechanical properties and finally their presentation to the operator by means of a haptic interface. The approach presents different technical difficulties and some solutions are proposed: the implementation of a fast Young’s modulus estimation algorithm, the implementation of a real time finite element model, and finally the implementation of a stiffness estimation approach in order to guarantee the system’s stability. The work is concluded with an experimental evaluation of the whole system.

  4. XPD Helicase Structures and Activities: Insights into the Cancer and Aging Phenotypes from XPD Mutations

    Tainer, John; Fan, Li; Fuss, Jill O.; Cheng, Quen J.; Arvai, Andrew S.; Hammel, Michal; Roberts, Victoria A.; Cooper, Priscilla K.; Tainer, John A.

    2008-06-02

    Mutations in XPD helicase, required for nucleotide excision repair (NER) as part of the transcription/repair complex TFIIH, cause three distinct phenotypes: cancer-prone xeroderma pigmentosum (XP), or aging disorders Cockayne syndrome (CS), and trichothiodystrophy (TTD). To clarify molecular differences underlying these diseases, we determined crystal structures of the XPD catalytic core from Sulfolobus acidocaldarius and measured mutant enzyme activities. Substrate-binding grooves separate adjacent Rad51/RecA-like helicase domains (HD1, HD2) and an arch formed by 4FeS and Arch domains. XP mutations map along the HD1 ATP-binding edge and HD2 DNA-binding channel and impair helicase activity essential for NER. XP/CS mutations both impair helicase activity and likely affect HD2 functional movement. TTD mutants lose or retain helicase activity but map to sites in all four domains expected to cause framework defects impacting TFIIH integrity. These results provide a foundation for understanding disease consequences of mutations in XPD and related 4Fe-4S helicases including FancJ.

  5. XPD Helicase Structures And Activities: Insights Into the Cancer And Aging Phenotypes From XPD Mutations

    Fan, L.; Fuss, J.O.; Cheng, Q.J.; Arvai, A.S.; Hammel, M.; Roberts, V.A.; Cooper, P.K.; Tainer, J.A.

    2009-05-18

    Mutations in XPD helicase, required for nucleotide excision repair (NER) as part of the transcription/repair complex TFIIH, cause three distinct phenotypes: cancer-prone xeroderma pigmentosum (XP), or aging disorders Cockayne syndrome (CS), and trichothiodystrophy (TTD). To clarify molecular differences underlying these diseases, we determined crystal structures of the XPD catalytic core from Sulfolobus acidocaldarius and measured mutant enzyme activities. Substrate-binding grooves separate adjacent Rad51/RecA-like helicase domains (HD1, HD2) and an arch formed by 4FeS and Arch domains. XP mutations map along the HD1 ATP-binding edge and HD2 DNA-binding channel and impair helicase activity essential for NER. XP/CS mutations both impair helicase activity and likely affect HD2 functional movement. TTD mutants lose or retain helicase activity but map to sites in all four domains expected to cause framework defects impacting TFIIH integrity. These results provide a foundation for understanding disease consequences of mutations in XPD and related 4Fe-4S helicases including FancJ.

  6. Crystal Structure of the Phage T4 Recombinase UvsX and Its Functional Interaction with the T4 SF2 Helicase UvsW

    Gajewski, Stefan; Webb, Michael R.; Galkin, Vitold; Egelman, Edward H.; Kreuzer, Kenneth N.; White, Stephen W. (Duke); (UV); (SJCH)

    2012-07-11

    Bacteriophage T4 provides an important model system for studying the mechanism of homologous recombination. We have determined the crystal structure of the T4 UvsX recombinase, and the overall architecture and fold closely resemble those of RecA, including a highly conserved ATP binding site. Based on this new structure, we reanalyzed electron microscopy reconstructions of UvsX-DNA filaments and docked the UvsX crystal structure into two different filament forms: a compressed filament generated in the presence of ADP and an elongated filament generated in the presence of ATP and aluminum fluoride. In these reconstructions, the ATP binding site sits at the protomer interface, as in the RecA filament crystal structure. However, the environment of the ATP binding site is altered in the two filament reconstructions, suggesting that nucleotide cannot be as easily accommodated at the protomer interface of the compressed filament. Finally, we show that the phage helicase UvsW completes the UvsX-promoted strand-exchange reaction, allowing the generation of a simple nicked circular product rather than complex networks of partially exchanged substrates.

  7. Inherited mutations in the helicase RTEL1 cause telomere dysfunction and Hoyeraal-Hreidarsson syndrome.

    Deng, Zhong; Glousker, Galina; Molczan, Aliah; Fox, Alan J; Lamm, Noa; Dheekollu, Jayaraju; Weizman, Orr-El; Schertzer, Michael; Wang, Zhuo; Vladimirova, Olga; Schug, Jonathan; Aker, Memet; Londoño-Vallejo, Arturo; Kaestner, Klaus H; Lieberman, Paul M; Tzfati, Yehuda

    2013-09-03

    Telomeres repress the DNA damage response at the natural chromosome ends to prevent cell-cycle arrest and maintain genome stability. Telomeres are elongated by telomerase in a tightly regulated manner to ensure a sufficient number of cell divisions throughout life, yet prevent unlimited cell division and cancer development. Hoyeraal-Hreidarsson syndrome (HHS) is characterized by accelerated telomere shortening and a broad range of pathologies, including bone marrow failure, immunodeficiency, and developmental defects. HHS-causing mutations have previously been found in telomerase and the shelterin component telomeric repeat binding factor 1 (TRF1)-interacting nuclear factor 2 (TIN2). We identified by whole-genome exome sequencing compound heterozygous mutations in four siblings affected with HHS, in the gene encoding the regulator of telomere elongation helicase 1 (RTEL1). Rtel1 was identified in mouse by its genetic association with telomere length. However, its mechanism of action and whether it regulates telomere length in human remained unknown. Lymphoblastoid cell lines obtained from a patient and from the healthy parents carrying heterozygous RTEL1 mutations displayed telomere shortening, fragility and fusion, and growth defects in culture. Ectopic expression of WT RTEL1 suppressed the telomere shortening and growth defect, confirming the causal role of the RTEL1 mutations in HHS and demonstrating the essential function of human RTEL1 in telomere protection and elongation. Finally, we show that human RTEL1 interacts with the shelterin protein TRF1, providing a potential recruitment mechanism of RTEL1 to telomeres.

  8. Inherited mutations in the helicase RTEL1 cause telomere dysfunction and Hoyeraal–Hreidarsson syndrome

    Deng, Zhong; Glousker, Galina; Molczan, Aliah; Fox, Alan J.; Lamm, Noa; Dheekollu, Jayaraju; Weizman, Orr-El; Schertzer, Michael; Wang, Zhuo; Vladimirova, Olga; Schug, Jonathan; Aker, Memet; Londoño-Vallejo, Arturo; Kaestner, Klaus H.; Lieberman, Paul M.; Tzfati, Yehuda

    2013-01-01

    Telomeres repress the DNA damage response at the natural chromosome ends to prevent cell-cycle arrest and maintain genome stability. Telomeres are elongated by telomerase in a tightly regulated manner to ensure a sufficient number of cell divisions throughout life, yet prevent unlimited cell division and cancer development. Hoyeraal–Hreidarsson syndrome (HHS) is characterized by accelerated telomere shortening and a broad range of pathologies, including bone marrow failure, immunodeficiency, and developmental defects. HHS-causing mutations have previously been found in telomerase and the shelterin component telomeric repeat binding factor 1 (TRF1)-interacting nuclear factor 2 (TIN2). We identified by whole-genome exome sequencing compound heterozygous mutations in four siblings affected with HHS, in the gene encoding the regulator of telomere elongation helicase 1 (RTEL1). Rtel1 was identified in mouse by its genetic association with telomere length. However, its mechanism of action and whether it regulates telomere length in human remained unknown. Lymphoblastoid cell lines obtained from a patient and from the healthy parents carrying heterozygous RTEL1 mutations displayed telomere shortening, fragility and fusion, and growth defects in culture. Ectopic expression of WT RTEL1 suppressed the telomere shortening and growth defect, confirming the causal role of the RTEL1 mutations in HHS and demonstrating the essential function of human RTEL1 in telomere protection and elongation. Finally, we show that human RTEL1 interacts with the shelterin protein TRF1, providing a potential recruitment mechanism of RTEL1 to telomeres. PMID:23959892

  9. Two mechanisms coordinate replication termination by the Escherichia coli Tus-Ter complex.

    Pandey, Manjula; Elshenawy, Mohamed M; Jergic, Slobodan; Takahashi, Masateru; Dixon, Nicholas E; Hamdan, Samir M; Patel, Smita S

    2015-07-13

    The Escherichia coli replication terminator protein (Tus) binds to Ter sequences to block replication forks approaching from one direction. Here, we used single molecule and transient state kinetics to study responses of the heterologous phage T7 replisome to the Tus-Ter complex. The T7 replisome was arrested at the non-permissive end of Tus-Ter in a manner that is explained by a composite mousetrap and dynamic clamp model. An unpaired C(6) that forms a lock by binding into the cytosine binding pocket of Tus was most effective in arresting the replisome and mutation of C(6) removed the barrier. Isolated helicase was also blocked at the non-permissive end, but unexpectedly the isolated polymerase was not, unless C(6) was unpaired. Instead, the polymerase was blocked at the permissive end. This indicates that the Tus-Ter mechanism is sensitive to the translocation polarity of the DNA motor. The polymerase tracking along the template strand traps the C(6) to prevent lock formation; the helicase tracking along the other strand traps the complementary G(6) to aid lock formation. Our results are consistent with the model where strand separation by the helicase unpairs the GC(6) base pair and triggers lock formation immediately before the polymerase can sequester the C(6) base. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

  10. Two mechanisms coordinate replication termination by the Escherichia coli Tus–Ter complex

    Pandey, Manjula

    2015-07-13

    The Escherichia coli replication terminator protein (Tus) binds to Ter sequences to block replication forks approaching from one direction. Here, we used single molecule and transient state kinetics to study responses of the heterologous phage T7 replisome to the Tus–Ter complex. The T7 replisome was arrested at the non-permissive end of Tus–Ter in a manner that is explained by a composite mousetrap and dynamic clamp model. An unpaired C(6) that forms a lock by binding into the cytosine binding pocket of Tus was most effective in arresting the replisome and mutation of C(6) removed the barrier. Isolated helicase was also blocked at the non-permissive end, but unexpectedly the isolated polymerase was not, unless C(6) was unpaired. Instead, the polymerase was blocked at the permissive end. This indicates that the Tus–Ter mechanism is sensitive to the translocation polarity of the DNA motor. The polymerase tracking along the template strand traps the C(6) to prevent lock formation; the helicase tracking along the other strand traps the complementary G(6) to aid lock formation. Our results are consistent with the model where strand separation by the helicase unpairs the GC(6) base pair and triggers lock formation immediately before the polymerase can sequester the C(6) base.

  11. Simulation based engineering in solid mechanics

    Rao, J S

    2017-01-01

    This book begins with a brief historical perspective of the advent of rotating machinery in 20th century Solid Mechanics and the development of the discipline of the Strength of Materials. High Performance Computing (HPC) and Simulation Based Engineering Science (SBES) have gradually replaced the conventional approach in Design bringing science directly into engineering without approximations. A recap of the required mathematical principles is given. The science of deformation, strain and stress at a point under the application of external traction loads is next presented. Only one-dimensional structures classified as Bars (axial loads), Rods (twisting loads) and Beams (bending loads) are considered in this book. The principal stresses and strains and von Mises stress and strain that used in design of structures are next presented. Lagrangian solution was used to derive the governing differential equations consistent with assumed deformation field and solution for deformations, strains and stresses were obtai...

  12. Unwinding after high salinity stress: Pea DNA helicase 45 over- expression in tobacco confers high salinity tolerance without affecting yield (abstract)

    Tuteja, N.

    2005-01-01

    Soil salinity is an increasing threat for agriculture and is a major factor in reducing plant productivity; therefore, it is necessary to obtain salinity-tolerant varieties. A typical characteristic of soil salinity is the induction of multiple stress- inducible genes. Some of the genes encoding osmolytes, ion channels or enzymes are able to confer salinity-tolerant phenotypes when transferred to sensitive plants. As salinity stress affects the cellular gene-expression machinery, it is evident that molecules involved in nucleic acid processing including helicases, are likely to be affected as well. DNA helicases unwind duplex DNA and are involved in replication, repair, recombination and transcription while RNA helicases unfold the secondary structures in RNA and are involved in transcription, ribosome biogenesis and translation initiation. We have earlier reported the isolation of a pea DNA helicase 45 (PDH45) that exhibits striking homology with eIF-4A (Plant J. 24:219-230,2000). Here we report that PDH45 mRNA is induced in pea seedlings in response to high salt and its over- expression driven by a constitutive CAMV-355-promoter in tobacco plants confers salinity tolerance, thus suggesting a new pathway for manipulating stress tolerance in crop plants. The T0 transgenic plants showed high-levels of PDH45 protein in normal and stress conditions, as compared to wild type (WT) plants. The T0 transgenics also showed tolerance to high salinity as tested by a leaf disc senescence assay. The T1 transgenics were able to grow to maturity and set normal viable seeds under continuous salinity stress, without any reduction in plant yield, in terms of seed weight. Measurement of Na/sup +/ ions in different parts of the plant showed higher accumulation in the old leaves and negligible in seeds of T1 transgenic lines as compared with the WT plants. The possible mechanism of salinity tolerance will be discussed. Over-expression of PDH45 provides a possible example of the

  13. The DEAD box helicase RDE-12 promotes amplification of RNAi in cytoplasmic foci in C. elegans.

    Yang, Huan; Vallandingham, Jim; Shiu, Philip; Li, Hua; Hunter, Craig P; Mak, Ho Yi

    2014-04-14

    RNAi is a potent mechanism for downregulating gene expression. Conserved RNAi pathway components are found in animals, plants, fungi, and other eukaryotes. In C. elegans, the RNAi response is greatly amplified by the synthesis of abundant secondary small interfering RNAs (siRNAs). Exogenous double-stranded RNA is processed by Dicer and RDE-1/Argonaute into primary siRNA that guides target mRNA recognition. The RDE-10/RDE-11 complex and the RNA-dependent RNA polymerase RRF-1 then engage the target mRNA for secondary siRNA synthesis. However, the molecular link between primary siRNA production and secondary siRNA synthesis remains largely unknown. Furthermore, it is unclear whether the subcellular sites for target mRNA recognition and degradation coincide with sites where siRNA synthesis and amplification occur. In the C. elegans germline, cytoplasmic P granules at the nuclear pores and perinuclear Mutator foci contribute to target mRNA surveillance and siRNA amplification, respectively. We report that RDE-12, a conserved phenylalanine-glycine (FG) domain-containing DEAD box helicase, localizes in P granules and cytoplasmic foci that are enriched in RSD-6 but are excluded from the Mutator foci. Our results suggest that RDE-12 promotes secondary siRNA synthesis by orchestrating the recruitment of RDE-10 and RRF-1 to primary siRNA-targeted mRNA in distinct cytoplasmic compartments. Copyright © 2014 Elsevier Ltd. All rights reserved.

  14. Functional Dynamics of Hexameric Helicase Probed by Hydrogen Exchange and Simulation

    Radou, Gaël; Dreyer, Frauke N.; Tuma, Roman; Paci, Emanuele

    2014-01-01

    The biological function of large macromolecular assemblies depends on their structure and their dynamics over a broad range of timescales; for this reason, it is a significant challenge to investigate these assemblies using conventional experimental techniques. One of the most promising experimental techniques is hydrogen-deuterium exchange detected by mass spectrometry. Here, we describe to our knowledge a new computational method for quantitative interpretation of deuterium exchange kinetics and apply it to a hexameric viral helicase P4 that unwinds and translocates RNA into a virus capsid at the expense of ATP hydrolysis. Room-temperature dynamics probed by a hundred nanoseconds of all-atom molecular dynamics simulations is sufficient to predict the exchange kinetics of most sequence fragments and provide a residue-level interpretation of the low-resolution experimental results. The strategy presented here is also a valuable tool to validate experimental data, e.g., assignments, and to probe mechanisms that cannot be observed by x-ray crystallography, or that occur over timescales longer than those that can be realistically simulated, such as the opening of the hexameric ring. PMID:25140434

  15. Helicase-primase inhibitor amenamevir for herpesvirus infection: Towards practical application for treating herpes zoster.

    Shiraki, K

    2017-11-01

    Valacyclovir and famciclovir enabled successful systemic therapy for treating herpes simplex virus (HSV) and varicella zoster virus (VZV) infection by their phosphorylation with viral thymidine kinase. Helicase-primase inhibitors (HPIs) inhibit the progression of the replication fork, an initial step in DNA synthesis to separate the double strand into two single strands. The HPIs amenamevir and pritelivir have a novel mechanism of action, once-daily administration with nonrenal excretory characteristics, and clinical efficacy for genital herpes. Amenamevir exhibits anti-VZV and anti-HSV activity while pritelivir only has anti-HSV activity. A clinical trial of amenamevir for herpes zoster has been completed, and amenamevir has been licensed and successfully used in 20,000 patients with herpes zoster so far in Japan. We have characterized the features of the antiviral action of amenamevir and, unlike acyclovir, the drug's antiviral activity is not influenced by the viral replication cycle. Amenamevir is opening a new era of antiherpes therapy. Copyright 2017 Clarivate Analytics.

  16. Cdt1 stabilizes an open MCM ring for helicase loading.

    Frigola, Jordi; He, Jun; Kinkelin, Kerstin; Pye, Valerie E; Renault, Ludovic; Douglas, Max E; Remus, Dirk; Cherepanov, Peter; Costa, Alessandro; Diffley, John F X

    2017-06-23

    ORC, Cdc6 and Cdt1 act together to load hexameric MCM, the motor of the eukaryotic replicative helicase, into double hexamers at replication origins. Here we show that Cdt1 interacts with MCM subunits Mcm2, 4 and 6, which both destabilizes the Mcm2-5 interface and inhibits MCM ATPase activity. Using X-ray crystallography, we show that Cdt1 contains two winged-helix domains in the C-terminal half of the protein and a catalytically inactive dioxygenase-related N-terminal domain, which is important for MCM loading, but not for subsequent replication. We used these structures together with single-particle electron microscopy to generate three-dimensional models of MCM complexes. These show that Cdt1 stabilizes MCM in a left-handed spiral open at the Mcm2-5 gate. We propose that Cdt1 acts as a brace, holding MCM open for DNA entry and bound to ATP until ORC-Cdc6 triggers ATP hydrolysis by MCM, promoting both Cdt1 ejection and MCM ring closure.

  17. RNases and Helicases in Gram-Positive Bacteria.

    Durand, Sylvain; Condon, Ciaran

    2018-04-01

    RNases are key enzymes involved in RNA maturation and degradation. Although they play a crucial role in all domains of life, bacteria, archaea, and eukaryotes have evolved with their own sets of RNases and proteins modulating their activities. In bacteria, these enzymes allow modulation of gene expression to adapt to rapidly changing environments. Today, >20 RNases have been identified in both Escherichia coli and Bacillus subtilis , the paradigms of the Gram-negative and Gram-positive bacteria, respectively. However, only a handful of these enzymes are common to these two organisms and some of them are essential to only one. Moreover, although sets of RNases can be very similar in closely related bacteria such as the Firmicutes Staphylococcus aureus and B. subtilis , the relative importance of individual enzymes in posttranscriptional regulation in these organisms varies. In this review, we detail the role of the main RNases involved in RNA maturation and degradation in Gram-positive bacteria, with an emphasis on the roles of RNase J1, RNase III, and RNase Y. We also discuss how other proteins such as helicases can modulate the RNA-degradation activities of these enzymes.

  18. Disintegration of cruciform and G-quadruplex structures during the course of helicase-dependent amplification (HDA).

    Li, Dawei; Lv, Bei; Zhang, Hao; Lee, Jasmine Yiqin; Li, Tianhu

    2015-04-15

    Unlike chemical damages on DNA, physical alterations of B-form of DNA occur commonly in organisms that serve as signals for specified cellular events. Although the modes of action for repairing of chemically damaged DNA have been well studied nowadays, the repairing mechanisms for physically altered DNA structures have not yet been understood. Our current in vitro studies show that both breakdown of stable non-B DNA structures and resumption of canonical B-conformation of DNA can take place during the courses of isothermal helicase-dependent amplification (HDA). The pathway that makes the non-B DNA structures repairable is presumably the relieving of the accumulated torsional stress that was caused by the positive supercoiling. Our new findings suggest that living organisms might have evolved this distinct and economical pathway for repairing their physically altered DNA structures. Copyright © 2015 Elsevier Ltd. All rights reserved.

  19. The C-terminal domain of the Bloom syndrome DNA helicase is essential for genomic stability

    Noonan James P

    2001-07-01

    Full Text Available Abstract Background Bloom syndrome is a rare cancer-prone disorder in which the cells of affected persons have a high frequency of somatic mutation and genomic instability. Bloom syndrome cells have a distinctive high frequency of sister chromatid exchange and quadriradial formation. BLM, the protein altered in BS, is a member of the RecQ DNA helicase family, whose members share an average of 40% identity in the helicase domain and have divergent N-terminal and C-terminal flanking regions of variable lengths. The BLM DNA helicase has been shown to localize to the ND10 (nuclear domain 10 or PML (promyelocytic leukemia nuclear bodies, where it associates with TOPIIIα, and to the nucleolus. Results This report demonstrates that the N-terminal domain of BLM is responsible for localization of the protein to the nuclear bodies, while the C-terminal domain directs the protein to the nucleolus. Deletions of the N-terminal domain of BLM have little effect on sister chromatid exchange frequency and chromosome stability as compared to helicase and C-terminal mutations which can increase SCE frequency and chromosome abnormalities. Conclusion The helicase activity and the C-terminal domain of BLM are critical for maintaining genomic stability as measured by the sister chromatid exchange assay. The localization of BLM into the nucleolus by the C-terminal domain appears to be more important to genomic stability than localization in the nuclear bodies.

  20. Unzippers, Resolvers and Sensors: A Structural and Functional Biochemistry Tale of RNA Helicases

    Ana Lúcia Leitão

    2015-01-01

    Full Text Available The centrality of RNA within the biological world is an irrefutable fact that currently attracts increasing attention from the scientific community. The panoply of functional RNAs requires the existence of specific biological caretakers, RNA helicases, devoted to maintain the proper folding of those molecules, resolving unstable structures. However, evolution has taken advantage of the specific position and characteristics of RNA helicases to develop new functions for these proteins, which are at the interface of the basic processes for transference of information from DNA to proteins. RNA helicases are involved in many biologically relevant processes, not only as RNA chaperones, but also as signal transducers, scaffolds of molecular complexes, and regulatory elements. Structural biology studies during the last decade, founded in X-ray crystallography, have characterized in detail several RNA-helicases. This comprehensive review summarizes the structural knowledge accumulated in the last two decades within this family of proteins, with special emphasis on the structure-function relationships of the most widely-studied families of RNA helicases: the DEAD-box, RIG-I-like and viral NS3 classes.

  1. Demonstration of helicase activity in the nonstructural protein, NSs, of the negative-sense RNA virus, groundnut bud necrosis virus.

    Bhushan, Lokesh; Abraham, Ambily; Choudhury, Nirupam Roy; Rana, Vipin Singh; Mukherjee, Sunil Kumar; Savithri, Handanahal Subbarao

    2015-04-01

    The nonstructural protein NSs, encoded by the S RNA of groundnut bud necrosis virus (GBNV) (genus Tospovirus, family Bunyaviridae) has earlier been shown to possess nucleic-acid-stimulated NTPase and 5' α phosphatase activity. ATP hydrolysis is an essential function of a true helicase. Therefore, NSs was tested for DNA helicase activity. The results demonstrated that GBNV NSs possesses bidirectional DNA helicase activity. An alanine mutation in the Walker A motif (K189A rNSs) decreased DNA helicase activity substantially, whereas a mutation in the Walker B motif resulted in a marginal decrease in this activity. The parallel loss of the helicase and ATPase activity in the K189A mutant confirms that NSs acts as a non-canonical DNA helicase. Furthermore, both the wild-type and K189A NSs could function as RNA silencing suppressors, demonstrating that the suppressor activity of NSs is independent of its helicase or ATPase activity. This is the first report of a true helicase from a negative-sense RNA virus.

  2. The roles of WRN and BLM RecQ helicases in the Alternative Lengthening of Telomeres.

    Mendez-Bermudez, Aaron; Hidalgo-Bravo, Alberto; Cotton, Victoria E; Gravani, Athanasia; Jeyapalan, Jennie N; Royle, Nicola J

    2012-11-01

    Approximately 10% of all cancers, but a higher proportion of sarcomas, use the recombination-based alternative lengthening of telomeres (ALT) to maintain telomeres. Two RecQ helicase genes, BLM and WRN, play important roles in homologous recombination repair and they have been implicated in telomeric recombination activity, but their precise roles in ALT are unclear. Using analysis of sequence variation present in human telomeres, we found that a WRN- ALT+ cell line lacks the class of complex telomere mutations attributed to inter-telomeric recombination in other ALT+ cell lines. This suggests that WRN facilitates inter-telomeric recombination when there are sequence differences between the donor and recipient molecules or that sister-telomere interactions are suppressed in the presence of WRN and this promotes inter-telomeric recombination. Depleting BLM in the WRN- ALT+ cell line increased the mutation frequency at telomeres and at the MS32 minisatellite, which is a marker of ALT. The absence of complex telomere mutations persisted in BLM-depleted clones, and there was a clear increase in sequence homogenization across the telomere and MS32 repeat arrays. These data indicate that BLM suppresses unequal sister chromatid interactions that result in excessive homogenization at MS32 and at telomeres in ALT+ cells.

  3. A rapid Salmonella detection method involving thermophilic helicase-dependent amplification and a lateral flow assay.

    Du, Xin-Jun; Zhou, Tian-Jiao; Li, Ping; Wang, Shuo

    2017-08-01

    Salmonella is a major foodborne pathogen that is widespread in the environment and can cause serious human and animal disease. Since conventional culture methods to detect Salmonella are time-consuming and laborious, rapid and accurate techniques to detect this pathogen are critically important for food safety and diagnosing foodborne illness. In this study, we developed a rapid, simple and portable Salmonella detection strategy that combines thermophilic helicase-dependent amplification (tHDA) with a lateral flow assay to provide a detection result based on visual signals within 90 min. Performance analyses indicated that the method had detection limits for DNA and pure cultured bacteria of 73.4-80.7 fg and 35-40 CFU, respectively. Specificity analyses showed no cross reactions with Escherichia coli, Staphylococcus aureus, Listeria monocytogenes, Enterobacter aerogenes, Shigella and Campylobacter jejuni. The results for detection in real food samples showed that 1.3-1.9 CFU/g or 1.3-1.9 CFU/mL of Salmonella in contaminated chicken products and infant nutritional cereal could be detected after 2 h of enrichment. The same amount of Salmonella in contaminated milk could be detected after 4 h of enrichment. This tHDA-strip can be used for the rapid detection of Salmonella in food samples and is particularly suitable for use in areas with limited equipment. Copyright © 2017 Elsevier Ltd. All rights reserved.

  4. Distinct functions of human RecQ helicases during DNA replication.

    Urban, Vaclav; Dobrovolna, Jana; Janscak, Pavel

    2017-06-01

    DNA replication is the most vulnerable process of DNA metabolism in proliferating cells and therefore it is tightly controlled and coordinated with processes that maintain genomic stability. Human RecQ helicases are among the most important factors involved in the maintenance of replication fork integrity, especially under conditions of replication stress. RecQ helicases promote recovery of replication forks being stalled due to different replication roadblocks of either exogenous or endogenous source. They prevent generation of aberrant replication fork structures and replication fork collapse, and are involved in proper checkpoint signaling. The essential role of human RecQ helicases in the genome maintenance during DNA replication is underlined by association of defects in their function with cancer predisposition. Copyright © 2016 Elsevier B.V. All rights reserved.

  5. DNA binding and unwinding by Hel308 helicase requires dual functions of a winged helix domain.

    Northall, Sarah J; Buckley, Ryan; Jones, Nathan; Penedo, J Carlos; Soultanas, Panos; Bolt, Edward L

    2017-09-01

    Hel308 helicases promote genome stability linked to DNA replication in archaea, and have homologues in metazoans. In the crystal structure of archaeal Hel308 bound to a tailed DNA duplex, core helicase domains encircle single-stranded DNA (ssDNA) in a "ratchet" for directional translocation. A winged helix domain (WHD) is also present, but its function is mysterious. We investigated the WHD in full-length Hel308, identifying that mutations in a solvent exposed α-helix resulted in reduced DNA binding and unwinding activities. When isolated from the rest of Hel308, the WHD protein alone bound to duplex DNA but not ssDNA, and DNA binding by WHD protein was abolished by the same mutations as were analyzed in full-length Hel308. Isolated WHD from a human Hel308 homologue (HelQ) also bound to duplex DNA. By disrupting the interface between the Hel308 WHD and a RecA-like domain, a topology typical of Ski2 helicases, we show that this is crucial for ATPase and helicase activities. The data suggest a model in which the WHD promotes activity of Hel308 directly, through binding to duplex DNA that is distinct from ssDNA binding by core helicase, and indirectly through interaction with the RecA-like domain. We propose how the WHD may contribute to ssDNA translocation, resulting in DNA helicase activity or in removal of other DNA bound proteins by "reeling" ssDNA. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. Cooperation of DNA-PKcs and WRN helicase in the maintenance of telomeric D-loops

    Kusumoto-Matsuo, Rika; Opresko, Patricia L; Ramsden, Dale

    2010-01-01

    Werner syndrome is an inherited human progeriod syndrome caused by mutations in the gene encoding the Werner Syndrome protein, WRN. It has both 3'-5' DNA helicase and exonuclease activities, and is suggested to have roles in many aspects of DNA metabolism, including DNA repair and telomere...... D-loop model substrate. In addition, the length of telomeric G-tails decreases in DNA-PKcs knockdown cells, and this phenotype is reversed by overexpression of WRN helicase. These results suggest that WRN and DNA-PKcs may cooperatively prevent G-tail shortening in vivo....

  7. Cyclosporin A associated helicase-like protein facilitates the association of hepatitis C virus RNA polymerase with its cellular cyclophilin B.

    Kengo Morohashi

    Full Text Available BACKGROUND: Cyclosporin A (CsA is well known as an immunosuppressive drug useful for allogeneic transplantation. It has been reported that CsA inhibits hepatitis C virus (HCV genome replication, which indicates that cellular targets of CsA regulate the viral replication. However, the regulation mechanisms of HCV replication governed by CsA target proteins have not been fully understood. PRINCIPAL FINDINGS: Here we show a chemical biology approach that elucidates a novel mechanism of HCV replication. We developed a phage display screening to investigate compound-peptide interaction and identified a novel cellular target molecule of CsA. This protein, named CsA associated helicase-like protein (CAHL, possessed RNA-dependent ATPase activity that was negated by treatment with CsA. The downregulation of CAHL in the cells resulted in a decrease of HCV genome replication. CAHL formed a complex with HCV-derived RNA polymerase NS5B and host-derived cyclophilin B (CyPB, known as a cellular cofactor for HCV replication, to regulate NS5B-CyPB interaction. CONCLUSIONS: We found a cellular factor, CAHL, as CsA associated helicase-like protein, which would form trimer complex with CyPB and NS5B of HCV. The strategy using a chemical compound and identifying its target molecule by our phage display analysis is useful to reveal a novel mechanism underlying cellular and viral physiology.

  8. Cyclosporin A associated helicase-like protein facilitates the association of hepatitis C virus RNA polymerase with its cellular cyclophilin B.

    Morohashi, Kengo; Sahara, Hiroeki; Watashi, Koichi; Iwabata, Kazuki; Sunoki, Takashi; Kuramochi, Kouji; Takakusagi, Kaori; Miyashita, Hiroki; Sato, Noriyuki; Tanabe, Atsushi; Shimotohno, Kunitada; Kobayashi, Susumu; Sakaguchi, Kengo; Sugawara, Fumio

    2011-04-29

    Cyclosporin A (CsA) is well known as an immunosuppressive drug useful for allogeneic transplantation. It has been reported that CsA inhibits hepatitis C virus (HCV) genome replication, which indicates that cellular targets of CsA regulate the viral replication. However, the regulation mechanisms of HCV replication governed by CsA target proteins have not been fully understood. Here we show a chemical biology approach that elucidates a novel mechanism of HCV replication. We developed a phage display screening to investigate compound-peptide interaction and identified a novel cellular target molecule of CsA. This protein, named CsA associated helicase-like protein (CAHL), possessed RNA-dependent ATPase activity that was negated by treatment with CsA. The downregulation of CAHL in the cells resulted in a decrease of HCV genome replication. CAHL formed a complex with HCV-derived RNA polymerase NS5B and host-derived cyclophilin B (CyPB), known as a cellular cofactor for HCV replication, to regulate NS5B-CyPB interaction. We found a cellular factor, CAHL, as CsA associated helicase-like protein, which would form trimer complex with CyPB and NS5B of HCV. The strategy using a chemical compound and identifying its target molecule by our phage display analysis is useful to reveal a novel mechanism underlying cellular and viral physiology.

  9. Identification of unique interactions between the flexible linker and the RecA-like domains of DEAD-box helicase Mss116

    Zhang, Yuan; Palla, Mirkó; Liao, Jung-Chi; Sun, Andrew

    2013-01-01

    DEAD-box RNA helicases are ATP-dependent proteins implicated in nearly all aspects of RNA metabolism. The yeast DEAD-box helicase Mss116 is unique in its functions of splicing group I and group II introns and activating mRNA translation, but the structural understanding of why it performs these unique functions remains unclear. Here we used sequence analysis and molecular dynamics simulation to identify residues in the flexible linker specific for yeast Mss116, potentially associated with its unique functions. We first identified residues that are 100% conserved in Mss116 of different species of the Saccharomycetaceae family. The amino acids of these conserved residues were then compared with the amino acids of the corresponding residue positions of other RNA helicases to identify residues that have distinct amino acids from other DEAD-box proteins. Four residues in the flexible linker, i.e. N334, E335, P336 and H339, are conserved and Mss116-specific. Molecular dynamics simulation was conducted for the wild-type Mss116 structure and mutant models to examine mutational effects of the linker on the conformational equilibrium. Relatively short MD simulation runs (within 20 ns) were enough for us to observe mutational effects, suggesting serious structural perturbations by these mutations. The mutation of E335 depletes the interactions between E335 and K95 in domain 1. The interactions between N334/P336 and N496/I497 of domain 2 are also abolished by mutation. Our results suggest that tight interactions between the Mss116-specific flexible linker and the two RecA-like domains may be mechanically required to crimp RNA for the unique RNA processes of yeast Mss116. (paper)

  10. Identification of unique interactions between the flexible linker and the RecA-like domains of DEAD-box helicase Mss116

    Zhang, Yuan; Palla, Mirkó; Sun, Andrew; Liao, Jung-Chi

    2013-09-01

    DEAD-box RNA helicases are ATP-dependent proteins implicated in nearly all aspects of RNA metabolism. The yeast DEAD-box helicase Mss116 is unique in its functions of splicing group I and group II introns and activating mRNA translation, but the structural understanding of why it performs these unique functions remains unclear. Here we used sequence analysis and molecular dynamics simulation to identify residues in the flexible linker specific for yeast Mss116, potentially associated with its unique functions. We first identified residues that are 100% conserved in Mss116 of different species of the Saccharomycetaceae family. The amino acids of these conserved residues were then compared with the amino acids of the corresponding residue positions of other RNA helicases to identify residues that have distinct amino acids from other DEAD-box proteins. Four residues in the flexible linker, i.e. N334, E335, P336 and H339, are conserved and Mss116-specific. Molecular dynamics simulation was conducted for the wild-type Mss116 structure and mutant models to examine mutational effects of the linker on the conformational equilibrium. Relatively short MD simulation runs (within 20 ns) were enough for us to observe mutational effects, suggesting serious structural perturbations by these mutations. The mutation of E335 depletes the interactions between E335 and K95 in domain 1. The interactions between N334/P336 and N496/I497 of domain 2 are also abolished by mutation. Our results suggest that tight interactions between the Mss116-specific flexible linker and the two RecA-like domains may be mechanically required to crimp RNA for the unique RNA processes of yeast Mss116.

  11. A Small Molecule Inhibitor of the BLM Helicase Modulates Chromosome Stability in Human Cells

    Nguyen, Giang Huong; Dexheimer, Thomas S; Rosenthal, Andrew S

    2013-01-01

    The Bloom's syndrome protein, BLM, is a member of the conserved RecQ helicase family. Although cell lines lacking BLM exist, these exhibit progressive genomic instability that makes distinguishing primary from secondary effects of BLM loss problematic. In order to be able to acutely disable BLM f...

  12. BLM helicase measures DNA unwound before switching strands and hRPA promotes unwinding reinitiation

    Yodh, J.G.; Stevens, B.C.; Kanagaraj, R.; Janščák, Pavel; Ha, T.

    2009-01-01

    Roč. 28, č. 4 (2009), s. 405-416 ISSN 0261-4189 Institutional research plan: CEZ:AV0Z50520514 Keywords : Bloom syndrome * FRET * helicase * hRPA * single molecule Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 8.993, year: 2009

  13. BLM helicase suppresses recombination at G-quadruplex motifs in transcribed genes

    van Wietmarschen, Niek; Merzouk, Sarra; Halsema, Nancy; Spierings, Diana C J; Guryev, Victor; Lansdorp, Peter M

    2018-01-01

    Bloom syndrome is a cancer predisposition disorder caused by mutations in the BLM helicase gene. Cells from persons with Bloom syndrome exhibit striking genomic instability characterized by excessive sister chromatid exchange events (SCEs). We applied single-cell DNA template strand sequencing

  14. Human RecQL4 helicase plays critical roles in prostate carcinogenesis

    Su, Yanrong; Meador, Jarah A; Calaf, Gloria M

    2010-01-01

    Prostate cancer is the second leading cause of cancer-associated deaths among men in the western countries. Here, we report that human RecQL4 helicase, which is implicated in the pathogenesis of a subset of cancer-prone Rothmund-Thomson syndrome, is highly elevated in metastatic prostate cancer c...

  15. The Helicase Activity of Hyperthermophilic Archaeal MCM is Enhanced at High Temperatures by Lysine Methylation.

    Xia, Yisui; Niu, Yanling; Cui, Jiamin; Fu, Yang; Chen, Xiaojiang S; Lou, Huiqiang; Cao, Qinhong

    2015-01-01

    Lysine methylation and methyltransferases are widespread in the third domain of life, archaea. Nevertheless, the effects of methylation on archaeal proteins wait to be defined. Here, we report that recombinant sisMCM, an archaeal homolog of Mcm2-7 eukaryotic replicative helicase, is methylated by aKMT4 in vitro. Mono-methylation of these lysine residues occurs coincidently in the endogenous sisMCM protein purified from the hyperthermophilic Sulfolobus islandicus cells as indicated by mass spectra. The helicase activity of mini-chromosome maintenance (MCM) is stimulated by methylation, particularly at temperatures over 70°C. The methylated MCM shows optimal DNA unwinding activity after heat-treatment between 76 and 82°C, which correlates well with the typical growth temperatures of hyperthermophilic Sulfolobus. After methylation, the half life of MCM helicase is dramatically extended at 80°C. The methylated sites are located on the accessible protein surface, which might modulate the intra- and inter- molecular interactions through changing the hydrophobicity and surface charge. Furthermore, the methylation-mimic mutants of MCM show heat resistance helicase activity comparable to the methylated MCM. These data provide the biochemical evidence that posttranslational modifications such as methylation may enhance kinetic stability of proteins under the elevated growth temperatures of hyperthermophilic archaea.

  16. MRE11 complex links RECQ5 helicase to sites of DNA damage

    Zheng, L.; Kanagaraj, R.; Mihaljevic, B.; Schwendener, S.; Sartori, A.A.; Gerrits, B.; Shevelev, Igor; Janščák, Pavel

    2009-01-01

    Roč. 37, č. 8 (2009), s. 2645-2657 ISSN 0305-1048 R&D Projects: GA ČR GA204/09/0565 Institutional research plan: CEZ:AV0Z50520514 Keywords : homologous recombination, * RECQ5 helicase * MRE11 * DNA repair Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 7.479, year: 2009

  17. TFIIH with inactive XPD helicase functions in transcription initiation but is defective in DNA repair

    G.S. Winkler (Sebastiaan); U. Fiedler; W. Vermeulen (Wim); F. Coin (Frédéric); R.D. Wood (Richard); H.T.M. Timmers (Marc); G. Weeda (Geert); J.H.J. Hoeijmakers (Jan); S.J. Araú jo; J-M. Egly (Jean-Marc)

    2000-01-01

    textabstractTFIIH is a multisubunit protein complex involved in RNA polymerase II transcription and nucleotide excision repair, which removes a wide variety of DNA lesions including UV-induced photoproducts. Mutations in the DNA-dependent ATPase/helicase subunits of TFIIH, XPB and

  18. FBH1 Helicase Disrupts RAD51 Filaments in Vitro and Modulates Homologous Recombination in Mammalian Cells

    Šimandlová, Jitka; Zagelbaum, J.; Payne, M.J.; Chu, W.K.; Shevelev, Igor; Hanada, K.; Chatterjee, S.; Reid, D.A.; Liu, Y.; Janščák, Pavel; Rothenberg, E.; Hickson, I.D.

    2013-01-01

    Roč. 288, č. 47 (2013), s. 34168-34180 ISSN 0021-9258 R&D Projects: GA ČR GAP305/10/0281 Institutional support: RVO:68378050 Keywords : DNA damage * DNA helicase * DNA recombination * DNA repair * DNA replication Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.600, year: 2013

  19. Interaction between the helicases genetically linked to Fanconi anemia group J and Bloom's syndrome

    Suhasini, Avvaru N; Rawtani, Nina A; Wu, Yuliang

    2011-01-01

    Bloom's syndrome (BS) and Fanconi anemia (FA) are autosomal recessive disorders characterized by cancer and chromosomal instability. BS and FA group J arise from mutations in the BLM and FANCJ genes, respectively, which encode DNA helicases. In this work, FANCJ and BLM were found to interact...

  20. DNA binding polarity, dimerization, and ATPase ring remodeling in the CMG helicase of the eukaryotic replisome

    Costa, Alessandro; Renault, Ludovic; Swuec, Paolo; Petojevic, Tatjana; Pesavento, James J; Ilves, Ivar; MacLellan-Gibson, Kirsty; Fleck, Roland A; Botchan, Michael R; Berger, James M

    2014-01-01

    The Cdc45/Mcm2-7/GINS (CMG) helicase separates DNA strands during replication in eukaryotes. How the CMG is assembled and engages DNA substrates remains unclear. Using electron microscopy, we have determined the structure of the CMG in the presence of ATPγS and a DNA duplex bearing a 3′ single-stranded tail. The structure shows that the MCM subunits of the CMG bind preferentially to single-stranded DNA, establishes the polarity by which DNA enters into the Mcm2-7 pore, and explains how Cdc45 helps prevent DNA from dissociating from the helicase. The Mcm2-7 subcomplex forms a cracked-ring, right-handed spiral when DNA and nucleotide are bound, revealing unexpected congruencies between the CMG and both bacterial DnaB helicases and the AAA+ motor of the eukaryotic proteasome. The existence of a subpopulation of dimeric CMGs establishes the subunit register of Mcm2-7 double hexamers and together with the spiral form highlights how Mcm2-7 transitions through different conformational and assembly states as it matures into a functional helicase. DOI: http://dx.doi.org/10.7554/eLife.03273.001 PMID:25117490

  1. Characteristics of mechanical alloying of Zn-Al-based alloys

    Zhu, Y.H.; Hong Kong Polytechnic; Perez Hernandez, A.; Lee, W.B.

    2001-01-01

    Three pure elemental powder mixtures of Zn-22%Al-18%Cu, Zn-5%Al-11%Cu, and Zn-27%Al-3%Cu (in wt.%) were mechanically alloyed by steel-ball milling processing. The mechanical alloying characteristics were investigated using X-ray diffraction, scanning electron microscopy, and transmission electron microscopy techniques. It was explored that mechanical alloying started with the formation of phases from pure elemental powders, and this was followed by mechanical milling-induced phase transformation. During mechanical alloying, phases stable at the higher temperatures formed at the near room temperature of milling. Nano-structure Zn-Al-based alloys were produced by mechanical alloying. (orig.)

  2. Mechanism-based drug exposure classification in pharmacoepidemiological studies

    Verdel, B.M.

    2010-01-01

    Mechanism-based classification of drug exposure in pharmacoepidemiological studies In pharmacoepidemiology and pharmacovigilance, the relation between drug exposure and clinical outcomes is crucial. Exposure classification in pharmacoepidemiological studies is traditionally based on

  3. Archaeal orthologs of Cdc45 and GINS form a stable complex that stimulates the helicase activity of MCM.

    Xu, Yuli; Gristwood, Tamzin; Hodgson, Ben; Trinidad, Jonathan C; Albers, Sonja-Verena; Bell, Stephen D

    2016-11-22

    The regulated recruitment of Cdc45 and GINS is key to activating the eukaryotic MCM(2-7) replicative helicase. We demonstrate that the homohexameric archaeal MCM helicase associates with orthologs of GINS and Cdc45 in vivo and in vitro. Association of these factors with MCM robustly stimulates the MCM helicase activity. In contrast to the situation in eukaryotes, archaeal Cdc45 and GINS form an extremely stable complex before binding MCM. Further, the archaeal GINS•Cdc45 complex contains two copies of Cdc45. Our analyses give insight into the function and evolution of the conserved core of the archaeal/eukaryotic replisome.

  4. RIG-I Helicase-Independent Pathway in Sendai Virus-Activated Dendritic Cells Is Critical for Preventing Lung Metastasis of AT6.3 Prostate Cancer

    Tomonori Kato

    2010-11-01

    Full Text Available We recently demonstrated highly efficient antitumor immunity against dermal tumors of B16F10 murine melanoma with the use of dendritic cells (DCs activated by replication-competent, as well as nontransmissible-type, recombinant Sendai viruses (rSeV, and proposed a new concept, “immunostimulatory virotherapy,” for cancer immunotherapy. However, there has been little information on the efficacies of thismethod: 1 inmore clinically relevant situations including metastatic diseases, 2 on other tumor types and other animal species, and 3 on the related molecular/cellular mechanisms. In this study, therefore, we investigated the efficacy of vaccinating DCs activated by fusion gene-deleted nontransmissible rSeV on a rat model of lung metastasis using a highly malignant subline of Dunning R-3327 prostate cancer, AT6.3. rSeV/dF-green fluorescent protein (GFP-activated bone marrow-derived DCs (rSeV/dF-GFP-DC, consistent with results previously observed in murine DCs. Vaccination of rSeV/dF-GFP-DC was highly effective at preventing lung metastasis after intravenous loading of R-3327 tumor cells, compared with the effects observed with immature DCs or lipopolysaccharide-activated DCs. Interestingly, neither CTL activity nor DC trafficking showed any apparent difference among groups. Notably, rSeV/dF-DCs expressing a dominant-negative mutant of retinoic acid-inducible gene I (RIG-I (rSeV/dF-RIGIC-DC, an RNA helicase that recognizes the rSeV genome for inducing type I interferons, largely lost the expression of proinflammatory cytokines without any impairment of antitumor activity. These results indicate the essential role of RIG-I-independent signaling on antimetastatic effect induced by rSeV-activated DCs and may provide important insights to DC-based immunotherapy for advanced malignancies.

  5. The Mechanism Of Auriculotherapy: A Case Report Based On The ...

    The Mechanism Of Auriculotherapy: A Case Report Based On The Fractal Structure Of Meridian System. ... The corresponding acupuncture mechanism of therapeutics has been attributed to the magneto-electric inductive effects of the meridian system via chaotic wave of nerve innervations and blood flow. Hence, based on ...

  6. Mechanically based generative laws of morphogenesis

    Beloussov, Lev V

    2008-01-01

    A deep (although at the first glance naïve) question which may be addressed to embryonic development is why during this process quite definite and accurately reproduced successions of precise and complicated shapes are taking place, or why, in several cases, the result of development is highly precise in spite of an extensive variability of intermediate stages. This problem can be attacked in two different ways. One of them, up to now just slightly employed, is to formulate robust macroscopic generative laws from which the observed successions of shapes could be derived. Another one, which dominates in modern embryology, regards the development as a succession of highly precise 'micropatterns', each of them arising due to the action of specific factors, having, as a rule, nothing in common with each other. We argue that the latter view contradicts a great bulk of firmly established data and gives no satisfactory answers to the main problems of development. Therefore we intend to follow the first way. By doing this, we regard developing embryos as self-organized systems transpierced by feedbacks among which we pay special attention to those linked with mechanical stresses (MS). We formulate a hypothesis of so-called MS hyper-restoration as a common basis for the developmentally important feedback loops. We present a number of examples confirming this hypothesis and use it for reconstructing prolonged chains of developmental events. Finally, we discuss the application of the same set of assumptions to the first steps of egg development and to the internal differentiation of embryonic cells

  7. Seismic metamaterials based on isochronous mechanical oscillators

    Finocchio, G., E-mail: gfinocchio@unime.it; Garescì, F.; Azzerboni, B. [Department of Electronic Engineering, Industrial Chemistry and Engineering, University of Messina, C.da di Dio, I-98166 Messina (Italy); Casablanca, O.; Chiappini, M. [Istituto Nazionale di Geofisica e Vulcanologia (INGV), Via Vigna Murata 605, 00143 Roma (Italy); Ricciardi, G. [Department of Civil, Informatic, Architectural, and Environmental Engineering and Applied Mathematics, C.da di Dio, I-98166 Messina (Italy); Alibrandi, U. [Department of Civil and Environmental Engineering, National University of Singapore, 1 Engineering Drive 2, Singapore 117576 (Singapore)

    2014-05-12

    This Letter introduces a seismic metamaterial (SM) composed by a chain of mass-in-mass system able to filter the S-waves of an earthquake. We included the effect of the SM into the mono dimensional model for the soil response analysis. The SM modifies the soil behavior and in presence of an internal damping the amplitude of the soil amplification function is reduced also in a region near the resonance frequency. This SM can be realized by a continuous structure with inside a 3d-matrix of isochronous oscillators based on a sphere rolling over a cycloidal trajectory.

  8. Antitumor Activity and Mechanism of Action of the Cyclopenta[b]benzofuran, Silvestrol

    Cencic, Regina; Carrier, Marilyn; Galicia-V?zquez, Gabriela; Bordeleau, Marie-Eve; Sukarieh, Rami; Bourdeau, Annie; Brem, Brigitte; Teodoro, Jose G.; Greger, Harald; Tremblay, Michel L.; Porco, John A.; Pelletier, Jerry

    2009-01-01

    Background Flavaglines are a family of natural products from the genus Aglaia that exhibit anti-cancer activity in vitro and in vivo and inhibit translation initiation. They have been shown to modulate the activity of eIF4A, the DEAD-box RNA helicase subunit of the eukaryotic initiation factor (eIF) 4F complex, a complex that stimulates ribosome recruitment during translation initiation. One flavagline, silvestrol, is capable of modulating chemosensitivity in a mechanism-based mouse model. Me...

  9. The roles of the Saccharomyces cerevisiae RecQ helicase SGS1 in meiotic genome surveillance.

    Amit Dipak Amin

    2010-11-01

    Full Text Available The Saccharomyces cerevisiae RecQ helicase Sgs1 is essential for mitotic and meiotic genome stability. The stage at which Sgs1 acts during meiosis is subject to debate. Cytological experiments showed that a deletion of SGS1 leads to an increase in synapsis initiation complexes and axial associations leading to the proposal that it has an early role in unwinding surplus strand invasion events. Physical studies of recombination intermediates implicate it in the dissolution of double Holliday junctions between sister chromatids.In this work, we observed an increase in meiotic recombination between diverged sequences (homeologous recombination and an increase in unequal sister chromatid events when SGS1 is deleted. The first of these observations is most consistent with an early role of Sgs1 in unwinding inappropriate strand invasion events while the second is consistent with unwinding or dissolution of recombination intermediates in an Mlh1- and Top3-dependent manner. We also provide data that suggest that Sgs1 is involved in the rejection of 'second strand capture' when sequence divergence is present. Finally, we have identified a novel class of tetrads where non-sister spores (pairs of spores where each contains a centromere marker from a different parent are inviable. We propose a model for this unusual pattern of viability based on the inability of sgs1 mutants to untangle intertwined chromosomes. Our data suggest that this role of Sgs1 is not dependent on its interaction with Top3. We propose that in the absence of SGS1 chromosomes may sometimes remain entangled at the end of pre-meiotic replication. This, combined with reciprocal crossing over, could lead to physical destruction of the recombined and entangled chromosomes. We hypothesise that Sgs1, acting in concert with the topoisomerase Top2, resolves these structures.This work provides evidence that Sgs1 interacts with various partner proteins to maintain genome stability throughout

  10. Optimization evaluation of cutting technology based on mechanical parts

    Wang, Yu

    2018-04-01

    The relationship between the mechanical manufacturing process and the carbon emission is studied on the basis of the process of the mechanical manufacturing process. The formula of carbon emission calculation suitable for mechanical manufacturing process is derived. Based on this, a green evaluation method for cold machining process of mechanical parts is proposed. The application verification and data analysis of the proposed evaluation method are carried out by an example. The results show that there is a great relationship between the mechanical manufacturing process data and carbon emissions.

  11. Access control mechanism of wireless gateway based on open flow

    Peng, Rong; Ding, Lei

    2017-08-01

    In order to realize the access control of wireless gateway and improve the access control of wireless gateway devices, an access control mechanism of SDN architecture which is based on Open vSwitch is proposed. The mechanism utilizes the features of the controller--centralized control and programmable. Controller send access control flow table based on the business logic. Open vSwitch helps achieve a specific access control strategy based on the flow table.

  12. Relationship between osteosarcoma and ionizing radiation hypersensitive human B lymphocyte cells lacking RecQL4 helicase

    Kohzaki, Masaoki; Moritake, Takashi; Okazaki, Ryuji; Ootsuyama, Akira

    2015-01-01

    Japanese society is now facing a transition period from aging society to super aging society. Concomitant with this situation, it is estimated that number of cancer patients and the requirement of less invasive Radiation Therapy (RT) for cancers will increase. Therefore, understanding of mechanisms without delay on second cancers caused by RT is indispensable. Osteosarcoma, an aggressive bone tumor frequently occurring 5% of cancers in young adult and children, increase statistically after RT for cancers. Although, mutation in p53, Rb and RecQL4 genes statistically relate with osteosarcoma incidence, precise mechanisms of osteosarcoma development by ionizing Radiation (IR) remain to be elucidated. Genome instability is one of the tumor promoting factors and we focused on RecQL4 in RecQ helicase family, which is involved in aging and cancer. We established RecQL4 knock-in human B lymphocyte Nalm-6 cells and found their hypersensitivity to IR, replication fork stall/collapses after IR. In this review, we summarize recently published studies on genetic cancer-predisposing syndrome and possible origins of bone cancers induced by IR. Then, we discuss what and how we address molecular mechanisms on osteosarcoma induced by IR in the future. (author)

  13. DndEi Exhibits Helicase Activity Essential for DNA Phosphorothioate Modification and ATPase Activity Strongly Stimulated by DNA Substrate with a GAAC/GTTC Motif.

    Zheng, Tao; Jiang, Pan; Cao, Bo; Cheng, Qiuxiang; Kong, Lingxin; Zheng, Xiaoqing; Hu, Qinghai; You, Delin

    2016-01-15

    Phosphorothioate (PT) modification of DNA, in which the non-bridging oxygen of the backbone phosphate group is replaced by sulfur, is governed by the DndA-E proteins in prokaryotes. To better understand the biochemical mechanism of PT modification, functional analysis of the recently found PT-modifying enzyme DndEi, which has an additional domain compared with canonical DndE, from Riemerella anatipestifer is performed in this study. The additional domain is identified as a DNA helicase, and functional deletion of this domain in vivo leads to PT modification deficiency, indicating an essential role of helicase activity in PT modification. Subsequent analysis reveals that the additional domain has an ATPase activity. Intriguingly, the ATPase activity is strongly stimulated by DNA substrate containing a GAAC/GTTC motif (i.e. the motif at which PT modifications occur in R. anatipestifer) when the additional domain and the other domain (homologous to canonical DndE) are co-expressed as a full-length DndEi. These results reveal that PT modification is a biochemical process with DNA strand separation and intense ATP hydrolysis. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  14. NS3 from Hepatitis C Virus Strain JFH-1 Is an Unusually Robust Helicase That Is Primed To Bind and Unwind Viral RNA

    Zhou, Ting; Ren, Xiaoming; Adams, Rebecca L.; Pyle, Anna Marie; Ou, J. -H. James

    2017-10-25

    Hepatitis C viruses (HCV) encode a helicase enzyme that is essential for viral replication and assembly (nonstructural protein 3 [NS3]). This helicase has become the focus of extensive basic research on the general helicase mechanism, and it is also of interest as a novel drug target. Despite the importance of this protein, mechanistic work on NS3 has been conducted almost exclusively on variants from HCV genotype 1. Our understanding of NS3 from the highly active HCV strains that are used to study HCV genetics and mechanism in cell culture (such as JFH-1) is lacking. We therefore set out to determine whether NS3 from the replicatively efficient genotype 2a strain JFH-1 displays novel functional or structural properties. Using biochemical assays for RNA binding and duplex unwinding, we show that JFH-1 NS3 binds RNA much more rapidly than the previously studied NS3 variants from genotype 1b. Unlike NS3 variants from other genotypes, JFH-1 NS3 binds RNA with high affinity in a functionally active form that is capable of immediately unwinding RNA duplexes without undergoing rate-limiting conformational changes that precede activation. Unlike other superfamily 2 (SF2) helicases, JFH-1 NS3 does not require long 3' overhangs, and it unwinds duplexes that are flanked by only a few nucleotides, as in the folded HCV genome. To understand the physical basis for this, we solved the crystal structure of JFH-1 NS3, revealing a novel conformation that contains an open, positively charged RNA binding cleft that is primed for productive interaction with RNA targets, potentially explaining robust replication by HCV JFH-1.

    IMPORTANCEGenotypes of HCV are as divergent as different types of flavivirus, and yet mechanistic features of HCV variants are presumed to be held in common. One of the most well-studied components of the HCV replication complex is a helicase known as nonstructural protein 3 (NS3). We set out to determine whether this important

  15. Understanding the mechanism of base development of HSQ

    Kim, Jihoon; Chao, Weilun; Griedel, Brian; Liang, Xiaogan; Lewis, Mark; Hilken, Dawn; Olynick, Deirdre

    2009-06-16

    We study the dissolution mechanism of HSQ (hydrogen silsesquioxane) in base solutions with the addition of chloride salts to elucidate the development mechanism. Reaction mechanisms are proposed based on the dissolution mechanism of quartz. Development kinetics points to two dose-dependent development mechanisms. Considering ion sizes, both hydrated and non-hydrated, and ion exchange, we propose that a combination of a surface dominated reaction at higher doses and a matrix dominated reaction at lower doses accounts for the high development contrast with a NaOH base/NaCl salt mixture. The interplay between the hydrated and non-hydrated ion size leads to higher contrast developers, such as tetramethyl ammonium hydroxide (TMAH) with NaCl.

  16. DNA secondary structure of the released strand stimulates WRN helicase action on forked duplexes without coordinate action of WRN exonuclease

    Ahn, Byungchan, E-mail: bbccahn@mail.ulsan.ac.kr [Department of Life Sciences, University of Ulsan, Ulsan (Korea, Republic of); Bohr, Vilhelm A. [Laboratory of Molecular Gerontology, Biomedical Research Center, National Institute on Aging, Baltimore, MD (United States)

    2011-08-12

    Highlights: {yields} In this study, we investigated the effect of a DNA secondary structure on the two WRN activities. {yields} We found that a DNA secondary structure of the displaced strand during unwinding stimulates WRN helicase without coordinate action of WRN exonuclease. {yields} These results imply that WRN helicase and exonuclease activities can act independently. -- Abstract: Werner syndrome (WS) is an autosomal recessive premature aging disorder characterized by aging-related phenotypes and genomic instability. WS is caused by mutations in a gene encoding a nuclear protein, Werner syndrome protein (WRN), a member of the RecQ helicase family, that interestingly possesses both helicase and exonuclease activities. Previous studies have shown that the two activities act in concert on a single substrate. We investigated the effect of a DNA secondary structure on the two WRN activities and found that a DNA secondary structure of the displaced strand during unwinding stimulates WRN helicase without coordinate action of WRN exonuclease. These results imply that WRN helicase and exonuclease activities can act independently, and we propose that the uncoordinated action may be relevant to the in vivo activity of WRN.

  17. Relativistic supersymmetric quantum mechanics based on Klein-Gordon equation

    Znojil, Miloslav

    2004-01-01

    Witten's the non-relativistic formalism of supersymmetric quantum mechanics was based on a factorization and partnership between Schroedinger equations. We show how it accommodates a transition to the partnership between relativistic Klein-Gordon equations

  18. Mechanics and model-based control of advanced engineering systems

    Irschik, Hans; Krommer, Michael

    2014-01-01

    Mechanics and Model-Based Control of Advanced Engineering Systems collects 32 contributions presented at the International Workshop on Advanced Dynamics and Model Based Control of Structures and Machines, which took place in St. Petersburg, Russia in July 2012. The workshop continued a series of international workshops, which started with a Japan-Austria Joint Workshop on Mechanics and Model Based Control of Smart Materials and Structures and a Russia-Austria Joint Workshop on Advanced Dynamics and Model Based Control of Structures and Machines. In the present volume, 10 full-length papers based on presentations from Russia, 9 from Austria, 8 from Japan, 3 from Italy, one from Germany and one from Taiwan are included, which represent the state of the art in the field of mechanics and model based control, with particular emphasis on the application of advanced structures and machines.

  19. Preparation and mechanical property of polymer-based biomaterials

    Zhang, P; Chen, G; Zheng, X F

    2010-01-01

    The porous polymer-based biomaterial has been synthesized from PLGA, dioxane and tricalcium phosphate (TCP) by low-temperature deposition process. The deformation behaviours and fracture mechanism of polymer-based biomaterials were investigated using the compression test and the finite element (FE) simulation. The results show that the stress-strain curve of compression process includes linear elastic stage I, platform stage II and densification stage III, and the fracture mechanism can be considered as brittle fracture.

  20. DNA unwinding by ASCC3 helicase is coupled to ALKBH3 dependent DNA alkylation repair and cancer cell proliferation

    Dango, Sebastian; Mosammaparast, Nima; Sowa, Mathew E.; Xiong, Li-Jun; Wu, Feizhen; Park, Keyjung; Rubin, Mark; Gygi, Steve; Harper, J. Wade; Shi, Yang

    2011-01-01

    Summary Demethylation by the AlkB dioxygenases represents an important mechanism for repair of N-alkylated nucleotides. However, little is known about their functions in mammalian cells. We report the purification of the ALKBH3 complex and demonstrate its association with the Activating Signal Co-integrator Complex (ASCC). ALKBH3 is overexpressed in various cancers, and both ALKBH3 and ASCC are important for alkylation damage resistance in these tumor cell lines. ASCC3, the largest subunit of ASCC, encodes a 3′-5′ DNA helicase, whose activity is crucial for the generation of single-stranded DNA upon which ALKBH3 preferentially functions for dealkylation. In cell lines that are dependent on ALKBH3 and ASCC3 for alkylation damage resistance, loss of ALKBH3 or ASCC3 leads to increased 3-methylcytosine and reduced cell proliferation, which correlates with pH2A.X and 53BP1 foci formation. Our data provide a molecular mechanism by which ALKBH3 collaborates with ASCC to maintain genomic integrity in a cell type specific manner. PMID:22055184

  1. DHX9 helicase is involved in preventing genomic instability induced by alternatively structured DNA in human cells.

    Jain, Aklank; Bacolla, Albino; Del Mundo, Imee M; Zhao, Junhua; Wang, Guliang; Vasquez, Karen M

    2013-12-01

    Sequences that have the capacity to adopt alternative (i.e. non-B) DNA structures in the human genome have been implicated in stimulating genomic instability. Previously, we found that a naturally occurring intra-molecular triplex (H-DNA) caused genetic instability in mammals largely in the form of DNA double-strand breaks. Thus, it is of interest to determine the mechanism(s) involved in processing H-DNA. Recently, we demonstrated that human DHX9 helicase preferentially unwinds inter-molecular triplex DNA in vitro. Herein, we used a mutation-reporter system containing H-DNA to examine the relevance of DHX9 activity on naturally occurring H-DNA structures in human cells. We found that H-DNA significantly increased mutagenesis in small-interfering siRNA-treated, DHX9-depleted cells, affecting mostly deletions. Moreover, DHX9 associated with H-DNA in the context of supercoiled plasmids. To further investigate the role of DHX9 in the recognition/processing of H-DNA, we performed binding assays in vitro and chromatin immunoprecipitation assays in U2OS cells. DHX9 recognized H-DNA, as evidenced by its binding to the H-DNA structure and enrichment at the H-DNA region compared with a control region in human cells. These composite data implicate DHX9 in processing H-DNA structures in vivo and support its role in the overall maintenance of genomic stability at sites of alternatively structured DNA.

  2. Mycobacterium smegmatis SftH exemplifies a distinctive clade of superfamily II DNA-dependent ATPases with 3′ to 5′ translocase and helicase activities

    Yakovleva, Lyudmila; Shuman, Stewart

    2012-01-01

    Bacterial DNA helicases are nucleic acid-dependent NTPases that play important roles in DNA replication, recombination and repair. We are interested in the DNA helicases of Mycobacteria, a genus of the phylum Actinobacteria, which includes the human pathogen Mycobacterium tuberculosis and its avirulent relative Mycobacterium smegmatis. Here, we identify and characterize M. smegmatis SftH, a superfamily II helicase with a distinctive domain structure, comprising an N-terminal NTPase domain and...

  3. A Container-based Trusted Multi-level Security Mechanism

    Li Xiao-Yong

    2017-01-01

    Full Text Available Multi-level security mechanism has been widely applied in the military, government, defense and other domains in which information is required to be divided by security-level. Through this type of security mechanism, users at different security levels are provided with information at corresponding security levels. Traditional multi-level security mechanism which depends on the safety of operating system finally proved to be not practical. We propose a container-based trusted multi-level security mechanism in this paper to improve the applicability of the multi-level mechanism. It guarantees multi-level security of the system through a set of multi-level security policy rules and trusted techniques. The technical feasibility and application scenarios are also discussed. The ease of realization, strong practical significance and low cost of our method will largely expand the application of multi-level security mechanism in real life.

  4. An improved anti-leech mechanism based on session identifier

    Zhang, Jianbiao; Zhu, Tong; Zhang, Han; Lin, Li

    2012-01-01

    With the rapid development of information technology and extensive requirement of network resource sharing, plenty of resource hotlinking phenomenons appear on the internet. The hotlinking problem not only harms the interests of legal websites but also leads to a great affection to fair internet environment. The anti-leech technique based on session identifier is highly secure, but the transmission of session identifier in plaintext form causes some security flaws. In this paper, a proxy hotlinking technique based on session identifier is introduced firstly to illustrate these security flaws; next, this paper proposes an improved anti-leech mechanism based on session identifier, the mechanism takes the random factor as the core and detects hotlinking request using a map table that contains random factor, user's information and time stamp; at last the paper analyzes the security of mechanism in theory. The result reveals that the improved mechanism has the merits of simple realization, high security and great flexibility.

  5. hSSB1 associates with and promotes stability of the BLM helicase

    O'BYRNE, KEN

    2017-01-01

    Background Maintenance of genome stability is critical in human cells. Mutations in or loss of genome stability pathways can lead to a number of pathologies including cancer. hSSB1 is a critical DNA repair protein functioning in the repair and signalling of stalled DNA replication forks, double strand DNA breaks and oxidised DNA lesions. The BLM helicase is central to the repair of both collapsed DNA replication forks and double strand DNA breaks by homologous recombination. Results In this s...

  6. The human RecQ helicases BLM and RECQL4 cooperate to preserve genome stability

    Singh, D.K.; Popuri, V.; Kulikowicz, T.; Shevelev, Igor; Ghosh, A.K.; Ramamoorthy, M.; Rossi, M.L.; Janščák, Pavel; Croteau, D.L.; Bohr, V.A.

    2012-01-01

    Roč. 40, č. 14 (2012), s. 6632-6648 ISSN 0305-1048 R&D Projects: GA ČR GAP305/10/0281 Grant - others:NIH(US) Z01-AG000726-17 Institutional research plan: CEZ:AV0Z50520514 Institutional support: RVO:68378050 Keywords : RecQ helicase * genome stability * BLM * RECQL4 Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 8.278, year: 2012

  7. A Co-Opted DEAD-Box RNA helicase enhances tombusvirus plus-strand synthesis.

    Nikolay Kovalev

    2012-02-01

    Full Text Available Replication of plus-strand RNA viruses depends on recruited host factors that aid several critical steps during replication. In this paper, we show that an essential translation factor, Ded1p DEAD-box RNA helicase of yeast, directly affects replication of Tomato bushy stunt virus (TBSV. To separate the role of Ded1p in viral protein translation from its putative replication function, we utilized a cell-free TBSV replication assay and recombinant Ded1p. The in vitro data show that Ded1p plays a role in enhancing plus-strand synthesis by the viral replicase. We also find that Ded1p is a component of the tombusvirus replicase complex and Ded1p binds to the 3'-end of the viral minus-stranded RNA. The data obtained with wt and ATPase deficient Ded1p mutants support the model that Ded1p unwinds local structures at the 3'-end of the TBSV (-RNA, rendering the RNA compatible for initiation of (+-strand synthesis. Interestingly, we find that Ded1p and glyceraldehyde-3-phosphate dehydrogenase (GAPDH, which is another host factor for TBSV, play non-overlapping functions to enhance (+-strand synthesis. Altogether, the two host factors enhance TBSV replication synergistically by interacting with the viral (-RNA and the replication proteins. In addition, we have developed an in vitro assay for Flock house virus (FHV, a small RNA virus of insects, that also demonstrated positive effect on FHV replicase activity by the added Ded1p helicase. Thus, two small RNA viruses, which do not code for their own helicases, seems to recruit a host RNA helicase to aid their replication in infected cells.

  8. The eIF4AIII RNA helicase is a critical determinant of human cytomegalovirus replication

    Ziehr, Ben; Lenarcic, Erik; Cecil, Chad; Moorman, Nathaniel J.

    2016-01-01

    Human cytomegalovirus (HCMV) was recently shown to encode a large number of spliced mRNAs. While the nuclear export of unspliced viral transcripts has been extensively studied, the role of host mRNA export factors in HCMV mRNA trafficking remains poorly defined. We found that the eIF4AIII RNA helicase, a component of the exon junction complex, was necessary for efficient virus replication. Depletion of eIF4AIII limited viral DNA accumulation, export of viral mRNAs from the nucleus, and the production of progeny virus. However eIF4AIII was dispensable for the association of viral transcripts with ribosomes. We found that pateamine A, a natural compound that inhibits both eIF4AI/II and eIF4AIII, has potent antiviral activity and inhibits HCMV replication throughout the virus lytic cycle. Our results demonstrate that eIF4AIII is required for efficient HCMV replication, and suggest that eIF4A family helicases may be a new class of targets for the development of host-directed antiviral therapeutics. - Highlights: • The host eIF4AIII RNA helicase is required for efficient HCMV replication. • Depleting eIF4AIII inhibited the nuclear export of HCMV mRNAs. • HCMV mRNAs did not require eIF4AIII to associate with polyribosomes. • The eIF4A family helicases may be new targets for host-directed antiviral drugs.

  9. Physical interaction of RECQ5 helicase with RAD51 facilitates its anti-recombinase activity

    Schwendener, S.; Raynard, S.; Paliwal, S.; Cheng, A.; Kanagaraj, R.; Shevelev, Igor; Stark, J.M.; Sung, P.; Janscak, P.

    2010-01-01

    Roč. 285, č. 21 (2010), s. 15739-15745 ISSN 0021-9258 Grant - others:NIH(US) R01CA120954; NIH(US) ES015632; SNSF(CH) 3100A0-116008 Institutional research plan: CEZ:AV0Z50520514 Keywords : DNA helicase * double-strand breaks * homologous recombination Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.328, year: 2010

  10. DEAD-box RNA helicase is dispensable for mitochondrial translation in Trypanosoma brucei

    Richterová, Lenka; Vávrová, Zuzana; Lukeš, Julius

    2011-01-01

    Roč. 127, č. 1 (2011), 300-303 ISSN 0014-4894 R&D Projects: GA ČR GA204/09/1667; GA MŠk LC07032; GA MŠk 2B06129 Institutional research plan: CEZ:AV0Z60220518 Keywords : Trypanosoma * Mitochondrial translation * RNA helicase * Cytochrome c oxidase * Mitochondrion Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.122, year: 2011

  11. The eIF4AIII RNA helicase is a critical determinant of human cytomegalovirus replication

    Ziehr, Ben; Lenarcic, Erik; Cecil, Chad; Moorman, Nathaniel J., E-mail: nmoorman@med.unc.edu

    2016-02-15

    Human cytomegalovirus (HCMV) was recently shown to encode a large number of spliced mRNAs. While the nuclear export of unspliced viral transcripts has been extensively studied, the role of host mRNA export factors in HCMV mRNA trafficking remains poorly defined. We found that the eIF4AIII RNA helicase, a component of the exon junction complex, was necessary for efficient virus replication. Depletion of eIF4AIII limited viral DNA accumulation, export of viral mRNAs from the nucleus, and the production of progeny virus. However eIF4AIII was dispensable for the association of viral transcripts with ribosomes. We found that pateamine A, a natural compound that inhibits both eIF4AI/II and eIF4AIII, has potent antiviral activity and inhibits HCMV replication throughout the virus lytic cycle. Our results demonstrate that eIF4AIII is required for efficient HCMV replication, and suggest that eIF4A family helicases may be a new class of targets for the development of host-directed antiviral therapeutics. - Highlights: • The host eIF4AIII RNA helicase is required for efficient HCMV replication. • Depleting eIF4AIII inhibited the nuclear export of HCMV mRNAs. • HCMV mRNAs did not require eIF4AIII to associate with polyribosomes. • The eIF4A family helicases may be new targets for host-directed antiviral drugs.

  12. Distinct functions of human RecQ helicases during DNA replication

    Urban, Václav; Dobrovolná, Jana; Janščák, Pavel

    2017-01-01

    Roč. 225, červen (2017), s. 20-26 ISSN 0301-4622 R&D Projects: GA ČR(CZ) GA14-05743S; GA MŠk LH14037 Institutional support: RVO:68378050 Keywords : DNA replication * Replication stress * RecQ helicases * Genomic instability * Cancer Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Biochemistry and molecular biology Impact factor: 2.402, year: 2016

  13. Relocalization of nuclear DNA helicase II during the growth period of bovine oocytes

    Baran, V.; Kovářová, Hana; Klíma, Jiří; Hozák, Pavel; Motlík, Jan

    2006-01-01

    Roč. 125, 1-2 (2006), s. 155-164 ISSN 0948-6143 R&D Projects: GA ČR GA523/03/0857 Grant - others:Slovenská Akademie věd(SK) VEGA 2/3065/23 Institutional research plan: CEZ:AV0Z50450515; CEZ:AV0Z50390512 Keywords : DNA helicase II * fibroblasts * oocytes Subject RIV: EB - Genetics ; Molecular Biology Impact factor : 3.220, year: 2006

  14. Senataxin, the ortholog of a yeast RNA helicase, is mutant in ataxia-ocular apraxia 2.

    Moreira, Maria-Céu; Klur, Sandra; Watanabe, Mitsunori; Németh, Andrea H; Le Ber, Isabelle; Moniz, José-Carlos; Tranchant, Christine; Aubourg, Patrick; Tazir, Meriem; Schöls, Lüdger; Pandolfo, Massimo; Schulz, Jörg B; Pouget, Jean; Calvas, Patrick; Shizuka-Ikeda, Masami; Shoji, Mikio; Tanaka, Makoto; Izatt, Louise; Shaw, Christopher E; M'Zahem, Abderrahim; Dunne, Eimear; Bomont, Pascale; Benhassine, Traki; Bouslam, Naïma; Stevanin, Giovanni; Brice, Alexis; Guimarães, João; Mendonça, Pedro; Barbot, Clara; Coutinho, Paula; Sequeiros, Jorge; Dürr, Alexandra; Warter, Jean-Marie; Koenig, Michel

    2004-03-01

    Ataxia-ocular apraxia 2 (AOA2) was recently identified as a new autosomal recessive ataxia. We have now identified causative mutations in 15 families, which allows us to clinically define this entity by onset between 10 and 22 years, cerebellar atrophy, axonal sensorimotor neuropathy, oculomotor apraxia and elevated alpha-fetoprotein (AFP). Ten of the fifteen mutations cause premature termination of a large DEAxQ-box helicase, the human ortholog of yeast Sen1p, involved in RNA maturation and termination.

  15. RTEL1: an essential helicase for telomere maintenance and the regulation of homologous recombination

    Uringa, Evert-Jan; Youds, Jillian L.; Lisaingo, Kathleen; Lansdorp, Peter M.; Boulton, Simon J.

    2010-01-01

    Telomere maintenance and DNA repair are crucial processes that protect the genome against instability. RTEL1, an essential iron–sulfur cluster-containing helicase, is a dominant factor that controls telomere length in mice and is required for telomere integrity. In addition, RTEL1 promotes synthesis-dependent strand annealing to direct DNA double-strand breaks into non-crossover outcomes during mitotic repair and in meiosis. Here, we review the role of RTEL1 in telomere maintenance and homolo...

  16. Archaeal MCM Proteins as an Analog for the Eukaryotic Mcm2–7 Helicase to Reveal Essential Features of Structure and Function

    Miller, Justin M.; Enemark, Eric J.

    2015-01-01

    In eukaryotes, the replicative helicase is the large multisubunit CMG complex consisting of the Mcm2–7 hexameric ring, Cdc45, and the tetrameric GINS complex. The Mcm2–7 ring assembles from six different, related proteins and forms the core of this complex. In archaea, a homologous MCM hexameric ring functions as the replicative helicase at the replication fork. Archaeal MCM proteins form thermostable homohexamers, facilitating their use as models of the eukaryotic Mcm2–7 helicase. Here we review archaeal MCM helicase structure and function and how the archaeal findings relate to the eukaryotic Mcm2–7 ring. PMID:26539061

  17. Escherichia coli and Neisseria gonorrhoeae UvrD helicase unwinds G4 DNA structures.

    Shukla, Kaustubh; Thakur, Roshan Singh; Ganguli, Debayan; Rao, Desirazu Narasimha; Nagaraju, Ganesh

    2017-10-18

    G-quadruplex (G4) secondary structures have been implicated in various biological processes, including gene expression, DNA replication and telomere maintenance. However, unresolved G4 structures impede replication progression which can lead to the generation of DNA double-strand breaks and genome instability. Helicases have been shown to resolve G4 structures to facilitate faithful duplication of the genome. Escherichia coli UvrD (EcUvrD) helicase plays a crucial role in nucleotide excision repair, mismatch repair and in the regulation of homologous recombination. Here, we demonstrate a novel role of E. coli and Neisseria gonorrhoeae UvrD in resolving G4 tetraplexes. EcUvrD and N gonorrhoeae UvrD were proficient in unwinding previously characterized tetramolecular G4 structures. Notably, EcUvrD was equally efficient in resolving tetramolecular and bimolecular G4 DNA that were derived from the potential G4-forming sequences from the genome of E. coli Interestingly, in addition to resolving intermolecular G4 structures, EcUvrD was robust in unwinding intramolecular G4 structures. These data for the first time provide evidence for the role of UvrD in the resolution of G4 structures, which has implications for the in vivo role of UvrD helicase in G4 DNA resolution and genome maintenance. © 2017 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

  18. The helicase domain of Polθ counteracts RPA to promote alt-NHEJ.

    Mateos-Gomez, Pedro A; Kent, Tatiana; Deng, Sarah K; McDevitt, Shane; Kashkina, Ekaterina; Hoang, Trung M; Pomerantz, Richard T; Sfeir, Agnel

    2017-12-01

    Mammalian polymerase theta (Polθ) is a multifunctional enzyme that promotes error-prone DNA repair by alternative nonhomologous end joining (alt-NHEJ). Here we present structure-function analyses that reveal that, in addition to the polymerase domain, Polθ-helicase activity plays a central role during double-strand break (DSB) repair. Our results show that the helicase domain promotes chromosomal translocations by alt-NHEJ in mouse embryonic stem cells and also suppresses CRISPR-Cas9- mediated gene targeting by homologous recombination (HR). In vitro assays demonstrate that Polθ-helicase activity facilitates the removal of RPA from resected DSBs to allow their annealing and subsequent joining by alt-NHEJ. Consistent with an antagonistic role for RPA during alt-NHEJ, inhibition of RPA1 enhances end joining and suppresses recombination. Taken together, our results reveal that the balance between HR and alt-NHEJ is controlled by opposing activities of Polθ and RPA, providing further insight into the regulation of repair-pathway choice in mammalian cells.

  19. Non-B DNA Secondary Structures and Their Resolution by RecQ Helicases

    Sudha Sharma

    2011-01-01

    Full Text Available In addition to the canonical B-form structure first described by Watson and Crick, DNA can adopt a number of alternative structures. These non-B-form DNA secondary structures form spontaneously on tracts of repeat sequences that are abundant in genomes. In addition, structured forms of DNA with intrastrand pairing may arise on single-stranded DNA produced transiently during various cellular processes. Such secondary structures have a range of biological functions but also induce genetic instability. Increasing evidence suggests that genomic instabilities induced by non-B DNA secondary structures result in predisposition to diseases. Secondary DNA structures also represent a new class of molecular targets for DNA-interactive compounds that might be useful for targeting telomeres and transcriptional control. The equilibrium between the duplex DNA and formation of multistranded non-B-form structures is partly dependent upon the helicases that unwind (resolve these alternate DNA structures. With special focus on tetraplex, triplex, and cruciform, this paper summarizes the incidence of non-B DNA structures and their association with genomic instability and emphasizes the roles of RecQ-like DNA helicases in genome maintenance by resolution of DNA secondary structures. In future, RecQ helicases are anticipated to be additional molecular targets for cancer chemotherapeutics.

  20. Helicase properties of the Escherichia coli UvrAb protein complex

    Oh, E.Y.; Grossman, L.

    1987-01-01

    The Escherichia coli UvrA protein has an associated ATPase activity with a turnover number affected by the presence of UvrB protein as well as by DNA. Specifically, the structure of DNA significantly influences the turnover rate of the UvrAB ATPase activity. Double-stranded DNA maximally activates the turnover rate 10-fold whereas single-stranded DNA maximally activates the turnover rate 20-fold, suggesting that the mode of interaction of UvrAB protein with different DNAs is distinctive. We have previously shown that the UvrAB protein complex, driven by the binding energy of ATP, can locally unwind supercoiled DNA. The nature of the DNA unwinding activity and single-stranded DNA activation of ATPase activity suggest potential helicase activity. In the presence of a number of helicase substrates, the UvrAB complex, indeed, manifests a strand-displacement activity-unwinding short duplexes and D-loop DNA, thereby generating component DNA structures. The energy for the activity is derived from ATP or dATP hydrolysis. Unlike the E. coli DnaB, the UvrAB helicase is sensitive to UV-induced photoproducts

  1. Isolation and Characterization of Pepper Genes Interacting with the CMV-P1 Helicase Domain.

    Yoomi Choi

    Full Text Available Cucumber mosaic virus (CMV is a destructive pathogen affecting Capsicum annuum (pepper production. The pepper Cmr1 gene confers resistance to most CMV strains, but is overcome by CMV-P1 in a process dependent on the CMV-P1 RNA1 helicase domain (P1 helicase. Here, to identify host factors involved in CMV-P1 infection in pepper, a yeast two-hybrid library derived from a C. annuum 'Bukang' cDNA library was screened, producing a total of 76 potential clones interacting with the P1 helicase. Beta-galactosidase filter lift assay, PCR screening, and sequencing analysis narrowed the candidates to 10 genes putatively involved in virus infection. The candidate host genes were silenced in Nicotiana benthamiana plants that were then inoculated with CMV-P1 tagged with the green fluorescent protein (GFP. Plants silenced for seven of the genes showed development comparable to N. benthamiana wild type, whereas plants silenced for the other three genes showed developmental defects including stunting and severe distortion. Silencing formate dehydrogenase and calreticulin-3 precursor led to reduced virus accumulation. Formate dehydrogenase-silenced plants showed local infection in inoculated leaves, but not in upper (systemic leaves. In the calreticulin-3 precursor-silenced plants, infection was not observed in either the inoculated or the upper leaves. Our results demonstrate that formate dehydrogenase and calreticulin-3 precursor are required for CMV-P1 infection.

  2. Staphylococcal SCCmec elements encode an active MCM-like helicase and thus may be replicative

    Mir-Sanchis, Ignacio; Roman, Christina A.; Misiura, Agnieszka; Pigli, Ying Z.; Boyle-Vavra, Susan; Rice , Phoebe A. (UC)

    2016-08-29

    Methicillin-resistant Staphylococcus aureus (MRSA) is a public-health threat worldwide. Although the mobile genomic island responsible for this phenotype, staphylococcal cassette chromosome (SCC), has been thought to be nonreplicative, we predicted DNA-replication-related functions for some of the conserved proteins encoded by SCC. We show that one of these, Cch, is homologous to the self-loading initiator helicases of an unrelated family of genomic islands, that it is an active 3'-to-5' helicase and that the adjacent ORF encodes a single-stranded DNA–binding protein. Our 2.9-Å crystal structure of intact Cch shows that it forms a hexameric ring. Cch, like the archaeal and eukaryotic MCM-family replicative helicases, belongs to the pre–sensor II insert clade of AAA+ ATPases. Additionally, we found that SCC elements are part of a broader family of mobile elements, all of which encode a replication initiator upstream of their recombinases. Replication after excision would enhance the efficiency of horizontal gene transfer.

  3. The Arabidopsis thaliana homolog of the helicase RTEL1 plays multiple roles in preserving genome stability.

    Recker, Julia; Knoll, Alexander; Puchta, Holger

    2014-12-01

    In humans, mutations in the DNA helicase Regulator of Telomere Elongation Helicase1 (RTEL1) lead to Hoyeraal-Hreidarsson syndrome, a severe, multisystem disorder. Here, we demonstrate that the RTEL1 homolog in Arabidopsis thaliana plays multiple roles in preserving genome stability. RTEL1 suppresses homologous recombination in a pathway parallel to that of the DNA translocase FANCM. Cytological analyses of root meristems indicate that RTEL1 is involved in processing DNA replication intermediates independently from FANCM and the nuclease MUS81. Moreover, RTEL1 is involved in interstrand and intrastrand DNA cross-link repair independently from FANCM and (in intrastrand cross-link repair) parallel to MUS81. RTEL1 contributes to telomere homeostasis; the concurrent loss of RTEL1 and the telomerase TERT leads to rapid, severe telomere shortening, which occurs much more rapidly than it does in the single-mutant line tert, resulting in developmental arrest after four generations. The double mutant rtel1-1 recq4A-4 exhibits massive growth defects, indicating that this RecQ family helicase, which is also involved in the suppression of homologous recombination and the repair of DNA lesions, can partially replace RTEL1 in the processing of DNA intermediates. The requirement for RTEL1 in multiple pathways to preserve genome stability in plants can be explained by its putative role in the destabilization of DNA loop structures, such as D-loops and T-loops. © 2014 American Society of Plant Biologists. All rights reserved.

  4. A new role for FBP21 as regulator of Brr2 helicase activity.

    Henning, Lisa M; Santos, Karine F; Sticht, Jana; Jehle, Stefanie; Lee, Chung-Tien; Wittwer, Malte; Urlaub, Henning; Stelzl, Ulrich; Wahl, Markus C; Freund, Christian

    2017-07-27

    Splicing of eukaryotic pre-mRNA is carried out by the spliceosome, which assembles stepwise on each splicing substrate. This requires the concerted action of snRNPs and non-snRNP accessory proteins, the functions of which are often not well understood. Of special interest are B complex factors that enter the spliceosome prior to catalytic activation and may alter splicing kinetics and splice site selection. One of these proteins is FBP21, for which we identified several spliceosomal binding partners in a yeast-two-hybrid screen, among them the RNA helicase Brr2. Biochemical and biophysical analyses revealed that an intrinsically disordered region of FBP21 binds to an extended surface of the C-terminal Sec63 unit of Brr2. Additional contacts in the C-terminal helicase cassette are required for allosteric inhibition of Brr2 helicase activity. Furthermore, the direct interaction between FBP21 and the U4/U6 di-snRNA was found to reduce the pool of unwound U4/U6 di-snRNA. Our results suggest FBP21 as a novel key player in the regulation of Brr2. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  5. Mechanical Design of Downhole Tractor Based on Two-Way Self-locking Mechanism

    Fang, Delei; Shang, Jianzhong; Luo, Zirong; Wu, Guoheng; Liu, Yiying

    2018-03-01

    Based on the technology of horizontal well tractor, a kind of downhole tractor was developed which can realize Two-Way self-locking function. Aiming at the needs of horizontal well logging to realize the target of small size, high traction and high reliability, the tractor selects unique heart-shaped CAM as the locking mechanism. The motion principle of telescopic downhole tractor, the design of mechanical structure and locking principle of the locking mechanism are all analyzed. The mathematical expressions of traction are obtained by mechanical analysis of parallel support rod in the locking mechanism. The force analysis and contour design of the heart-shaped CAM are performed, which can lay the foundation for the development of tractor prototype.

  6. Perceived benefits of adopting Standard – Based pricing mechanism for mechanical and electrical services installations

    Ganiyu Amuda Yusuf; Sarajul Fikri Mohamed

    2014-01-01

    Cost is an important measure of project success and clients will expect a reliable forecast at the early stage of construction projects to inform their business decision. This study was undertaken to investigate the current practices in managing cost of mechanical and electrical (M&E) services in buildings. The perceptions of practitioners on the benefits of adopting Standard – Based Pricing Mechanism for M&E services as used for building fabrics and finishes was ascertained. The meth...

  7. Shape memory-based actuators and release mechanisms therefrom

    Vaidyanathan, Rajan (Inventor); Snyder, Daniel W. (Inventor); Schoenwald, David K. (Inventor); Lam, Nhin S. (Inventor); Watson, Daniel S. (Inventor); Krishnan, Vinu B. (Inventor); Noebe, Ronald D. (Inventor)

    2012-01-01

    SM-based actuators (110) and release mechanisms (100) therefrom and systems (500) including one or more release mechanisms (100). The actuators (110) comprise a SM member (118) and a deformable member (140) mechanically coupled to the SM member (118) which deforms upon a shape change of the SM member triggered by a phase transition of the SM member. A retaining element (160) is mechanically coupled to the deformable member (140), wherein the retaining element (160) moves upon the shape change. Release mechanism (100) include an actuator, a rotatable mechanism (120) including at least one restraining feature (178) for restraining rotational movement of the retaining element (160) before the shape change, and at least one spring (315) that provides at least one locked spring-loaded position when the retaining element is in the restraining feature and at least one released position that is reached when the retaining element is in a position beyond the restraining feature (178). The rotatable mechanism (120) includes at least one load-bearing protrusion (310). A hitch (400) is for mechanically coupling to the load, wherein the hitch is supported on the load bearing protrusion (310) when the rotatable mechanism is in the locked spring-loaded position.

  8. Project-Based Laboratory Experiences in Mechanical Engineering

    Narendra Sharma

    2011-12-01

    Full Text Available In this paper we describe project-based laboratories in Mechanical Engineering designed to provide semester-long team experiences which mimic the real life industrial processes of design, development, testing and optimization. The labs are focused on courses at the sophomore level and thus require special attention to constraints of student backgrounds and experience. This paper describes laboratory projects in Dynamics and Fluid Mechanics.

  9. Mechanical properties of soldered joints of niobium base alloys

    Grishin, V.L.

    1980-01-01

    Mechanical properties of soldered joints of niobium alloys widely distributed in industry: VN3, VN4, VN5A, VN5AE, VN5AEP etc., 0.6-1.2 mm thick are investigated. It is found out that the usage of zirconium-vanadium, titanium-tantalum solders for welding niobium base alloys permits to obtain soldered joints with satisfactory mechanical properties at elevated temperatures

  10. X-ray structure of the pestivirus NS3 helicase and its conformation in solution.

    Tortorici, M Alejandra; Duquerroy, Stéphane; Kwok, Jane; Vonrhein, Clemens; Perez, Javier; Lamp, Benjamin; Bricogne, Gerard; Rümenapf, Till; Vachette, Patrice; Rey, Félix A

    2015-04-01

    Pestiviruses form a genus in the Flaviviridae family of small enveloped viruses with a positive-sense single-stranded RNA genome. Viral replication in this family requires the activity of a superfamily 2 RNA helicase contained in the C-terminal domain of nonstructural protein 3 (NS3). NS3 features two conserved RecA-like domains (D1 and D2) with ATPase activity, plus a third domain (D3) that is important for unwinding nucleic acid duplexes. We report here the X-ray structure of the pestivirus NS3 helicase domain (pNS3h) at a 2.5-Å resolution. The structure deviates significantly from that of NS3 of other genera in the Flaviviridae family in D3, as it contains two important insertions that result in a narrower nucleic acid binding groove. We also show that mutations in pNS3h that rescue viruses from which the core protein is deleted map to D3, suggesting that this domain may be involved in interactions that facilitate particle assembly. Finally, structural comparisons of the enzyme in different crystalline environments, together with the findings of small-angle X-ray-scattering studies in solution, show that D2 is mobile with respect to the rest of the enzyme, oscillating between closed and open conformations. Binding of a nonhydrolyzable ATP analog locks pNS3h in a conformation that is more compact than the closest apo-form in our crystals. Together, our results provide new insight and bring up new questions about pNS3h function during pestivirus replication. Although pestivirus infections impose an important toll on the livestock industry worldwide, little information is available about the nonstructural proteins essential for viral replication, such as the NS3 helicase. We provide here a comparative structural and functional analysis of pNS3h with respect to its orthologs in other viruses of the same family, the flaviviruses and hepatitis C virus. Our studies reveal differences in the nucleic acid binding groove that could have implications for understanding the

  11. Symmetry-based reciprocity: evolutionary constraints on a proximate mechanism.

    Campennì, Marco; Schino, Gabriele

    2016-01-01

    Background. While the evolution of reciprocal cooperation has attracted an enormous attention, the proximate mechanisms underlying the ability of animals to cooperate reciprocally are comparatively neglected. Symmetry-based reciprocity is a hypothetical proximate mechanism that has been suggested to be widespread among cognitively unsophisticated animals. Methods. We developed two agent-based models of symmetry-based reciprocity (one relying on an arbitrary tag and the other on interindividual proximity) and tested their ability both to reproduce significant emergent features of cooperation in group living animals and to promote the evolution of cooperation. Results. Populations formed by agents adopting symmetry-based reciprocity showed differentiated "social relationships" and a positive correlation between cooperation given and received: two common aspects of animal cooperation. However, when reproduction and selection across multiple generations were added to the models, agents adopting symmetry-based reciprocity were outcompeted by selfish agents that never cooperated. Discussion. In order to evolve, hypothetical proximate mechanisms must be able to stand competition from alternative strategies. While the results of our simulations require confirmation using analytical methods, we provisionally suggest symmetry-based reciprocity is to be abandoned as a possible proximate mechanism underlying the ability of animals to reciprocate cooperative interactions.

  12. Genome-wide comparative in silico analysis of the RNA helicase gene family in Zea mays and Glycine max: a comparison with Arabidopsis and Oryza sativa.

    Xu, Ruirui; Zhang, Shizhong; Huang, Jinguang; Zheng, Chengchao

    2013-01-01

    RNA helicases are enzymes that are thought to unwind double-stranded RNA molecules in an energy-dependent fashion through the hydrolysis of NTP. RNA helicases are associated with all processes involving RNA molecules, including nuclear transcription, editing, splicing, ribosome biogenesis, RNA export, and organelle gene expression. The involvement of RNA helicase in response to stress and in plant growth and development has been reported previously. While their importance in Arabidopsis and Oryza sativa has been partially studied, the function of RNA helicase proteins is poorly understood in Zea mays and Glycine max. In this study, we identified a total of RNA helicase genes in Arabidopsis and other crop species genome by genome-wide comparative in silico analysis. We classified the RNA helicase genes into three subfamilies according to the structural features of the motif II region, such as DEAD-box, DEAH-box and DExD/H-box, and different species showed different patterns of alternative splicing. Secondly, chromosome location analysis showed that the RNA helicase protein genes were distributed across all chromosomes with different densities in the four species. Thirdly, phylogenetic tree analyses identified the relevant homologs of DEAD-box, DEAH-box and DExD/H-box RNA helicase proteins in each of the four species. Fourthly, microarray expression data showed that many of these predicted RNA helicase genes were expressed in different developmental stages and different tissues under normal growth conditions. Finally, real-time quantitative PCR analysis showed that the expression levels of 10 genes in Arabidopsis and 13 genes in Zea mays were in close agreement with the microarray expression data. To our knowledge, this is the first report of a comparative genome-wide analysis of the RNA helicase gene family in Arabidopsis, Oryza sativa, Zea mays and Glycine max. This study provides valuable information for understanding the classification and putative functions of

  13. EM structure of a helicase-loader complex depicting a 6:2 binding sub-stoichiometry from Geobacillus kaustophilus HTA426

    Lin, Yen-Chen [Institute of Molecular Biology, Academia Sinica, Taipei 115, Taiwan (China); Naveen, Vankadari [Institute of Molecular Biology, Academia Sinica, Taipei 115, Taiwan (China); Molecular Cell Biology, Taiwan International Graduate Program, Institute of Molecular Biology, Academia Sinica, and Graduate Institute of Life Sciences, National Defense Medical Center, Taipei, Taiwan (China); Hsiao, Chwan-Deng, E-mail: hsiao@gate.sinica.edu.tw [Institute of Molecular Biology, Academia Sinica, Taipei 115, Taiwan (China); Molecular Cell Biology, Taiwan International Graduate Program, Institute of Molecular Biology, Academia Sinica, and Graduate Institute of Life Sciences, National Defense Medical Center, Taipei, Taiwan (China)

    2016-04-22

    During DNA replication, bacterial helicase is recruited as a complex in association with loader proteins to unwind the parental duplex. Previous structural studies have reported saturated 6:6 helicase-loader complexes with different conformations. However, structural information on the sub-stoichiometric conformations of these previously-documented helicase-loader complexes remains elusive. Here, with the aid of single particle electron-microscopy (EM) image reconstruction, we present the Geobacillus kaustophilus HTA426 helicase-loader (DnaC-DnaI) complex with a 6:2 binding stoichiometry in the presence of ATPγS. In the 19 Å resolution EM map, the undistorted and unopened helicase ring holds a robust loader density above the C-terminal RecA-like domain. Meanwhile, the path of the central DNA binding channel appears to be obstructed by the reconstructed loader density, implying its potential role as a checkpoint conformation to prevent the loading of immature complex onto DNA. Our data also reveals that the bound nucleotides and the consequently induced conformational changes in the helicase hexamer are essential for active association with loader proteins. These observations provide fundamental insights into the formation of the helicase-loader complex in bacteria that regulates the DNA replication process. - Highlights: • Helicase-loader complex structure with 6:2 sub-stoichiometry is resolved by EM. • Helicase hexamer in 6:2 sub-stoichiometry is constricted and un-opened. • 6:2 binding ratio of helicase-loader complex could act as a DNA loading checkpoint. • Nucleotides stabilize helicase-loader complex at low protein concentrations.

  14. EM structure of a helicase-loader complex depicting a 6:2 binding sub-stoichiometry from Geobacillus kaustophilus HTA426

    Lin, Yen-Chen; Naveen, Vankadari; Hsiao, Chwan-Deng

    2016-01-01

    During DNA replication, bacterial helicase is recruited as a complex in association with loader proteins to unwind the parental duplex. Previous structural studies have reported saturated 6:6 helicase-loader complexes with different conformations. However, structural information on the sub-stoichiometric conformations of these previously-documented helicase-loader complexes remains elusive. Here, with the aid of single particle electron-microscopy (EM) image reconstruction, we present the Geobacillus kaustophilus HTA426 helicase-loader (DnaC-DnaI) complex with a 6:2 binding stoichiometry in the presence of ATPγS. In the 19 Å resolution EM map, the undistorted and unopened helicase ring holds a robust loader density above the C-terminal RecA-like domain. Meanwhile, the path of the central DNA binding channel appears to be obstructed by the reconstructed loader density, implying its potential role as a checkpoint conformation to prevent the loading of immature complex onto DNA. Our data also reveals that the bound nucleotides and the consequently induced conformational changes in the helicase hexamer are essential for active association with loader proteins. These observations provide fundamental insights into the formation of the helicase-loader complex in bacteria that regulates the DNA replication process. - Highlights: • Helicase-loader complex structure with 6:2 sub-stoichiometry is resolved by EM. • Helicase hexamer in 6:2 sub-stoichiometry is constricted and un-opened. • 6:2 binding ratio of helicase-loader complex could act as a DNA loading checkpoint. • Nucleotides stabilize helicase-loader complex at low protein concentrations.

  15. Quantum control mechanism analysis through field based Hamiltonian encoding

    Mitra, Abhra; Rabitz, Herschel

    2006-01-01

    Optimal control of quantum dynamics in the laboratory is proving to be increasingly successful. The control fields can be complex, and the mechanisms by which they operate have often remained obscure. Hamiltonian encoding (HE) has been proposed as a method for understanding mechanisms in quantum dynamics. In this context mechanism is defined in terms of the dominant quantum pathways leading to the final state of the controlled system. HE operates by encoding a special modulation into the Hamiltonian and decoding its signature in the dynamics to determine the dominant pathway amplitudes. Earlier work encoded the modulation directly into the Hamiltonian operators. This present work introduces the alternative scheme of field based HE, where the modulation is encoded into the control field and not directly into the Hamiltonian operators. This distinct form of modulation yields a new perspective on mechanism and is computationally faster than the earlier approach. Field based encoding is also an important step towards a laboratory based algorithm for HE as it is the only form of encoding that may be experimentally executed. HE is also extended to cover systems with noise and uncertainty and finally, a hierarchical algorithm is introduced to reveal mechanism in a stepwise fashion of ever increasing detail as desired. This new hierarchical algorithm is an improvement over earlier approaches to HE where the entire mechanism was determined in one stroke. The improvement comes from the use of less complex modulation schemes, which leads to fewer evaluations of Schroedinger's equation. A number of simulations are presented on simple systems to illustrate the new field based encoding technique for mechanism assessment

  16. Comprehensive Protein Interactome Analysis of a Key RNA Helicase: Detection of Novel Stress Granule Proteins

    Rebecca Bish

    2015-07-01

    Full Text Available DDX6 (p54/RCK is a human RNA helicase with central roles in mRNA decay and translation repression. To help our understanding of how DDX6 performs these multiple functions, we conducted the first unbiased, large-scale study to map the DDX6-centric protein-protein interactome using immunoprecipitation and mass spectrometry. Using DDX6 as bait, we identify a high-confidence and high-quality set of protein interaction partners which are enriched for functions in RNA metabolism and ribosomal proteins. The screen is highly specific, maximizing the number of true positives, as demonstrated by the validation of 81% (47/58 of the RNA-independent interactors through known functions and interactions. Importantly, we minimize the number of indirect interaction partners through use of a nuclease-based digestion to eliminate RNA. We describe eleven new interactors, including proteins involved in splicing which is an as-yet unknown role for DDX6. We validated and characterized in more detail the interaction of DDX6 with Nuclear fragile X mental retardation-interacting protein 2 (NUFIP2 and with two previously uncharacterized proteins, FAM195A and FAM195B (here referred to as granulin-1 and granulin-2, or GRAN1 and GRAN2. We show that NUFIP2, GRAN1, and GRAN2 are not P-body components, but re-localize to stress granules upon exposure to stress, suggesting a function in translation repression in the cellular stress response. Using a complementary analysis that resolved DDX6’s multiple complex memberships, we further validated these interaction partners and the presence of splicing factors. As DDX6 also interacts with the E3 SUMO ligase TIF1β, we tested for and observed a significant enrichment of sumoylation amongst DDX6’s interaction partners. Our results represent the most comprehensive screen for direct interaction partners of a key regulator of RNA life cycle and localization, highlighting new stress granule components and possible DDX6 functions

  17. Interactive Roles of DNA Helicases and Translocases with the Single-Stranded DNA Binding Protein RPA in Nucleic Acid Metabolism.

    Awate, Sanket; Brosh, Robert M

    2017-06-08

    Helicases and translocases use the energy of nucleoside triphosphate binding and hydrolysis to unwind/resolve structured nucleic acids or move along a single-stranded or double-stranded polynucleotide chain, respectively. These molecular motors facilitate a variety of transactions including replication, DNA repair, recombination, and transcription. A key partner of eukaryotic DNA helicases/translocases is the single-stranded DNA binding protein Replication Protein A (RPA). Biochemical, genetic, and cell biological assays have demonstrated that RPA interacts with these human molecular motors physically and functionally, and their association is enriched in cells undergoing replication stress. The roles of DNA helicases/translocases are orchestrated with RPA in pathways of nucleic acid metabolism. RPA stimulates helicase-catalyzed DNA unwinding, enlists translocases to sites of action, and modulates their activities in DNA repair, fork remodeling, checkpoint activation, and telomere maintenance. The dynamic interplay between DNA helicases/translocases and RPA is just beginning to be understood at the molecular and cellular levels, and there is still much to be learned, which may inform potential therapeutic strategies.

  18. SECURE SERVICE DISCOVERY BASED ON PROBE PACKET MECHANISM FOR MANETS

    S. Pariselvam

    2015-03-01

    Full Text Available In MANETs, Service discovery process is always considered to be crucial since they do not possess a centralized infrastructure for communication. Moreover, different services available through the network necessitate varying categories. Hence, a need arises for devising a secure probe based service discovery mechanism to reduce the complexity in providing the services to the network users. In this paper, we propose a Secure Service Discovery Based on Probe Packet Mechanism (SSDPPM for identifying the DoS attack in MANETs, which depicts a new approach for estimating the level of trust present in each and every routing path of a mobile ad hoc network by using probe packets. Probing based service discovery mechanisms mainly identifies a mobile node’s genuineness using a test packet called probe that travels the entire network for the sake of computing the degree of trust maintained between the mobile nodes and it’s attributed impact towards the network performance. The performance of SSDPPM is investigated through a wide range of network related parameters like packet delivery, throughput, Control overhead and total overhead using the version ns-2.26 network simulator. This mechanism SSDPPM, improves the performance of the network in an average by 23% and 19% in terms of packet delivery ratio and throughput than the existing service discovery mechanisms available in the literature.

  19. Mechanism of Strain Rate Effect Based on Dislocation Theory

    Kun, Qin; Shi-Sheng, Hu; Li-Ming, Yang

    2009-01-01

    Based on dislocation theory, we investigate the mechanism of strain rate effect. Strain rate effect and dislocation motion are bridged by Orowan's relationship, and the stress dependence of dislocation velocity is considered as the dynamics relationship of dislocation motion. The mechanism of strain rate effect is then investigated qualitatively by using these two relationships although the kinematics relationship of dislocation motion is absent due to complicated styles of dislocation motion. The process of strain rate effect is interpreted and some details of strain rate effect are adequately discussed. The present analyses agree with the existing experimental results. Based on the analyses, we propose that strain rate criteria rather than stress criteria should be satisfied when a metal is fully yielded at a given strain rate. (condensed matter: structure, mechanical and thermal properties)

  20. Transient Dynamic Mechanical Analysis of Resilin-based Elastomeric Hydrogels

    Li, Linqing; Kiick, Kristi

    2014-04-01

    The outstanding high-frequency properties of emerging resilin-like polypeptides (RLPs) have motivated their development for vocal fold tissue regeneration and other applications. Recombinant RLP hydrogels show efficient gelation, tunable mechanical properties, and display excellent extensibility, but little has been reported about their transient mechanical properties. In this manuscript, we describe the transient mechanical behavior of new RLP hydrogels investigated via both sinusoidal oscillatory shear deformation and uniaxial tensile testing. Oscillatory stress relaxation and creep experiments confirm that RLP-based hydrogels display significantly reduced stress relaxation and improved strain recovery compared to PEG-based control hydrogels. Uniaxial tensile testing confirms the negligible hysteresis, reversible elasticity and superior resilience (up to 98%) of hydrated RLP hydrogels, with Young’s modulus values that compare favorably with those previously reported for resilin and that mimic the tensile properties of the vocal fold ligament at low strain (engineering applications, of a range of RLP hydrogels.

  1. Mechanical and physical properties of agro-based fiberboard

    S. Lee; T.F. Shupe; C.Y. Hse

    2006-01-01

    In order to better utilize agricultural fibers as an alternative resource for composite panels, several variables were investigated to improve mechanical and physical properties of agm-based fiberboard. This study focused on the effect of fiber morphology, slenderness ratios (UD), and fiber mixing combinations on panel properties. The panel construction types were also...

  2. Perceived benefits of adopting Standard – Based pricing mechanism for mechanical and electrical services installations

    Ganiyu Amuda Yusuf

    2014-06-01

    Full Text Available Cost is an important measure of project success and clients will expect a reliable forecast at the early stage of construction projects to inform their business decision. This study was undertaken to investigate the current practices in managing cost of mechanical and electrical (M&E services in buildings. The perceptions of practitioners on the benefits of adopting Standard – Based Pricing Mechanism for M&E services as used for building fabrics and finishes was ascertained. The methodology adopted for the study was semi – structure interview and questionnaire survey.  Inferential statistics technique was used to analyse the data collected. The results revealed that, M&E services tender documents are often based on lump sum contract. Practitioners are of the opinion that the adoption of Standard – Based Pricing Mechanism (SBPM could enhance the quality of M&E services price forecast; ensure active post contract cost monitoring and control; encourage collaborative working relationship; enhance efficient whole life cycle cost management; improve risk management and facilitate efficient tendering process. The study suggested the development of local Standard Method of Measurement for M&E services and proposed strategies to facilitate the adoption of SBPM as basis for forecasting contract price of mechanical and electrical services in buildings.

  3. Multiscale experimental mechanics of hierarchical carbon-based materials.

    Espinosa, Horacio D; Filleter, Tobin; Naraghi, Mohammad

    2012-06-05

    Investigation of the mechanics of natural materials, such as spider silk, abalone shells, and bone, has provided great insight into the design of materials that can simultaneously achieve high specific strength and toughness. Research has shown that their emergent mechanical properties are owed in part to their specific self-organization in hierarchical molecular structures, from nanoscale to macroscale, as well as their mixing and bonding. To apply these findings to manmade materials, researchers have devoted significant efforts in developing a fundamental understanding of multiscale mechanics of materials and its application to the design of novel materials with superior mechanical performance. These efforts included the utilization of some of the most promising carbon-based nanomaterials, such as carbon nanotubes, carbon nanofibers, and graphene, together with a variety of matrix materials. At the core of these efforts lies the need to characterize material mechanical behavior across multiple length scales starting from nanoscale characterization of constituents and their interactions to emerging micro- and macroscale properties. In this report, progress made in experimental tools and methods currently used for material characterization across multiple length scales is reviewed, as well as a discussion of how they have impacted our current understanding of the mechanics of hierarchical carbon-based materials. In addition, insight is provided into strategies for bridging experiments across length scales, which are essential in establishing a multiscale characterization approach. While the focus of this progress report is in experimental methods, their concerted use with theoretical-computational approaches towards the establishment of a robust material by design methodology is also discussed, which can pave the way for the development of novel materials possessing unprecedented mechanical properties. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  4. Emerging Importance of Helicases in Plant Stress Tolerance: Characterization of Oryza sativa Repair Helicase XPB2 Promoter and Its Functional Validation in Tobacco under Multiple Stresses.

    Raikwar, Shailendra; Srivastava, Vineet K; Gill, Sarvajeet S; Tuteja, Renu; Tuteja, Narendra

    2015-01-01

    Genetic material always remains at the risk of spontaneous or induced damage which challenges the normal functioning of DNA molecule, thus, DNA repair is vital to protect the organisms against genetic damage. Helicases, the unique molecular motors, are emerged as prospective molecules to engineer stress tolerance in plants and are involved in nucleic acid metabolism including DNA repair. The repair helicase, XPB is an evolutionary conserved protein present in different organisms, including plants. Availability of few efficient promoters for gene expression in plants provoked us to study the promoter of XPB for better understanding of gene regulation under stress conditions. Here, we report the in silico analysis of novel stress inducible promoter of Oryza sativa XPB2 (OsXPB2). The in vivo validation of functionality/activity of OsXPB2 promoter under abiotic and hormonal stress conditions was performed by Agrobacterium-mediated transient assay in tobacco leaves using OsXPB2::GUS chimeric construct. The present research revealed that OsXPB2 promoter contains cis-elements accounting for various abiotic stresses (salt, dehydration, or cold) and hormone (Auxin, ABA, or MeJA) induced GUS expression/activity in the promoter-reporter assay. The promoter region of OsXPB2 contains CACG, GTAACG, CACGTG, CGTCA CCGCCGCGCT cis acting-elements which are reported to be salt, dehydration, cold, MeJA, or ABA responsive, respectively. Functional analysis was done by Agrobacterium-mediated transient assay using agroinfiltration in tobacco leaves, followed by GUS staining and fluorescence quantitative analyses. The results revealed high induction of GUS activity under multiple abiotic stresses as compared to mock treated control. The present findings suggest that OsXPB2 promoter is a multi-stress inducible promoter and has potential applications in sustainable crop production under abiotic stresses by regulating desirable pattern of gene expression.

  5. Development of Reverse Transcription Thermostable Helicase-Dependent DNA Amplification for the Detection of Tomato Spotted Wilt Virus.

    Wu, Xinghai; Chen, Chanfa; Xiao, Xizhi; Deng, Ming Jun

    2016-11-01

    A protocol for the reverse transcription-helicase-dependent amplification (RT-HDA) of isothermal DNA was developed for the detection of tomato spotted wilt virus (TSWV). Specific primers, which were based on the highly conserved region of the N gene sequence in TSWV, were used for the amplification of virus's RNA. The LOD of RT-HDA, reverse transcriptase-loop-mediated isothermal amplification (RT-LAMP), and reverse transcriptase-polymerase chain reaction (RT-PCR) assays were conducted using 10-fold serial dilution of RNA eluates. TSWV sensitivity in RT-HDA and RT-LAMP was 4 pg RNA compared with 40 pg RNA in RT-PCR. The specificity of RT-HDA for TSWV was high, showing no cross-reactivity with other tomato and Tospovirus viruses including cucumber mosaic virus (CMV), tomato black ring virus (TBRV), tomato mosaic virus (ToMV), or impatiens necrotic spot virus (INSV). The RT-HDA method is effective for the detection of TSWV in plant samples and is a potential tool for early and rapid detection of TSWV.

  6. Conserved helicase domain of human RecQ4 is required for strand annealing-independent DNA unwinding

    Rossi, Marie L; Ghosh, Avik K; Kulikowicz, Tomasz

    2010-01-01

    Humans have five members of the well conserved RecQ helicase family: RecQ1, Bloom syndrome protein (BLM), Werner syndrome protein (WRN), RecQ4, and RecQ5, which are all known for their roles in maintaining genome stability. BLM, WRN, and RecQ4 are associated with premature aging and cancer...... provide the first evidence that human RecQ4's unwinding is independent of strand annealing, and that it does not require the presence of excess ssDNA. Moreover, we demonstrate that a point mutation of the conserved lysine in the Walker A motif abolished helicase activity, implying that not the N...... activities and protein partners of RecQ4 are conserved with those of the other RecQ helicases....

  7. Zirconia based dental ceramics: structure, mechanical properties, biocompatibility and applications.

    Gautam, Chandkiram; Joyner, Jarin; Gautam, Amarendra; Rao, Jitendra; Vajtai, Robert

    2016-12-06

    Zirconia (ZrO 2 ) based dental ceramics have been considered to be advantageous materials with adequate mechanical properties for the manufacturing of medical devices. Due to its very high compression strength of 2000 MPa, ZrO 2 can resist differing mechanical environments. During the crack propagation on the application of stress on the surface of ZrO 2 , a crystalline modification diminishes the propagation of cracks. In addition, zirconia's biocompatibility has been studied in vivo, leading to the observation of no adverse response upon the insertion of ZrO 2 samples into the bone or muscle. In vitro experimentation has exhibited the absence of mutations and good viability of cells cultured on this material leading to the use of ZrO 2 in the manufacturing of hip head prostheses. The mechanical properties of zirconia fixed partial dentures (FPDs) have proven to be superior to other ceramic/composite restorations and hence leading to their significant applications in implant supported rehabilitations. Recent developments were focused on the synthesis of zirconia based dental materials. More recently, zirconia has been introduced in prosthetic dentistry for the fabrication of crowns and fixed partial dentures in combination with computer aided design/computer aided manufacturing (CAD/CAM) techniques. This systematic review covers the results of past as well as recent scientific studies on the properties of zirconia based ceramics such as their specific compositions, microstructures, mechanical strength, biocompatibility and other applications in dentistry.

  8. Ultrasound-based testing of tendon mechanical properties

    Seynnes, O R; Bojsen-Møller, J.; Albracht, K

    2015-01-01

    In the past 20 years, the use of ultrasound-based methods has become a standard approach to measure tendon mechanical properties in vivo. Yet the multitude of methodological approaches adopted by various research groups probably contribute to the large variability of reported values. The technique......, or signal synchronization; and 2) in physiological considerations related to the viscoelastic behavior or length measurements of tendons. Hence, the purpose of the present review is to assess and discuss the physiological and technical aspects connected to in vivo testing of tendon mechanical properties...

  9. Gating mechanisms of mechanosensitive channels of large conductance, I: a continuum mechanics-based hierarchical framework.

    Chen, Xi; Cui, Qiang; Tang, Yuye; Yoo, Jejoong; Yethiraj, Arun

    2008-07-01

    A hierarchical simulation framework that integrates information from molecular dynamics (MD) simulations into a continuum model is established to study the mechanical response of mechanosensitive channel of large-conductance (MscL) using the finite element method (FEM). The proposed MD-decorated FEM (MDeFEM) approach is used to explore the detailed gating mechanisms of the MscL in Escherichia coli embedded in a palmitoyloleoylphosphatidylethanolamine lipid bilayer. In Part I of this study, the framework of MDeFEM is established. The transmembrane and cytoplasmic helices are taken to be elastic rods, the loops are modeled as springs, and the lipid bilayer is approximated by a three-layer sheet. The mechanical properties of the continuum components, as well as their interactions, are derived from molecular simulations based on atomic force fields. In addition, analytical closed-form continuum model and elastic network model are established to complement the MDeFEM approach and to capture the most essential features of gating. In Part II of this study, the detailed gating mechanisms of E. coli-MscL under various types of loading are presented and compared with experiments, structural model, and all-atom simulations, as well as the analytical models established in Part I. It is envisioned that such a hierarchical multiscale framework will find great value in the study of a variety of biological processes involving complex mechanical deformations such as muscle contraction and mechanotransduction.

  10. The Q Motif Is Involved in DNA Binding but Not ATP Binding in ChlR1 Helicase.

    Hao Ding

    Full Text Available Helicases are molecular motors that couple the energy of ATP hydrolysis to the unwinding of structured DNA or RNA and chromatin remodeling. The conversion of energy derived from ATP hydrolysis into unwinding and remodeling is coordinated by seven sequence motifs (I, Ia, II, III, IV, V, and VI. The Q motif, consisting of nine amino acids (GFXXPXPIQ with an invariant glutamine (Q residue, has been identified in some, but not all helicases. Compared to the seven well-recognized conserved helicase motifs, the role of the Q motif is less acknowledged. Mutations in the human ChlR1 (DDX11 gene are associated with a unique genetic disorder known as Warsaw Breakage Syndrome, which is characterized by cellular defects in genome maintenance. To examine the roles of the Q motif in ChlR1 helicase, we performed site directed mutagenesis of glutamine to alanine at residue 23 in the Q motif of ChlR1. ChlR1 recombinant protein was overexpressed and purified from HEK293T cells. ChlR1-Q23A mutant abolished the helicase activity of ChlR1 and displayed reduced DNA binding ability. The mutant showed impaired ATPase activity but normal ATP binding. A thermal shift assay revealed that ChlR1-Q23A has a melting point value similar to ChlR1-WT. Partial proteolysis mapping demonstrated that ChlR1-WT and Q23A have a similar globular structure, although some subtle conformational differences in these two proteins are evident. Finally, we found ChlR1 exists and functions as a monomer in solution, which is different from FANCJ, in which the Q motif is involved in protein dimerization. Taken together, our results suggest that the Q motif is involved in DNA binding but not ATP binding in ChlR1 helicase.

  11. Structural and functional analysis of the human spliceosomal DEAD-box helicase Prp28

    Möhlmann, Sina [Georg-August-University Göttingen, Justus-von-Liebig Weg 11, 37077 Göttingen (Germany); Mathew, Rebecca [Max-Planck-Institute for Biophysical Chemistry, Am Fassberg, 37077 Göttingen (Germany); Neumann, Piotr; Schmitt, Andreas [Georg-August-University Göttingen, Justus-von-Liebig Weg 11, 37077 Göttingen (Germany); Lührmann, Reinhard [Max-Planck-Institute for Biophysical Chemistry, Am Fassberg, 37077 Göttingen (Germany); Ficner, Ralf, E-mail: rficner@uni-goettingen.de [Georg-August-University Göttingen, Justus-von-Liebig Weg 11, 37077 Göttingen (Germany)

    2014-06-01

    The crystal structure of the helicase domain of the human spliceosomal DEAD-box protein Prp28 was solved by SAD. The binding of ADP and ATP by Prp28 was studied biochemically and analysed with regard to the crystal structure. The DEAD-box protein Prp28 is essential for pre-mRNA splicing as it plays a key role in the formation of an active spliceosome. Prp28 participates in the release of the U1 snRNP from the 5′-splice site during association of the U5·U4/U6 tri-snRNP, which is a crucial step in the transition from a pre-catalytic spliceosome to an activated spliceosome. Here, it is demonstrated that the purified helicase domain of human Prp28 (hPrp28ΔN) binds ADP, whereas binding of ATP and ATPase activity could not be detected. ATP binding could not be observed for purified full-length hPrp28 either, but within an assembled spliceosomal complex hPrp28 gains ATP-binding activity. In order to understand the structural basis for the ATP-binding deficiency of isolated hPrp28, the crystal structure of hPrp28ΔN was determined at 2.0 Å resolution. In the crystal the helicase domain adopts a wide-open conformation, as the two RecA-like domains are extraordinarily displaced from the productive ATPase conformation. Binding of ATP is hindered by a closed conformation of the P-loop, which occupies the space required for the γ-phosphate of ATP.

  12. In vivo mapping of the functional regions of the DEAD-box helicase Vasa

    Mehrnoush Dehghani

    2015-03-01

    Full Text Available The maternally expressed Drosophila melanogaster DEAD-box helicase Vasa (Vas is necessary for many cellular and developmental processes, including specification of primordial germ cells (pole cells, posterior patterning of the embryo, piRNA-mediated repression of transposon-encoded mRNAs, translational activation of gurken (grk mRNA, and completion of oogenesis itself. Vas protein accumulates in the perinuclear nuage in nurse cells soon after their specification, and then at stage 10 Vas translocates to the posterior pole plasm of the oocyte. We produced a series of transgenic constructs encoding eGFP-Vas proteins carrying mutations affecting different regions of the protein, and analyzed in vivo which Vas functions each could support. We identified novel domains in the N- and C-terminal regions of the protein that are essential for localization, transposon repression, posterior patterning, and pole cell specification. One such functional region, the most C-terminal seven amino acids, is specific to Vas orthologues and is thus critical to distinguishing Vas from other closely related DEAD-box helicases. Surprisingly, we also found that many eGFP-Vas proteins carrying mutations that would be expected to abrogate DEAD-box helicase function localized to the nuage and posterior pole, and retained the capacity to support oogenesis, although they did not function in embryonic patterning, pole cell specification, grk activation, or transposon repression. We conclude from these experiments that Vas, a multifunctional protein, uses different domains and different molecular associations to carry out its various cellular and developmental roles.

  13. Functional interaction between Smad, CREB binding protein, and p68 RNA helicase

    Warner, Dennis R.; Bhattacherjee, Vasker; Yin, Xiaolong; Singh, Saurabh; Mukhopadhyay, Partha; Pisano, M. Michele; Greene, Robert M.

    2004-01-01

    The transforming growth factors β control a diversity of biological processes including cellular proliferation, differentiation, apoptosis, and extracellular matrix production, and are critical effectors of embryonic patterning and development, including that of the orofacial region. TGFβ superfamily members signal through specific cell surface receptors that phosphorylate the cytoplasmic Smad proteins, resulting in their translocation to the nucleus and interaction with promoters of TGFβ-responsive genes. Subsequent alterations in transcription are cell type-specific and dependent on recruitment to the Smad/transcription factor complex of coactivators, such as CBP and p300, or corepressors, such as c-ski and SnoN. Since the affinity of Smads for DNA is generally low, additional accessory proteins that facilitate Smad/DNA binding are required, and are often cell- and tissue-specific. In order to identify novel Smad 3 binding proteins in developing orofacial tissue, a yeast two hybrid assay was employed in which the MH2 domain of Smad 3 was used to screen an expression library derived from mouse embryonic orofacial tissue. The RNA helicase, p68, was identified as a unique Smad binding protein, and the specificity of the interaction was confirmed through various in vitro and in vivo assays. Co-expression of Smad 3 and a CBP-Gal4 DNA binding domain fusion protein in a Gal4-luciferase reporter assay resulted in increased TGFβ-stimulated reporter gene transcription. Moreover, co-expression of p68 RNA helicase along with Smad 3 and CBP-Gal4 resulted in synergistic activation of Gal4-luciferase reporter expression. Collectively, these data indicate that the RNA helicase, p68, can directly interact with Smad 3 resulting in formation of a transcriptionally active ternary complex containing Smad 3, p68, and CBP. This offers a means of enhancing TGFβ-mediated cellular responses in developing orofacial tissue

  14. Fracture mechanics based life assessment in petrochemical plants

    Norasiah Ab Kasim; Abd Nassir Ibrahim; Ab Razak Hamzah; Shukri Mohd

    2004-01-01

    The increasing use of thick walled pressure vessels in petrochemical plants operating at high pressure under severe service conditions could lead to catastrophic failure. In the Malaysian Institute for Nuclear Technology Research (MINT), initial efforts are underway to apply fracture mechanics approach for assessment of significance of defects detected during periodic in service inspection (ISI) of industrial plants. This paper outlines the integrity management strategy based on fracture mechanics and proposes a new procedure for life assessment of petrochemical plants based on ASME Boiler and Pressure Vessel Code, Section XI, BSI PD 6493:1991, BSI 6539:1994, BSI Standard 7910:1999 and API 579:2000. Essential relevant data required for the assessment is listed. Several methods available for determination of fracture toughness are reviewed with limitations in their application to petrochemical plants. A new non destructive method for determination of fracture toughness based on hardness testing and normalized key roughness curve is given. Results of fracture mechanics based life assessment conducted for 100 mm thick ammonia converter of Ni r o steel and 70 mm thick plat forming reactor vessel of ASTM A 38 7 grade B steel in operational fertilizer and petroleum refining plants are presented. (Author)

  15. Toward a Mechanism-Based Approach to Pain Diagnosis.

    Vardeh, Daniel; Mannion, Richard J; Woolf, Clifford J

    2016-09-01

    The past few decades have witnessed a huge leap forward in our understanding of the mechanistic underpinnings of pain, in normal states where it helps protect from injury, and also in pathological states where pain evolves from a symptom reflecting tissue injury to become the disease itself. However, despite these scientific advances, chronic pain remains extremely challenging to manage clinically. Although the number of potential treatment targets has grown substantially and a strong case has been made for a mechanism-based and individualized approach to pain therapy, arguably clinicians are not much more advanced now than 20 years ago, in their capacity to either diagnose or effectively treat their patients. The gulf between pain research and pain management is as wide as ever. We are still currently unable to apply an evidence-based approach to chronic pain management that reflects mechanistic understanding, and instead, clinical practice remains an empirical and often unsatisfactory journey for patients, whose individual response to treatment cannot be predicted. In this article we take a common and difficult to treat pain condition, chronic low back pain, and use its presentation in clinical practice as a framework to highlight what is known about pathophysiological pain mechanisms and how we could potentially detect these to drive rational treatment choice. We discuss how present methods of assessment and management still fall well short, however, of any mechanism-based or precision medicine approach. Nevertheless, substantial improvements in chronic pain management could be possible if a more strategic and coordinated approach were to evolve, one designed to identify the specific mechanisms driving the presenting pain phenotype. We present an analysis of such an approach, highlighting the major problems in identifying mechanisms in patients, and develop a framework for a pain diagnostic ladder that may prove useful in the future, consisting of successive

  16. Yeast as a model system to study RecQ helicase function

    Ashton, Thomas M; Hickson, Ian David

    2010-01-01

    Mutations in the highly conserved RecQ helicase, BLM, cause the rare cancer predisposition disorder, Bloom's syndrome. The orthologues of BLM in Saccharomyces cerevisiae and Schizosaccharomyces pombe are SGS1 and rqh1(+), respectively. Studies in these yeast species have revealed a plethora...... of roles for the Sgs1 and Rqh1 proteins in repair of double strand breaks, restart of stalled replication forks, processing of aberrant intermediates that arise during meiotic recombination, and maintenance of telomeres. In this review, we focus on the known roles of Sgs1 and Rqh1 and how studies in yeast...

  17. RTEL1: an essential helicase for telomere maintenance and the regulation of homologous recombination.

    Uringa, Evert-Jan; Youds, Jillian L; Lisaingo, Kathleen; Lansdorp, Peter M; Boulton, Simon J

    2011-03-01

    Telomere maintenance and DNA repair are crucial processes that protect the genome against instability. RTEL1, an essential iron-sulfur cluster-containing helicase, is a dominant factor that controls telomere length in mice and is required for telomere integrity. In addition, RTEL1 promotes synthesis-dependent strand annealing to direct DNA double-strand breaks into non-crossover outcomes during mitotic repair and in meiosis. Here, we review the role of RTEL1 in telomere maintenance and homologous recombination and discuss models linking RTEL1's enzymatic activity to its function in telomere maintenance and DNA repair.

  18. Mechanism of high-temperature resistant water-base mud

    Luo, P

    1981-01-01

    Based on experiments, the causes and laws governing the changes in the performance of water-base mud under high temperature are analyzed, and the requisites and mechanism of treating agents resisting high temperature are discussed. Ways and means are sought for inhibiting, delaying and making use of the effect of high temperature on the performance of mud, while new ideas and systematic views have been expressed on the preparation of treating agents and set-up of a high temperature resistant water-base mud system. High temperature dispersion and high temperature surface inactivation of clay in the mud, as well as their effect and method of utilization are reviewed. Subjects also touched upon include degradation and cross-linking of the high-temperature resistant treating agents, their use and effect. Based on the above, the preparation of a water-base and system capable of resisting 180 to 250/sup 0/C is recommended.

  19. The helicase and ATPase activities of RECQL4 are compromised by mutations reported in three human patients

    Jensen, Martin Borch; Dunn, Christopher A; Keijzers, Guido

    2012-01-01

    RECQL4 is one of five members of the human RecQ helicase family, and is implicated in three syndromes displaying accelerating aging, developmental abnormalities and a predisposition to cancer. In this study, we purified three variants of RECQL4 carrying previously reported patient mutations....... These three mutant proteins were analyzed for the known biochemical activities of RECQL4: DNA binding, unwinding of duplex DNA, ATP hydrolysis and annealing of simplex DNA. Further, the mutant proteins were evaluated for stability and recruitment to sites of laser-induced DNA damage. One mutant was helicase...

  20. Mechanisms of time-based figure-ground segregation.

    Kandil, Farid I; Fahle, Manfred

    2003-11-01

    Figure-ground segregation can rely on purely temporal information, that is, on short temporal delays between positional changes of elements in figure and ground (Kandil, F.I. & Fahle, M. (2001) Eur. J. Neurosci., 13, 2004-2008). Here, we investigate the underlying mechanisms by measuring temporal segregation thresholds for various kinds of motion cues. Segregation can rely on monocular first-order motion (based on luminance modulation) and second-order motion cues (contrast modulation) with a high temporal resolution of approximately 20 ms. The mechanism can also use isoluminant motion with a reduced temporal resolution of 60 ms. Figure-ground segregation can be achieved even at presentation frequencies too high for human subjects to inspect successive frames individually. In contrast, when stimuli are presented dichoptically, i.e. separately to both eyes, subjects are unable to perceive any segregation, irrespective of temporal frequency. We propose that segregation in these displays is detected by a mechanism consisting of at least two stages. On the first level, standard motion or flicker detectors signal local positional changes (flips). On the second level, a segregation mechanism combines the local activities of the low-level detectors with high temporal precision. Our findings suggest that the segregation mechanism can rely on monocular detectors but not on binocular mechanisms. Moreover, the results oppose the idea that segregation in these displays is achieved by motion detectors of a higher order (motion-from-motion), but favour mechanisms sensitive to short temporal delays even without activation of higher-order motion detectors.

  1. A calibration mechanism based on worm drive for space telescope

    Chong, Yaqin; Li, Chuang; Xia, Siyu; Zhong, Peifeng; Lei, Wang

    2017-08-01

    In this paper, a new type of calibration mechanism based on worm drive is presented for a space telescope. This calibration mechanism based on worm drive has the advantages of compact size and self-lock. The mechanism mainly consists of thirty-six LEDs as the light source for flat calibration, a diffuse plate, a step motor, a worm gear reducer and a potentiometer. As the main part of the diffuse plate, a PTFE tablet is mounted in an aluminum alloy frame. The frame is fixed on the shaft of the worm gear, which is driven by the step motor through the worm. The shaft of the potentiometer is connected to that of the worm gear to measure the rotation angle of the diffuse plate through a flexible coupler. Firstly, the calibration mechanism is designed, which includes the LEDs assembly design, the worm gear reducer design and the diffuse plate assembly design. The counterweight blocks and two end stops are also designed for the diffuse plate assembly. Then a modal analysis with finite element method for the diffuse plate assembly is completed.

  2. Mycobacterium tuberculosis UvrB Is a Robust DNA-Stimulated ATPase That Also Possesses Structure-Specific ATP-Dependent DNA Helicase Activity.

    Thakur, Manoj; Kumar, Mohan B J; Muniyappa, K

    2016-10-18

    Much is known about the Escherichia coli nucleotide excision repair (NER) pathway; however, very little is understood about the proteins involved and the molecular mechanism of NER in mycobacteria. In this study, we show that Mycobacterium tuberculosis UvrB (MtUvrB), which exists in solution as a monomer, binds to DNA in a structure-dependent manner. A systematic examination of MtUvrB substrate specificity reveals that it associates preferentially with single-stranded DNA, duplexes with 3' or 5' overhangs, and linear duplex DNA with splayed arms. Whereas E. coli UvrB (EcUvrB) binds weakly to undamaged DNA and has no ATPase activity, MtUvrB possesses intrinsic ATPase activity that is greatly stimulated by both single- and double-stranded DNA. Strikingly, we found that MtUvrB, but not EcUvrB, possesses the DNA unwinding activity characteristic of an ATP-dependent DNA helicase. The helicase activity of MtUvrB proceeds in the 3' to 5' direction and is strongly modulated by a nontranslocating 5' single-stranded tail, indicating that in addition to the translocating strand it also interacts with the 5' end of the substrate. The fraction of DNA unwound by MtUvrB decreases significantly as the length of the duplex increases: it fails to unwind duplexes longer than 70 bp. These results, on one hand, reveal significant mechanistic differences between MtUvrB and EcUvrB and, on the other, support an alternative role for UvrB in the processing of key DNA replication intermediates. Altogether, our findings provide insights into the catalytic functions of UvrB and lay the foundation for further understanding of the NER pathway in M. tuberculosis.

  3. Structure of a Novel DNA-binding Domain of Helicase-like Transcription Factor (HLTF) and Its Functional Implication in DNA Damage Tolerance.

    Hishiki, Asami; Hara, Kodai; Ikegaya, Yuzu; Yokoyama, Hideshi; Shimizu, Toshiyuki; Sato, Mamoru; Hashimoto, Hiroshi

    2015-05-22

    HLTF (helicase-like transcription factor) is a yeast RAD5 homolog found in mammals. HLTF has E3 ubiquitin ligase and DNA helicase activities, and plays a pivotal role in the template-switching pathway of DNA damage tolerance. HLTF has an N-terminal domain that has been designated the HIRAN (HIP116 and RAD5 N-terminal) domain. The HIRAN domain has been hypothesized to play a role in DNA binding; however, the structural basis of, and functional evidence for, the HIRAN domain in DNA binding has remained unclear. Here we show for the first time the crystal structure of the HIRAN domain of human HLTF in complex with DNA. The HIRAN domain is composed of six β-strands and two α-helices, forming an OB-fold structure frequently found in ssDNA-binding proteins, including in replication factor A (RPA). Interestingly, this study reveals that the HIRAN domain interacts with not only with a single-stranded DNA but also with a duplex DNA. Furthermore, the structure unexpectedly clarifies that the HIRAN domain specifically recognizes the 3'-end of DNA. These results suggest that the HIRAN domain functions as a sensor to the 3'-end of the primer strand at the stalled replication fork and that the domain facilitates fork regression. HLTF is recruited to a damaged site through the HIRAN domain at the stalled replication fork. Furthermore, our results have implications for the mechanism of template switching. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  4. A statistical mechanical interpretation of algorithmic information theory: Total statistical mechanical interpretation based on physical argument

    Tadaki, Kohtaro

    2010-01-01

    The statistical mechanical interpretation of algorithmic information theory (AIT, for short) was introduced and developed by our former works [K. Tadaki, Local Proceedings of CiE 2008, pp. 425-434, 2008] and [K. Tadaki, Proceedings of LFCS'09, Springer's LNCS, vol. 5407, pp. 422-440, 2009], where we introduced the notion of thermodynamic quantities, such as partition function Z(T), free energy F(T), energy E(T), statistical mechanical entropy S(T), and specific heat C(T), into AIT. We then discovered that, in the interpretation, the temperature T equals to the partial randomness of the values of all these thermodynamic quantities, where the notion of partial randomness is a stronger representation of the compression rate by means of program-size complexity. Furthermore, we showed that this situation holds for the temperature T itself, which is one of the most typical thermodynamic quantities. Namely, we showed that, for each of the thermodynamic quantities Z(T), F(T), E(T), and S(T) above, the computability of its value at temperature T gives a sufficient condition for T is an element of (0,1) to satisfy the condition that the partial randomness of T equals to T. In this paper, based on a physical argument on the same level of mathematical strictness as normal statistical mechanics in physics, we develop a total statistical mechanical interpretation of AIT which actualizes a perfect correspondence to normal statistical mechanics. We do this by identifying a microcanonical ensemble in the framework of AIT. As a result, we clarify the statistical mechanical meaning of the thermodynamic quantities of AIT.

  5. Towards a mechanism-based approach to pain diagnosis

    Vardeh, Daniel

    2016-01-01

    The last few decades have witnessed a huge leap forward in our understanding of the mechanistic underpinnings of pain, both in normal states where it helps protect from injury, and in pathological states where pain evolves from a symptom reflecting tissue injury to become the disease itself. However, despite these scientific advances, chronic pain remains extremely challenging to manage clinically. While the number of potential treatment targets has grown substantially and a strong case has been made for a mechanism-based and individualized approach to pain therapy, arguably clinicians are not much more advanced now than 20 years ago, in their capacity to either diagnose or effectively treat their patients. The gulf between pain research and pain management is as wide as ever. We are still currently unable to apply an evidence-based approach to chronic pain management that reflects mechanistic understanding, and instead, clinical practice remains an empirical and often unsatisfactory journey for patients, whose individual response to treatment cannot be predicted. Here we take a common and difficult to treat pain condition, chronic low back pain, and use its presentation in clinical practice as a framework to highlight what is known about pathophysiological pain mechanisms and how we could potentially detect these to drive rational treatment choice. We discuss how present methods of assessment and management still fall well short, however, of any mechanism-based or precision-medicine approach. Nevertheless, substantial improvements in chronic pain management could be possible if a more strategic and coordinated approach were to evolve, one designed to identify the specific mechanisms driving the presenting pain phenotype. We present an analysis of such an approach, highlighting the major problems in identifying mechanisms in patients, and develop a framework for a pain diagnostic ladder that may prove useful in the future, consisting of successive identification of

  6. Mechanisms of Base Substitution Mutagenesis in Cancer Genomes

    Albino Bacolla

    2014-03-01

    Full Text Available Cancer genome sequence data provide an invaluable resource for inferring the key mechanisms by which mutations arise in cancer cells, favoring their survival, proliferation and invasiveness. Here we examine recent advances in understanding the molecular mechanisms responsible for the predominant type of genetic alteration found in cancer cells, somatic single base substitutions (SBSs. Cytosine methylation, demethylation and deamination, charge transfer reactions in DNA, DNA replication timing, chromatin status and altered DNA proofreading activities are all now known to contribute to the mechanisms leading to base substitution mutagenesis. We review current hypotheses as to the major processes that give rise to SBSs and evaluate their relative relevance in the light of knowledge acquired from cancer genome sequencing projects and the study of base modifications, DNA repair and lesion bypass. Although gene expression data on APOBEC3B enzymes provide support for a role in cancer mutagenesis through U:G mismatch intermediates, the enzyme preference for single-stranded DNA may limit its activity genome-wide. For SBSs at both CG:CG and YC:GR sites, we outline evidence for a prominent role of damage by charge transfer reactions that follow interactions of the DNA with reactive oxygen species (ROS and other endogenous or exogenous electron-abstracting molecules.

  7. Mechanisms of base substitution mutagenesis in cancer genomes.

    Bacolla, Albino; Cooper, David N; Vasquez, Karen M

    2014-03-05

    Cancer genome sequence data provide an invaluable resource for inferring the key mechanisms by which mutations arise in cancer cells, favoring their survival, proliferation and invasiveness. Here we examine recent advances in understanding the molecular mechanisms responsible for the predominant type of genetic alteration found in cancer cells, somatic single base substitutions (SBSs). Cytosine methylation, demethylation and deamination, charge transfer reactions in DNA, DNA replication timing, chromatin status and altered DNA proofreading activities are all now known to contribute to the mechanisms leading to base substitution mutagenesis. We review current hypotheses as to the major processes that give rise to SBSs and evaluate their relative relevance in the light of knowledge acquired from cancer genome sequencing projects and the study of base modifications, DNA repair and lesion bypass. Although gene expression data on APOBEC3B enzymes provide support for a role in cancer mutagenesis through U:G mismatch intermediates, the enzyme preference for single-stranded DNA may limit its activity genome-wide. For SBSs at both CG:CG and YC:GR sites, we outline evidence for a prominent role of damage by charge transfer reactions that follow interactions of the DNA with reactive oxygen species (ROS) and other endogenous or exogenous electron-abstracting molecules.

  8. Material recognition based on thermal cues: Mechanisms and applications.

    Ho, Hsin-Ni

    2018-01-01

    Some materials feel colder to the touch than others, and we can use this difference in perceived coldness for material recognition. This review focuses on the mechanisms underlying material recognition based on thermal cues. It provides an overview of the physical, perceptual, and cognitive processes involved in material recognition. It also describes engineering domains in which material recognition based on thermal cues have been applied. This includes haptic interfaces that seek to reproduce the sensations associated with contact in virtual environments and tactile sensors aim for automatic material recognition. The review concludes by considering the contributions of this line of research in both science and engineering.

  9. The activation mechanism of Fe-based olefin metathesis catalysts

    Poater, Albert

    2014-08-01

    Density functional theory calculations have been used to describe the first turnover for olefin metathesis reaction of a homogenous Fe-based catalyst bearing a N-heterocyclic carbene ligand with methoxyethene as a substrate. Equal to conventional Ru-based catalysts, the activation of its Fe congener occurs through a dissociative mechanism, however with a more exothermic reaction energy profile. Predicted upper energy barriers were calculated to be on average ∼2 kcal/mol more beneficial for Fe catalyzed metathesis. Overall, this present computational study emphasises on advantages of Fe-based metathesis and gives a potential recipe for the design of an efficient Fe-based olefin metathesis catalysts. © 2014 Elsevier B.V.

  10. Waterbomb base: a symmetric single-vertex bistable origami mechanism

    Hanna, Brandon H; Lund, Jason M; Magleby, Spencer P; Howell, Larry L; Lang, Robert J

    2014-01-01

    The origami waterbomb base is a single-vertex bistable origami mechanism that has unique properties which may prove useful in a variety of applications. It also shows promise as a test bed for smart materials and actuation because of its straightforward geometry and multiple phases of motion, ranging from simple to more complex. This study develops a quantitative understanding of the symmetric waterbomb base's kinetic behavior. This is done by completing kinematic and potential energy analyses to understand and predict bistable behavior. A physical prototype is constructed and tested to validate the results of the analyses. Finite element and virtual work analyses based on the prototype are used to explore the locations of the stable equilibrium positions and the force–deflection response. The model results are verified through comparisons to measurements on a physical prototype. The resulting models describe waterbomb base behavior and provide an engineering tool for application development. (paper)

  11. The activation mechanism of Fe-based olefin metathesis catalysts

    Poater, Albert; Pump, Eva; Vummaleti, Sai V. C.; Cavallo, Luigi

    2014-01-01

    Density functional theory calculations have been used to describe the first turnover for olefin metathesis reaction of a homogenous Fe-based catalyst bearing a N-heterocyclic carbene ligand with methoxyethene as a substrate. Equal to conventional Ru-based catalysts, the activation of its Fe congener occurs through a dissociative mechanism, however with a more exothermic reaction energy profile. Predicted upper energy barriers were calculated to be on average ∼2 kcal/mol more beneficial for Fe catalyzed metathesis. Overall, this present computational study emphasises on advantages of Fe-based metathesis and gives a potential recipe for the design of an efficient Fe-based olefin metathesis catalysts. © 2014 Elsevier B.V.

  12. Micro-mechanics based damage mechanics for 3D Orthogonal Woven Composites: Experiment and Numerical Modelling

    Saleh, Mohamed Nasr

    2016-01-08

    Damage initiation and evolution of three-dimensional (3D) orthogonal woven carbon fibre composite (3DOWC) is investigated experimentally and numerically. Meso-scale homogenisation of the representative volume element (RVE) is utilised to predict the elastic properties, simulate damage initiation and evolution when loaded in tension. The effect of intra-yarns transverse cracking and shear diffused damage on the in-plane transverse modulus and shear modulus is investigated while one failure criterion is introduced to simulate the matrix damage. The proposed model is based on two major assumptions. First, the effect of the binder yarns, on the in-plane properties, is neglected, so the 3DOWC unit cell can be approximated as a (0o/90o) cross-ply laminate. Second, a micro-mechanics based damage approach is used at the meso-scale, so damage indicators can be correlated, explicitly, to the density of cracks within the material. Results from the simulated RVE are validated against experimental results along the warp (0o direction) and weft (90o direction). This approach paves the road for more predictive models as damage evolution laws are obtained from micro mechanical considerations and rely on few well-defined material parameters. This largely differs from classical damage mechanics approaches in which the evolution law is obtained by retrofitting experimental observations.

  13. Micro-mechanics based damage mechanics for 3D Orthogonal Woven Composites: Experiment and Numerical Modelling

    Saleh, Mohamed Nasr; Lubineau, Gilles; Potluri, Prasad; Withers, Philip; Soutis, Constantinos

    2016-01-01

    Damage initiation and evolution of three-dimensional (3D) orthogonal woven carbon fibre composite (3DOWC) is investigated experimentally and numerically. Meso-scale homogenisation of the representative volume element (RVE) is utilised to predict the elastic properties, simulate damage initiation and evolution when loaded in tension. The effect of intra-yarns transverse cracking and shear diffused damage on the in-plane transverse modulus and shear modulus is investigated while one failure criterion is introduced to simulate the matrix damage. The proposed model is based on two major assumptions. First, the effect of the binder yarns, on the in-plane properties, is neglected, so the 3DOWC unit cell can be approximated as a (0o/90o) cross-ply laminate. Second, a micro-mechanics based damage approach is used at the meso-scale, so damage indicators can be correlated, explicitly, to the density of cracks within the material. Results from the simulated RVE are validated against experimental results along the warp (0o direction) and weft (90o direction). This approach paves the road for more predictive models as damage evolution laws are obtained from micro mechanical considerations and rely on few well-defined material parameters. This largely differs from classical damage mechanics approaches in which the evolution law is obtained by retrofitting experimental observations.

  14. A prototype of behavior selection mechanism based on emotion

    Zhang, Guofeng; Li, Zushu

    2007-12-01

    In bionic methodology rather than in design methodology more familiar with, summarizing the psychological researches of emotion, we propose the biologic mechanism of emotion, emotion selection role in creature evolution and a anima framework including emotion similar to the classical control structure; and consulting Prospect Theory, build an Emotion Characteristic Functions(ECF) that computer emotion; two more emotion theories are added to them that higher emotion is preferred and middle emotion makes brain run more efficiently, emotional behavior mechanism comes into being. A simulation of proposed mechanism are designed and carried out on Alife Swarm software platform. In this simulation, a virtual grassland ecosystem is achieved where there are two kinds of artificial animals: herbivore and preyer. These artificial animals execute four types of behavior: wandering, escaping, finding food, finding sex partner in their lives. According the theories of animal ethnology, escaping from preyer is prior to other behaviors for its existence, finding food is secondly important behavior, rating is third one and wandering is last behavior. In keeping this behavior order, based on our behavior characteristic function theory, the specific functions of emotion computing are built of artificial autonomous animals. The result of simulation confirms the behavior selection mechanism.

  15. Time dependent mechanical modeling for polymers based on network theory

    Billon, Noëlle [MINES ParisTech, PSL-Research University, CEMEF – Centre de mise en forme des matériaux, CNRS UMR 7635, CS 10207 rue Claude Daunesse 06904 Sophia Antipolis Cedex (France)

    2016-05-18

    Despite of a lot of attempts during recent years, complex mechanical behaviour of polymers remains incompletely modelled, making industrial design of structures under complex, cyclic and hard loadings not totally reliable. The non linear and dissipative viscoelastic, viscoplastic behaviour of those materials impose to take into account non linear and combined effects of mechanical and thermal phenomena. In this view, a visco-hyperelastic, viscoplastic model, based on network description of the material has recently been developed and designed in a complete thermodynamic frame in order to take into account those main thermo-mechanical couplings. Also, a way to account for coupled effects of strain-rate and temperature was suggested. First experimental validations conducted in the 1D limit on amorphous rubbery like PMMA in isothermal conditions led to pretty goods results. In this paper a more complete formalism is presented and validated in the case of a semi crystalline polymer, a PA66 and a PET (either amorphous or semi crystalline) are used. Protocol for identification of constitutive parameters is described. It is concluded that this new approach should be the route to accurately model thermo-mechanical behaviour of polymers using a reduced number of parameters of some physical meaning.

  16. Task-Based Method for Designing Underactuated Mechanisms

    Shoichiro Kamada

    2012-03-01

    Full Text Available In this paper we introduce a task-based method for designing underactuated multi-joint prosthetic hands for specific grasping tasks. The designed robotic hands or prosthetic hands contain fewer independent actuators than joints. We chose a few specific grasping tasks that are frequently repeated in everyday life and analysed joint motions of the hand during the completion of each task and the level of participation of each joint. The information was used for the synthesis of dedicated underactuated mechanisms that can operate in a low dimensional task coordinate space. We propose two methods for reducing the actuators' number. The kinematic parameters of the synthesized mechanism are determined by using a numerical approach. In this study the joint angles of the synthesized hand are considered as linearly dependent on the displacements of the actuators. We introduced a special error index that allowed us to compare the original trajectory and the trajectory performed by the synthesized mechanism, and to select the kinematic parameters of the new kinematic structure as a way to reduce the error. The approach allows the design of simple gripper mechanisms with good accuracy for the preliminary defined tasks.

  17. Computer-based mechanical design of overhead lines

    Rusinaru, D.; Bratu, C.; Dinu, R. C.; Manescu, L. G.

    2016-02-01

    Beside the performance, the safety level according to the actual standards is a compulsory condition for distribution grids’ operation. Some of the measures leading to improvement of the overhead lines reliability ask for installations’ modernization. The constraints imposed to the new lines components refer to the technical aspects as thermal stress or voltage drop, and look for economic efficiency, too. The mechanical sizing of the overhead lines is after all an optimization problem. More precisely, the task in designing of the overhead line profile is to size poles, cross-arms and stays and locate poles along a line route so that the total costs of the line's structure to be minimized and the technical and safety constraints to be fulfilled.The authors present in this paper an application for the Computer-Based Mechanical Design of the Overhead Lines and the features of the corresponding Visual Basic program, adjusted to the distribution lines. The constraints of the optimization problem are adjusted to the existing weather and loading conditions of Romania. The outputs of the software application for mechanical design of overhead lines are: the list of components chosen for the line: poles, cross-arms, stays; the list of conductor tension and forces for each pole, cross-arm and stay for different weather conditions; the line profile drawings.The main features of the mechanical overhead lines design software are interactivity, local optimization function and high-level user-interface

  18. Mechanical Seal Opening Condition Monitoring Based on Acoustic Emission Technology

    Erqing Zhang

    2014-06-01

    Full Text Available Since the measurement of mechanical sealing film thickness and just-lift-off time is very difficult, the sealing film condition monitoring method based on acoustic emission signal is proposed. The mechanical seal acoustic emission signal present obvious characteristics of time-varying nonlinear and pulsating. In this paper, the acoustic emission signal is used to monitor the seal end faces just-lift-off time and friction condition. The acoustic emission signal is decomposed by empirical mode decomposition into a series of intrinsic mode function with independent characteristics of different time scales and different frequency band. The acoustic emission signal only generated by end faces friction is obtained by eliminating the false intrinsic mode function components. The correlation coefficient of acoustic emission signal and Multi-scale Laplace Wavelet is calculated. It is proved that the maximum frequency (8000 Hz of the correlation coefficient is appeared at the spindle speed of 300 rpm. And at this time (300 rpm the end faces have just lifted off. By a set of mechanical oil seal running test, it is demonstrated that this method could accurately identify mechanical seal end faces just-lift-off time and friction condition.

  19. Germline mutations of regulator of telomere elongation helicase 1, RTEL1, in Dyskeratosis congenita.

    Ballew, Bari J; Yeager, Meredith; Jacobs, Kevin; Giri, Neelam; Boland, Joseph; Burdett, Laurie; Alter, Blanche P; Savage, Sharon A

    2013-04-01

    Dyskeratosis congenita (DC) is an inherited bone marrow failure and cancer predisposition syndrome caused by aberrant telomere biology. The classic triad of dysplastic nails, abnormal skin pigmentation, and oral leukoplakia is diagnostic of DC, but substantial clinical heterogeneity exists; the clinically severe variant Hoyeraal Hreidarsson syndrome (HH) also includes cerebellar hypoplasia, severe immunodeficiency, enteropathy, and intrauterine growth retardation. Germline mutations in telomere biology genes account for approximately one-half of known DC families. Using exome sequencing, we identified mutations in RTEL1, a helicase with critical telomeric functions, in two families with HH. In the first family, two siblings with HH and very short telomeres inherited a premature stop codon from their mother who has short telomeres. The proband from the second family has HH and inherited a premature stop codon in RTEL1 from his father and a missense mutation from his mother, who also has short telomeres. In addition, inheritance of only the missense mutation led to very short telomeres in the proband's brother. Targeted sequencing identified a different RTEL1 missense mutation in one additional DC proband who has bone marrow failure and short telomeres. Both missense mutations affect the helicase domain of RTEL1, and three in silico prediction algorithms suggest that they are likely deleterious. The nonsense mutations both cause truncation of the RTEL1 protein, resulting in loss of the PIP box; this may abrogate an important protein-protein interaction. These findings implicate a new telomere biology gene, RTEL1, in the etiology of DC.

  20. Characterization of the Caenorhabditis elegans HIM-6/BLM helicase: unwinding recombination intermediates.

    Jung, Hana; Lee, Jin A; Choi, Seoyoon; Lee, Hyunwoo; Ahn, Byungchan

    2014-01-01

    Mutations in three human RecQ genes are implicated in heritable human syndromes. Mutations in BLM, a RecQ gene, cause Bloom syndrome (BS), which is characterized by short stature, cancer predisposition, and sensitivity to sunlight. BLM is a RecQ DNA helicase that, with interacting proteins, is able to dissolve various DNA structures including double Holliday junctions. A BLM ortholog, him-6, has been identified in Caenorhabditis elegans, but little is known about its enzymatic activities or its in vivo roles. By purifying recombinant HIM-6 and performing biochemical assays, we determined that the HIM-6 has DNA-dependent ATPase activity HIM-6 and helicase activity that proceeds in the 3'-5' direction and needs at least five 3' overhanging nucleotides. HIM-6 is also able to unwind DNA structures including D-loops and Holliday junctions. Worms with him-6 mutations were defective in recovering the cell cycle arrest after HU treatment. These activities strongly support in vivo roles for HIM-6 in processing recombination intermediates.

  1. The Application of Problem-Based Learning in Mechanical Engineering

    Putra, Z. A.; Dewi, M.

    2018-02-01

    The course of Technology and Material Testing prepare students with the ability to do a variety of material testing in the study of mechanical engineering. Students find it difficult to understand the materials to make them unable to carry out the material testing in accordance with the purpose of study. This happens because they knowledge is not adequately supported by the competence to find and construct learning experience. In this study, quasy experiment research method with pre-post-test with control group design was used. The subjects of the study were students divided in two groups; control and experiment with twenty-two students in each group. Study result: their grades showed no difference in between the pre-test or post-test in control group, but the difference in grade existed between the pre-test and post-test in experiment group. Yet, there is no significant difference in the study result on both groups. The researcher recommend that it is necessary to develop Problem-Based Learning that suits need analysis on D3 Program for Mechanical Engineering Department at the State University of Padang, to ensure the compatibility between Model of Study and problems and need. This study aims to analyze how Problem-Based Learning effects on the course of Technology and Material Testing for the students of D3 Program of Mechanical Engineering of the State University of Padang.

  2. Forcefields based molecular modeling on the mechanical and physical properties of emeraldine base polyaniline

    Chen, X.; Yuan, C.A.; Wong, K.Y.; Zhang, G.Q.

    2010-01-01

    Molecular dynamics (MD) and molecular mechanical (MM) analysis are carried out to provide reliable and accurate model for emeraldine base polyaniline. This study validate the forcefields and model with the physical and mechanical properties of the polyaniline. The temperature effects on non-bond

  3. Analysis of Environmental Law Enforcement Mechanism Based on Economic Principle

    Cao, Hongjun; Shao, Haohao; Cai, Xuesen

    2017-11-01

    Strengthening and improving the environmental law enforcement mechanism is an important way to protect the ecological environment. This paper is based on economical principles, we did analysis of the marginal management costs by using Pigou means and the marginal transaction costs by using Coase means vary with the quantity growth of pollutant discharge Enterprises. We analyzed all this information, then we got the conclusion as follows. In the process of strengthening the environmental law enforcement mechanism, firstly, we should fully mobilize all aspects of environmental law enforcement, such as legislative bodies and law enforcement agencies, public welfare organizations, television, newspapers, enterprises, people and so on, they need to form a reasonable and organic structure system; then we should use various management means, such as government regulation, legal sanctions, fines, persuasion and denounce, they also need to form an organic structural system.

  4. Contract-Based Incentive Mechanism for Mobile Crowdsourcing Networks

    Nan Zhao

    2017-09-01

    Full Text Available Mobile crowdsourcing networks (MCNs are a promising method of data collecting and processing by leveraging the mobile devices’ sensing and computing capabilities. However, because of the selfish characteristics of the service provider (SP and mobile users (MUs, crowdsourcing participants only aim to maximize their own benefits. This paper investigates the incentive mechanism between the above two parties to create mutual benefits. By modeling MCNs as a labor market, a contract-based crowdsourcing model with moral hazard is proposed under the asymmetric information scenario. In order to incentivize the potential MUs to participate in crowdsourcing tasks, the optimization problem is formulated to maximize the SP’s utility by jointly examining the crowdsourcing participants’ risk preferences. The impact of crowdsourcing participants’ attitudes of risks on the incentive mechanism has been studied analytically and experimentally. Numerical simulation results demonstrate the effectiveness of the proposed contract design scheme for the crowdsourcing incentive.

  5. Mechanism-based Enzyme Inactivators of Phytosterol Biosynthesis

    W. David Nes

    2004-03-01

    Full Text Available Current progress on the mechanism and substrate recognition by sterol methyl transferase (SMT, the role of mechanism-based inactivators, other inhibitors of SMT action to probe catalysis and phytosterol synthesis is reported. SMT is a membrane-bound enzyme which catalyzes the coupled C-methylation-deprotonation reaction of sterol acceptor molecules generating the 24-alkyl sterol side chains of fungal ergosterol and plant sitosterol. This C-methylation step can be rate-limiting in the post-lanosterol (fungal or post-cycloartenol (plant pathways. A series of sterol analogs designed to impair SMT activity irreversibly have provided deep insight into the C-methylation reaction and topography of the SMT active site and as reviewed provide leads for the development of antifungal agents.

  6. A security mechanism based on evolutionary game in fog computing.

    Sun, Yan; Lin, Fuhong; Zhang, Nan

    2018-02-01

    Fog computing is a distributed computing paradigm at the edge of the network and requires cooperation of users and sharing of resources. When users in fog computing open their resources, their devices are easily intercepted and attacked because they are accessed through wireless network and present an extensive geographical distribution. In this study, a credible third party was introduced to supervise the behavior of users and protect the security of user cooperation. A fog computing security mechanism based on human nervous system is proposed, and the strategy for a stable system evolution is calculated. The MATLAB simulation results show that the proposed mechanism can reduce the number of attack behaviors effectively and stimulate users to cooperate in application tasks positively.

  7. Mechanical properties of tannin-based rigid foams undergoing compression

    Celzard, A., E-mail: Alain.Celzard@enstib.uhp-nancy.fr [Institut Jean Lamour - UMR CNRS 7198, CNRS - Nancy-Universite - UPV-Metz, Departement Chimie et Physique des Solides et des Surfaces, ENSTIB, 27 rue du Merle Blanc, BP 1041, 88051 Epinal cedex 9 (France); Zhao, W. [Institut Jean Lamour - UMR CNRS 7198, CNRS - Nancy-Universite - UPV-Metz, Departement Chimie et Physique des Solides et des Surfaces, ENSTIB, 27 rue du Merle Blanc, BP 1041, 88051 Epinal cedex 9 (France); Pizzi, A. [ENSTIB-LERMAB, Nancy-University, 27 rue du Merle Blanc, BP 1041, 88051 Epinal cedex 9 (France); Fierro, V. [Institut Jean Lamour - UMR CNRS 7198, CNRS - Nancy-Universite - UPV-Metz, Departement Chimie et Physique des Solides et des Surfaces, ENSTIB, 27 rue du Merle Blanc, BP 1041, 88051 Epinal cedex 9 (France)

    2010-06-25

    The mechanical properties of a new class of extremely lightweight tannin-based materials, namely organic foams and their carbonaceous counterparts are detailed. Scaling laws are shown to describe correctly the observed behaviour. Information about the mechanical characteristics of the elementary forces acting within these solids is derived. It is suggested that organic materials present a rather bending-dominated behaviour and are partly plastic. On the contrary, carbon foams obtained by pyrolysis of the former present a fracture-dominated behaviour and are purely brittle. These conclusions are supported by the differences in the exponent describing the change of Young's modulus as a function of relative density, while that describing compressive strength is unchanged. Features of the densification strain also support such conclusions. Carbon foams of very low density may absorb high energy when compressed, making them valuable materials for crash protection.

  8. A security mechanism based on evolutionary game in fog computing

    Yan Sun

    2018-02-01

    Full Text Available Fog computing is a distributed computing paradigm at the edge of the network and requires cooperation of users and sharing of resources. When users in fog computing open their resources, their devices are easily intercepted and attacked because they are accessed through wireless network and present an extensive geographical distribution. In this study, a credible third party was introduced to supervise the behavior of users and protect the security of user cooperation. A fog computing security mechanism based on human nervous system is proposed, and the strategy for a stable system evolution is calculated. The MATLAB simulation results show that the proposed mechanism can reduce the number of attack behaviors effectively and stimulate users to cooperate in application tasks positively.

  9. Human SUV3 helicase regulates growth rate of the HeLa cells and can localize in the nucleoli.

    Szewczyk, Maciej; Fedoryszak-Kuśka, Natalia; Tkaczuk, Katarzyna; Dobrucki, Jurek; Waligórska, Agnieszka; Stępień, Piotr P

    2017-01-01

    The human SUV3 helicase (SUV3, hSUV3, SUPV3L1) is a DNA/RNA unwinding enzyme belonging to the class of DexH-box helicases. It localizes predominantly in the mitochondria, where it forms an RNA-degrading complex called mitochondrial degradosome with exonuclease PNP (polynucleotide phosphorylase). Association of this complex with the polyA polymerase can modulate mitochondrial polyA tails. Silencing of the SUV3 gene was shown to inhibit the cell cycle and to induce apoptosis in human cell lines. However, since small amounts of the SUV3 helicase were found in the cell nuclei, it was not clear whether the observed phenotypes of SUV3 depletion were of mitochondrial or nuclear origin. In order to answer this question we have designed gene constructs able to inhibit the SUV3 activity exclusively in the cell nuclei. The results indicate that the observed growth rate impairment upon SUV3 depletion is due to its nuclear function(s). Unexpectedly, overexpression of the nuclear-targeted wild-type copies of the SUV3 gene resulted in a higher growth rate. In addition, we demonstrate that the SUV3 helicase can be found in the HeLa cell nucleoli, but it is not detectable in the DNA-repair foci. Our results indicate that the nucleolar-associated human SUV3 protein is an important factor in regulation of the cell cycle.

  10. Regulation of gene expression by the BLM helicase correlates with the presence of G-quadruplex DNA motifs

    Nguyen, Giang Huong; Tang, Weiliang; Robles, Ana I

    2014-01-01

    Bloom syndrome is a rare autosomal recessive disorder characterized by genetic instability and cancer predisposition, and caused by mutations in the gene encoding the Bloom syndrome, RecQ helicase-like (BLM) protein. To determine whether altered gene expression might be responsible for pathologic...

  11. Molecular architecture of the recombinant human MCM2-7 helicase in complex with nucleotides and DNA

    Boskovic, Jasminka; Bragado-Nilsson, Elisabeth; Saligram Prabhakar, Bhargav

    2016-01-01

    DNA replication is a key biological process that involves different protein complexes whose assembly is rigorously regulated in a successive order. One of these complexes is a replicative hexameric helicase, the MCM complex, which is essential for the initiation and elongation phases of replicati...

  12. Ufd1-Npl4 Recruit Cdc48 for Disassembly of Ubiquitylated CMG Helicase at the End of Chromosome Replication

    Marija Maric

    2017-03-01

    Full Text Available Disassembly of the Cdc45-MCM-GINS (CMG DNA helicase is the key regulated step during DNA replication termination in eukaryotes, involving ubiquitylation of the Mcm7 helicase subunit, leading to a disassembly process that requires the Cdc48 “segregase”. Here, we employ a screen to identify partners of budding yeast Cdc48 that are important for disassembly of ubiquitylated CMG helicase at the end of chromosome replication. We demonstrate that the ubiquitin-binding Ufd1-Npl4 complex recruits Cdc48 to ubiquitylated CMG. Ubiquitylation of CMG in yeast cell extracts is dependent upon lysine 29 of Mcm7, which is the only detectable site of ubiquitylation both in vitro and in vivo (though in vivo other sites can be modified when K29 is mutated. Mutation of K29 abrogates in vitro recruitment of Ufd1-Npl4-Cdc48 to the CMG helicase, supporting a model whereby Ufd1-Npl4 recruits Cdc48 to ubiquitylated CMG at the end of chromosome replication, thereby driving the disassembly reaction.

  13. RNA helicase DDX3 is a regulatory subunit of casein kinase 1 in Wnt-beta-catenin signaling

    Cruciat, C.M.; Dolde, C.; de Groot, R.E.; Ohkawara, B.; Reinhard, C.; Korswagen, H.C.; Niehrs, C.

    2013-01-01

    Casein kinase 1 (CK1) members play key roles in numerous biological processes. They are considered "rogue" kinases, because their enzymatic activity appears unregulated. Contrary to this notion, we have identified the DEAD-box RNA helicase DDX3 as a regulator of the Wnt-beta-catenin network, where

  14. Mechanism of chain formation in nanofluid based MR fluids

    Patel, Rajesh

    2011-01-01

    Mechanism of structure formation in bidispersed colloids is important for its physical and optical properties. It is microscopically observed that the mechanism of chain formation in magnetic nanofluid based magnetorheological (MR) fluid is quite different from that in the conventional MR fluid. Under the application of magnetic field the magnetic nanoparticles are filled inside the structural microcavities formed due to the association of large magnetic particles, and some of the magnetic nanoparticles are attached at the end of the chains formed by the large particles. The dipolar energy of the large particles in a magnetic nanofluid matrix becomes effective magnetic permeability (μ eff ) times smaller than that of the neutral medium. Inclusion of magnetic nanoparticles (∼10 nm) with large magnetic particles (∼3-5 μm) restricts the aggregation of large particles, which causes the field induced phase separation in MR fluids. Hence, nanofluid based MR fluids are more stable than conventional MR fluids, which subsequently increase their application potentiality. - Research highlights: → In bidispersed magnetic colloids nanoparticles are attached at the end of the chains formed by the large particles. → Inclusion of magnetic nanoparticles (∼10 nm) with large magnetic particles (∼3-5 m) restricts the aggregation of large particles. → Nanofluid based MR fluids are more stable than conventional MR fluids.

  15. Video-based problems in introductory mechanics physics courses

    Gröber, Sebastian; Klein, Pascal; Kuhn, Jochen

    2014-01-01

    Introductory mechanics physics courses at the transition from school to university are a challenge for students. They are faced with an abrupt and necessary increase of theoretical content and requirements on their conceptual understanding of phyiscs. In order to support this transition we replaced part of the mandatory weekly theory-based paper-and-pencil problems with video analysis problems of equal content and level of difficulty. Video-based problems (VBP) are a new problem format for teaching physics from a linked sequence of theoretical and video-based experimental tasks. Experimental tasks are related to the well-known concept of video motion analysis. This introduction of an experimental part in recitations allows the establishment of theory–experiment interplay as well as connections between physical content and context fields such as nature, technique, everyday life and applied physics by conducting model-and context-related experiments. Furthermore, laws and formulas as predominantly representative forms are extended by the use of diagrams and vectors. In this paper we give general reasons for this approach, describe the structure and added values of VBP, and show that they cover a relevant part of mechanics courses at university. Emphasis is put on theory–experiment interplay as a structural added value of VBP to promote students' construction of knowledge and conceptual understanding. (paper)

  16. Emerging importance of helicases in plant stress tolerance: characterization of Oryza sativa repair helicase XPB2 promoter and its functional validation in tobacco under multiple stresses

    Shailendra eRaikwar

    2015-12-01

    Full Text Available Genetic material always remains at the risk of spontaneous or induced damage which challenges the normal functioning of DNA molecule, thus, DNA repair is vital to protect the organisms against genetic damage. DNA hHelicases, the unique molecular motors, are emerged as potentialprospective molecules to engineer stress tolerance in plants and are involved in a variety of DNA nucleic acid metabolismc processes including DNA repair. The DNA repair helicase, OsXPB2 is an evolutionary conserved protein present in different organisms, including plants. Availability of few efficient promoters for gene expression in plants provoked us to study the promoter of XPB for better understanding of gene regulation under stress The analysis of promoter sequence from plant genome is important in understanding the gene regulation. Hereconditions. Here, we report the in silico analysis of novel stress inducible promoter of rice Oryza sativa OsXPB2 (OsXPB2. gene is reported. The in vivo validation of functionality/activity of novel stress inducible promoter of rice OsXPB2 gene promoter under abiotic and hormonal stress conditions was performed by Agrobacterium-mediated transient assay in tobacco leaves using OsXPB2::GUS chimeric construct. Our resultsThe present research revealed that OsXPB2 promoter contains cis-elements accounting for various abiotic stresses (salt, dehydration or cold and hormone (Auxin, ABA or MeJA induced GUS expression/activity in the promoter-reporter assay. The promoter region of OsXPB2 contains CACG, GTAACG, CACGTG, CGTCA CCGCCGCGCT cis acting-elements which are reported to be salt, dehydration, cold, MeJA or ABA responsive, respectively. Functional analysis was done by Agrobacterium-transient assays using agroinfiltration in tobacco leaves, followed by GUS staining and fluorescence quantitative analyses. The results revealed high induction of GUS activity under multiple abiotic stresses as compared to mock treated control. The present

  17. Development of Numerical Analysis Techniques Based on Damage Mechanics and Fracture Mechanics

    Chang, Yoon Suk; Lee, Dock Jin; Choi, Shin Beom; Kim, Sun Hye; Cho, Doo Ho; Lee, Hyun Boo

    2010-04-01

    The scatter of measured fracture toughness data and transferability problems among different crack configurations as well as geometry and loading conditions are major obstacles for application of fracture mechanics. To address these issues, recently, concerns on the local approach employing reliable micro-mechanical damage models are being increased again in connection with a progress of computational technology. In the present research, as part of development of fracture mechanical evaluation model for material degradation of reactor pressure boundary, several investigations on fracture behaviors were carried out. Especially, a numerical scheme to determine key parameters consisting both cleavage and ductile fracture estimate models was changed efficiently by incorporating a genetic algorithm. Also, with regard to the well-known master curve, newly reported methods such as bimodal master curve, randomly inhomogeneous master curve and single point estimation were reviewed to deal with homogeneous and inhomogeneous material characteristics. A series of preliminary finite element analyses was conducted to examine the element size effect on micro-mechanical models. Then, a new thickness correction equation was derived from parametric three-dimensional numerical simulations, which was founded on the current test standard, ASTM E1921, but could lead to get more realistic fracture toughness values. As a result, promising modified master curves as well as fracture toughness diagrams to convert data between pre-cracked V-notched and compact tension specimens were generated. Moreover, a user-subroutine in relation to GTN(Gurson-Tvergaard-Needleman) model was made by adopting Hill's 48 yield potential theory. By applying GTN model combined with the subroutine to small punch specimens, the effect of inhomogeneous properties on fracture behaviors of miniature specimens was confirmed. Therefore, it is anticipated that the aforementioned enhanced research results can be utilized

  18. Microscopic machining mechanism of polishing based on vibrations of liquid

    Huang, Z G; Guo, Z N; Chen, X; Yu, Z Q; Yu, T M; Lee, W B

    2007-01-01

    A molecular dynamics method has been applied to study the mechanism of polishing based on vibrations of liquid. Movements of polishing particles and formations of impact dents are simulated and discussed. The abrasive effect between particle and machined substrate is evaluated empirically. Polishing qualities, including roughness and fractal character under multiple impacts, are obtained by numerical methods. Results show that the particle will vibrate and roll viscously on the substrate. Press, tear and self-organization effects will be responsible for the formation of impact dents. Simulation results are compared with experimental data to verify the conclusions

  19. Parametric Design and Mechanical Analysis of Beams based on SINOVATION

    Xu, Z. G.; Shen, W. D.; Yang, D. Y.; Liu, W. M.

    2017-07-01

    In engineering practice, engineer needs to carry out complicated calculation when the loads on the beam are complex. The processes of analysis and calculation take a lot of time and the results are unreliable. So VS2005 and ADK are used to develop a software for beams design based on the 3D CAD software SINOVATION with C ++ programming language. The software can realize the mechanical analysis and parameterized design of various types of beams and output the report of design in HTML format. Efficiency and reliability of design of beams are improved.

  20. Giant Glial Cell: New Insight Through Mechanism-Based Modeling

    Postnov, D. E.; Ryazanova, L. S.; Brazhe, Nadezda

    2008-01-01

    The paper describes a detailed mechanism-based model of a tripartite synapse consisting of P- and R-neurons together with a giant glial cell in the ganglia of the medical leech (Hirudo medicinalis), which is a useful object for experimental studies in situ. We describe the two main pathways...... of the glial cell activation: (1) via IP3 production and Ca2+ release from the endoplasmic reticulum and (2) via increase of the extracellular potassium concentration, glia depolarization, and opening of voltage-dependent Ca2+ channels. We suggest that the second pathway is the more significant...

  1. Marine Ecological Environment Management Based on Ecological Compensation Mechanisms

    Qunzhen Qu

    2016-12-01

    Full Text Available The level of marine environmental management is a key factor in the successful implementation of marine power strategies. The improvement in management levels of marine environments requires innovation in marine management. In other words, the transformation of marine environmental management into marine ecological environment management must be done in order to achieve sustainable development of the marine economy. As an environmental economic policy that combines both administrative and market measures, ecological compensation mechanisms have significant advantages in marine ecological environment management. Based on the study of the current development of ecological compensation mechanisms in China, through the analysis of the connotation of marine ecological civilization, existing marine ecological protection practices and marine environmental management methods, this paper posits that the current marine ecological environment management in China should be established on the basis of ecological compensation mechanisms. At present, a lack of laws and regulations for overall marine ecological environment management is the key factor restricting the practice of marine ecological environment management. Therefore, it is necessary to explore the current path of marine ecological environment management in China from the perspective of the construction of legal system of ecological compensation law, the establishment of ecological compensation fees, ecological taxes and ecological compensation fund systems, and the clear status for a marine ecological management and supervision body.

  2. Actuation mechanisms of carbon nanotube-based architectures

    Geier, Sebastian; Mahrholz, Thorsten; Wierach, Peter; Sinapius, Michael

    2016-04-01

    State of the art smart materials such as piezo ceramics or electroactive polymers cannot feature both, mechanical stiffness and high active strain. Moreover, properties like low density, high mechanical stiffness and high strain at the same time driven by low energy play an increasingly important role for their future application. Carbon nanotubes (CNT), show this behavior. Their active behavior was observed 1999 the first time using paper-like mats made of CNT. Therefore the CNT-papers are electrical charged within an electrolyte thus forming a double- layer. The measured deflection of CNT material is based on the interaction between the charged high surface area formed by carbon nanotubes and ions provided by the electrolyte. Although CNT-papers have been extensively analyzed as well at the macro-scale as nano-scale there is still no generally accepted theory for the actuation mechanism. This paper focuses on investigations of the actuation mechanisms of CNT-papers in comparison to vertically aligned CNT-arrays. One reason of divergent results found in literature might be attributed to different types of CNT samples. While CNT-papers represent architectures of short CNTs which need to bridge each other to form the dimensions of the sample, the continuous CNTs of the array feature a length of almost 3 mm, along which the experiments are carried out. Both sample types are tested within an actuated tensile test set-up under different conditions. While the CNT-papers are tested in water-based electrolytes with comparably small redox-windows the hydrophobic CNT-arrays are tested in ionic liquids with comparatively larger redox-ranges. Furthermore an in-situ micro tensile test within an SEM is carried out to prove the optimized orientation of the MWCNTs as result of external load. It was found that the performance of CNT-papers strongly depends on the test conditions. However, the CNT-arrays are almost unaffected by the conditions showing active response at negative

  3. Regulator of telomere elongation helicase 1 (RTEL1) rs6010620 polymorphism contribute to increased risk of glioma.

    Zhao, Wei; Bian, Yusong; Zhu, Wei; Zou, Peng; Tang, Guotai

    2014-06-01

    Regulator of telomere elongation helicase 1 (RTEL1) is critical for genome stability and tumor avoidance. Many studies have reported the associations of RTEL1 rs6010620 with glioma risk, but individually published results were inconclusive. This meta-analysis was performed to quantitatively summarize the evidence for such a relationship. The PubMed, Embase, and Web of Science were systematically searched to identify relevant studies. The odds ratio (OR) and 95 % confidence interval (95 % CI) were computed to estimate the strength of the association using a fixed or random effects model. Ten studies were eligible for meta-analysis including data on glioma with 6,490 cases and 9,288 controls. Overall, there was a significant association between RTEL1 rs6010620 polymorphism and glioma risk in all four genetic models (GG vs. AA: OR=1.87, 95 % CI=1.60-2.18, P heterogeneity=0.552; GA vs. AA: OR=1.30, 95 % CI=1.16-1.46, P heterogeneity=0.495; dominant model-GG+GA vs. AA: OR=1.46, 95 % CI=1.31-1.63, P heterogeneity=0.528; recessive model-GG vs. GA+AA: OR=1.36, 95 % CI=1.27-1.46, P heterogeneity=0.093). Subgroup analyses by ethnicity showed that RTEL1 rs6010620 polymorphism resulted in a higher risk of glioma among both Asians and Caucasians. In the stratified analysis by ethnicity and source of controls, significantly increased risk was observed for Asians and Europeans in all genetic models, population-based studies in all genetic models, and hospital-based studies in three genetic models (heterozygote comparison, homozygote comparison, and dominant model). Our meta-analysis suggested that RTEL1 rs6010620 polymorphism is likely to be associated with increased glioma risk, which lends further biological plausibility to these findings.

  4. Fe-based composite materials with improved mechanical properties

    Werniewicz, Katarzyna [IFW Dresden, P.O. Box 270116, D-01171 Dresden (Germany); Warsaw University of Technology, Faculty of Materials Science and Engineering, Warsaw (Poland); Kuehn, Uta; Mattern, Norbert; Eckert, Juergen; Schultz, Ludwig [IFW Dresden, P.O. Box 270116, D-01171 Dresden (Germany); Kulik, Tadeusz [Warsaw University of Technology, Faculty of Materials Science and Engineering, Warsaw (Poland)

    2008-07-01

    Following a previous study by the authors two new compositions (Fe{sub 89.0}Cr{sub 5.5}Mo{sub 5.5}){sub 91}C{sub 9} and (Fe{sub 89.0}Cr{sub 5.5}Mo{sub 5.5}){sub 83}C{sub 17} have been developed with the aim of improving the ductility of Fe{sub 65.5}Cr{sub 4}Mo{sub 4}Ga{sub 4}P{sub 12}C{sub 5}B{sub 5.5} bulk metallic glass (BMG). In contrast to the alloys in that study, the recently prepared Fe-based materials are Ga-free. It was expected that the variations in the composition will lead to the changes in the phase formation and, hence, in the mechanical response of the investigated alloys. It was recognized that in-situ formed Fe-based composites show superior plasticity ({epsilon}{sub pl}{approx}37%) for the alloy with lower C content and ({epsilon}{sub pl}{approx}4%) for the alloy with higher C content compared to monolithic glass ({epsilon}{sub pl}{approx}0.2%). Furthermore, on the basis of present as well as previous investigations it has been shown that the Ga addition is beneficial for the plasticity of these Fe-based alloys. It was observed that the (Fe{sub 89.0}Cr{sub 5.5}Mo{sub 5.5}){sub 83}C{sub 17} alloy exhibits a significantly smaller fracture strain ({epsilon}{sub f}{approx}5%) compared to its Ga-containing counterpart ({epsilon}{sub f}{approx}16%). Therefore, it can be concluded that appropriate alloying additions are crucial in enhancing the mechanical properties of the complex Fe-based materials developed here.

  5. Maintainability design and evaluation of mechanical systems based on tribology

    Wani, M.F.; Gandhi, O.P.

    2002-01-01

    Maintainability of mechanical systems based on tribology is suggested and evaluated in this paper. Tribo-features of mechanical systems, which characterise maintainability are identified and are modelled in terms of tribo-maintainability digraph. The nodes in the digraph represent the tribo-features and edges represent the degree of influence among the features. A matrix, one to one representation of the digraph, is defined to develop system maintainability expression (SPF-t) based on the tribology. It is also useful in comparing the various design alternatives from tribo-maintainability point of view. Maintainability is evaluated from the tribo-maintainability index, obtained from SPF-t (i.e. permanent of the matrix) by substituting the numerical values of the features and their interdependence. A higher value of the index implies better maintainability of the systems. The proposed methodology also guides designers in enhancing the maintainability of a system by appropriately incorporating tribo-features. An example to illustrate the methodology is also presented

  6. A damage mechanics based general purpose interface/contact element

    Yan, Chengyong

    Most of the microelectronics packaging structures consist of layered substrates connected with bonding materials, such as solder or epoxy. Predicting the thermomechanical behavior of these multilayered structures is a challenging task in electronic packaging engineering. In a layered structure the most complex part is always the interfaces between the strates. Simulating the thermo-mechanical behavior of such interfaces, is the main theme of this dissertation. The most commonly used solder material, Pb-Sn alloy, has a very low melting temperature 180sp°C, so that the material demonstrates a highly viscous behavior. And, creep usually dominates the failure mechanism. Hence, the theory of viscoplasticity is adapted to describe the constitutive behavior. In a multilayered assembly each layer has a different coefficient of thermal expansion. Under thermal cycling, due to heat dissipated from circuits, interfaces and interconnects experience low cycle fatigue. Presently, the state-of-the art damage mechanics model used for fatigue life predictions is based on Kachanov (1986) continuum damage model. This model uses plastic strain as a damage criterion. Since plastic strain is a stress path dependent value, the criterion does not yield unique damage values for the same state of stress. In this dissertation a new damage evolution equation based on the second law of thermodynamic is proposed. The new criterion is based on the entropy of the system and it yields unique damage values for all stress paths to the final state of stress. In the electronics industry, there is a strong desire to develop fatigue free interconnections. The proposed interface/contact element can also simulate the behavior of the fatigue free Z-direction thin film interconnections as well as traditional layered interconnects. The proposed interface element can simulate behavior of a bonded interface or unbonded sliding interface, also called contact element. The proposed element was verified against

  7. An EV Charging Scheduling Mechanism Based on Price Negotiation

    Baocheng Wang

    2018-05-01

    Full Text Available Scheduling EV user’s charging behavior based on charging price and applying renewable energy resources are the effective methods to release the load pressure of power grids brought about by the large-scale popularity of electric vehicles (EVs. This paper presents a novel approach for EV charging scheduling based on price negotiation. Firstly, the EV charging system framework based on price negotiation and renewable energy resources is discussed. Secondly, the price negotiation model is presented, including the initial price models and the conditions of transactions. Finally, an EV charging scheduling mechanism based on price negotiation (CSM-PN, including the price adjustment strategies of both the operator and EV users is proposed to seek a final transaction during multi-round price negotiation. Simulation results show that this novel approach can effectively improve the charging station operator’s income, reduce the EV users’ costs, and balance the load of the power grid while improving the efficiency of the EV charging system.

  8. Gallium nitride-based micro-opto-electro-mechanical systems

    Stonas, Andreas Robert

    Gallium Nitride and its associated alloys InGaN and AlGaN have many material properties that are highly desirable for micro-electro-mechanical systems (MEMS), and more specifically micro-opto-electro-mechanical systems (MOEMS). The group III-nitrides are tough, stiff, optically transparent, direct bandgap, chemically inert, highly piezoelectric, and capable of functioning at high temperatures. There is currently no other semiconductor system that possesses all of these properties. Taken together, these attributes make the nitrides prime candidates not only for creating new versions of existing device structures, but also for creating entirely unique devices which combine these properties in novel ways. Unfortunately, their chemical resiliency also makes the group III-nitrides extraordinarily difficult to shape into devices. In particular, until this research, no undercut etch technology existed that could controllably separate a selected part of a MEMS device from its sapphire or silicon carbide substrate. This has effectively prevented GaN-based MEMS from being developed. This dissertation describes how this fabrication obstacle was overcome by a novel etching geometry (bandgap-selective backside-illuminated photoelectochemical (BS-BIPEC) etching) and its resulting morphologies. Several gallium-nitride based MEMS devices were created, actuated, and modelled, including cantilevers and membranes. We describe in particular our pursuit of one of the many novel device elements that is possible only in this material system: a transducer that uses an externally applied strain to dynamically change the optical transition energy of a quantum well. While the device objective of a dynamically tunable quantum well was not achieved, we have demonstrated sufficient progress to believe that such a device will be possible soon. We have observed a shift (5.5meV) of quantum well transition energies in released structures, and we have created structures that can apply large biaxial

  9. Effect of addition of organo clay on mechanical properties and dynamic-mechanical based TPV

    Honorato, Luciana R.; Silva, Adriana A.; Soares, Bluma G.; Soares, Ketly P.

    2011-01-01

    The effect of organophilic clay on the mechanical and dynamical-mechanical properties of thermoplastic elastomers based on polypropylene (PP) and nitrile rubber (NBR) was investigated. The addition of clay was performed from a master batch prepared by a solution intercalation of NBR inside the clay galleries. Since the PP/NBR blend is highly incompatible, PP functionalized with maleic anhydride (PP-g-MA) and carboxylated NBR (XNBR) were employed as compatibilizing system together with triethylene-tetramine (TETA) used as coupling agent. The addition of Clay inside the elastomeric phase of the TPV resulted in a significant decrease of the elongation at break without changes on the tensile strength. The presence of clay also promoted a slight increase of the storage modulus and the glass transition temperature. The small angle X ray scattering confirmed the high dispersion of clay inside the TPV. Analysis of light scattering small angle (SAXS) confirmed the high dispersion of clay in the matrix of the TPV. (author)

  10. FBH1 helicase disrupts RAD51 filaments in vitro and modulates homologous recombination in mammalian cells

    Simandlova, Jitka; Zagelbaum, Jennifer; Payne, Miranda J

    2013-01-01

    Efficient repair of DNA double strand breaks and interstrand cross-links requires the homologous recombination (HR) pathway, a potentially error-free process that utilizes a homologous sequence as a repair template. A key player in HR is RAD51, the eukaryotic ortholog of bacterial RecA protein. RAD......51 can polymerize on DNA to form a nucleoprotein filament that facilitates both the search for the homologous DNA sequences and the subsequent DNA strand invasion required to initiate HR. Because of its pivotal role in HR, RAD51 is subject to numerous positive and negative regulatory influences...... filaments on DNA through its ssDNA translocase function. Consistent with this, a mutant mouse embryonic stem cell line with a deletion in the FBH1 helicase domain fails to limit RAD51 chromatin association and shows hyper-recombination. Our data are consistent with FBH1 restraining RAD51 DNA binding under...

  11. RTEL1 is a replisome-associated helicase that promotes telomere and genome-wide replication.

    Vannier, Jean-Baptiste; Sandhu, Sumit; Petalcorin, Mark I R; Wu, Xiaoli; Nabi, Zinnatun; Ding, Hao; Boulton, Simon J

    2013-10-11

    Regulator of telomere length 1 (RTEL1) is an essential DNA helicase that disassembles telomere loops (T loops) and suppresses telomere fragility to maintain the integrity of chromosome ends. We established that RTEL1 also associates with the replisome through binding to proliferating cell nuclear antigen (PCNA). Mouse cells disrupted for the RTEL1-PCNA interaction (PIP mutant) exhibited accelerated senescence, replication fork instability, reduced replication fork extension rates, and increased origin usage. Although T-loop disassembly at telomeres was unaffected in the mutant cells, telomere replication was compromised, leading to fragile sites at telomeres. RTEL1-PIP mutant mice were viable, but loss of the RTEL1-PCNA interaction accelerated the onset of tumorigenesis in p53-deficient mice. We propose that RTEL1 plays a critical role in both telomere and genome-wide replication, which is crucial for genetic stability and tumor avoidance.

  12. Association between regulator of telomere elongation helicase1 (RTEL1) gene and HAPE risk

    Rong, Hao; He, Xue; Zhu, Linhao; Zhu, Xikai; Kang, Longli; Wang, Li; He, Yongjun; Yuan, Dongya; Jin, Tianbo

    2017-01-01

    Abstract High altitude pulmonary edema (HAPE) is a paradigm of pulmonary edema. Mutations in regulator of telomere elongation helicase1 (RTEL1) represent an important contributor to risk for pulmonary fibrosis. However, little information is found about the association between RTEL1 and HAPE risk. The present study was undertaken to tentatively explore the potential relation between single-nucleotide polymorphisms (SNPs) in RTEL1 and HAPE risk in Chinese Han population. A total of 265 HAPE patients and 303 healthy controls were included in our case-control study. Four SNPs in RTEL1 were selected and genotyped using the Sequenom MassARRAY method. Odds ratios (ORs) and 95% confidence intervals (95% CIs) were calculated by unconditional logistic regression with adjustment for gender and age. All P values were Bonferroni corrected, and statistical significance was set at P RTEL1 and a decreased risk HAPE in the Chinese population. The results need further confirmation. PMID:28953687

  13. Mechanical properties of nanostructured nickel based superalloy Inconel 718

    Mukhtarov, Sh; Ermachenko, A, E-mail: shamil@anrb.r [Institute for Metals Superplasticity Problems RAS, 39, Khalturina, Ufa, 450001 (Russian Federation)

    2010-07-01

    This paper will describe the investigations of a nanostructured (NS) state of nickel based INCONEL alloy 718. This structure was generated in bulk semiproducts by severe plastic deformation (SPD) via multiple isothermal forging (MIF) of a coarse-grained alloy. The initial structure consisted of {gamma}-phase grains with disperse precipitations of {gamma}{sup -}phase in the forms of discs, 50-75 nm in diameter and 20 nm in thickness. The MIF generated structures possess a large quantity of non-coherent plates and rounded precipitations of {delta}-phase, primarily along grain boundaries. In the duplex ({gamma}+{delta}) structure the grains have high dislocation density and a large number of nonequilibrium boundaries. Investigations to determine mechanical properties of the alloy in a nanostructured state were carried out. Nanocrystalline Inconel 718 (80 nm) possesses a very high room-temperature strength after SPD. Microcrystalline (MC) and NS states of the alloy were subjected to strengthening thermal treatment, and the obtained results were compared in order to determine their mechanical properties at room and elevated temperatures.

  14. Failure Mechanism of Rock Bridge Based on Acoustic Emission Technique

    Guoqing Chen

    2015-01-01

    Full Text Available Acoustic emission (AE technique is widely used in various fields as a reliable nondestructive examination technology. Two experimental tests were carried out in a rock mechanics laboratory, which include (1 small scale direct shear tests of rock bridge with different lengths and (2 large scale landslide model with locked section. The relationship of AE event count and record time was analyzed during the tests. The AE source location technology and comparative analysis with its actual failure model were done. It can be found that whether it is small scale test or large scale landslide model test, AE technique accurately located the AE source point, which reflected the failure generation and expansion of internal cracks in rock samples. Large scale landslide model with locked section test showed that rock bridge in rocky slope has typical brittle failure behavior. The two tests based on AE technique well revealed the rock failure mechanism in rocky slope and clarified the cause of high speed and long distance sliding of rocky slope.

  15. Photoprotection in Plants Optical Screening-based Mechanisms

    Solovchenko, Alexei

    2010-01-01

    Optical screening of excessive and potentially harmful solar radiation is an important photoprotective mechanism, though it has received much less attention in comparison with other systems preventing photooxidative damage to photoautotrophic organisms. This photoprotection in the form of screening appears to be especially important for juvenile and senescing plants as well as under environmental stresses—i.e. in situations where the efficiency of enzymatic ROS elimination, DNA repair and other ‘classical’ photoprotective systems could be impaired. This book represents an attempt to develop an integral view of optical screening-based photoprotection in microalgae and higher plants. Towards this end, the key groups of pigments involved in the screening of ultraviolet and visible components of solar radiation in microalgae and higher plants, and the patterns of their accumulation and distribution within plant cells and tissues, are described. Special attention is paid to the manifestations of screening pi...

  16. Study on Mine Emergency Mechanism based on TARP and ICS

    Xi, Jian; Wu, Zongzhi

    2018-01-01

    By analyzing the experiences and practices of mine emergency in China and abroad, especially the United States and Australia, normative principle, risk management principle and adaptability principle of constructing mine emergency mechanism based on Trigger Action Response Plans (TARP) and Incident Command System (ICS) are summarized. Classification method, framework, flow and subject of TARP and ICS which are suitable for the actual situation of domestic mine emergency are proposed. The system dynamics model of TARP and ICS is established. The parameters such as evacuation ratio, response rate, per capita emergency capability and entry rate of rescuers are set up. By simulating the operation process of TARP and ICS, the impact of these parameters on the emergency process are analyzed, which could provide a reference and basis for building emergency capacity, formulating emergency plans and setting up action plans in the emergency process.

  17. Principle-Based Ethics and the Clean Development Mechanism

    Bay, Ingrid; Oughton, Deborah

    2003-01-01

    Search for common values can go a long way resolving international political and social differences, and represents an area where ethical evaluation can offer an input to international legislation. This paper argues that a principle-based, normative approach should be applied to evaluation of Clean Development Mechanism (CDM) projects and the goal of sustainable development (SD). Primafacie ethical principles to promote weil-being, dignity and justice can be defined and supported from international agreements on sustainable developments. To promote coherency, these principles should then be specifled according to particular CDM-projects, to produce a set of norms that covers: 1) the different dimension to sustainable development, 2) the distribution of costs and benefits across time and place, and 3) consequences for different affected parties, including future generations and the environment

  18. Mechanical Stability of Flexible Graphene-Based Displays.

    Anagnostopoulos, George; Pappas, Panagiotis-Nektarios; Li, Zheling; Kinloch, Ian A; Young, Robert J; Novoselov, Kostya S; Lu, Ching Yu; Pugno, Nicola; Parthenios, John; Galiotis, Costas; Papagelis, Konstantinos

    2016-08-31

    The mechanical behavior of a prototype touch panel display, which consists of two layers of CVD graphene embedded into PET films, is investigated in tension and under contact-stress dynamic loading. In both cases, laser Raman spectroscopy was employed to assess the stress transfer efficiency of the embedded graphene layers. The tensile behavior was found to be governed by the "island-like" microstructure of the CVD graphene, and the stress transfer efficiency was dependent on the size of graphene "islands" but also on the yielding behavior of PET at relatively high strains. Finally, the fatigue tests, which simulate real operation conditions, showed that the maximum temperature gradient developed at the point of "finger" contact after 80 000 cycles does not exceed the glass transition temperature of the PET matrix. The effect of these results on future product development and the design of new graphene-based displays are discussed.

  19. Mechanical Stability of Flexible Graphene-Based Displays

    2016-01-01

    The mechanical behavior of a prototype touch panel display, which consists of two layers of CVD graphene embedded into PET films, is investigated in tension and under contact-stress dynamic loading. In both cases, laser Raman spectroscopy was employed to assess the stress transfer efficiency of the embedded graphene layers. The tensile behavior was found to be governed by the “island-like” microstructure of the CVD graphene, and the stress transfer efficiency was dependent on the size of graphene “islands” but also on the yielding behavior of PET at relatively high strains. Finally, the fatigue tests, which simulate real operation conditions, showed that the maximum temperature gradient developed at the point of “finger” contact after 80 000 cycles does not exceed the glass transition temperature of the PET matrix. The effect of these results on future product development and the design of new graphene-based displays are discussed. PMID:27494211

  20. Trajectory data privacy protection based on differential privacy mechanism

    Gu, Ke; Yang, Lihao; Liu, Yongzhi; Liao, Niandong

    2018-05-01

    In this paper, we propose a trajectory data privacy protection scheme based on differential privacy mechanism. In the proposed scheme, the algorithm first selects the protected points from the user’s trajectory data; secondly, the algorithm forms the polygon according to the protected points and the adjacent and high frequent accessed points that are selected from the accessing point database, then the algorithm calculates the polygon centroids; finally, the noises are added to the polygon centroids by the differential privacy method, and the polygon centroids replace the protected points, and then the algorithm constructs and issues the new trajectory data. The experiments show that the running time of the proposed algorithms is fast, the privacy protection of the scheme is effective and the data usability of the scheme is higher.

  1. Retinitis Pigmentosa Mutations in Bad Response to Refrigeration 2 (Brr2) Impair ATPase and Helicase Activity.

    Ledoux, Sarah; Guthrie, Christine

    2016-06-03

    Brr2 is an RNA-dependent ATPase required to unwind the U4/U6 snRNA duplex during spliceosome assembly. Mutations within the ratchet helix of the Brr2 RNA binding channel result in a form of degenerative human blindness known as retinitis pigmentosa (RP). The biochemical consequences of these mutations on Brr2's RNA binding, helicase, and ATPase activity have not yet been characterized. Therefore, we identified the largest construct of Brr2 that is soluble in vitro, which truncates the first 247 amino acids of the N terminus (Δ247-Brr2), to characterize the effects of the RP mutations on Brr2 activity. The Δ247-Brr2 RP mutants exhibit a gradient of severity of weakened RNA binding, reduced helicase activity, and reduced ATPase activity compared with wild type Δ247-Brr2. The globular C-terminal Jab1/Mpn1-like domain of Prp8 increases the ability of Δ247-Brr2 to bind the U4/U6 snRNA duplex at high pH and increases Δ247-Brr2's RNA-dependent ATPase activity and the extent of RNA unwinding. However, this domain of Prp8 does not differentially affect the Δ247-Brr2 RP mutants compared with the wild type Δ247-Brr2. When stimulated by Prp8, wild type Δ247-Brr2 is able to unwind long stable duplexes in vitro, and even the RP mutants capable of binding RNA with tight affinity are incapable of fully unwinding short duplex RNAs. Our data suggest that the RP mutations within the ratchet helix impair Brr2 translocation through RNA helices. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  2. Human regulator of telomere elongation helicase 1 (RTEL1) is required for the nuclear and cytoplasmic trafficking of pre-U2 RNA

    Schertzer , Michael; Jouravleva , Karina; Perderiset , Mylène; Dingli , Florent; Loew , Damarys; Le Guen , Tangui; Bardoni , Barbara; De Villartay , Jean-Pierre; Revy , Patrick; Londono-Vallejo , Arturo

    2015-01-01

    International audience; Hoyeraal-Hreidarsson syndrome (HHS) is a severe form of Dyskeratosis congenita characterized by developmental defects, bone marrow failure and im-munodeficiency and has been associated with telom-ere dysfunction. Recently, mutations in Regulator of Telomere ELongation helicase 1 (RTEL1), a helicase first identified in Mus musculus as being responsible for the maintenance of long telomeres, have been identified in several HHS patients. Here we show that RTEL1 is require...

  3. Identification and Biochemical Characterization of Halisulfate 3 and Suvanine as Novel Inhibitors of Hepatitis C Virus NS3 Helicase from a Marine Sponge

    Atsushi Furuta

    2014-01-01

    Full Text Available Hepatitis C virus (HCV is an important etiological agent that is responsible for the development of chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma. HCV nonstructural protein 3 (NS3 helicase is a possible target for novel drug development due to its essential role in viral replication. In this study, we identified halisulfate 3 (hal3 and suvanine as novel NS3 helicase inhibitors, with IC50 values of 4 and 3 µM, respectively, from a marine sponge by screening extracts of marine organisms. Both hal3 and suvanine inhibited the ATPase, RNA binding, and serine protease activities of NS3 helicase with IC50 values of 8, 8, and 14 µM, and 7, 3, and 34 µM, respectively. However, the dengue virus (DENV NS3 helicase, which shares a catalytic core (consisting mainly of ATPase and RNA binding sites with HCV NS3 helicase, was not inhibited by hal3 and suvanine, even at concentrations of 100 µM. Therefore, we conclude that hal3 and suvanine specifically inhibit HCV NS3 helicase via an interaction with an allosteric site in NS3 rather than binding to the catalytic core. This led to the inhibition of all NS3 activities, presumably by inducing conformational changes.

  4. SAFOD Brittle Microstructure and Mechanics Knowledge Base (BM2KB)

    Babaie, Hassan A.; Broda Cindi, M.; Hadizadeh, Jafar; Kumar, Anuj

    2013-07-01

    Scientific drilling near Parkfield, California has established the San Andreas Fault Observatory at Depth (SAFOD), which provides the solid earth community with short range geophysical and fault zone material data. The BM2KB ontology was developed in order to formalize the knowledge about brittle microstructures in the fault rocks sampled from the SAFOD cores. A knowledge base, instantiated from this domain ontology, stores and presents the observed microstructural and analytical data with respect to implications for brittle deformation and mechanics of faulting. These data can be searched on the knowledge base‧s Web interface by selecting a set of terms (classes, properties) from different drop-down lists that are dynamically populated from the ontology. In addition to this general search, a query can also be conducted to view data contributed by a specific investigator. A search by sample is done using the EarthScope SAFOD Core Viewer that allows a user to locate samples on high resolution images of core sections belonging to different runs and holes. The class hierarchy of the BM2KB ontology was initially designed using the Unified Modeling Language (UML), which was used as a visual guide to develop the ontology in OWL applying the Protégé ontology editor. Various Semantic Web technologies such as the RDF, RDFS, and OWL ontology languages, SPARQL query language, and Pellet reasoning engine, were used to develop the ontology. An interactive Web application interface was developed through Jena, a java based framework, with AJAX technology, jsp pages, and java servlets, and deployed via an Apache tomcat server. The interface allows the registered user to submit data related to their research on a sample of the SAFOD core. The submitted data, after initial review by the knowledge base administrator, are added to the extensible knowledge base and become available in subsequent queries to all types of users. The interface facilitates inference capabilities in the

  5. A continuum mechanics-based musculo-mechanical model for esophageal transport

    Kou, Wenjun; Griffith, Boyce E.; Pandolfino, John E.; Kahrilas, Peter J.; Patankar, Neelesh A.

    2017-11-01

    In this work, we extend our previous esophageal transport model using an immersed boundary (IB) method with discrete fiber-based structural model, to one using a continuum mechanics-based model that is approximated based on finite elements (IB-FE). To deal with the leakage of flow when the Lagrangian mesh becomes coarser than the fluid mesh, we employ adaptive interaction quadrature points to deal with Lagrangian-Eulerian interaction equations based on a previous work (Griffith and Luo [1]). In particular, we introduce a new anisotropic adaptive interaction quadrature rule. The new rule permits us to vary the interaction quadrature points not only at each time-step and element but also at different orientations per element. This helps to avoid the leakage issue without sacrificing the computational efficiency and accuracy in dealing with the interaction equations. For the material model, we extend our previous fiber-based model to a continuum-based model. We present formulations for general fiber-reinforced material models in the IB-FE framework. The new material model can handle non-linear elasticity and fiber-matrix interactions, and thus permits us to consider more realistic material behavior of biological tissues. To validate our method, we first study a case in which a three-dimensional short tube is dilated. Results on the pressure-displacement relationship and the stress distribution matches very well with those obtained from the implicit FE method. We remark that in our IB-FE case, the three-dimensional tube undergoes a very large deformation and the Lagrangian mesh-size becomes about 6 times of Eulerian mesh-size in the circumferential orientation. To validate the performance of the method in handling fiber-matrix material models, we perform a second study on dilating a long fiber-reinforced tube. Errors are small when we compare numerical solutions with analytical solutions. The technique is then applied to the problem of esophageal transport. We use two

  6. Mechanisms for adsorption of organic bases on hydrated smectite surfaces

    Laird, D.A.; Fleming, P.D.

    1999-08-01

    The environmental fate of anthropogenic organic bases introduced to soils and sediments, either deliberately as pesticides or inadvertently as contaminants, depends, to a large extent, on reactions between those compounds and the surfaces of soil mineral and organic constituents. Mechanisms by which organic bases are adsorbed on hydrated smectite surfaces were investigated. Three Ca-saturated reference smectites (Otay, SPV, and Panther Creek) were dispersed in distilled water containing 5 {micro}mol of pyridine or 3-butylpyridine. The pH was adjusted to between 7.5 and 3 using 0.01 M HCl. After a 2-h equilibration, the amounts of pyridine or 3-butylpyridine adsorbed on the clay and the amount of Ca desorbed from the clay were determined. Negligible amounts of pyridine were adsorbed by the Ca-smectites in the neutral systems (pH > 7); however, most of the added pyridine was adsorbed on the smectites in the acidified systems (pH < 5). Equivalent amounts of Ca{sup 2+} were desorbed from the clays, indicating that pyridine was adsorbed as a protonated species by cation exchange. By contrast, 40 to 90% of added 3-butylpyridine was adsorbed on the smectites at neutral pHs, whereas only small amounts of Ca{sup 2+} were desorbed. The results suggest that 3-butylpyridine is initially retained by hydrophobic bonding between the alkyl side chain of the molecule and hydrophobic nanosites located between the charge sites on smectite surfaces. Surface acidity catalyzed protonation 1 to 1.5 pH units above the pK{sub a} of the bases.

  7. Mediating mechanisms of a military Web-based alcohol intervention.

    Williams, Jason; Herman-Stahl, Mindy; Calvin, Sara L; Pemberton, Michael; Bradshaw, Michael

    2009-03-01

    This study explored the mediating mechanisms of two Web-based alcohol interventions in a sample of active duty United States military personnel. Personnel were recruited from eight bases and received the Drinker's Check-Up (N=1483), Alcohol Savvy (N=688), or served as controls (N=919). The interventions drew on motivational interviewing and social learning theory and targeted multiple mediators including social norms, perceived risks and benefits, readiness to change, and coping strategies. Baseline data were collected prior to the intervention and follow-up data on alcohol consumption were gathered 1 month and 6 months after program completion. Two mediation models were examined: (1) a longitudinal two-wave model with outcomes and mediators assessed concurrently at the 1-month follow-up; and (2) a three-wave model in which the causal chain was fully lagged. Results indicated strong support for the role of perceived descriptive norms in transmitting the effects of the Drinker's Check-Up, with consistent mediation across the majority of alcohol outcome measures for both the concurrent and fully lagged mediation models. These results suggest that web-based interventions that are effective in lowering perceived norms about the frequency and quantity of drinking may be a viable strategy for reducing alcohol consumption in military populations. The results did not support program mediation by the other targeted variables, indicating the need for future research on the effective components of alcohol interventions. The mediation models also suggest reasons why program effects were not found for some outcomes or were different across programs.

  8. A review on mechanical properties of magnesium based nano composites

    Tarafder, Nilanjan; Prasad, M. Lakshmi Vara

    2018-04-01

    A review was done on Magnesium (Mg) based composite materials reinforced with different nano particles such as TiO2, Cu, Y2O3, SiC, ZrO2 and Al2O3. TiO2 and Al2O3 nanoparticles were synthesised by melt deposition process. Cu, Y2O3, SiC and ZrO2 nanoparticles were synthesised by powder metallurgy process. Composite microstructural characteristics shows that the nano-size reinforcements are uniformly distributed in the composite matrix and also minimum porosity with solid interfacial integrity. The mechanical properties showed yield strength improvement by 0.2 percentage and Ultimate tensile strength (UTS) was also improved for all the nano-particles. But UTS was adversely affected with TiO2 reinforcement while ductility was increased. With Cu reinforcement elastic modulus, hardness and fracture resistance increased and improved the co-efficient of thermal expansion (CTE) of Mg based matrix. By Y2O3 reinforcement hardness, fracture resistance was improved and ductility reached maximum by 0.22 volume percentage of Y2O3 and decreased with succeeding increase in Y2O3 reinforcement. The readings exposed that mechanical properties were gathered from the composite comprising 2.0 weight percentage of Y2O3. Ductility and fracture resistance increased with ZrO2 reinforcement in Mg matrix. Using Al2O3 as reinforcement in Mg composite matrix hardness, elastic modulus and ductility was increased but porosity reduced with well interfacial integrity. Dissipation of energy in the form of damping capacity was resolved by classical vibration theory. The result showed that an increasing up to 0.4 volume percentage alumina content increases the damping capacity up to 34 percent. In another sample, addition of 2 weight percentage nano-Al2O3 particles showed big possibility in reducing CTE from 27.9-25.9×10-6 K-1 in Magnesium, tensile and yield strength amplified by 40MPa. In another test, Mg/1.1Al2O3 nanocomposite was manufactured by solidification process followed by hot extrusion

  9. Market-based mechanisms for biodiversity conservation: a review of existing schemes and an outline for a global mechanism

    Alvarado Quesada, I.; Hein, L.G.; Weikard, H.P.

    2014-01-01

    Continuous decline of biodiversity over the past decades suggests that efforts to decrease biodiversity loss have been insufficient. One option to deal with this problem is the use of market-based mechanisms for biodiversity conservation. Several studies have analysed such mechanisms individually,

  10. The DEAD-Box RNA Helicase DDX3 Interacts with m6A RNA Demethylase ALKBH5

    Abdullah Shah

    2017-01-01

    Full Text Available DDX3 is a member of the family of DEAD-box RNA helicases. DDX3 is a multifaceted helicase and plays essential roles in key biological processes such as cell cycle, stress response, apoptosis, and RNA metabolism. In this study, we found that DDX3 interacted with ALKBH5, an m6A RNA demethylase. The ATP domain of DDX3 and DSBH domain of ALKBH5 were indispensable to their interaction with each other. Furthermore, DDX3 could modulate the demethylation of mRNAs. We also showed that DDX3 regulated the methylation status of microRNAs and there was an interaction between DDX3 and AGO2. The dynamics of m6A RNA modification is still a field demanding further investigation, and here, we add a link by showing that RNA demethylation can be regulated by proteins such as DDX3.

  11. SAD-3, a Putative Helicase Required for Meiotic Silencing by Unpaired DNA, Interacts with Other Components of the Silencing Machinery

    Hammond, Thomas M.; Xiao, Hua; Boone, Erin C.; Perdue, Tony D.; Pukkila, Patricia J.; Shiu, Patrick K. T.

    2011-01-01

    In Neurospora crassa, genes lacking a pairing partner during meiosis are suppressed by a process known as meiotic silencing by unpaired DNA (MSUD). To identify novel MSUD components, we have developed a high-throughput reverse-genetic screen for use with the N. crassa knockout library. Here we describe the screening method and the characterization of a gene (sad-3) subsequently discovered. SAD-3 is a putative helicase required for MSUD and sexual spore production. It exists in a complex with other known MSUD proteins in the perinuclear region, a center for meiotic silencing activity. Orthologs of SAD-3 include Schizosaccharomyces pombe Hrr1, a helicase required for RNAi-induced heterochromatin formation. Both SAD-3 and Hrr1 interact with an RNA-directed RNA polymerase and an Argonaute, suggesting that certain aspects of silencing complex formation may be conserved between the two fungal species. PMID:22384347

  12. Mutations of the RTEL1 Helicase in a Hoyeraal-Hreidarsson Syndrome Patient Highlight the Importance of the ARCH Domain.

    Jullien, Laurent; Kannengiesser, Caroline; Kermasson, Laetitia; Cormier-Daire, Valérie; Leblanc, Thierry; Soulier, Jean; Londono-Vallejo, Arturo; de Villartay, Jean-Pierre; Callebaut, Isabelle; Revy, Patrick

    2016-05-01

    The DNA helicase RTEL1 participates in telomere maintenance and genome stability. Biallelic mutations in the RTEL1 gene account for the severe telomere biology disorder characteristic of the Hoyeraal-Hreidarsson syndrome (HH). Here, we report a HH patient (P4) carrying two novel compound heterozygous mutations in RTEL1: a premature stop codon (c.949A>T, p.Lys317*) and an intronic deletion leading to an exon skipping and an in-frame deletion of 25 amino-acids (p.Ile398_Lys422). P4's cells exhibit short and dysfunctional telomeres similarly to other RTEL1-deficient patients. 3D structure predictions indicated that the p.Ile398_Lys422 deletion affects a part of the helicase ARCH domain, which lines the pore formed with the core HD and the iron-sulfur cluster domains and is highly specific of sequences from the eukaryotic XPD family members. © 2016 WILEY PERIODICALS, INC.

  13. Technology Transfer of Isotopes-Based Assay: Strategies and Mechanisms

    Tabbada, R.S.D.C.; Rañada, M.L.O.; Mendoza, A.D.L.; Panganiban, R.; Castañeda, S.S.; Sombrito, E.Z.; Arcamo, S.V.R.

    2015-01-01

    Receptor Binding Assay for Paralytic Shellfish Poisoning (PSP RBA) is an isotope-based assay for detection and quantification of PSP toxins in seafood. It was established in the Philippines through a national program based on the recommendations of the Expert Mission sent by the International Atomic Energy Agency (IAEA). Through the said program, the Philippines Nuclear Research Institute (PNRI) was able to put up an RBA facility and develop expertise. Advantages of the technique against Mouse Bioassay (MBA) and high-performance Liquid Chromatography (HPLC) methods were are established. RBA is being utilized by some developed countries as screening method for Harmful Algal Bloom (HAB) Monitoring. However, it was not immediately adopted by the national HAB regulatory body for the following reasons: (1) acceptance of RBA as an official national method of analysis for PSP, (2) logistics and financial concerns in building up and maintaining a RBA facility, (3) considerations on the use of radioactive materials. To address these issues, the Philippines Council for Agriculture, Aquatic and Natural Resources Research and Development (PCAARRD) approved a Grants-In-Aid Project to initiate and to facilitate the transfer of the RBA technology to the monitoring and regulatory body. The project has two major objectives: capacity building and technology transfer. The capacity building focuses on human resources development of HAB monitoring personnel, specifically training on RBA and on the use of radioactive materials. On the other hand, the technology transfer deals with assistance that PNRI may render in establishing the new RBA facility and over-all know-how of the project. In this is poster, the mechanisms and strategies being undertaken by PNRI, in collaboration with the regulatory and monitoring body, to address the limitation of transferring a technology that utilizes radioactive materials including the technical difficulties are presented and discussed. (author)

  14. Mechanisms of acid-base regulation in peritoneal dialysis.

    Sow, Amadou; Morelle, Johann; Hautem, Nicolas; Bettoni, Carla; Wagner, Carsten A; Devuyst, Olivier

    2017-11-22

    Peritoneal dialysis (PD) contributes to restore acid-base homeostasis in patients with end-stage renal disease. The transport pathways for buffers and carbon dioxide (CO2) across the peritoneal membrane remain poorly understood. Combining well-established PD protocols, whole-body plethysmography and renal function studies in mice, we investigated molecular mechanisms of acid-base regulation in PD, including the potential role of the water channel aquaporin-1 (AQP1). After instillation in peritoneal cavity, the pH of acidic dialysis solutions increased within minutes to rapidly equilibrate with blood pH, whereas the neutral pH of biocompatible solutions remained constant. Predictions from the three-pore model of peritoneal transport suggested that local production of HCO3- accounts at least in part for the changes in intraperitoneal pH observed with acidic solutions. Carbonic anhydrase (CA) isoforms were evidenced in the peritoneal membrane and their inhibition with acetazolamide significantly decreased local production of HCO3- and delayed changes in intraperitoneal pH. On the contrary, genetic deletion of AQP1 had no effect on peritoneal transport of buffers and diffusion of CO2. Besides intraperitoneal modifications, the use of acidic dialysis solutions enhanced acid excretion both at pulmonary and renal levels. These findings suggest that changes in intraperitoneal pH during PD are mediated by bidirectional buffer transport and by CA-mediated production of HCO3- in the membrane. The use of acidic solutions enhances acid excretion through respiratory and renal responses, which should be considered in patients with renal failure. © The Author 2017. Published by Oxford University Press on behalf of ERA-EDTA. All rights reserved.

  15. Community-based research as a mechanism to reduce ...

    Racial and ethnic minority communities, including American Indian and Alaska Natives, have been disproportionately impacted by environmental pollution and contamination. This includes siting and location of point sources of pollution, legacies of contamination of drinking and recreational water, and mining, military and agricultural impacts. As a result, both quantity and quality of culturally important subsistence resources are diminished, contributing to poor nutrition and obesity, and overall reductions in quality of life and life expectancy. Climate change is adding to these impacts on Native American communities (Wildcat 2013), variably causing drought, increased flooding and forced relocation (Maldonado et al. 2013), affecting Tribal water resources (Cozzetto et al. 2013), traditional foods (Lynn et al. 2013; Gautam et al. 2013), forests and forest resources (Voggesser et al. 2013) and Tribal health (Donatuto et al 2014; Doyle et al. 2013). This article will highlight several extramural research projects supported by the United States Environmental Protection Agency (USEPA) Science to Achieve Results (STAR) Tribal environmental research grants as a mechanism to address the environmental health inequities and disparities faced by Tribal communities (USEPA, 2014a, www.epa.gov/ncer/tribalresearch). The Tribal Research portfolio has focused on addressing tribal environmental health risks through community based participatory research. Specifically, the STA

  16. Relative gravimeter prototype based on micro electro mechanical system

    Rozy, A. S. A.; Nugroho, H. A.; Yusuf, M.

    2018-03-01

    This research to make gravity measurement system by utilizing micro electro mechanical system based sensor in Gal order. System design consists of three parts, design of hardware, software, and interface. The design of the hardware include of designing the sensor design to measure the value of a stable gravity acceleration. The ADXL345 and ADXL335 sensors are tuned to obtain stable measurements. The design of the instrumentation system the next stage by creating a design to integrate between the sensor, microcontroller, and GPS. The design of programming algorithm is done with Arduino IDE software. The interface design uses a 20x4 LCD display to display the gravity acceleration value and store data on the storage media. The system uses a box made of iron and plate leveling to minimize measurement errors. The sensor test shows the ADXL345 sensor has a more stable value. The system is examined by comparing with gravity measurement of gravimeter A-10 results in Bandung observation post. The result of system test resulted the average of system correction value equal to 0.19 Gal. The system is expected to use for mineral exploration, water supply analyze, and earthquake precursor.

  17. Essential and distinct roles of the F-box and helicase domains of Fbh1 in DNA damage repair

    Shinagawa Hideo

    2008-03-01

    Full Text Available Abstract Background DNA double-strand breaks (DSBs are induced by exogenous insults such as ionizing radiation and chemical exposure, and they can also arise as a consequence of stalled or collapsed DNA replication forks. Failure to repair DSBs can lead to genomic instability or cell death and cancer in higher eukaryotes. The Schizosaccharomyces pombe fbh1 gene encodes an F-box DNA helicase previously described to play a role in the Rhp51 (an orthologue of S. cerevisiae RAD51-dependent recombinational repair of DSBs. Fbh1 fused to GFP localizes to discrete nuclear foci following DNA damage. Results To determine the functional roles of the highly conserved F-box and helicase domains, we have characterized fbh1 mutants carrying specific mutations in these domains. We show that the F-box mutation fbh1-fb disturbs the nuclear localization of Fbh1, conferring an fbh1 null-like phenotype. Moreover, nuclear foci do not form in fbh1-fb cells with DNA damage even if Fbh1-fb is targeted to the nucleus by fusion to a nuclear localization signal sequence. In contrast, the helicase mutation fbh1-hl causes the accumulation of Fbh1 foci irrespective of the presence of DNA damage and confers damage sensitivity greater than that conferred by the null allele. Additional mutation of the F-box alleviates the hypermorphic phenotype of the fbh1-hl mutant. Conclusion These results suggest that the F-box and DNA helicase domains play indispensable but distinct roles in Fbh1 function. Assembly of the SCFFbh1 complex is required for both the nuclear localization and DNA damage-induced focus formation of Fbh1 and is therefore prerequisite for the Fbh1 recombination function.

  18. DEAD-Box RNA Helicases are among the Constituents of the Tobacco Pollen mRNA Storing Bodies

    Hafidh, Said; Potěšil, D.; Zdráhal, Z.; Honys, David

    2013-01-01

    Roč. 1, č. 3 (2013) ISSN 2329-9029 R&D Projects: GA ČR GPP501/11/P321; GA ČR(CZ) GAP501/11/1462; GA MŠk(CZ) ED1.1.00/02.0068; GA MŠk(CZ) LD13049 Institutional support: RVO:61389030 Keywords : Translation * mRNA storage * RNA helicase Subject RIV: EB - Genetics ; Molecular Biology

  19. Structural Studies of RNA Helicases Involved in Eukaryotic Pre-mRNA Splicing, Ribosome Biogenesis, and Translation Initiation

    He, Yangzi

    and ligates the neighbouring exons to generate mature mRNAs. Prp43 is an RNA helicase of the DEAH/RHA family. In yeast, once mRNAs are released, Prp43 catalyzes the disassembly of spliceosomes. The 18S, 5.8S and 25S rRNAs are transcribed as a single polycistronic transcript—the 35S pre......-rRNA. It is nucleolytically cleaved and chemically modified to generate mature rRNAs, which assemble with ribosomal proteins to form the ribosome. Prp43 is required for the processing of the 18S rRNA. Using X-ray crystallography, I determined a high resolution structure of Prp43 bound to ADP, the first structure of a DEAH....../RHA helicase. It defined the conserved structural features of all DEAH/RHA helicases, and unveiled a novel nucleotide binding site. Additionally a preliminary low resolution structure of a ternary complex comprising Prp43, a non-hydrolyzable ATP analogue, and a single-stranded RNA, was obtained. The ribosome...

  20. Zebrafish P54 RNA helicases are cytoplasmic granule residents that are required for development and stress resilience

    Cecilia Zampedri

    2016-10-01

    Full Text Available Stress granules are cytoplasmic foci that directly respond to the protein synthesis status of the cell. Various environmental insults, such as oxidative stress or extreme heat, block protein synthesis; consequently, mRNA will stall in translation, and stress granules will immediately form and become enriched with mRNAs. P54 DEAD box RNA helicases are components of RNA granules such as P-bodies and stress granules. We studied the expression, in cytoplasmic foci, of both zebrafish P54 RNA helicases (P54a and P54b during development and found that they are expressed in cytoplasmic granules under both normal conditions and stress conditions. In zebrafish embryos exposed to heat shock, some proportion of P54a and P54b helicases move to larger granules that exhibit the properties of genuine stress granules. Knockdown of P54a and/or P54b in zebrafish embryos produces developmental abnormalities restricted to the posterior trunk; further, these embryos do not form stress granules, and their survival upon exposure to heat-shock conditions is compromised. Our observations fit the model that cells lacking stress granules have no resilience or ability to recover once the stress has ended, indicating that stress granules play an essential role in the way organisms adapt to a changing environment.

  1. The Arabidopsis thaliana Homolog of the Helicase RTEL1 Plays Multiple Roles in Preserving Genome Stability[C][W

    Recker, Julia; Knoll, Alexander; Puchta, Holger

    2014-01-01

    In humans, mutations in the DNA helicase Regulator of Telomere Elongation Helicase1 (RTEL1) lead to Hoyeraal-Hreidarsson syndrome, a severe, multisystem disorder. Here, we demonstrate that the RTEL1 homolog in Arabidopsis thaliana plays multiple roles in preserving genome stability. RTEL1 suppresses homologous recombination in a pathway parallel to that of the DNA translocase FANCM. Cytological analyses of root meristems indicate that RTEL1 is involved in processing DNA replication intermediates independently from FANCM and the nuclease MUS81. Moreover, RTEL1 is involved in interstrand and intrastrand DNA cross-link repair independently from FANCM and (in intrastrand cross-link repair) parallel to MUS81. RTEL1 contributes to telomere homeostasis; the concurrent loss of RTEL1 and the telomerase TERT leads to rapid, severe telomere shortening, which occurs much more rapidly than it does in the single-mutant line tert, resulting in developmental arrest after four generations. The double mutant rtel1-1 recq4A-4 exhibits massive growth defects, indicating that this RecQ family helicase, which is also involved in the suppression of homologous recombination and the repair of DNA lesions, can partially replace RTEL1 in the processing of DNA intermediates. The requirement for RTEL1 in multiple pathways to preserve genome stability in plants can be explained by its putative role in the destabilization of DNA loop structures, such as D-loops and T-loops. PMID:25516598

  2. Controlling chaos based on an adaptive adjustment mechanism

    Zheng Yongai

    2006-01-01

    In this paper, we extend the ideas and techniques developed by Huang [Huang W. Stabilizing nonlinear dynamical systems by an adaptive adjustment mechanism. Phys Rev E 2000;61:R1012-5] for controlling discrete-time chaotic system using adaptive adjustment mechanism to continuous-time chaotic system. Two control approaches, namely adaptive adjustment mechanism (AAM) and modified adaptive adjustment mechanism (MAAM), are investigated. In both case sufficient conditions for the stabilization of chaotic systems are given analytically. The simulation results on Chen chaotic system have verified the effectiveness of the proposed techniques

  3. Fabrication of nano piezoelectric based vibration accelerometer for mechanical sensing

    Murugan, S.; Prasad, M. V. N.; Jayakumar, K.

    2016-05-01

    An electromechanical sensor unit has been fabricated using nano PZT embedded in PVDF polymer. Such a polymer nano composite has been used as vibration sensor element and sensitivity, detection of mechanical vibration, and linearity measurements have been investigated. It is found from its performance, that this nano composite sensor is suitable for mechanical sensing applications.

  4. Competency-Based Materials for the Florida Automotive Mechanics Curriculum

    Goodson, Ludy; And Others

    1978-01-01

    Describes Florida's new automotive mechanics curriculum, an individualized, self-paced learning sequence that combines text material, review exercises and actual work activities. Development of the materials, including incorporation of Florida's V-TECS catalog of performance objectives in auto mechanics, is described. A field-test experience of a…

  5. Mechanism-Based Design of Green Oxidation Catalysts

    Rybak-Akimova, Elena [Tufts Univ., Medford, MA (United States)

    2015-03-16

    situation. Growing families of synthetic iron complexes that resemble active sites of metalloenzymes produce metal-based intermediates (rather than hydroxyl radicals) in reactions with oxygen donors. These complexes are very promising for selective oxygen and peroxide activation. In order to understand the mechanisms of metal-based small molecule activation, kinetically competent metal-oxygen intermediates must be identified. One of the grand challenges identified by the Department of Energy workshop "Catalysis for Energy" is understanding mechanisms and dynamics of catalyzed reactions. The research summarized herein focuses on detailed characterization of the formation and reactivity of various iron-peroxo- and iron-oxo intermediates that are involved in catalysis. Rates of rapid reactions were studied at low temperatures by a specialized technique termed cryogenic stopped-flow spectrophotometry. These measurements identified reaction conditions which favor the formation of catalytically competent oxidants. Chemical structures of reactive complexes was determined, and new, efficient catalysts for hydrocarbon oxidation were synthesized. Importantly, these catalysts are selective, they promote oxidation of hydrocarbons at a specific site. The catalysts are also efficient and robust, hundreds of cycles of substrate oxidation occur within minutes at room temperature. Furthermore, they enable utilization of environmentally friendly oxidants, such as hydrogen peroxide, which produces water as the only byproduct. Mechanistic insights uncovered the role of various acid-containing additives in catalytic oxidations. Proton delivery to the active catalytic sites facilitated oxidations, similarly to the catalytic pathways in metal-containing enzymes. Under certain conditions, two metals in one complex can act in concert, modeling the reactivity of a bacterial enzyme which converts methane into methanol. In related studies, a family of nickel complexes that react with carbon dioxide at

  6. Model-based PEEP optimisation in mechanical ventilation

    Chiew Yeong Shiong

    2011-12-01

    Full Text Available Abstract Background Acute Respiratory Distress Syndrome (ARDS patients require mechanical ventilation (MV for breathing support. Patient-specific PEEP is encouraged for treating different patients but there is no well established method in optimal PEEP selection. Methods A study of 10 patients diagnosed with ALI/ARDS whom underwent recruitment manoeuvre is carried out. Airway pressure and flow data are used to identify patient-specific constant lung elastance (Elung and time-variant dynamic lung elastance (Edrs at each PEEP level (increments of 5cmH2O, for a single compartment linear lung model using integral-based methods. Optimal PEEP is estimated using Elung versus PEEP, Edrs-Pressure curve and Edrs Area at minimum elastance (maximum compliance and the inflection of the curves (diminishing return. Results are compared to clinically selected PEEP values. The trials and use of the data were approved by the New Zealand South Island Regional Ethics Committee. Results Median absolute percentage fitting error to the data when estimating time-variant Edrs is 0.9% (IQR = 0.5-2.4 and 5.6% [IQR: 1.8-11.3] when estimating constant Elung. Both Elung and Edrs decrease with PEEP to a minimum, before rising, and indicating potential over-inflation. Median Edrs over all patients across all PEEP values was 32.2 cmH2O/l [IQR: 26.1-46.6], reflecting the heterogeneity of ALI/ARDS patients, and their response to PEEP, that complicates standard approaches to PEEP selection. All Edrs-Pressure curves have a clear inflection point before minimum Edrs, making PEEP selection straightforward. Model-based selected PEEP using the proposed metrics were higher than clinically selected values in 7/10 cases. Conclusion Continuous monitoring of the patient-specific Elung and Edrs and minimally invasive PEEP titration provide a unique, patient-specific and physiologically relevant metric to optimize PEEP selection with minimal disruption of MV therapy.

  7. Bohmian mechanics with complex action: A new trajectory-based formulation of quantum mechanics

    Goldfarb, Yair; Degani, Ilan; Tannor, David J.

    2006-01-01

    In recent years there has been a resurgence of interest in Bohmian mechanics as a numerical tool because of its local dynamics, which suggest the possibility of significant computational advantages for the simulation of large quantum systems. However, closer inspection of the Bohmian formulation reveals that the nonlocality of quantum mechanics has not disappeared--it has simply been swept under the rug into the quantum force. In this paper we present a new formulation of Bohmian mechanics in which the quantum action, S, is taken to be complex. This leads to a single equation for complex S, and ultimately complex x and p but there is a reward for this complexification - a significantly higher degree of localization. The quantum force in the new approach vanishes for Gaussian wave packet dynamics, and its effect on barrier tunneling processes is orders of magnitude lower than that of the classical force. In fact, the current method is shown to be a rigorous extension of generalized Gaussian wave packet dynamics to give exact quantum mechanics. We demonstrate tunneling probabilities that are in virtually perfect agreement with the exact quantum mechanics down to 10 -7 calculated from strictly localized quantum trajectories that do not communicate with their neighbors. The new formulation may have significant implications for fundamental quantum mechanics, ranging from the interpretation of non-locality to measures of quantum complexity

  8. A conserved helicase processivity factor is needed for conjugation and replication of an integrative and conjugative element.

    Jacob Thomas

    Full Text Available Integrative and conjugative elements (ICEs are agents of horizontal gene transfer and have major roles in evolution and acquisition of new traits, including antibiotic resistances. ICEs are found integrated in a host chromosome and can excise and transfer to recipient bacteria via conjugation. Conjugation involves nicking of the ICE origin of transfer (oriT by the ICE-encoded relaxase and transfer of the nicked single strand of ICE DNA. For ICEBs1 of Bacillus subtilis, nicking of oriT by the ICEBs1 relaxase NicK also initiates rolling circle replication. This autonomous replication of ICEBs1 is critical for stability of the excised element in growing cells. We found a conserved and previously uncharacterized ICE gene that is required for conjugation and replication of ICEBs1. Our results indicate that this gene, helP (formerly ydcP, encodes a helicase processivity factor that enables the host-encoded helicase PcrA to unwind the double-stranded ICEBs1 DNA. HelP was required for both conjugation and replication of ICEBs1, and HelP and NicK were the only ICEBs1 proteins needed for replication from ICEBs1 oriT. Using chromatin immunoprecipitation, we measured association of HelP, NicK, PcrA, and the host-encoded single-strand DNA binding protein Ssb with ICEBs1. We found that NicK was required for association of HelP and PcrA with ICEBs1 DNA. HelP was required for association of PcrA and Ssb with ICEBs1 regions distal, but not proximal, to oriT, indicating that PcrA needs HelP to progress beyond nicked oriT and unwind ICEBs1. In vitro, HelP directly stimulated the helicase activity of the PcrA homologue UvrD. Our findings demonstrate that HelP is a helicase processivity factor needed for efficient unwinding of ICEBs1 for conjugation and replication. Homologues of HelP and PcrA-type helicases are encoded on many known and putative ICEs. We propose that these factors are essential for ICE conjugation, replication, and genetic stability.

  9. A probabilistic approach to quantum mechanics based on 'tomograms'

    Caponigro, M.; Mancini, S.; Man'ko, V.I.

    2006-01-01

    It is usually believed that a picture of Quantum Mechanics in terms of true probabilities cannot be given due to the uncertainty relations. Here we discuss a tomographic approach to quantum states that leads to a probability representation of quantum states. This can be regarded as a classical-like formulation of quantum mechanics which avoids the counterintuitive concepts of wave function and density operator. The relevant concepts of quantum mechanics are then reconsidered and the epistemological implications of such approach discussed. (Abstract Copyright [2006], Wiley Periodicals, Inc.)

  10. Mechanical strenght and niobium and niobium-base alloys substructures

    Monteiro, W.A.; Andrade, A.H.P. de

    1986-01-01

    Niobium and some of its alloys have been used in several fields of technological applications such as the aerospace, chemical and nuclear industries. This is due to its excelent mechanical stringth at high temperatures and reasonable ductility at low temperatures. In this work, we review the main features of the relationship mechanical strength - substructure in niobium and its alloys, taking into account the presence of impurities, the influence of initial thermal and thermo - mechanical treatments as well as the irradiation by energetic particles. (Author) [pt

  11. Yeast Srs2 Helicase Promotes Redistribution of Single-Stranded DNA-Bound RPA and Rad52 in Homologous Recombination Regulation

    Luisina De Tullio

    2017-10-01

    Full Text Available Srs2 is a super-family 1 helicase that promotes genome stability by dismantling toxic DNA recombination intermediates. However, the mechanisms by which Srs2 remodels or resolves recombination intermediates remain poorly understood. Here, single-molecule imaging is used to visualize Srs2 in real time as it acts on single-stranded DNA (ssDNA bound by protein factors that function in recombination. We demonstrate that Srs2 is highly processive and translocates rapidly (∼170 nt per second in the 3′→5′ direction along ssDNA saturated with replication protein A (RPA. We show that RPA is evicted from DNA during the passage of Srs2. Remarkably, Srs2 also readily removes the recombination mediator Rad52 from RPA-ssDNA and, in doing so, promotes rapid redistribution of both Rad52 and RPA. These findings have important mechanistic implications for understanding how Srs2 and related nucleic acid motor proteins resolve potentially pathogenic nucleoprotein intermediates.

  12. Yeast Srs2 Helicase Promotes Redistribution of Single-Stranded DNA-Bound RPA and Rad52 in Homologous Recombination Regulation.

    De Tullio, Luisina; Kaniecki, Kyle; Kwon, Youngho; Crickard, J Brooks; Sung, Patrick; Greene, Eric C

    2017-10-17

    Srs2 is a super-family 1 helicase that promotes genome stability by dismantling toxic DNA recombination intermediates. However, the mechanisms by which Srs2 remodels or resolves recombination intermediates remain poorly understood. Here, single-molecule imaging is used to visualize Srs2 in real time as it acts on single-stranded DNA (ssDNA) bound by protein factors that function in recombination. We demonstrate that Srs2 is highly processive and translocates rapidly (∼170 nt per second) in the 3'→5' direction along ssDNA saturated with replication protein A (RPA). We show that RPA is evicted from DNA during the passage of Srs2. Remarkably, Srs2 also readily removes the recombination mediator Rad52 from RPA-ssDNA and, in doing so, promotes rapid redistribution of both Rad52 and RPA. These findings have important mechanistic implications for understanding how Srs2 and related nucleic acid motor proteins resolve potentially pathogenic nucleoprotein intermediates. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

  13. Identification of human genes involved in cellular responses to ionizing radiation: molecular and cellular studies of gene encoding the p68 helicase in mammalian cells

    Menaa, F.

    2003-12-01

    Cells submitted to genotoxic factors -like IR- activate several and important mechanisms such as repair, cell cycle arrest or 'apoptosis' to maintain genetic integrity. So, the damaged cells will induce many and different genes. The human transcriptome analysis by 'SSH' method in a human breast carcinoma cell line MCF7 γ-irradiated versus not irradiated, allowed to identify about one hundred genes. Among of these genes, we have focused our study on a radio-induced gene encoding the p68 helicase. In the conditions of irradiation used, our results show that the kinetic and the regulation of this gene expression differs between the nature of radiations used. Indeed, in γ-irradiated mammalian cells, ATM, a protein kinase activated by DSB and IR, is required to induce quickly P68 gene via the important transcription factor p53 stabilized by IR. In the case of UVC-irradiated cells, the P68 gene induction is late and the intracellular signalling pathway that lead to this induction is independent from the p53 protein. Finally, we show that the p68 protein under-expression is responsible for an increased radiosensitivity of MCF7 cells. Consequently, we can postulate that the p68 protein is involved in cellular responses to radiations to reduce the increased radiosensitivity of cells exposed to γ-rays. (author)

  14. Minichromosome maintenance helicase paralog MCM9 is dispensible for DNA replication but functions in germ-line stem cells and tumor suppression.

    Hartford, Suzanne A; Luo, Yunhai; Southard, Teresa L; Min, Irene M; Lis, John T; Schimenti, John C

    2011-10-25

    Effective DNA replication is critical to the health and reproductive success of organisms. The six MCM2-7 proteins, which form the replicative helicase, are essential for high-fidelity replication of the genome. Many eukaryotes have a divergent paralog, MCM9, that was reported to be essential for loading MCM2-7 onto replication origins in the Xenopus oocyte extract system. To address the in vivo role of mammalian MCM9, we created and analyzed the phenotypes of mice with various mutations in Mcm9 and an intronic DNA replication-related gene Asf1a. Ablation of Mcm9 was compatible with cell proliferation and mouse viability, showing that it is nonessential for MCM2-7 loading or DNA replication. Mcm9 mutants underwent p53-independent embryonic germ-cell depletion in both sexes, with males also exhibiting defective spermatogonial stem-cell renewal. MCM9-deficient cells had elevated genomic instability and defective cell cycle reentry following replication stress, and mutant animals were prone to sex-specific cancers, most notably hepatocellular carcinoma in males. The phenotypes of mutant mice and cells suggest that MCM9 evolved a specialized but nonessential role in DNA replication or replication-linked quality-control mechanisms that are especially important for germ-line stem cells, and also for tumor suppression and genome maintenance in the soma.

  15. Innovative Mechanism of Rural Organization Based on Self-Organization

    Wang, Xing jin; Gao, Bing

    2011-01-01

    The paper analyzes the basic situation for the formation of innovative rural organizations with the form of self-organization; revels the features of self-organization, including the four aspects of openness of rural organization, innovation of rural organization is far away from equilibrium, the non-linear response mechanism of rural organization innovation and the random rise and fall of rural organization innovation. The evolution mechanism of rural organization innovation is reveled accor...

  16. Radiolysis of nucleosides in aqueous solutions: base liberation by the base attack mechanism

    Fujita, S.

    1984-01-01

    On the radiolysis of uridine and some other nucleosides in aqueous solution, a pH-dependent liberation of uracil or the corresponding base was found. e - sub(aq) and HOsup(anion radicals) 2 gave no freed bases, although many oxidizing radicals, including OH, Clsup(anion radicals) 2 , Brsup(anion radicals) 2 , (CNS)sup(anion radicals) 2 and SOsup(anion radicals) 4 , did cause the release of unaltered bases, depending on the pH of the solutions. The base yields were generally high at pH >= 11, with the exception of SOsup(anion radicals) 4 , which gave a rather high yield of uracil (from uridine) even in the pH region of - , present at high pH as the dissociated form of OH, may act partly as an oxidizing radical. A plausible mechanism of 3 1 -radical formation is discussed. (author)

  17. Antiviral drug resistance and helicase-primase inhibitors of herpes simplex virus.

    Field, Hugh J; Biswas, Subhajit

    2011-02-01

    A new class of chemical inhibitors has been discovered that interferes with the process of herpesvirus DNA replication. To date, the majority of useful herpesvirus antivirals are nucleoside analogues that block herpesvirus DNA replication by targeting the DNA polymerase. The new helicase-primase inhibitors (HPI) target a different enzyme complex that is also essential for herpesvirus DNA replication. This review will place the HPI in the context of previous work on the nucleoside analogues. Several promising highly potent HPI will be described with a particular focus on the identification of drug-resistance mutations. Several HPI have good pharmacological profiles and are now at the outset of phase II clinical trials. Provided there are no safety issues to stop their progress, this new class of compound will be a major advance in the herpesvirus antiviral field. Furthermore, HPI are likely to have a major impact on the therapy and prevention of herpes simplex virus and varicella zoster in both immunocompetent and immunocompromised patients alone or in combination with current nucleoside analogues. The possibility of acquired drug-resistance to HPI will then become an issue of great practical importance. Copyright © 2010 Elsevier Ltd. All rights reserved.

  18. DNA replication restart and cellular dynamics of Hef helicase/nuclease protein in Haloferax volcanii.

    Lestini, Roxane; Delpech, Floriane; Myllykallio, Hannu

    2015-11-01

    Understanding how frequently spontaneous replication arrests occur and how archaea deal with these arrests are very interesting and challenging research topics. Here we will described how genetic and imaging studies have revealed the central role of the archaeal helicase/nuclease Hef belonging to the XPF/MUS81/FANCM family of endonucleases in repair of arrested replication forks. Special focus will be on description of a recently developed combination of genetic and imaging tools to study the dynamic localization of a functional Hef::GFP (Green Fluorescent Protein) fusion protein in the living cells of halophilic archaea Haloferax volcanii. As Archaea provide an excellent and unique model for understanding how DNA replication is regulated to allow replication of a circular DNA molecule either from single or multiple replication origins, we will also summarize recent studies that have revealed peculiar features regarding DNA replication, particularly in halophilic archaea. We strongly believe that fundamental knowledge of our on-going studies will shed light on the evolutionary history of the DNA replication machinery and will help to establish general rules concerning replication restart and the key role of recombination proteins not only in bacteria, yeast and higher eukaryotes but also in archaea. Copyright © 2015 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

  19. Mutation and Methylation Analysis of the Chromodomain-Helicase-DNA Binding 5 Gene in Ovarian Cancer

    Kylie L. Gorringe

    2008-11-01

    Full Text Available Chromodomain, helicase, DNA binding 5 (CHD5 is a member of a subclass of the chromatin remodeling Swi/Snf proteins and has recently been proposed as a tumor suppressor in a diverse range of human cancers. We analyzed all 41 coding exons of CHD5 for somatic mutations in 123 primary ovarian cancers as well as 60 primary breast cancers using high-resolution melt analysis. We also examined methylation of the CHD5 promoter in 48 ovarian cancer samples by methylation-specific single-stranded conformation polymorphism and bisulfite sequencing. In contrast to previous studies, no mutations were identified in the breast cancers, but somatic heterozygous missense mutations were identified in 3 of 123 ovarian cancers. We identified promoter methylation in 3 of 45 samples with normal CHD5 and in 2 of 3 samples with CHD5 mutation, suggesting these tumors may have biallelic inactivation of CHD5. Hemizygous copy number loss at CHD5 occurred in 6 of 85 samples as assessed by single nucleotide polymorphism array. Tumors with CHD5 mutation or methylation were more likely to have mutation of KRAS or BRAF (P = .04. The aggregate frequency of CHD5 haploinsufficiency or inactivation is 16.2% in ovarian cancer. Thus, CHD5 may play a role as a tumor suppressor gene in ovarian cancer; however, it is likely that there is another target of the frequent copy number neutral loss of heterozygosity observed at 1p36.

  20. Helicase Dependent Isothermal Amplification of DNA and RNA using Self-Avoiding Molecular Recognition Systems

    Yang, Zunyi; McLendon, Chris; Hutter, Daniel; Bradley, Kevin M.; Hoshika, Shuichi; Frye, Carole; Benner, Steven A.

    2015-01-01

    Assays that target DNA or RNA (xNA) are highly sensitive, as small amounts of xNA can be amplified by PCR. Unfortunately, PCR is inconvenient in low resource environments, requiring equipment and power that may not be available in these environments. However, isothermal procedures that avoid thermal cycling are often confounded by primer dimers, off-target priming, and other artifacts. Here, we show how a “self avoiding molecular recognition system” (SAMRS) eliminates these artifacts to give clean amplicons in a helicase-dependent isothermal amplification (SAMRS-HDA). We also show that incorporating SAMRS into the 3′-ends of primers facilitates the design and screening of primers for HDA assays. Finally, we show that SAMRS-HDA can be twofold multiplexed, something difficult to achieve with HDA using standard primers. This shows that SAMRS-HDA is a more versatile approach than standard HDA with a broader applicability for xNA-targeted diagnostics and research. PMID:25953623

  1. Phosphopeptide binding by Sld3 links Dbf4-dependent kinase to MCM replicative helicase activation.

    Deegan, Tom D; Yeeles, Joseph Tp; Diffley, John Fx

    2016-05-02

    The initiation of eukaryotic DNA replication requires the assembly of active CMG (Cdc45-MCM-GINS) helicases at replication origins by a set of conserved and essential firing factors. This process is controlled during the cell cycle by cyclin-dependent kinase (CDK) and Dbf4-dependent kinase (DDK), and in response to DNA damage by the checkpoint kinase Rad53/Chk1. Here we show that Sld3, previously shown to be an essential CDK and Rad53 substrate, is recruited to the inactive MCM double hexamer in a DDK-dependent manner. Sld3 binds specifically to DDK-phosphorylated peptides from two MCM subunits (Mcm4, 6) and then recruits Cdc45. MCM mutants that cannot bind Sld3 or Sld3 mutants that cannot bind phospho-MCM or Cdc45 do not support replication. Moreover, phosphomimicking mutants in Mcm4 and Mcm6 bind Sld3 without DDK and facilitate DDK-independent replication. Thus, Sld3 is an essential "reader" of DDK phosphorylation, integrating signals from three distinct protein kinase pathways to coordinate DNA replication during S phase. © 2016 The Authors. Published under the terms of the CC BY 4.0 license.

  2. Translational control by the DEAD Box RNA helicase belle regulates ecdysone-triggered transcriptional cascades.

    Robert J Ihry

    Full Text Available Steroid hormones act, through their respective nuclear receptors, to regulate target gene expression. Despite their critical role in development, physiology, and disease, however, it is still unclear how these systemic cues are refined into tissue-specific responses. We identified a mutation in the evolutionarily conserved DEAD box RNA helicase belle/DDX3 that disrupts a subset of responses to the steroid hormone ecdysone during Drosophila melanogaster metamorphosis. We demonstrate that belle directly regulates translation of E74A, an ets transcription factor and critical component of the ecdysone-induced transcriptional cascade. Although E74A mRNA accumulates to abnormally high levels in belle mutant tissues, no E74A protein is detectable, resulting in misregulation of E74A-dependent ecdysone response genes. The accumulation of E74A mRNA in belle mutant salivary glands is a result of auto-regulation, fulfilling a prediction made by Ashburner nearly 40 years ago. In this model, Ashburner postulates that, in addition to regulating secondary response genes, protein products of primary response genes like E74A also inhibit their own ecdysone-induced transcription. Moreover, although ecdysone-triggered transcription of E74A appears to be ubiquitous during metamorphosis, belle-dependent translation of E74A mRNA is spatially restricted. These results demonstrate that translational control plays a critical, and previously unknown, role in refining transcriptional responses to the steroid hormone ecdysone.

  3. Mechanical Testing of Carbon Based Woven Thermal Protection Materials

    Pham, John; Agrawal, Parul; Arnold, James O.; Peterson, Keith; Venkatapathy, Ethiraj

    2013-01-01

    Three Dimensional Woven thermal protection system (TPS) materials are one of the enabling technologies for mechanically deployable hypersonic decelerator systems. These materials have been shown capable of serving a dual purpose as TPS and as structural load bearing members during entry and descent operations. In order to ensure successful structural performance, it is important to characterize the mechanical properties of these materials prior to and post exposure to entry-like heating conditions. This research focuses on the changes in load bearing capacity of woven TPS materials after being subjected to arcjet simulations of entry heating. Preliminary testing of arcjet tested materials [1] has shown a mechanical degradation. However, their residual strength is significantly more than the requirements for a mission to Venus [2]. A systematic investigation at the macro and microstructural scales is reported here to explore the potential causes of this degradation. The effects of heating on the sizing (an epoxy resin coating used to reduce friction and wear during fiber handling) are discussed as one of the possible causes for the decrease in mechanical properties. This investigation also provides valuable guidelines for margin policies for future mechanically deployable entry systems.

  4. Mechanics, waves and thermodynamics an example-based approach

    Jain, Sudhir Ranjan

    2016-01-01

    The principles of classical physics, though superseded in specific fields by such theories as quantum mechanics and general relativity, are still of great importance in a broad range of applications. The book presents fundamental concepts of classical physics in a coherent and logical manner. It discusses important topics including the mechanics of a single particle, kinetic theory, oscillations and waves. Topics including the kinetic theory of gases, thermodynamics and statistical mechanics are discussed, which are normally not present in the books on classical physics. The fundamental concepts of energy, momentum, mass and entropy are explained with examples. Discussion on concepts of thermodynamics is presented along with the simplified explanation on Caratheodory's axioms. It covers chapters on wave motion and statistical physics, useful for the graduate students. Each concept is supported with real-life applications on several concepts including impulse and collision, Bernoulli's equation, and friction.

  5. Mycobacterium smegmatis Lhr Is a DNA-dependent ATPase and a 3'-to-5' DNA translocase and helicase that prefers to unwind 3'-tailed RNA:DNA hybrids.

    Ordonez, Heather; Shuman, Stewart

    2013-05-17

    We are interested in the distinctive roster of helicases of Mycobacterium, a genus of the phylum Actinobacteria that includes the human pathogen Mycobacterium tuberculosis and its avirulent relative Mycobacterium smegmatis. Here, we identify and characterize M. smegmatis Lhr as the exemplar of a novel clade of superfamily II helicases, by virtue of its biochemical specificities and signature domain organization. Lhr is a 1507-amino acid monomeric nucleic acid-dependent ATPase that uses the energy of ATP hydrolysis to drive unidirectional 3'-to-5' translocation along single strand DNA and to unwind duplexes en route. The ATPase is more active in the presence of calcium than magnesium. ATP hydrolysis is triggered by either single strand DNA or single strand RNA, yet the apparent affinity for a DNA activator is 11-fold higher than for an RNA strand of identical size and nucleobase sequence. Lhr is 8-fold better at unwinding an RNA:DNA hybrid than it is at displacing a DNA:DNA duplex of identical nucleobase sequence. The truncated derivative Lhr-(1-856) is an autonomous ATPase, 3'-to-5' translocase, and RNA:DNA helicase. Lhr-(1-856) is 100-fold better RNA:DNA helicase than DNA:DNA helicase. Lhr homologs are found in bacteria representing eight different phyla, being especially prevalent in Actinobacteria (including M. tuberculosis) and Proteobacteria (including Escherichia coli).

  6. Content-Based Image Retrieval Based on Electromagnetism-Like Mechanism

    Hamid A. Jalab

    2013-01-01

    Full Text Available Recently, many researchers in the field of automatic content-based image retrieval have devoted a remarkable amount of research looking for methods to retrieve the best relevant images to the query image. This paper presents a novel algorithm for increasing the precision in content-based image retrieval based on electromagnetism optimization technique. The electromagnetism optimization is a nature-inspired technique that follows the collective attraction-repulsion mechanism by considering each image as an electrical charge. The algorithm is composed of two phases: fitness function measurement and electromagnetism optimization technique. It is implemented on a database with 8,000 images spread across 80 classes with 100 images in each class. Eight thousand queries are fired on the database, and the overall average precision is computed. Experimental results of the proposed approach have shown significant improvement in the retrieval performance in regard to precision.

  7. Hybrid magnetic mechanism for active locomotion based on inchworm motion

    Kim, Sung Hoon; Hashi, Shuichiro; Ishiyama, Kazushi

    2013-01-01

    Magnetic robots have been studied in the past. Insect-type micro-robots are used in various biomedical applications; researchers have developed inchworm micro-robots for endoscopic use. A biological inchworm has a looping locomotion gait. However, most inchworm micro-robots depend on a general bending, or bellows, motion. In this paper, we introduce a new robotic mechanism using magnetic force and torque control in a rotating magnetic field for a looping gait. The proposed robot is controlled by the magnetic torque, attractive force, and body mechanisms (two stoppers, flexible body, and different frictional legs). The magnetic torque generates a general bending motion. In addition, the attractive force and body mechanisms produce the robot’s looping motion within a rotating magnetic field and without the use of an algorithm for field control. We verified the device’s performance and analyzed the motion through simulations and various experiments. The robot mechanism can be applied to active locomotion for various medical robots, such as wireless endoscopes. (technical note)

  8. Innovative Mechanism of Rural Organization Based on Self-Organization

    2011-01-01

    The paper analyzes the basic situation of the formation of innovative rural organizations with the form of self-organization;reveals the features of self-organization,including the four aspects of openness of rural organization,innovation of rural organization far away from equilibrium,the non-linear response mechanism of rural organization innovation and the random rise and fall of rural organization innovation.The evolution mechanism of rural organization innovation is revealed according to the growth stage,the ideal stage,the decline and the fall stage.The paper probes into the basic restriction mechanism of the self-organization evaluation of rural organization from three aspects,including target recognition,path dependence and knowledge sharing.The basic measures on cultivating the innovative mechanism of rural organization are put forward.Firstly,constructing the dissipative structure of rural organization innovation;secondly,cultivating the dynamic study capability of rural organization innovation system;thirdly,selecting the step-by-step evolution strategy of rural organization innovation system.

  9. Mechanism-Based Causal Reasoning in Young Children

    Buchanan, David W.; Sobel, David M.

    2011-01-01

    The hypothesis that children develop an understanding of causal mechanisms was tested across 3 experiments. In Experiment 1 (N = 48), preschoolers had to choose as efficacious either a cause that had worked in the past, but was now disconnected from its effect, or a cause that had failed to work previously, but was now connected. Four-year-olds…

  10. Designing Competency-Based Materials for the Automotive Mechanics Curriculum

    Richardson, Roger L.

    1977-01-01

    Describes the Career Education Center's (Florida State University) development of the "Automotive Mechanics Catalog" (a job inventory noting performance objectives for specific occupational programs), using the Vocational-Technical Education Consortium of States (V-TECS) model. Also describes the development of curriculum materials using…

  11. Measuring Clearance Mechanics Based on Dynamic Leg Length

    Khamis, Sam; Danino, Barry; Hayek, Shlomo; Carmeli, Eli

    2018-01-01

    The aim of this study was to quantify clearance mechanics during gait. Seventeen children diagnosed with hemiplegic cerebral palsy underwent a three-dimensional gait analysis evaluation. Dynamic leg lengths were measured from the hip joint center to the heel, to the ankle joint center and to the forefoot throughout the gait cycle. Significant…

  12. Structure-based membrane dome mechanism for Piezo mechanosensitivity.

    Guo, Yusong R; MacKinnon, Roderick

    2017-12-12

    Mechanosensitive ion channels convert external mechanical stimuli into electrochemical signals for critical processes including touch sensation, balance, and cardiovascular regulation. The best understood mechanosensitive channel, MscL, opens a wide pore, which accounts for mechanosensitive gating due to in-plane area expansion. Eukaryotic Piezo channels have a narrow pore and therefore must capture mechanical forces to control gating in another way. We present a cryo-EM structure of mouse Piezo1 in a closed conformation at 3.7Å-resolution. The channel is a triskelion with arms consisting of repeated arrays of 4-TM structural units surrounding a pore. Its shape deforms the membrane locally into a dome. We present a hypothesis in which the membrane deformation changes upon channel opening. Quantitatively, membrane tension will alter gating energetics in proportion to the change in projected area under the dome. This mechanism can account for highly sensitive mechanical gating in the setting of a narrow, cation-selective pore. © 2017, Guo et al.

  13. Designing remote web-based mechanical-volumetric flow meter ...

    Today, in water and wastewater industry a lot of mechanical-volumetric flow meters are used for the navigation of the produced water and the data of these flow meters, due to use in a wide geographical range, is done physically and by in person presence. All this makes reading the data costly and, in some cases, due to ...

  14. Could humicity affect the mechanical properties of carbon based coatings?

    Sobota, Jaroslav; Grossman, Jan; Vyskočil, J.; Novák, R.; Fořt, Tomáš; Vítů, T.; Dupák, Libor

    2010-01-01

    Roč. 104, č. 15 (2010), s. 375-377 ISSN 0009-2770 Institutional research plan: CEZ:AV0Z20650511 Keywords : carbon * mechanical properties * humidity * fracture toughness of hard thin coatings Subject RIV: JI - Composite Materials Impact factor: 0.620, year: 2010

  15. Mechanics

    Hartog, J P Den

    1961-01-01

    First published over 40 years ago, this work has achieved the status of a classic among introductory texts on mechanics. Den Hartog is known for his lively, discursive and often witty presentations of all the fundamental material of both statics and dynamics (and considerable more advanced material) in new, original ways that provide students with insights into mechanical relationships that other books do not always succeed in conveying. On the other hand, the work is so replete with engineering applications and actual design problems that it is as valuable as a reference to the practicing e

  16. 'Fingerprints' of vertebral trauma - a unifying concept based on mechanisms

    Daffner, R.H.; Deeb, Z.L.; Rothfus, W.E.

    1986-01-01

    Vertebral fractures, like fractures in the peripheral skeleton, occur in predictable and reproducible patterns that are related to the kind of force applied to the affected bone. The same force applied to the cervical, thoracic, or lumbar column will result in injuries which appear quite similar. A review of 621 injuries to the vertebral column revealed that there are basically four mechanisms of injury: flexion, extension, shearing, and torque (rotation). These injuries may occur by themselves or in combination with one another. The severity and extent of damage produced by any mechanism is dependent upon the incident force, the position of the patient at the time of injury, and the velocity of the patient. Thus, there is a pattern of recognizable signs which span the spectrum from mild soft tissue damage to severe skeletal and ligamentous disruption. These patterns are termed the 'fingerprints' of the injury, and this presentation illustrates the four basic types of vertebral injury producing those 'fingerprints'. (orig.)

  17. Mechanical properties of green composites based on thermoplastic starch

    Fornes, F.; Sánchez-Nácher, L.; Fenollar, O.; Boronat, T.; Garcia-Sanoguera, D.

    2010-06-01

    The present work is focused on study of "green composites" elaborated from thermoplastic starch (TPS) as polymer matrix and a fiber from natural origin (rush) as reinforced fiber. The effect of the fiber content has been studied by means of the mechanical properties. The composite resulting presents a lack of interaction between matrix and fiber that represents a performance decrease. However the biodegradability behavior of the resulting composite raise this composite as useful an industrial level.

  18. Artificial Neural Network Based Mission Planning Mechanism for Spacecraft

    Li, Zhaoyu; Xu, Rui; Cui, Pingyuan; Zhu, Shengying

    2018-04-01

    The ability to plan and react fast in dynamic space environments is central to intelligent behavior of spacecraft. For space and robotic applications, many planners have been used. But it is difficult to encode the domain knowledge and directly use existing techniques such as heuristic to improve the performance of the application systems. Therefore, regarding planning as an advanced control problem, this paper first proposes an autonomous mission planning and action selection mechanism through a multiple layer perceptron neural network approach to select actions in planning process and improve efficiency. To prove the availability and effectiveness, we use autonomous mission planning problems of the spacecraft, which is a sophisticated system with complex subsystems and constraints as an example. Simulation results have shown that artificial neural networks (ANNs) are usable for planning problems. Compared with the existing planning method in EUROPA, the mechanism using ANNs is more efficient and can guarantee stable performance. Therefore, the mechanism proposed in this paper is more suitable for planning problems of spacecraft that require real time and stability.

  19. A Snapshot-Based Mechanism for Celestial Orientation.

    El Jundi, Basil; Foster, James J; Khaldy, Lana; Byrne, Marcus J; Dacke, Marie; Baird, Emily

    2016-06-06

    In order to protect their food from competitors, ball-rolling dung beetles detach a piece of dung from a pile, shape it into a ball, and roll it away along a straight path [1]. They appear to rely exclusively on celestial compass cues to maintain their bearing [2-8], but the mechanism that enables them to use these cues for orientation remains unknown. Here, we describe the orientation strategy that allows dung beetles to use celestial cues in a dynamic fashion. We tested the underlying orientation mechanism by presenting beetles with a combination of simulated celestial cues (sun, polarized light, and spectral cues). We show that these animals do not rely on an innate prediction of the natural geographical relationship between celestial cues, as other navigating insects seem to [9, 10]. Instead, they appear to form an internal representation of the prevailing celestial scene, a "celestial snapshot," even if that scene represents a physical impossibility for the real sky. We also find that the beetles are able to maintain their bearing with respect to the presented cues only if the cues are visible when the snapshot is taken. This happens during the "dance," a behavior in which the beetle climbs on top of its ball and rotates about its vertical axis [11]. This strategy for reading celestial signals is a simple but efficient mechanism for straight-line orientation. Copyright © 2016 Elsevier Ltd. All rights reserved.

  20. Nanomechanical microcantilever operated in vibration modes with use of RNA aptamer as receptor molecules for label-free detection of HCV helicase.

    Hwang, Kyo Seon; Lee, Sang-Myung; Eom, Kilho; Lee, Jeong Hoon; Lee, Yoon-Sik; Park, Jung Ho; Yoon, Dae Sung; Kim, Tae Song

    2007-11-30

    We report the nanomechanical microcantilevers operated in vibration modes (oscillation) with use of RNA aptamers as receptor molecules for label-free detection of hepatitis C virus (HCV) helicase. The nanomechanical detection principle is that the ligand-receptor binding on the microcantilever surface induces the dynamic response change of microcantilevers. We implemented the label-free detection of HCV helicase in the low concentration as much as 100 pg/ml from measuring the dynamic response change of microcantilevers. Moreover, from the recent studies showing that the ligand-receptor binding generates the surface stress on the microcantilever, we estimate the surface stress, on the oscillating microcantilevers, induced by ligand-receptor binding, i.e. binding between HCV helicase and RNA aptamer. In this article, it is suggested that the oscillating microcantilevers with use of RNA aptamers as receptor molecules may enable one to implement the sensitive label-free detection of very small amount of small-scale proteins.

  1. Mechanical properties of denture base resins: An evaluation

    Pooran Chand

    2011-01-01

    Conclusion: The heat cure denture base material D (Trevalon "HI" was the strongest and C (Trevalon was the weakest among all materials used in this study. The study showed that the deflection of various denture base resins (A to D increases proportionately with the increase in load.

  2. The Processing and Mechanical Properties of High Temperature/ High Performance Composites. Mechanism-Based Constitutive Laws and Design. Book 1

    1994-03-01

    NTIS CRA&I IDT!C TAB TABLE OF CONTENTS U.11 nulce Distf ib,.tion, I Availab~iity Cojes EXECUTIVE SUMMARY Dist spc :BOOKh1 MECHANISM-BASED...unit cylinders (H=D), but rather by slightly squat cylinders. This is in contrast to the results of Bao et al. [1] in which unit cylinders resulted in

  3. Mycobacterium tuberculosis DinG is a structure-specific helicase that unwinds G4 DNA: implications for targeting G4 DNA as a novel therapeutic approach.

    Thakur, Roshan Singh; Desingu, Ambika; Basavaraju, Shivakumar; Subramanya, Shreelakshmi; Rao, Desirazu N; Nagaraju, Ganesh

    2014-09-05

    The significance of G-quadruplexes and the helicases that resolve G4 structures in prokaryotes is poorly understood. The Mycobacterium tuberculosis genome is GC-rich and contains >10,000 sequences that have the potential to form G4 structures. In Escherichia coli, RecQ helicase unwinds G4 structures. However, RecQ is absent in M. tuberculosis, and the helicase that participates in G4 resolution in M. tuberculosis is obscure. Here, we show that M. tuberculosis DinG (MtDinG) exhibits high affinity for ssDNA and ssDNA translocation with a 5' → 3' polarity. Interestingly, MtDinG unwinds overhangs, flap structures, and forked duplexes but fails to unwind linear duplex DNA. Our data with DNase I footprinting provide mechanistic insights and suggest that MtDinG is a 5' → 3' polarity helicase. Notably, in contrast to E. coli DinG, MtDinG catalyzes unwinding of replication fork and Holliday junction structures. Strikingly, we find that MtDinG resolves intermolecular G4 structures. These data suggest that MtDinG is a multifunctional structure-specific helicase that unwinds model structures of DNA replication, repair, and recombination as well as G4 structures. We finally demonstrate that promoter sequences of M. tuberculosis PE_PGRS2, mce1R, and moeB1 genes contain G4 structures, implying that G4 structures may regulate gene expression in M. tuberculosis. We discuss these data and implicate targeting G4 structures and DinG helicase in M. tuberculosis could be a novel therapeutic strategy for culminating the infection with this pathogen. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  4. CMG helicase and DNA polymerase ε form a functional 15-subunit holoenzyme for eukaryotic leading-strand DNA replication.

    Langston, Lance D; Zhang, Dan; Yurieva, Olga; Georgescu, Roxana E; Finkelstein, Jeff; Yao, Nina Y; Indiani, Chiara; O'Donnell, Mike E

    2014-10-28

    DNA replication in eukaryotes is asymmetric, with separate DNA polymerases (Pol) dedicated to bulk synthesis of the leading and lagging strands. Pol α/primase initiates primers on both strands that are extended by Pol ε on the leading strand and by Pol δ on the lagging strand. The CMG (Cdc45-MCM-GINS) helicase surrounds the leading strand and is proposed to recruit Pol ε for leading-strand synthesis, but to date a direct interaction between CMG and Pol ε has not been demonstrated. While purifying CMG helicase overexpressed in yeast, we detected a functional complex between CMG and native Pol ε. Using pure CMG and Pol ε, we reconstituted a stable 15-subunit CMG-Pol ε complex and showed that it is a functional polymerase-helicase on a model replication fork in vitro. On its own, the Pol2 catalytic subunit of Pol ε is inefficient in CMG-dependent replication, but addition of the Dpb2 protein subunit of Pol ε, known to bind the Psf1 protein subunit of CMG, allows stable synthesis with CMG. Dpb2 does not affect Pol δ function with CMG, and thus we propose that the connection between Dpb2 and CMG helps to stabilize Pol ε on the leading strand as part of a 15-subunit leading-strand holoenzyme we refer to as CMGE. Direct binding between Pol ε and CMG provides an explanation for specific targeting of Pol ε to the leading strand and provides clear mechanistic evidence for how strand asymmetry is maintained in eukaryotes.

  5. Mechanism of action of mRNA-based vaccines.

    Iavarone, Carlo; O'hagan, Derek T; Yu, Dong; Delahaye, Nicolas F; Ulmer, Jeffrey B

    2017-09-01

    The present review summarizes the growing body of work defining the mechanisms of action of this exciting new vaccine technology that should allow rational approaches in the design of next generation mRNA vaccines. Areas covered: Bio-distribution of mRNA, localization of antigen production, role of the innate immunity, priming of the adaptive immune response, route of administration and effects of mRNA delivery systems. Expert commentary: In the last few years, the development of RNA vaccines had a fast growth, the rising number of proof will enable rational approaches to improving the effectiveness and safety of this modern class of medicine.

  6. A PLL based automated magnetron tuning mechanism for electron accelerators

    Khan, A M; Mahfooz, Mohammed; Sanjeev, Ganesh

    2008-01-01

    In this paper we report on a control system developed to tune the magnetron frequency to get the maximum beam pulse in a Microtron (an electron accelerator facility at Mangalore University). The control system so designed consists of a Microcontroller Unit (MCU), a Phase Locked Loop (PLL) and a Digital to Analog Converter (DAC) to track and tune the magnetron frequency. A PLL is used to track the deviation of the magnetron output frequency, and by monitoring the reflected wave voltage level, the microcontroller unit tunes the magnetron with the help of a tuner mechanism connected through a stepper motor.

  7. A PLL based automated magnetron tuning mechanism for electron accelerators

    Khan, A M; Mahfooz, Mohammed [Dept. of Electronics, Mangalore University, Mangalagangotri, Karnataka State, India - 574 199 (India); Sanjeev, Ganesh [Microtron Centre, Mangalore University, Mangalagangotri, Karnataka State, India - 574 199 (India)], E-mail: mahfooz_81@yahoo.com

    2008-09-15

    In this paper we report on a control system developed to tune the magnetron frequency to get the maximum beam pulse in a Microtron (an electron accelerator facility at Mangalore University). The control system so designed consists of a Microcontroller Unit (MCU), a Phase Locked Loop (PLL) and a Digital to Analog Converter (DAC) to track and tune the magnetron frequency. A PLL is used to track the deviation of the magnetron output frequency, and by monitoring the reflected wave voltage level, the microcontroller unit tunes the magnetron with the help of a tuner mechanism connected through a stepper motor.

  8. ASPECTS REGARDING MECHANICAL PROCESSING OF STEELS FOR MAGNETS, NDFEB BASED

    MELANIA TĂMAŞ

    2015-05-01

    Full Text Available This paper presents experimental research concerning the behavior on cutting by turning of steels for magnets NdFeB based. In this context, cutting by rough turning of steels magnet neodymium and boron ferrite based were performed. Turning processing with the values of the cutting parameters recommended by European Union and Sandvik Coromant company rules, taking into account the chemical composition of the processed material and cutting tool were performed. The tables have been drawn up with different values of the cutting parameters. By comparing these data it can be concluded that steels for permanent magnets, NdFeB based have acceptable behavior, the process by rough turning. A full assessment of the optimization of the cutting by turning process of steels for permanent magnet, NdFeB base will result from subsequent experimental research that will take into account the wear of cutting tools and quality (roughness of processed surface.

  9. Mcm3 replicative helicase mutation impairs neuroblast proliferation and memory in Drosophila.

    Blumröder, R; Glunz, A; Dunkelberger, B S; Serway, C N; Berger, C; Mentzel, B; de Belle, J S; Raabe, T

    2016-09-01

    In the developing Drosophila brain, a small number of neural progenitor cells (neuroblasts) generate in a co-ordinated manner a high variety of neuronal cells by integration of temporal, spatial and cell-intrinsic information. In this study, we performed the molecular and phenotypic characterization of a structural brain mutant called small mushroom bodies (smu), which was isolated in a screen for mutants with altered brain structure. Focusing on the mushroom body neuroblast lineages we show that failure of neuroblasts to generate the normal number of mushroom body neurons (Kenyon cells) is the major cause of the smu phenotype. In particular, the premature loss of mushroom body neuroblasts caused a pronounced effect on the number of late-born Kenyon cells. Neuroblasts showed no obvious defects in processes controlling asymmetric cell division, but generated less ganglion mother cells. Cloning of smu uncovered a single amino acid substitution in an evolutionarily conserved protein interaction domain of the Minichromosome maintenance 3 (Mcm3) protein. Mcm3 is part of the multimeric Cdc45/Mcm/GINS (CMG) complex, which functions as a helicase during DNA replication. We propose that at least in the case of mushroom body neuroblasts, timely replication is not only required for continuous proliferation but also for their survival. The absence of Kenyon cells in smu reduced learning and early phases of conditioned olfactory memory. Corresponding to the absence of late-born Kenyon cells projecting to α'/β' and α/β lobes, smu is profoundly defective in later phases of persistent memory. © 2016 John Wiley & Sons Ltd and International Behavioural and Neural Genetics Society.

  10. Frequency of Werner helicase 1367 polymorphism and age-related morbidity in an elderly Brazilian population

    M.A.C. Smith

    2005-07-01

    Full Text Available Werner syndrome (WS is a premature aging disease caused by a mutation in the WRN gene. The gene was identified in 1996 and its product acts as a DNA helicase and exonuclease. Some specific WRN polymorphic variants were associated with increased risk for cardiovascular diseases. The identification of genetic polymorphisms as risk factors for complex diseases affecting older people can improve their prevention, diagnosis and prognosis. We investigated WRN codon 1367 polymorphism in 383 residents in a district of the city of São Paulo, who were enrolled in an Elderly Brazilian Longitudinal Study. Their mean age was 79.70 ± 5.32 years, ranging from 67 to 97. This population was composed of 262 females (68.4% and 121 males (31.6% of European (89.2%, Japanese (3.3%, Middle Eastern (1.81%, and mixed and/or other origins (5.7%. There are no studies concerning this polymorphism in Brazilian population. These subjects were evaluated clinically every two years. The major health problems and morbidities affecting this cohort were cardiovascular diseases (21.7%, hypertension (83.7%, diabetes (63.3%, obesity (41.23%, dementia (8.0%, depression (20.0%, and neoplasia (10.8%. Their prevalence is similar to some urban elderly Brazilian samples. DNA was isolated from blood cells, amplified by PCR and digested with PmaCI. Allele frequencies were 0.788 for the cysteine and 0.211 for the arginine. Genotype distributions were within that expected for the Hardy-Weinberg equilibrium. Female gender was associated with hypertension and obesity. Logistic regression analysis did not detect significant association between the polymorphism and morbidity. These findings confirm those from Europeans and differ from Japanese population.

  11. Antiulcer properties of fruits and vegetables: A mechanism based perspective.

    Harsha, Choudhary; Banik, Kishore; Bordoloi, Devivasha; Kunnumakkara, Ajaikumar B

    2017-10-01

    Gastric ulcer is the damage caused to mucosal layer of the stomach under the action of various factors like high levels of acid and pepsin, invasion by Helicobacter pylori, etc. Although most cases have been controlled and the rate of ulcer occurrence has reduced over the last few decades, gastric ulcer still holds a prime concern today. A range of palliative medicines comprising proton pump inhibitors, H2 receptor antagonists, COX-2 inhibitors (coxibs) is widely in use and patients have also been administered with acid suppression therapies. But these remedies aggravate the condition of patients causing severe side effects, or rather impart temporary relief. Therefore, it is highly imperative to develop safe and effective therapies for the treatment of gastric ulcer. Nature provides us various fruits and vegetables that can combat gastric ulcer through multiple mechanisms; predominantly via antioxidant, anti-inflammatory, antisecretory, antimicrobial, anticholinergic and cytoprotective activity, inhibition of small intestinal propulsion etc. Various phytochemicals from fruits and vegetables such as phenolics, flavonoids, tannins and saponins play a vital role in the prevention and cure of gastric ulcer. This review is a compendium of all fruits and vegetables known for their profound antiulcer effect and their underlying mechanisms of action. Copyright © 2017 Elsevier Ltd. All rights reserved.

  12. Epoxy Resin Based Composites, Mechanical and Tribological Properties: A Review

    S.A. Bello

    2015-12-01

    Full Text Available High fuel consumption by automobile and aerospace vehicles built from legacy alloys has been a great challenge to global design and material engineers. This has called for researches into material development for the production of lighter materials of the same or even superior mechanical properties to the existing materials in this area of applications. This forms a part of efforts to achieve the global vision 2025 i.e to reduce the fuel consumption by automobile and aerospace vehicles by at least 75 %. Many researchers have identified advanced composites as suitable materials in this regard. Among the common matrices used for the development of advanced composites, epoxy resin has attained a dominance among its counterparts because of its excellent properties including chemical, thermal and electrical resistance properties, mechanical properties and dimensional stability. This review is a reflection of the extensive study on the currently ongoing research aimed at development of epoxy resin hybrid nanocomposites for engineering applications. In this paper, brief explanation has been given to different terms related to the research work and also, some previous works (in accordance with materials within authors’ reach in the area of the ongoing research have been reported.

  13. Epidemiological bases and molecular mechanisms linking obesity, diabetes, and cancer.

    Gutiérrez-Salmerón, María; Chocarro-Calvo, Ana; García-Martínez, José Manuel; de la Vieja, Antonio; García-Jiménez, Custodia

    2017-02-01

    The association between diabetes and cancer was hypothesized almost one century ago. Today, a vast number of epidemiological studies support that obese and diabetic populations are more likely to experience tissue-specific cancers, but the underlying molecular mechanisms remain unknown. Obesity, diabetes, and cancer share many hormonal, immune, and metabolic changes that may account for the relationship between diabetes and cancer. In addition, antidiabetic treatments may have an impact on the occurrence and course of some cancers. Moreover, some anticancer treatments may induce diabetes. These observations aroused a great controversy because of the ethical implications and the associated commercial interests. We report an epidemiological update from a mechanistic perspective that suggests the existence of many common and differential individual mechanisms linking obesity and type 1 and 2 diabetes mellitus to certain cancers. The challenge today is to identify the molecular links responsible for this association. Classification of cancers by their molecular signatures may facilitate future mechanistic and epidemiological studies. Copyright © 2016 SEEN. Publicado por Elsevier España, S.L.U. All rights reserved.

  14. Adaptive mechanism-based congestion control for networked systems

    Liu, Zhi; Zhang, Yun; Chen, C. L. Philip

    2013-03-01

    In order to assure the communication quality in network systems with heavy traffic and limited bandwidth, a new ATRED (adaptive thresholds random early detection) congestion control algorithm is proposed for the congestion avoidance and resource management of network systems. Different to the traditional AQM (active queue management) algorithms, the control parameters of ATRED are not configured statically, but dynamically adjusted by the adaptive mechanism. By integrating with the adaptive strategy, ATRED alleviates the tuning difficulty of RED (random early detection) and shows a better control on the queue management, and achieve a more robust performance than RED under varying network conditions. Furthermore, a dynamic transmission control protocol-AQM control system using ATRED controller is introduced for the systematic analysis. It is proved that the stability of the network system can be guaranteed when the adaptive mechanism is finely designed. Simulation studies show the proposed ATRED algorithm achieves a good performance in varying network environments, which is superior to the RED and Gentle-RED algorithm, and providing more reliable service under varying network conditions.

  15. Beyond reverse pharmacology: Mechanism-based screening of Ayurvedic drugs.

    Lele, R D

    2010-10-01

    This paper reviews the pharmacology of Indian medicinal plants, starting with the historical background of European work on the subject beginning as early as the 17th century, and tracing its history through the work of Sen and Bose in the 1930's, and Vakhil's historic 1949 paper on Sarpaghanda. The often crucial role of patient feedback in early discoveries is highlighted, as is the time lag between proof of pharmacological action and identification of the active principle, and subsequent elucidation of mechanism of action. In the case of Indian plants in the 20th century this process sometimes took almost 50 years. Reserpine and its mechanisms are given in detail, and its current relevance to public health discussed. The foundation of present day methods of pharmacology is briefly presented so the complexity of methods used to identify properties of Ayurveda derived drugs like forskolin and baicalein, and their bioavailability, may be better appreciated. Ayurveda derived anti-oxidants and their levels of action, immuno-modulators, particularly with respect to the NF-kB pathway and its implications for cancer control, are all considered. The example of curcumin derived from turmeric is explained in more detail, because of its role in cancer prevention. Finally, the paper emphasizes the importance of Ayurveda's concepts of rasayana as a form of dietary chemo-prevention; the significance of ahar, diet, in Ayurveda's aspiration to prevent disease and restore health thus becomes clear. Understood in this light, Ayurveda may transcend pharmacology as a treatment paradigm.

  16. microRNAs targeting DEAD-box helicases are involved in salinity stress response in rice (Oryza sativa L.

    Macovei Anca

    2012-10-01

    Full Text Available Abstract Background Rice (Oryza sativa L., one of the most important food crop in the world, is considered to be a salt-sensitive crop. Excess levels of salt adversely affect all the major metabolic activities, including cell wall damage, cytoplasmic lysis and genomic stability. In order to cope with salt stress, plants have evolved high degrees of developmental plasticity, including adaptation via cascades of molecular networks and changes in gene expression profiles. Posttranscriptional regulation, through the activity of microRNAs, also plays an important role in the plant response to salinity conditions. MicroRNAs are small endogenous RNAs that modulate gene expression and are involved in the most essential physiological processes, including plant development and adaptation to environmental changes. Results In the present study, we investigated the expression profiles of osa-MIR414, osa-MIR408 and osa-MIR164e along with their targeted genes, under salinity stress conditions in wild type and transgenic rice plants ectopically expressing the PDH45 (Pea DNA Helicase gene. The present miRNAs were predicted to target the OsABP (ATP-Binding Protein, OsDSHCT (DOB1/SK12/helY-like DEAD-box Helicase and OsDBH (DEAD-Box Helicase genes, included in the DEAD-box helicase family. An in silico characterization of the proteins was performed and the miRNAs predicted targets were validated by RLM-5′RACE. The qRT-PCR analysis showed that the OsABP, OsDBH and OsDSHCT genes were up-regulated in response to 100 and 200 mM NaCl treatments. The present study also highlighted an increased accumulation of the gene transcripts in wild type plants, with the exception of the OsABP mRNA which showed the highest level (15.1-fold change compared to control in the transgenic plants treated with 200 mM NaCl. Salinity treatments also affected the expression of osa-MIR414, osa-MIR164e and osa-MIR408, found to be significantly down-regulated, although the changes in mi

  17. RECQ5 Helicase Cooperates with MUS81 Endonuclease in Processing Stalled Replication Forks at Common Fragile Sites during Mitosis

    Di Marco, Stefano; Hasanova, Zdenka; Kanagaraj, Radhakrishnan

    2017-01-01

    The MUS81-EME1 endonuclease cleaves late replication intermediates at common fragile sites (CFSs) during early mitosis to trigger DNA-repair synthesis that ensures faithful chromosome segregation. Here, we show that these DNA transactions are promoted by RECQ5 DNA helicase in a manner dependent...... on its Ser727 phosphorylation by CDK1. Upon replication stress, RECQ5 associates with CFSs in early mitosis through its physical interaction with MUS81 and promotes MUS81-dependent mitotic DNA synthesis. RECQ5 depletion or mutational inactivation of its ATP-binding site, RAD51-interacting domain...

  18. Human RECQ5 helicase promotes repair of DNA double-strand breaks by synthesis-dependent strand annealing

    Paliwal, S.; Kanagaraj, R.; Sturzenegger, A.; Burdová, Kamila; Janščák, Pavel

    2014-01-01

    Roč. 42, č. 4 (2014), s. 2380-2390 ISSN 0305-1048 R&D Projects: GA ČR GA204/09/0565; GA ČR GAP305/10/0281 Grant - others:Swiss National Science Foundation(CH) 31003A-129747; Swiss National Science Foundation(CH) 31003A_146206 Institutional support: RVO:68378050 Keywords : Human RECQ5 helicase * DNA double-strand breaks * mitotic homologous recombination Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 9.112, year: 2014

  19. Design and fabrication of a eccentric wheels based motorised alignment mechanism for cylindrical accelerator components

    Mundra, G.; Jain, V.; Karmarkar, Mangesh; Kotaiah, S.

    2006-01-01

    Precision alignment mechanisms with long term stability are required for accelerator components. For some of the components motorised and remotely operable alignment mechanism are required. An eccentric wheel mechanism based alignment system is very much suitable for such application. One such alignment system is designed, a prototype is machined/fabricated for SFDTL type accelerating structure and preliminary trial experiments have been done. (author)

  20. Bismuth oxide based ceramics with improved electrical and mechanical properties: Part II. Structural and mechanical properties

    Kruidhof, H.; Seshan, Kulathuiyer; van de Velde, G.M.H.; de Vries, K.J.; Burggraaf, A.J.

    1988-01-01

    Coprecipitation as a method of preparation for bismuth oxides based ceramics yields relatively strong and machineable materials in comparison with the solid state reaction. Compositions within the system (1−x)Bi2O3|xEr2O3 containing up to twenty five mole percent of erbium oxide show a slow

  1. Density functional theory and quantum mechanics/molecular mechanics study of cysteine protease inhibition by nitrile-based inhibitors.

    Sam P De Visser

    2013-12-01

    Full Text Available Cysteine protease enzymes are important for human physiology and catalyze key protein degradation pathways. These enzymes react via a nucleophilic reaction mechanism that involves a cysteine residue and the proton of a proximal histidine. Particularly efficient inhibitors of these enzymes are nitrile-based, however, the details of the catalytic reaction mechanism currently are poorly understood. To gain further insight into the inhibition of these molecules, we have performed a combined density functional theory and quantum mechanics/molecular mechanics study on the reaction of a nitrile-based inhibitor with the enzyme active site amino acids. We show here that small perturbations to the inhibitor structure can have dramatic effects on the catalysis and inhibition processes. Thus, we investigated a range of inhibitor templates and show that specific structural changes reduce the inhibitory efficiency by several orders of magnitude. Moreover, as the reaction takes place on a polar surface, we find strong differences between the DFT and QM/MM calculated energetics. In particular, the DFT model led to dramatic distortions from the starting structure and the convergence to a structure that would not fit the enzyme active site. In the subsequent QM/MM study we investigated the use of mechanical versus electronic embedding on the kinetics, thermodynamics and geometries along the reaction mechanism. We find minor effects on the kinetics of the reaction but large geometric and thermodynamics differences as a result of inclusion of electronic embedding corrections. The work here highlights the importance of model choice in the investigation of this biochemical reaction mechanism.

  2. Binding of DEAD-box helicase Dhh1 to the 5'-untranslated region of ASH1 mRNA represses localized translation of ASH1 in yeast cells.

    Zhang, Qianjun; Meng, Xiuhua; Li, Delin; Chen, Shaoyin; Luo, Jianmin; Zhu, Linjie; Singer, Robert H; Gu, Wei

    2017-06-09

    Local translation of specific mRNAs is regulated by dynamic changes in their subcellular localization, and these changes are due to complex mechanisms controlling cytoplasmic mRNA transport. The budding yeast Saccharomyces cerevisiae is well suited to studying these mechanisms because many of its transcripts are transported from the mother cell to the budding daughter cell. Here, we investigated the translational control of ASH1 mRNA after transport and localization. We show that although ASH1 transcripts were translated after they reached the bud tip, some mRNAs were bound by the RNA-binding protein Puf6 and were non-polysomal. We also found that the DEAD-box helicase Dhh1 complexed with the untranslated ASH1 mRNA and Puf6. Loss of Dhh1 affected local translation of ASH1 mRNA and resulted in delocalization of ASH1 transcript in the bud. Forcibly shifting the non-polysomal ASH1 mRNA into polysomes was associated with Dhh1 dissociation. We further demonstrated that Dhh1 is not recruited to ASH1 mRNA co-transcriptionally, suggesting that it could bind to ASH1 mRNA within the cytoplasm. Of note, Dhh1 bound to the 5'-UTR of ASH1 mRNA and inhibited its translation in vitro These results suggest that after localization to the bud tip, a portion of the localized ASH1 mRNA becomes translationally inactive because of binding of Dhh1 and Puf6 to the 5'- and 3'-UTRs of ASH1 mRNA. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  3. DNA helicase HIM-6/BLM both promotes MutSγ-dependent crossovers and antagonizes MutSγ-independent interhomolog associations during caenorhabditis elegans meiosis.

    Schvarzstein, Mara; Pattabiraman, Divya; Libuda, Diana E; Ramadugu, Ajit; Tam, Angela; Martinez-Perez, Enrique; Roelens, Baptiste; Zawadzki, Karl A; Yokoo, Rayka; Rosu, Simona; Severson, Aaron F; Meyer, Barbara J; Nabeshima, Kentaro; Villeneuve, Anne M

    2014-09-01

    Meiotic recombination is initiated by the programmed induction of double-strand DNA breaks (DSBs), lesions that pose a potential threat to the genome. A subset of the DSBs induced during meiotic prophase become designated to be repaired by a pathway that specifically yields interhomolog crossovers (COs), which mature into chiasmata that temporarily connect the homologs to ensure their proper segregation at meiosis I. The remaining DSBs must be repaired by other mechanisms to restore genomic integrity prior to the meiotic divisions. Here we show that HIM-6, the Caenorhabditis elegans ortholog of the RecQ family DNA helicase BLM, functions in both of these processes. We show that him-6 mutants are competent to load the MutSγ complex at multiple potential CO sites, to generate intermediates that fulfill the requirements of monitoring mechanisms that enable meiotic progression, and to accomplish and robustly regulate CO designation. However, recombination events at a subset of CO-designated sites fail to mature into COs and chiasmata, indicating a pro-CO role for HIM-6/BLM that manifests itself late in the CO pathway. Moreover, we find that in addition to promoting COs, HIM-6 plays a role in eliminating and/or preventing the formation of persistent MutSγ-independent associations between homologous chromosomes. We propose that HIM-6/BLM enforces biased outcomes of recombination events to ensure that both (a) CO-designated recombination intermediates are reliably resolved as COs and (b) other recombination intermediates reliably mature into noncrossovers in a timely manner. Copyright © 2014 by the Genetics Society of America.

  4. The curvature calculation mechanism based on simple cell model.

    Yu, Haiyang; Fan, Xingyu; Song, Aiqi

    2017-07-20

    A conclusion has not yet been reached on how exactly the human visual system detects curvature. This paper demonstrates how orientation-selective simple cells can be used to construct curvature-detecting neural units. Through fixed arrangements, multiple plurality cells were constructed to simulate curvature cells with a proportional output to their curvature. In addition, this paper offers a solution to the problem of narrow detection range under fixed resolution by selecting an output value under multiple resolution. Curvature cells can be treated as concrete models of an end-stopped mechanism, and they can be used to further understand "curvature-selective" characteristics and to explain basic psychophysical findings and perceptual phenomena in current studies.

  5. A microcontroller based tuning mechanism for the magnetron

    Khan, A.M.; Mahfooz, M.; Hanumaiah, B.; Ganesh; Siddappa, K.

    2006-01-01

    In this paper we report on a control system developed to tune the magnetron frequency to get the maximum beam pulse in the microtron (electron accelerator facility at Mangalore University). The control system so designed consists of a microcontroller, a phase locked loop (PLL) and a digital to analog converter (DAC) to control the magnetron frequency. The voltage value given by the microcontroller through the DAC decides the reference frequency. The PLL gives the error voltage whenever there is difference between the reference and the magnetron frequencies. The microcontroller unit tracks the error voltage and tunes the magnetron with the help of a tuner mechanism connected through a stepper motor. The microcontroller also monitors the beam current level and accordingly adjusts the reference frequency to successfully tune the magnetron. (author)

  6. Mechanisms of Action of Probiotics based on Bacillus subtilis

    A.V. Savustyanenko

    2016-04-01

    Full Text Available The bacterium B.subtilis is one of the most promi­sing probiotics studied in recent decades. Mechanisms of its probiotic action are associated with the synthesis of antimicrobial agents, increasing of non-specific and specific immunity, stimulation of growth of normal microflora of the intestine and the releasing of digestive enzymes. B.subtilis releases ribosomally synthesized peptides, non-ribosomally synthesized peptides and non-peptide substances with a broad spectrum of antimicrobial activity covering Gram-positive, Gram-negative bacteria, viruses and fungi. Resistance to these antimicrobial agents is rare. Enhancement of non-specific immunity is associated with macrophage activation and the release of pro-inflammatory cytokines from them, increasing of barrier function of the intestinal mucosa, releasing of vitamins and amino acids (including essential ones. Enhancement of specific immunity manifests by activation of T- and B-lymphocytes and the release from the latter of immunoglobulins — IgG and IgA. B.subtilis stimulates the growth of normal intestinal flora, in particular, bacteria of the genus Lactobacillus and Bifidobacterium. Furthermore, probiotic increases the diversity of intestinal microflora. Probiotic secretes all major digestive enzymes to the intestinal lumen: amylases, lipases, proteases, pectinases and cellulases. In addition to digestion, these enzymes destroy antinutritional factors and allergenic substances contained in the food. These mechanisms of action make reasonable the use of B.subtilis in the combination therapy to treat intestinal infections; prevention of respiratory infections during the cold season; prevention of antibiotic-associated diarrhea; for the correction of food digestion and movement impairments of various origin (errors in the diet, changes in the diet, diseases of the gastrointestinal tract, disorders of the autonomic nervous system, etc.. B.subtilis does not usually cause side effects. This

  7. Beyond reverse pharmacology: Mechanism-based screening of Ayurvedic drugs

    R D Lele

    2010-01-01

    Full Text Available This paper reviews the pharmacology of Indian medicinal plants, starting with the historical background of European work on the subject beginning as early as the 17th century, and tracing its history through the work of Sen and Bose in the 1930′s, and Vakhil′s historic 1949 paper on Sarpaghanda. The often crucial role of patient feedback in early discoveries is highlighted, as is the time lag between proof of pharmacological action and identification of the active principle, and subsequent elucidation of mechanism of action. In the case of Indian plants in the 20th century this process sometimes took almost 50 years. Reserpine and its mechanisms are given in detail, and its current relevance to public health discussed. The foundation of present day methods of pharmacology is briefly presented so the complexity of methods used to identify properties of Ayurveda derived drugs like forskolin and baicalein, and their bioavailability, may be better appreciated. Ayurveda derived anti-oxidants and their levels of action, immuno-modulators, particularly with respect to the NF-kB pathway and its implications for cancer control, are all considered. The example of curcumin derived from turmeric is explained in more detail, because of its role in cancer prevention. Finally, the paper emphasizes the importance of Ayurveda′s concepts of rasayana as a form of dietary chemo-prevention; the significance of ahar, diet, in Ayurveda′s aspiration to prevent disease and restore health thus becomes clear. Understood in this light, Ayurveda may transcend pharmacology as a treatment paradigm.

  8. Toxicogenomics-based identification of mechanisms for direct immunotoxicity.

    Shao, Jia; Katika, Madhumohan R; Schmeits, Peter C J; Hendriksen, Peter J M; van Loveren, Henk; Peijnenburg, Ad A C M; Volger, Oscar L

    2013-10-01

    Compounds with direct immunotoxic properties, including metals, mycotoxins, agricultural pesticides, and industrial chemicals, form potential human health risks due to exposure through food, drinking water, and the environment. Insights into the mechanisms of action are currently lacking for the majority of these direct immunotoxicants. Therefore, the present work aimed to gain insights into the molecular mechanisms underlying direct immunotoxicity. To this end, we assessed in vitro the effects of 31 test compounds on the transcriptome of the human Jurkat T-cell line. These compounds included direct immunotoxicants, immunosuppressive drugs with different mode of actions, and nonimmunotoxic control chemicals. Pathway analysis of the microarray data allowed us to identify canonical pathways and Gene Ontology processes that were transcriptionally regulated in common by immunotoxicants (1) with structural similarities, such as tributyltin chloride and tributyltin oxide that activated the retinoic acid/X receptor signaling pathway and (2) without structural similarities, such as As2O3, dibutyltin chloride, diazinon, MeHg, ochratoxin A (OTA), S9-treated OTA, S9-treated cyclophosphamide, and S9-treated benzo[a]pyrene, which activated unfolded protein response, and FTY720, lindane, and propanil, which activated the cholesterol biosynthesis pathway. In addition, processes uniquely affected by individual immunotoxicants were identified, such as the induction of Notch receptor signaling and the downregulation of acute-phase response genes by OTA. These findings were validated by quantitative real-time PCR analysis of genes involved in these processes. Our study indicated that diverse modes of action are involved in direct immunotoxicity and that a set of pathways or genes, rather than one single gene, can be used to screen compounds for direct immunotoxicity.

  9. Base excision repair mechanisms and relevance to cancer susceptibility

    Dogliotti, E.; Wilson, S.H.

    2009-01-01

    The base excision repair (BER) pathway is considered the predominant DNA repair system in mammalian cells for eliminating small DNA lesions generated at DNA bases either exogenously by environmental agents or endogenously by normal cellular metabolic processes (e.g. production of oxyradical species, alkylating agents, etc). The main goal of this project is the understanding of the involvement of BER in genome stability and in particular in sporadic cancer development associated with inflammation such as gastric cancer (GC). A major risk factor of GC is the infection by Helicobacter pylori, which causes oxidative stress. Oxidative DNA damage is mainly repaired by BER

  10. Mechanism of cadmium ion removal by base treated juniper fiber

    Soo-Hong Min; J.K. Park; James S. Han; Eun Woo Shin

    2003-01-01

    Pinyon juniper, Juniperus Monosperma, is a small-diameter and underutilized (SDU) lignocellulosic material. Evaluated were efficacy of base-treated juniper fiber (BTJF) sample for cadmium (Cd 2+ ) sorption and the viability of juniper fiber as a sorbent for the removal of Cd 2+ from water. Fourier Transform Infrared Spectroscopy (FTIR) analysis indicated that...

  11. Mechanical properties of wood-based composite materials

    Zhiyong Cai; Robert J. Ross

    2010-01-01

    The term composite is used to describe any wood material bonded together with adhesives. The current product mix ranges from fiberboard to laminated beams and components. In this chapter, wood-based composite materials are classified into the following categories: panel products (plywood, oriented strandboard (OSB), particleboard, fiberboard, medium-density fiberboard...

  12. A screw-based dynamic balancing approach, applied to a 5-bar mechanism

    de Jong, Jan Johannes; van Dijk, Johannes; Herder, Justus Laurens; Lenarcic, Jadran; Merlet, Jean-Pierre

    2016-01-01

    Dynamic balancing aims to reduce or eliminate the shaking base reaction forces and moments of mechanisms, in order to minimize vibration and wear. The derivation of the dynamic balance conditions requires significant algebraic effort, even for simple mechanisms. In this study, a screw-based

  13. Antitumor Mechanisms of Curcumae Rhizoma Based on Network Pharmacology

    Yan-Hua Bi

    2018-01-01

    Full Text Available Curcumae Rhizoma, a traditional Chinese medication, is commonly used in both traditional treatment and modern clinical care. Its anticancer effects have attracted a great deal of attention, but the mechanisms of action remain obscure. In this study, we screened for the active compounds of Curcumae Rhizoma using a drug-likeness approach. Candidate protein targets with functions related to cancer were predicted by reverse docking and then checked by manual search of the PubMed database. Potential target genes were uploaded to the GeneMANIA server and DAVID 6.8 database for analysis. Finally, compound-target, target-pathway, and compound-target-pathway networks were constructed using Cytoscape 3.3. The results revealed that the anticancer activity of Curcumae Rhizoma potentially involves 13 active compounds, 33 potential targets, and 31 signaling pathways, thus constituting a “multiple compounds, multiple targets, and multiple pathways” network corresponding to the concept of systematic actions in TCM. These findings provide an overview of the anticancer action of Curcumae Rhizoma from a network perspective, as well as setting an example for future studies of other materials used in TCM.

  14. VISCOSE BASED MAGNETIC YARNS – PHYSICAL AND MECHANICAL CHARACTERIZATION

    GROSU Marian-Cătălin

    2017-05-01

    Full Text Available In the context of the rapid growth in the number of electrical and electronic devices and accessories that emit electromagnetic energy in different frequency bands we present and characterize here several magnetic functionalized viscose twisted yarns. A 100% viscose twisted staple yarn was covered through an in-house developed process with a polymeric solution containing micrometric sized barium hexaferrite magnetic powder. The in-house developed process allows deposition of micrometric thickness polymeric paste layer on the yarn surface. Barium hexaferrite is a hard magnetic material exhibiting high chemical stability and corrosion resistivity, relatively large saturation and residual magnetization and microwave absorbing properties. Five different percentages of the magnetic powder in the polymer solution were used, i.e. ranging from 15 wt% to 45 wt%. Physical characterization shows a very good adherence between the highly hygroscopic viscose staple fibers and the polymeric solution that contains polyvinyl acetate and polyurethane as binders. SEM images evidenced the fact that the polymeric solution penetrated more than 1/3 of the yarn diameter. The concentration of magnetic powder in the polymeric solution has a direct influence on the coating amount, diameter and density. The mechanical characterization of the coated yarns revealed that the breaking force is increasing with increasing magnetic powder content up to o certain value and then decreased because the magnetic layer became stiffer. At the same time, the elongation at brake is decreasing.

  15. A damage mechanics based approach to structural deterioration and reliability

    Bhattcharya, B.; Ellingwood, B. [Johns Hopkins Univ., Baltimore, MD (United States). Dept. of Civil Engineering

    1998-02-01

    Structural deterioration often occurs without perceptible manifestation. Continuum damage mechanics defines structural damage in terms of the material microstructure, and relates the damage variable to the macroscopic strength or stiffness of the structure. This enables one to predict the state of damage prior to the initiation of a macroscopic flaw, and allows one to estimate residual strength/service life of an existing structure. The accumulation of damage is a dissipative process that is governed by the laws of thermodynamics. Partial differential equations for damage growth in terms of the Helmholtz free energy are derived from fundamental thermodynamical conditions. Closed-form solutions to the equations are obtained under uniaxial loading for ductile deformation damage as a function of plastic strain, for creep damage as a function of time, and for fatigue damage as function of number of cycles. The proposed damage growth model is extended into the stochastic domain by considering fluctuations in the free energy, and closed-form solutions of the resulting stochastic differential equation are obtained in each of the three cases mentioned above. A reliability analysis of a ring-stiffened cylindrical steel shell subjected to corrosion, accidental pressure, and temperature is performed.

  16. Silicon-based tracking system: Mechanical engineering and design

    Miller, W.O.; Gamble, M.T.; Thompson, T.C.; Woloshun, K.A.; Reid, R.S.; Hanlon, J.A.; Michaud, F.D.; Dransfield, G.D.; Ziock, H.J.; Palounek, A.P.

    1992-01-01

    The Silicon Tracking System (STS) is composed of silicon strip detectors arranged by both in a cylindrical array and an array of flat panels about the interaction region. The cylindrical array is denoted the central region and the flat panel arrays, which are normal to the beam axis, we denoted the forward regions. The overall length of the silicon array is 5.16 m and the maximum diameter is 0.93 m. The Silicon Tracking System Conceptual Design Report, should be consulted for the body of analysis performed to quantify the present design concept. For the STS to achieve its physics goals, the mechanical structures and services must support 17 m 2 of silicon detectors and stabilize their positions to within 5 μm, uniformly cool the detector the system to O degrees C and at the same time potentially remove up to 13 kW of waste heat generated by the detector electronics, provide up to 3400 A of current to supply the 6.5 million electronics channels, and supply of control and data transmission lines for those channels. These objectives must be achieved in a high ionizing radiation environment, using virtually no structural mass and only low-Z materials. The system must be maintainable during its 10 year operating life

  17. BlenX-based compositional modeling of complex reaction mechanisms

    Judit Zámborszky

    2010-02-01

    Full Text Available Molecular interactions are wired in a fascinating way resulting in complex behavior of biological systems. Theoretical modeling provides a useful framework for understanding the dynamics and the function of such networks. The complexity of the biological networks calls for conceptual tools that manage the combinatorial explosion of the set of possible interactions. A suitable conceptual tool to attack complexity is compositionality, already successfully used in the process algebra field to model computer systems. We rely on the BlenX programming language, originated by the beta-binders process calculus, to specify and simulate high-level descriptions of biological circuits. The Gillespie's stochastic framework of BlenX requires the decomposition of phenomenological functions into basic elementary reactions. Systematic unpacking of complex reaction mechanisms into BlenX templates is shown in this study. The estimation/derivation of missing parameters and the challenges emerging from compositional model building in stochastic process algebras are discussed. A biological example on circadian clock is presented as a case study of BlenX compositionality.

  18. Mechanism based evaluation of materials behavior and reference curves

    Toerroenen, K.; Saario, T.; Wallin, K.; Forsten, J.

    1984-01-01

    The safety assessment of nuclear pressure vessels and piping requires a quantitative estimation of defect growth by stable and unstable manner during service. This estimation is essential for determining whether the defect detected during inspection should be repaired or whether the size of the defect even after its expected growth is small enough to leave the integrity of the vessel unaffected. The most important stable defect growth mechanism is that of environmentally assisted cyclic crack growth. Recent results indicate that it is markedly affected by sulfur content and/or manganese sulfide morphology and distribution. This implies that an essential improvement in component safety has been gained by currently applied steelmaking practices, which result in extra low sulfur content, generally below 0.01 wt%, and in round shape and small size of inclusions, through, e.g., calcium treatment, hence considerably reducing the effect of environment on crack growth rate. This further implies that the ASME Section XI reference curves for environmentally accelerated cyclic crack growth are conservative for steels produced by current steelmaking practices. (orig./WL)

  19. A damage mechanics based approach to structural deterioration and reliability

    Bhattcharya, B.; Ellingwood, B.

    1998-02-01

    Structural deterioration often occurs without perceptible manifestation. Continuum damage mechanics defines structural damage in terms of the material microstructure, and relates the damage variable to the macroscopic strength or stiffness of the structure. This enables one to predict the state of damage prior to the initiation of a macroscopic flaw, and allows one to estimate residual strength/service life of an existing structure. The accumulation of damage is a dissipative process that is governed by the laws of thermodynamics. Partial differential equations for damage growth in terms of the Helmholtz free energy are derived from fundamental thermodynamical conditions. Closed-form solutions to the equations are obtained under uniaxial loading for ductile deformation damage as a function of plastic strain, for creep damage as a function of time, and for fatigue damage as function of number of cycles. The proposed damage growth model is extended into the stochastic domain by considering fluctuations in the free energy, and closed-form solutions of the resulting stochastic differential equation are obtained in each of the three cases mentioned above. A reliability analysis of a ring-stiffened cylindrical steel shell subjected to corrosion, accidental pressure, and temperature is performed

  20. Mechanical and Electrochemical Performance of Graphene-Based Flexible Supercapacitors

    2014-08-01

    pulling apart as the supercapacitor is bent. The degradation of ionic liquid based supercapacitors due to water permeation into the package also needs to...electrodes are assembled with an intervening frame of FEP-coated Kapton, c) electrodes are heat sealed face to face on three sides, d) the polypropylene ... liquid in thick Kapton/FEP, c) propylene carbonate in thick Kapton, d) water in thick Kapton covered with aluminum tape, e) water in thick Kapton/FEP

  1. Reliability-based sensitivity of mechanical components with arbitrary distribution parameters

    Zhang, Yi Min; Yang, Zhou; Wen, Bang Chun; He, Xiang Dong; Liu, Qiaoling

    2010-01-01

    This paper presents a reliability-based sensitivity method for mechanical components with arbitrary distribution parameters. Techniques from the perturbation method, the Edgeworth series, the reliability-based design theory, and the sensitivity analysis approach were employed directly to calculate the reliability-based sensitivity of mechanical components on the condition that the first four moments of the original random variables are known. The reliability-based sensitivity information of the mechanical components can be accurately and quickly obtained using a practical computer program. The effects of the design parameters on the reliability of mechanical components were studied. The method presented in this paper provides the theoretic basis for the reliability-based design of mechanical components

  2. Hot isostatic pressing of single-crystal nickel-base superalloys: Mechanism of pore closure and effect on Mechanical properties

    Epishin Alexander I.

    2014-01-01

    Full Text Available Pore annihilation was investigated in the single-crystal nickel-base superalloy CMSX-4. HIP tests at 1288 °C/103 MPa were interrupted at different times, then the specimens were investigated by TEM, metallography and density measurements. The kinetics of pore annihilation was determined. The pore closure mechanism was identified as plastic deformation on the octahedral slip systems. A model describing the kinetics of pore closure has been developed on the base of crystal plasticity and large strain theory. Mechanical tests with the superalloy CMSX-4 and the Ru-containing superalloy VGM4 showed, that HIP significantly increases the fatigue life at low temperatures but has no effect on creep strength.

  3. Physical-Mechanisms Based Reliability Analysis For Emerging Technologies

    2017-05-05

    irradiation effects on deep level states in Mg-doped p-type GaN grown by ammonia-based molecular beam epitaxy," Appl. Phys. Lett., vol. 106, article...Chan, A. Raman, and M. Turowski, "State and angular dependence of single-event upsets in an asymmetric RC-hardened SRAM using deep trench capacitors...Trinczek, E. W. Blackmore, S. J. Wen , R. Wong, B. Narasimham, J. A. Pellish and H. Puchner, "The contribution of low-energy protons to the total

  4. A Cluster Based Group Signature Mechanism For Secure Vanet Communication

    Navjot Kaur

    2015-08-01

    Full Text Available Vehicular adhoc network is one of the recent area of research to administer safety to human lives controlling of messages and in disposal of messages to users and passengers. VANETs allows communication of moving vehicular nodes. Movement of nodes leads in changing network size and scenario. Whenever a new node joins the network there is a threat of malicious node attack. So we need an environment that is secure and trust worthy. Therefore a new cluster based secure technique is proposed where cluster head is responsible for providing communication between the vehicular nodes. Performance parameters used in this paper are message drop ratio packet delay ratio and verification time.

  5. Double-sided auction mechanism design in electricity based on maximizing social welfare

    Zou Xiaoyan

    2009-01-01

    An efficient electricity double-sided auction mechanism should control market power and enhance the social welfare of the electricity market. Based on this goal, the paper designs a new double-sided auction mechanism. In the new mechanism, the social welfare contribution of each participant plays a pivotal role, because this contribution is the critical factor in market clearing, payment settling, and transaction matching rules. In particular, each winner of the auction can gain transfer payments according to his contribution to social welfare in the electricity market, and this gives the mechanism the ability to control the market power of some participants. At the same time, this mechanism ensures that the market organizer balances his budget. We then conduct a theoretical and empirical analysis based on the Spanish electricity market. Both of the results show that compared to the uniform-pricing mechanism, the new mechanism can reduce market power of participants and enhance the social welfare of the electricity market.

  6. Development of a skeletal multi-component fuel reaction mechanism based on decoupling methodology

    Mohan, Balaji; Tay, Kun Lin; Yang, Wenming; Chua, Kian Jon

    2015-01-01

    Highlights: • A compact multi-component skeletal reaction mechanism was developed. • Combined bio-diesel and PRF mechanism was proposed. • The mechanism consists of 68 species and 183 reactions. • Well validated against ignition delay times, flame speed and engine results. - Abstract: A new coupled bio-diesel surrogate and primary reference fuel (PRF) oxidation skeletal mechanism has been developed. The bio-diesel surrogate sub-mechanism consists of oxidation sub-mechanisms of Methyl decanoate (MD), Methyl 9-decenoate (MD9D) and n-Heptane fuel components. The MD and MD9D are chosen to represent the saturated and unsaturated methyl esters respectively in bio-diesel fuels. Then, a reduced iso-Octane oxidation sub-mechanism is added to the bio-diesel surrogate sub-mechanism. Then, all the sub-mechanisms are integrated to a reduced C_2–C_3 mechanism, detailed H_2/CO/C_1 mechanism and reduced NO_x mechanism based on decoupling methodology. The final mechanism consisted of 68 species and 183 reactions. The mechanism was well validated with shock-tube ignition delay times, laminar flame speed and 3D engine simulations.

  7. An ATR-dependent function for the Ddx19 RNA helicase in nuclear R-loop metabolism.

    Hodroj, Dana; Recolin, Bénédicte; Serhal, Kamar; Martinez, Susan; Tsanov, Nikolay; Abou Merhi, Raghida; Maiorano, Domenico

    2017-05-02

    Coordination between transcription and replication is crucial in the maintenance of genome integrity. Disturbance of these processes leads to accumulation of aberrant DNA:RNA hybrids (R-loops) that, if unresolved, generate DNA damage and genomic instability. Here we report a novel, unexpected role for the nucleopore-associated mRNA export factor Ddx19 in removing nuclear R-loops formed upon replication stress or DNA damage. We show, in live cells, that Ddx19 transiently relocalizes from the nucleopore to the nucleus upon DNA damage, in an ATR/Chk1-dependent manner, and that Ddx19 nuclear relocalization is required to clear R-loops. Ddx19 depletion induces R-loop accumulation, proliferation-dependent DNA damage and defects in replication fork progression. Further, we show that Ddx19 resolves R-loops in vitro via its helicase activity. Furthermore, mutation of a residue phosphorylated by Chk1 in Ddx19 disrupts its interaction with Nup214 and allows its nuclear relocalization. Finally, we show that Ddx19 operates in resolving R-loops independently of the RNA helicase senataxin. Altogether these observations put forward a novel, ATR-dependent function for Ddx19 in R-loop metabolism to preserve genome integrity in mammalian cells. © 2017 The Authors.

  8. TbPIF5 is a Trypanosoma brucei mitochondrial DNA helicase involved in processing of minicircle Okazaki fragments.

    Beiyu Liu

    2009-09-01

    Full Text Available Trypanosoma brucei's mitochondrial genome, kinetoplast DNA (kDNA, is a giant network of catenated DNA rings. The network consists of a few thousand 1 kb minicircles and several dozen 23 kb maxicircles. Here we report that TbPIF5, one of T. brucei's six mitochondrial proteins related to Saccharomyces cerevisiae mitochondrial DNA helicase ScPIF1, is involved in minicircle lagging strand synthesis. Like its yeast homolog, TbPIF5 is a 5' to 3' DNA helicase. Together with other enzymes thought to be involved in Okazaki fragment processing, TbPIF5 localizes in vivo to the antipodal sites flanking the kDNA. Minicircles in wild type cells replicate unidirectionally as theta-structures and are unusual in that Okazaki fragments are not joined until after the progeny minicircles have segregated. We now report that overexpression of TbPIF5 causes premature removal of RNA primers and joining of Okazaki fragments on theta structures. Further elongation of the lagging strand is blocked, but the leading strand is completed and the minicircle progeny, one with a truncated H strand (ranging from 0.1 to 1 kb, are segregated. The minicircles with a truncated H strand electrophorese on an agarose gel as a smear. This replication defect is associated with kinetoplast shrinkage and eventual slowing of cell growth. We propose that TbPIF5 unwinds RNA primers after lagging strand synthesis, thus facilitating processing of Okazaki fragments.

  9. Insights into the Structure of Dimeric RNA Helicase CsdA and Indispensable Role of Its C-Terminal Regions.

    Xu, Ling; Wang, Lijun; Peng, Junhui; Li, Fudong; Wu, Lijie; Zhang, Beibei; Lv, Mengqi; Zhang, Jiahai; Gong, Qingguo; Zhang, Rongguang; Zuo, Xiaobing; Zhang, Zhiyong; Wu, Jihui; Tang, Yajun; Shi, Yunyu

    2017-12-05

    CsdA has been proposed to be essential for the biogenesis of ribosome and gene regulation after cold shock. However, the structure of CsdA and the function of its long C-terminal regions are still unclear. Here, we solved all of the domain structures of CsdA and found two previously uncharacterized auxiliary domains: a dimerization domain (DD) and an RNA-binding domain (RBD). Small-angle X-ray scattering experiments helped to track the conformational flexibilities of the helicase core domains and C-terminal regions. Biochemical assays revealed that DD is indispensable for stabilizing the CsdA dimeric structure. We also demonstrate for the first time that CsdA functions as a stable dimer at low temperature. The C-terminal regions are critical for RNA binding and efficient enzymatic activities. CsdA_RBD could specifically bind to the regions with a preference for single-stranded G-rich RNA, which may help to bring the helicase core to unwind the adjacent duplex. Copyright © 2017 Elsevier Ltd. All rights reserved.

  10. Cdc45 (cell division cycle protein 45) guards the gate of the Eukaryote Replisome helicase stabilizing leading strand engagement

    Petojevic, Tatjana; Pesavento, James J.; Costa, Alessandro; Liang, Jingdan; Wang, Zhijun; Berger, James M.; Botchan, Michael R.

    2015-01-01

    DNA replication licensing is now understood to be the pathway that leads to the assembly of double hexamers of minichromosome maintenance (Mcm2–7) at origin sites. Cell division control protein 45 (Cdc45) and GINS proteins activate the latent Mcm2–7 helicase by inducing allosteric changes through binding, forming a Cdc45/Mcm2-7/GINS (CMG) complex that is competent to unwind duplex DNA. The CMG has an active gate between subunits Mcm2 and Mcm5 that opens and closes in response to nucleotide binding. The consequences of inappropriate Mcm2/5 gate actuation and the role of a side channel formed between GINS/Cdc45 and the outer edge of the Mcm2–7 ring for unwinding have remained unexplored. Here we uncover a novel function for Cdc45. Cross-linking studies trace the path of the DNA with the CMG complex at a fork junction between duplex and single strands with the bound CMG in an open or closed gate conformation. In the closed state, the lagging strand does not pass through the side channel, but in the open state, the leading strand surprisingly interacts with Cdc45. Mutations in the recombination protein J fold of Cdc45 that ablate this interaction diminish helicase activity. These data indicate that Cdc45 serves as a shield to guard against occasional slippage of the leading strand from the core channel. PMID:25561522

  11. The N-terminal domain of human DNA helicase Rtel1 contains a redox active iron-sulfur cluster.

    Landry, Aaron P; Ding, Huangen

    2014-01-01

    Human telomere length regulator Rtel1 is a superfamily II DNA helicase and is essential for maintaining proper length of telomeres in chromosomes. Here we report that the N-terminal domain of human Rtel1 (RtelN) expressed in Escherichia coli cells produces a protein that contains a redox active iron-sulfur cluster with the redox midpoint potential of -248 ± 10 mV (pH 8.0). The iron-sulfur cluster in RtelN is sensitive to hydrogen peroxide and nitric oxide, indicating that reactive oxygen/nitrogen species may modulate the DNA helicase activity of Rtel1 via modification of its iron-sulfur cluster. Purified RtelN retains a weak binding affinity for the single-stranded (ss) and double-stranded (ds) DNA in vitro. However, modification of the iron-sulfur cluster by hydrogen peroxide or nitric oxide does not significantly affect the DNA binding activity of RtelN, suggesting that the iron-sulfur cluster is not directly involved in the DNA interaction in the N-terminal domain of Rtel1.

  12. The N-Terminal Domain of Human DNA Helicase Rtel1 Contains a Redox Active Iron-Sulfur Cluster

    Aaron P. Landry

    2014-01-01

    Full Text Available Human telomere length regulator Rtel1 is a superfamily II DNA helicase and is essential for maintaining proper length of telomeres in chromosomes. Here we report that the N-terminal domain of human Rtel1 (RtelN expressed in Escherichia coli cells produces a protein that contains a redox active iron-sulfur cluster with the redox midpoint potential of −248 ± 10 mV (pH 8.0. The iron-sulfur cluster in RtelN is sensitive to hydrogen peroxide and nitric oxide, indicating that reactive oxygen/nitrogen species may modulate the DNA helicase activity of Rtel1 via modification of its iron-sulfur cluster. Purified RtelN retains a weak binding affinity for the single-stranded (ss and double-stranded (ds DNA in vitro. However, modification of the iron-sulfur cluster by hydrogen peroxide or nitric oxide does not significantly affect the DNA binding activity of RtelN, suggesting that the iron-sulfur cluster is not directly involved in the DNA interaction in the N-terminal domain of Rtel1.

  13. Major depressive disorder: mechanism-based prescribing for personalized medicine

    Saltiel PF

    2015-03-01

    Full Text Available Philip F Saltiel,1 Daniel I Silvershein2 1Department of Psychiatry, New York University School of Medicine/Langone Medical Center New York University Behavioral Health Programs, New York University Pearl Barlow Center for Memory Evaluation and Treatment, New York, NY, USA; 2Department of Medicine, New York University School of Medicine/Langone Medical Center, New York, NY, USA Abstract: Individual patients with depression present with unique symptom clusters – before, during, and even after treatment. The prevalence of persistent, unresolved symptoms and their contribution to patient functioning and disease progression emphasize the importance of finding the right treatment choice at the onset and the utility of switching medications based on suboptimal responses. Our primary goal as clinicians is to improve patient function and quality of life. In fact, feelings of well-being and the return to premorbid levels of functioning are frequently rated by patients as being more important than symptom relief. However, functional improvements often lag behind resolution of mood, attributed in large part to persistent and functionally impairing symptoms – namely, fatigue, sleep/wake disturbance, and cognitive dysfunction. Thus, patient outcomes can be optimized by deconstructing each patient’s depressive profile to its component symptoms and specifically targeting those domains that differentially limit patient function. This article will provide an evidence-based framework within which clinicians may tailor pharmacotherapy to patient symptomatology for improved treatment outcomes. Keywords: MDD, tailored pharmacotherapy, patient-specific profile, individualized pharmacotherapy

  14. Mechanics

    Chester, W

    1979-01-01

    When I began to write this book, I originally had in mind the needs of university students in their first year. May aim was to keep the mathematics simple. No advanced techniques are used and there are no complicated applications. The emphasis is on an understanding of the basic ideas and problems which require expertise but do not contribute to this understanding are not discussed. How­ ever, the presentation is more sophisticated than might be considered appropri­ ate for someone with no previous knowledge of the subject so that, although it is developed from the beginning, some previous acquaintance with the elements of the subject would be an advantage. In addition, some familiarity with element­ ary calculus is assumed but not with the elementary theory of differential equations, although knowledge of the latter would again be an advantage. It is my opinion that mechanics is best introduced through the motion of a particle, with rigid body problems left until the subject is more fully developed. Howev...

  15. Personal Computer (PC) based image processing applied to fluid mechanics

    Cho, Y.-C.; Mclachlan, B. G.

    1987-01-01

    A PC based image processing system was employed to determine the instantaneous velocity field of a two-dimensional unsteady flow. The flow was visualized using a suspension of seeding particles in water, and a laser sheet for illumination. With a finite time exposure, the particle motion was captured on a photograph as a pattern of streaks. The streak pattern was digitized and processed using various imaging operations, including contrast manipulation, noise cleaning, filtering, statistical differencing, and thresholding. Information concerning the velocity was extracted from the enhanced image by measuring the length and orientation of the individual streaks. The fluid velocities deduced from the randomly distributed particle streaks were interpolated to obtain velocities at uniform grid points. For the interpolation a simple convolution technique with an adaptive Gaussian window was used. The results are compared with a numerical prediction by a Navier-Stokes computation.

  16. Market Mechanism Design for Renewable Energy based on Risk Theory

    Yang, Wu; Bo, Wang; Jichun, Liu; Wenjiao, Zai; Pingliang, Zeng; Haobo, Shi

    2018-02-01

    Generation trading between renewable energy and thermal power is an efficient market means for transforming supply structure of electric power into sustainable development pattern. But the trading is hampered by the output fluctuations of renewable energy and the cost differences between renewable energy and thermal power at present. In this paper, the external environmental cost (EEC) is defined and the EEC is introduced into the generation cost. At same time, the incentive functions of renewable energy and low-emission thermal power are designed, which are decreasing functions of EEC. On these bases, for the market risks caused by the random variability of EEC, the decision-making model of generation trading between renewable energy and thermal power is constructed according to the risk theory. The feasibility and effectiveness of the proposed model are verified by simulation results.

  17. From evidence based medicine to mechanism based medicine. Reviewing the role of pharmacogenetics.

    Wilffert, Bob; Swen, Jesse; Mulder, Hans; Touw, Daan; Maitland-Van der Zee, Anke-Hilse; Deneer, Vera

    2013-06-01

    The translation of evidence based medicine to a specific patient presents a considerable challenge. We present by means of the examples nortriptyline, tramadol, clopidogrel, coumarins, abacavir and antipsychotics the discrepancy between available pharmacogenetic information and its implementation in daily clinical practice. Literature review. A mechanism based approach may be helpful to personalize medicine for the individual patient to which pharmacogenetics may contribute significantly. The lack of consistency in what we accept in bioequivalence and in pharmacogenetics of drug metabolising enzymes is discussed and illustrated with the example of nortriptyline. The impact of pharmacogenetics on examples like tramadol, clopidogrel, coumarins and abacavir is described. Also the present status of the polymorphisms of 5-HT2A and C receptors in antipsychotic-induced weight gain is presented as a pharmacodynamic example with until now a greater distance to clinical implementation. The contribution of pharmacogenetics to tailor-made pharmacotherapy, which especially might be of value for patients deviating from the average, has not yet reached the position it seems to deserve.

  18. [Interactions of DNA bases with individual water molecules. Molecular mechanics and quantum mechanics computation results vs. experimental data].

    Gonzalez, E; Lino, J; Deriabina, A; Herrera, J N F; Poltev, V I

    2013-01-01

    To elucidate details of the DNA-water interactions we performed the calculations and systemaitic search for minima of interaction energy of the systems consisting of one of DNA bases and one or two water molecules. The results of calculations using two force fields of molecular mechanics (MM) and correlated ab initio method MP2/6-31G(d, p) of quantum mechanics (QM) have been compared with one another and with experimental data. The calculations demonstrated a qualitative agreement between geometry characteristics of the most of local energy minima obtained via different methods. The deepest minima revealed by MM and QM methods correspond to water molecule position between two neighbor hydrophilic centers of the base and to the formation by water molecule of hydrogen bonds with them. Nevertheless, the relative depth of some minima and peculiarities of mutual water-base positions in' these minima depend on the method used. The analysis revealed insignificance of some differences in the results of calculations performed via different methods and the importance of other ones for the description of DNA hydration. The calculations via MM methods enable us to reproduce quantitatively all the experimental data on the enthalpies of complex formation of single water molecule with the set of mono-, di-, and trimethylated bases, as well as on water molecule locations near base hydrophilic atoms in the crystals of DNA duplex fragments, while some of these data cannot be rationalized by QM calculations.

  19. Non-Fourier based thermal-mechanical tissue damage prediction for thermal ablation.

    Li, Xin; Zhong, Yongmin; Smith, Julian; Gu, Chengfan

    2017-01-02

    Prediction of tissue damage under thermal loads plays important role for thermal ablation planning. A new methodology is presented in this paper by combing non-Fourier bio-heat transfer, constitutive elastic mechanics as well as non-rigid motion of dynamics to predict and analyze thermal distribution, thermal-induced mechanical deformation and thermal-mechanical damage of soft tissues under thermal loads. Simulations and comparison analysis demonstrate that the proposed methodology based on the non-Fourier bio-heat transfer can account for the thermal-induced mechanical behaviors of soft tissues and predict tissue thermal damage more accurately than classical Fourier bio-heat transfer based model.

  20. Nuclear Export of Pre-Ribosomal Subunits Requires Dbp5, but Not as an RNA-Helicase as for mRNA Export.

    Neumann, Bettina; Wu, Haijia; Hackmann, Alexandra; Krebber, Heike

    2016-01-01

    The DEAD-box RNA-helicase Dbp5/Rat8 is known for its function in nuclear mRNA export, where it displaces the export receptor Mex67 from the mRNA at the cytoplasmic side of the nuclear pore complex (NPC). Here we show that Dbp5 is also required for the nuclear export of both pre-ribosomal subunits. Yeast temperature-sensitive dbp5 mutants accumulate both ribosomal particles in their nuclei. Furthermore, Dbp5 genetically and physically interacts with known ribosomal transport factors such as Nmd3. Similar to mRNA export we show that also for ribosomal transport Dbp5 is required at the cytoplasmic side of the NPC. However, unlike its role in mRNA export, Dbp5 does not seem to undergo its ATPase cycle for this function, as ATPase-deficient dbp5 mutants that selectively inhibit mRNA export do not affect ribosomal transport. Furthermore, mutants of GLE1, the ATPase stimulating factor of Dbp5, show no major ribosomal export defects. Consequently, while Dbp5 uses its ATPase cycle to displace the export receptor Mex67 from the translocated mRNAs, Mex67 remains bound to ribosomal subunits upon transit to the cytoplasm, where it is detectable on translating ribosomes. Therefore, we propose a model, in which Dbp5 supports ribosomal transport by capturing ribosomal subunits upon their cytoplasmic appearance at the NPC, possibly by binding export factors such as Mex67. Thus, our findings reveal that although different ribonucleoparticles, mRNAs and pre-ribosomal subunits, use shared export factors, they utilize different transport mechanisms.

  1. Polarizable Atomic Multipole-based Molecular Mechanics for Organic Molecules.

    Ren, Pengyu; Wu, Chuanjie; Ponder, Jay W

    2011-10-11

    An empirical potential based on permanent atomic multipoles and atomic induced dipoles is reported for alkanes, alcohols, amines, sulfides, aldehydes, carboxylic acids, amides, aromatics and other small organic molecules. Permanent atomic multipole moments through quadrupole moments have been derived from gas phase ab initio molecular orbital calculations. The van der Waals parameters are obtained by fitting to gas phase homodimer QM energies and structures, as well as experimental densities and heats of vaporization of neat liquids. As a validation, the hydrogen bonding energies and structures of gas phase heterodimers with water are evaluated using the resulting potential. For 32 homo- and heterodimers, the association energy agrees with ab initio results to within 0.4 kcal/mol. The RMS deviation of hydrogen bond distance from QM optimized geometry is less than 0.06 Å. In addition, liquid self-diffusion and static dielectric constants computed from molecular dynamics simulation are consistent with experimental values. The force field is also used to compute the solvation free energy of 27 compounds not included in the parameterization process, with a RMS error of 0.69 kcal/mol. The results obtained in this study suggest the AMOEBA force field performs well across different environments and phases. The key algorithms involved in the electrostatic model and a protocol for developing parameters are detailed to facilitate extension to additional molecular systems.

  2. Forming mechanism of Te-based conductive-bridge memories

    Mendes, M. Kazar; Martinez, E.; Marty, A.; Veillerot, M.; Yamashita, Y.; Gassilloud, R.; Bernard, M.; Renault, O.; Barrett, N.

    2018-02-01

    We investigated origins of the resistivity change during the forming of ZrTe/Al2O3 based conductive-bridge resistive random access memories. Non-destructive hard X-ray photoelectron spectroscopy was used to investigate redox processes with sufficient depth sensitivity. Results highlighted the reduction of alumina correlated to the oxidation of zirconium at the interface between the solid electrolyte and the active electrode. In addition the resistance switching caused a decrease of Zr-Te bonds and an increase of elemental Te showing an enrichment of tellurium at the ZrTe/Al2O3 interface. XPS depth profiling using argon clusters ion beam confirmed the oxygen diffusion towards the top electrode. A four-layer capacitor model showed an increase of both the ZrO2 and AlOx interfacial layers, confirming the redox process located at the ZrTe/Al2O3 interface. Oxygen vacancies created in the alumina help the filament formation by acting as preferential conductive paths. This study provides a first direct evidence of the physico-chemical phenomena involved in resistive switching of such devices.

  3. Mycobacterium smegmatis SftH exemplifies a distinctive clade of superfamily II DNA-dependent ATPases with 3' to 5' translocase and helicase activities.

    Yakovleva, Lyudmila; Shuman, Stewart

    2012-08-01

    Bacterial DNA helicases are nucleic acid-dependent NTPases that play important roles in DNA replication, recombination and repair. We are interested in the DNA helicases of Mycobacteria, a genus of the phylum Actinobacteria, which includes the human pathogen Mycobacterium tuberculosis and its avirulent relative Mycobacterium smegmatis. Here, we identify and characterize M. smegmatis SftH, a superfamily II helicase with a distinctive domain structure, comprising an N-terminal NTPase domain and a C-terminal DUF1998 domain (containing a putative tetracysteine metal-binding motif). We show that SftH is a monomeric DNA-dependent ATPase/dATPase that translocates 3' to 5' on single-stranded DNA and has 3' to 5' helicase activity. SftH homologs are found in bacteria representing 12 different phyla, being especially prevalent in Actinobacteria (including M. tuberculosis). SftH homologs are evident in more than 30 genera of Archaea. Among eukarya, SftH homologs are present in plants and fungi.

  4. A temperature-sensitive allele of a putative mRNA splicing helicase down-regulates many cell wall genes and causes radial swelling in Arabidopsis thaliana.

    Howles, Paul A; Gebbie, Leigh K; Collings, David A; Varsani, Arvind; Broad, Ronan C; Ohms, Stephen; Birch, Rosemary J; Cork, Ann H; Arioli, Tony; Williamson, Richard E

    2016-05-01

    The putative RNA helicase encoded by the Arabidopsis gene At1g32490 is a homolog of the yeast splicing RNA helicases Prp2 and Prp22. We isolated a temperature-sensitive allele (rsw12) of the gene in a screen for root radial swelling mutants. Plants containing this allele grown at the restrictive temperature showed weak radial swelling, were stunted with reduced root elongation, and contained reduced levels of cellulose. The role of the protein was further explored by microarray analysis. By using both fold change cutoffs and a weighted gene coexpression network analysis (WGCNA) to investigate coexpression of genes, we found that the radial swelling phenotype was not linked to genes usually associated with primary cell wall biosynthesis. Instead, the mutation has strong effects on expression of secondary cell wall related genes. Many genes potentially associated with secondary walls were present in the most significant WGCNA module, as were genes coding for arabinogalactans and proteins with GPI anchors. The proportion of up-regulated genes that possess introns in rsw12 was above that expected if splicing was unrelated to the activity of the RNA helicase, suggesting that the helicase does indeed play a role in splicing in Arabidopsis. The phenotype may be due to a change in the expression of one or more genes coding for cell wall proteins.

  5. The Smc5/6 complex regulates the yeast Mph1 helicase at RNA-DNA hybrid-mediated DNA damage

    Lafuente-Barquero, Juan; Luke-Glaser, Sarah; Graf, Marco

    2017-01-01

    of Fanconi anemia protein M (FANCM), is required for cell viability in the absence of RNase H enzymes. The integrity of the Mph1 helicase domain is crucial to prevent the accumulation of RNA-DNA hybrids and RNA-DNA hybrid-dependent DNA damage, as determined by Rad52 foci. Mph1 forms foci when RNA-DNA hybrids...

  6. Arabidopsis RecQsim, a plant-specific member of the RecQ helicase family, can suppress the MMS hypersensitivity of the yeast sgs1 mutant

    Bagherieh-Najjar, MB; de Vries, OMH; Kroon, JTM; Wright, EL; Elborough, KM; Hille, J; Dijkwel, PP

    The Arabidopsis genome contains seven genes that belong to the RecQ family of ATP-dependent DNA helicases. RecQ members in Saccharomyces cerevisiae (SGS1) and man (WRN, BLM and RecQL4) are involved in DNA recombination, repair and genome stability maintenance, but little is known about the function

  7. Unique Helicase Determinants in the Essential Conjugative TraI Factor from Salmonella enterica Serovar Typhimurium Plasmid pCU1

    McLaughlin, Krystle J.; Nash, Rebekah P.; Redinbo, Mathew R. (UNC)

    2014-06-16

    The widespread development of multidrug-resistant bacteria is a major health emergency. Conjugative DNA plasmids, which harbor a wide range of antibiotic resistance genes, also encode the protein factors necessary to orchestrate the propagation of plasmid DNA between bacterial cells through conjugative transfer. Successful conjugative DNA transfer depends on key catalytic components to nick one strand of the duplex DNA plasmid and separate the DNA strands while cell-to-cell transfer occurs. The TraI protein from the conjugative Salmonella plasmid pCU1 fulfills these key catalytic roles, as it contains both single-stranded DNA-nicking relaxase and ATP-dependent helicase domains within a single, 1,078-residue polypeptide. In this work, we unraveled the helicase determinants of Salmonella pCU1 TraI through DNA binding, ATPase, and DNA strand separation assays. TraI binds DNA substrates with high affinity in a manner influenced by nucleic acid length and the presence of a DNA hairpin structure adjacent to the nick site. TraI selectively hydrolyzes ATP, and mutations in conserved helicase motifs eliminate ATPase activity. Surprisingly, the absence of a relatively short (144-residue) domain at the extreme C terminus of the protein severely diminishes ATP-dependent strand separation. Collectively, these data define the helicase motifs of the conjugative factor TraI from Salmonella pCU1 and reveal a previously uncharacterized C-terminal functional domain that uncouples ATP hydrolysis from strand separation activity.

  8. DNA Sequences Proximal to Human Mitochondrial DNA Deletion Breakpoints Prevalent in Human Disease Form G-quadruplexes, a Class of DNA Structures Inefficiently Unwound by the Mitochondrial Replicative Twinkle Helicase*

    Bharti, Sanjay Kumar; Sommers, Joshua A.; Zhou, Jun; Kaplan, Daniel L.; Spelbrink, Johannes N.; Mergny, Jean-Louis; Brosh, Robert M.

    2014-01-01

    Mitochondrial DNA deletions are prominent in human genetic disorders, cancer, and aging. It is thought that stalling of the mitochondrial replication machinery during DNA synthesis is a prominent source of mitochondrial genome instability; however, the precise molecular determinants of defective mitochondrial replication are not well understood. In this work, we performed a computational analysis of the human mitochondrial genome using the “Pattern Finder” G-quadruplex (G4) predictor algorithm to assess whether G4-forming sequences reside in close proximity (within 20 base pairs) to known mitochondrial DNA deletion breakpoints. We then used this information to map G4P sequences with deletions characteristic of representative mitochondrial genetic disorders and also those identified in various cancers and aging. Circular dichroism and UV spectral analysis demonstrated that mitochondrial G-rich sequences near deletion breakpoints prevalent in human disease form G-quadruplex DNA structures. A biochemical analysis of purified recombinant human Twinkle protein (gene product of c10orf2) showed that the mitochondrial replicative helicase inefficiently unwinds well characterized intermolecular and intramolecular G-quadruplex DNA substrates, as well as a unimolecular G4 substrate derived from a mitochondrial sequence that nests a deletion breakpoint described in human renal cell carcinoma. Although G4 has been implicated in the initiation of mitochondrial DNA replication, our current findings suggest that mitochondrial G-quadruplexes are also likely to be a source of instability for the mitochondrial genome by perturbing the normal progression of the mitochondrial replication machinery, including DNA unwinding by Twinkle helicase. PMID:25193669

  9. Mechanical modeling for magnetorheological elastomer isolators based on constitutive equations and electromagnetic analysis

    Wang, Qi; Dong, Xufeng; Li, Luyu; Ou, Jinping

    2018-06-01

    As constitutive models are too complicated and existing mechanical models lack universality, these models are beyond satisfaction for magnetorheological elastomer (MRE) devices. In this article, a novel universal method is proposed to build concise mechanical models. Constitutive model and electromagnetic analysis were applied in this method to ensure universality, while a series of derivations and simplifications were carried out to obtain a concise formulation. To illustrate the proposed modeling method, a conical MRE isolator was introduced. Its basic mechanical equations were built based on equilibrium, deformation compatibility, constitutive equations and electromagnetic analysis. An iteration model and a highly efficient differential equation editor based model were then derived to solve the basic mechanical equations. The final simplified mechanical equations were obtained by re-fitting the simulations with a novel optimal algorithm. In the end, verification test of the isolator has proved the accuracy of the derived mechanical model and the modeling method.

  10. Effect of Al and Y2O3 on Mechanical Properties in Mechanically Alloyed Nanograin Ni-Based Alloys.

    Kim, Chung Seok; Kim, Il-Ho

    2015-08-01

    The effects of aluminum and Y2O3 on the mechanical properties in nano grain Ni-based alloys have been investigated. The test specimens are prepared by mechanical alloying at an Ar atmosphere. The addition of Y2O3 and Al may cause an increase in the tensile strength at room temperature, 400 °C and 600 °C. However, it was confirmed that the increase of tensile strength at room temperature and 400 °C was predominantly caused by addition of Y2O3, while that at 600 °C was mainly due to addition of Al. These results can be attributed to the dispersion strengthening of Y2O3, preventing the formation of Cr2O3 and the change of fracture mode at 600 °C by the addition of Al.

  11. A QM/MM–Based Computational Investigation on the Catalytic Mechanism of Saccharopine Reductase

    Almasi, Joel N.; Bushnell, Eric A.C.; Gauld, James W.

    2011-01-01

    Saccharopine reductase from Magnaporthe grisea, an NADPH-containing enzyme in the α-aminoadipate pathway, catalyses the formation of saccharopine, a precursor to L-lysine, from the substrates glutamate and α-aminoadipate-δ-semialdehyde. Its catalytic mechanism has been investigated using quantum mechanics/molecular mechanics (QM/MM) ONIOM-based approaches. In particular, the overall catalytic pathway has been elucidated and the effects of electron correlation and the anisotropic polar protein...

  12. Silicon based mechanic-photonic wavelength converter for infrared photo-detection

    Rudnitsky, Arkady; Agdarov, Sergey; Gulitsky, Konstantin; Zalevsky, Zeev

    2017-06-01

    In this paper we present a new concept to realize a mechanic-photonic wavelength converter in silicon chip by construction of nanorods and by modulating the input illumination at temporal frequency matched to the mechanic resonance of the nanorods. The use case is to realize an infrared photo detector in silicon which is not based on absorption but rather on the mechanical interaction of the nanorods with the incoming illumination.

  13. Market-Based Coordination and Auditing Mechanisms for Self-Interested Multi-Robot Systems

    Ham, MyungJoo

    2009-01-01

    We propose market-based coordinated task allocation mechanisms, which allocate complex tasks that require synchronized and collaborated services of multiple robot agents to robot agents, and an auditing mechanism, which ensures proper behaviors of robot agents by verifying inter-agent activities, for self-interested, fully-distributed, and…

  14. Resonance-Based Sparse Signal Decomposition and its Application in Mechanical Fault Diagnosis: A Review.

    Huang, Wentao; Sun, Hongjian; Wang, Weijie

    2017-06-03

    Mechanical equipment is the heart of industry. For this reason, mechanical fault diagnosis has drawn considerable attention. In terms of the rich information hidden in fault vibration signals, the processing and analysis techniques of vibration signals have become a crucial research issue in the field of mechanical fault diagnosis. Based on the theory of sparse decomposition, Selesnick proposed a novel nonlinear signal processing method: resonance-based sparse signal decomposition (RSSD). Since being put forward, RSSD has become widely recognized, and many RSSD-based methods have been developed to guide mechanical fault diagnosis. This paper attempts to summarize and review the theoretical developments and application advances of RSSD in mechanical fault diagnosis, and to provide a more comprehensive reference for those interested in RSSD and mechanical fault diagnosis. Followed by a brief introduction of RSSD's theoretical foundation, based on different optimization directions, applications of RSSD in mechanical fault diagnosis are categorized into five aspects: original RSSD, parameter optimized RSSD, subband optimized RSSD, integrated optimized RSSD, and RSSD combined with other methods. On this basis, outstanding issues in current RSSD study are also pointed out, as well as corresponding instructional solutions. We hope this review will provide an insightful reference for researchers and readers who are interested in RSSD and mechanical fault diagnosis.

  15. KINEMATICS OF A MECHANISM WITH STOPPINGS, BASED ON A HYPOCYCLOID WITH 6 BRANCHES

    Liliana LUCA

    2010-10-01

    Full Text Available They are studied the positions, velocities and accelerations of a mechanism based on planetary gear and a RRP dyad. The mechanism ensures the plunger stop on a certain subinterval of the cycle, although the leading element is rotating. Because of the approximation of the hypocycloid shortened arc with six branches, by a circle arc, errors occur.

  16. A constitutive model of nanocrystalline metals based on competing grain boundary and grain interior deformation mechanisms

    Gurses, Ercan; El Sayed, Tamer S.

    2011-01-01

    In this work, a viscoplastic constitutive model for nanocrystalline metals is presented. The model is based on competing grain boundary and grain interior deformation mechanisms. In particular, inelastic deformations caused by grain boundary

  17. Nonlinear Lyapunov-based boundary control of distributed heat transfer mechanisms in membrane distillation plant

    Eleiwi, Fadi; Laleg-Kirati, Taous-Meriem

    2015-01-01

    This paper presents a nonlinear Lyapunov-based boundary control for the temperature difference of a membrane distillation boundary layers. The heat transfer mechanisms inside the process are modeled with a 2D advection-diffusion equation. The model

  18. Research on structures, mechanical properties, and mechanical responses of TKX-50 and TKX-50 based PBX with molecular dynamics.

    Ma, Song; Li, Yajin; Li, Yang; Luo, Yunjun

    2016-02-01

    To improve the practicality and safety of a novel explosive dihydroxylamm onium 5,5'-bis (tetrazole)-1,1'-diolate (TKX-50), polyvinylidene difluoride (PVDF) and polychlorotrifluoroe-thylene (PCTFE) were respectively added to the TKX-50, forming the polymer-bonded explosives (PBX). Interfacial and mechanical properties of PBX were investigated through molecular dynamics (MD) method, desensitizing mechanisms of fluorine-polymers for TKX-50 were researched by compression and bulk shear simulations. Results show that the binding energies (E bind ) between polymers (PVDF or PCTFE) and TKX-50 surfaces all rank in order of (011) > (100) > (010), shorter interatomic distance and the resulted higher potentials lead to higher E bind on TKX-50/PVDF interfaces than that on PCTFE/TKX-50 interfaces. Compared with TKX-50, the ductility of PBX is improved due to the isotropic mechanical property and flexibility of fluorine-polymers especially the PCTFE. Desensitizing effect of fluorine-polymers for TKX-50 is found under loading condition, which is attributed to the enhanced compressibility and buffer capacity against external pressure in compression, as well as the improved lubricity to reduce the sliding potentials in bulk shear process. Graphical Abstract Comparisons of the internal stress and slide potentials of the novel explosive,TKX-50 and its based PBX. Desensitizing effects can be found by the adding of fluorine-polymers, it owes to their better flexibility and lubricity as well as the amorphous nature.

  19. Design of the incentive mechanism in electricity auction market based on the signaling game theory

    Liu, Zhen; Zhang, Xiliang; Lieu, Jenny

    2010-01-01

    At present, designing a proper bidding mechanism to decrease the generators' market power is considered to be one of the key approaches to deepen the reform of the electricity market. Based on the signaling game theory, the paper analyzes the main electricity bidding mechanisms in the electricity auction markets and considers the degree of information disturbance as an important factor for evaluating bidding mechanisms. Under the above studies, an incentive electricity bidding mechanism defined as the Generator Semi-randomized Matching (GSM) mechanism is proposed. In order to verify the new bidding mechanism, this paper uses the Swarm platform to develop a simulation model based on the multi-agents. In the simulation model, the generators and purchasers use the partly superior study strategy to adjust their price and their electricity quantity. Then, the paper examines a simulation experiment of the GSM bidding mechanism and compares it to a simulation of the High-Low Matching (HLM) bidding mechanism. According to the simulation results, several conclusions can be drawn when comparing the proposed GSM bidding mechanism to the equilibrium state of HLM: the clearing price decreases, the total transaction volume increases, the profits of electricity generators decreases, and the overall benefits of purchasers increases. Index Terms - signaling game; semi-randomized matching; high-low match. (author)

  20. Interfacial mechanisms of heterogeneous Fenton reactions catalyzed by iron-based materials: A review.

    He, Jie; Yang, Xiaofang; Men, Bin; Wang, Dongsheng

    2016-01-01

    The heterogeneous Fenton reaction can generate highly reactive hydroxyl radicals (OH) from reactions between recyclable solid catalysts and H2O2 at acidic or even circumneutral pH. Hence, it can effectively oxidize refractory organics in water or soils and has become a promising environmentally friendly treatment technology. Due to the complex reaction system, the mechanism behind heterogeneous Fenton reactions remains unresolved but fascinating, and is crucial for understanding Fenton chemistry and the development and application of efficient heterogeneous Fenton technologies. Iron-based materials usually possess high catalytic activity, low cost, negligible toxicity and easy recovery, and are a superior type of heterogeneous Fenton catalysts. Therefore, this article reviews the fundamental but important interfacial mechanisms of heterogeneous Fenton reactions catalyzed by iron-based materials. OH, hydroperoxyl radicals/superoxide anions (HO2/O2(-)) and high-valent iron are the three main types of reactive oxygen species (ROS), with different oxidation reactivity and selectivity. Based on the mechanisms of ROS generation, the interfacial mechanisms of heterogeneous Fenton systems can be classified as the homogeneous Fenton mechanism induced by surface-leached iron, the heterogeneous catalysis mechanism, and the heterogeneous reaction-induced homogeneous mechanism. Different heterogeneous Fenton systems catalyzed by characteristic iron-based materials are comprehensively reviewed. Finally, related future research directions are also suggested. Copyright © 2015. Published by Elsevier B.V.

  1. HTLV-1 Tax plugs and freezes UPF1 helicase leading to nonsense-mediated mRNA decay inhibition.

    Fiorini, Francesca; Robin, Jean-Philippe; Kanaan, Joanne; Borowiak, Malgorzata; Croquette, Vincent; Le Hir, Hervé; Jalinot, Pierre; Mocquet, Vincent

    2018-01-30

    Up-Frameshift Suppressor 1 Homolog (UPF1) is a key factor for nonsense-mediated mRNA decay (NMD), a cellular process that can actively degrade mRNAs. Here, we study NMD inhibition during infection by human T-cell lymphotropic virus type I (HTLV-1) and characterise the influence of the retroviral Tax factor on UPF1 activity. Tax interacts with the central helicase core domain of UPF1 and might plug the RNA channel of UPF1, reducing its affinity for nucleic acids. Furthermore, using a single-molecule approach, we show that the sequential interaction of Tax with a RNA-bound UPF1 freezes UPF1: this latter is less sensitive to the presence of ATP and shows translocation defects, highlighting the importance of this feature for NMD. These mechanistic insights reveal how HTLV-1 hijacks the central component of NMD to ensure expression of its own genome.

  2. RECQ HELICASE RECQL4 PARTICIPATES IN NON-HOMOLOGOUS END JOINING AND INTERACTS WITH THE KU COMPLEX

    Shamanna, Raghavendra A; Singh, Dharmendra Kumar; Lu, Huiming

    2014-01-01

    -irradiation and resulted in accumulation of 53BP1 foci after irradiation, indicating defects in the processing of DSB. We find that RECQL4 interacts with the Ku70/Ku80 heterodimer, part of the DNA-dependent protein kinase (DNA-PK) complex, via its N-terminal domain. Further, RECQL4 stimulates higher order DNA binding...... of Ku70/Ku80 to a blunt end DNA substrate. Taken together, these results implicate that RECQL4 participates in the NHEJ pathway of DSB repair via a functional interaction with the Ku70/Ku80 complex. This is the first study to provide both in vitro and in vivo evidence for a role of a RecQ helicase...

  3. Association between regulator of telomere elongation helicase1 (RTEL1) gene and HAPE risk: A case-control study.

    Rong, Hao; He, Xue; Zhu, Linhao; Zhu, Xikai; Kang, Longli; Wang, Li; He, Yongjun; Yuan, Dongya; Jin, Tianbo

    2017-09-01

    High altitude pulmonary edema (HAPE) is a paradigm of pulmonary edema. Mutations in regulator of telomere elongation helicase1 (RTEL1) represent an important contributor to risk for pulmonary fibrosis. However, little information is found about the association between RTEL1 and HAPE risk. The present study was undertaken to tentatively explore the potential relation between single-nucleotide polymorphisms (SNPs) in RTEL1 and HAPE risk in Chinese Han population. A total of 265 HAPE patients and 303 healthy controls were included in our case-control study. Four SNPs in RTEL1 were selected and genotyped using the Sequenom MassARRAY method. Odds ratios (ORs) and 95% confidence intervals (95% CIs) were calculated by unconditional logistic regression with adjustment for gender and age. All P values were Bonferroni corrected, and statistical significance was set at P RTEL1 and a decreased risk HAPE in the Chinese population. The results need further confirmation.

  4. A monolithic glass chip for active single-cell sorting based on mechanical phenotyping.

    Faigle, Christoph; Lautenschläger, Franziska; Whyte, Graeme; Homewood, Philip; Martín-Badosa, Estela; Guck, Jochen

    2015-03-07

    The mechanical properties of biological cells have long been considered as inherent markers of biological function and disease. However, the screening and active sorting of heterogeneous populations based on serial single-cell mechanical measurements has not been demonstrated. Here we present a novel monolithic glass chip for combined fluorescence detection and mechanical phenotyping using an optical stretcher. A new design and manufacturing process, involving the bonding of two asymmetrically etched glass plates, combines exact optical fiber alignment, low laser damage threshold and high imaging quality with the possibility of several microfluidic inlet and outlet channels. We show the utility of such a custom-built optical stretcher glass chip by measuring and sorting single cells in a heterogeneous population based on their different mechanical properties and verify sorting accuracy by simultaneous fluorescence detection. This offers new possibilities of exact characterization and sorting of small populations based on rheological properties for biological and biomedical applications.

  5. Mechanism-based classification of pain for physical therapy management in palliative care: A clinical commentary

    Senthil P Kumar

    2011-01-01

    Full Text Available Pain relief is a major goal for palliative care in India so much that most palliative care interventions necessarily begin first with pain relief. Physical therapists play an important role in palliative care and they are regarded as highly proficient members of a multidisciplinary healthcare team towards management of chronic pain. Pain necessarily involves three different levels of classification-based upon pain symptoms, pain mechanisms and pain syndromes. Mechanism-based treatments are most likely to succeed compared to symptomatic treatments or diagnosis-based treatments. The objective of this clinical commentary is to update the physical therapists working in palliative care, on the mechanism-based classification of pain and its interpretation, with available therapeutic evidence for providing optimal patient care using physical therapy. The paper describes the evolution of mechanism-based classification of pain, the five mechanisms (central sensitization, peripheral neuropathic, nociceptive, sympathetically maintained pain and cognitive-affective are explained with recent evidence for physical therapy treatments for each of the mechanisms.

  6. Quality prediction modeling for sintered ores based on mechanism models of sintering and extreme learning machine based error compensation

    Tiebin, Wu; Yunlian, Liu; Xinjun, Li; Yi, Yu; Bin, Zhang

    2018-06-01

    Aiming at the difficulty in quality prediction of sintered ores, a hybrid prediction model is established based on mechanism models of sintering and time-weighted error compensation on the basis of the extreme learning machine (ELM). At first, mechanism models of drum index, total iron, and alkalinity are constructed according to the chemical reaction mechanism and conservation of matter in the sintering process. As the process is simplified in the mechanism models, these models are not able to describe high nonlinearity. Therefore, errors are inevitable. For this reason, the time-weighted ELM based error compensation model is established. Simulation results verify that the hybrid model has a high accuracy and can meet the requirement for industrial applications.

  7. Mutations in the putative zinc-binding motif of UL52 demonstrate a complex interdependence between the UL5 and UL52 subunits of the human herpes simplex virus type 1 helicase/primase complex.

    Chen, Yan; Carrington-Lawrence, Stacy D; Bai, Ping; Weller, Sandra K

    2005-07-01

    Herpes simplex virus type 1 (HSV-1) encodes a heterotrimeric helicase-primase (UL5/8/52) complex. UL5 contains seven motifs found in helicase superfamily 1, and UL52 contains conserved motifs found in primases. The contributions of each subunit to the biochemical activities of the complex, however, remain unclear. We have previously demonstrated that a mutation in the putative zinc finger at UL52 C terminus abrogates not only primase but also ATPase, helicase, and DNA-binding activities of a UL5/UL52 subcomplex, indicating a complex interdependence between the two subunits. To test this hypothesis and to further investigate the role of the zinc finger in the enzymatic activities of the helicase-primase, a series of mutations were constructed in this motif. They differed in their ability to complement a UL52 null virus: totally defective, partial complementation, and potentiating. In this study, four of these mutants were studied biochemically after expression and purification from insect cells infected with recombinant baculoviruses. All mutants show greatly reduced primase activity. Complementation-defective mutants exhibited severe defects in ATPase, helicase, and DNA-binding activities. Partially complementing mutants displayed intermediate levels of these activities, except that one showed a wild-type level of helicase activity. These data suggest that the UL52 zinc finger motif plays an important role in the activities of the helicase-primase complex. The observation that mutations in UL52 affected helicase, ATPase, and DNA-binding activities indicates that UL52 binding to DNA via the zinc finger may be necessary for loading UL5. Alternatively, UL5 and UL52 may share a DNA-binding interface.

  8. Lift vs. drag based mechanisms for vertical force production in the smallest flying insects.

    Jones, S K; Laurenza, R; Hedrick, T L; Griffith, B E; Miller, L A

    2015-11-07

    We used computational fluid dynamics to determine whether lift- or drag-based mechanisms generate the most vertical force in the flight of the smallest insects. These insects fly at Re on the order of 4-60 where viscous effects are significant. Detailed quantitative data on the wing kinematics of the smallest insects is not available, and as a result both drag- and lift-based strategies have been suggested as the mechanisms by which these insects stay aloft. We used the immersed boundary method to solve the fully-coupled fluid-structure interaction problem of a flexible wing immersed in a two-dimensional viscous fluid to compare three idealized hovering kinematics: a drag-based stroke in the vertical plane, a lift-based stroke in the horizontal plane, and a hybrid stroke on a tilted plane. Our results suggest that at higher Re, a lift-based strategy produces more vertical force than a drag-based strategy. At the Re pertinent to small insect hovering, however, there is little difference in performance between the two strategies. A drag-based mechanism of flight could produce more vertical force than a lift-based mechanism for insects at Re<5; however, we are unaware of active fliers at this scale. Copyright © 2015 Elsevier Ltd. All rights reserved.

  9. A time-based admission control mechanism for IEEE 802.11 ad Hoc networks

    Costa, Luís Henrique M. K.; Cerveira, Carlos Rodrigo

    2006-01-01

    This paper presents a time-based admission control mechanism (TAC) for IEEE 802.11 ad hoc networks. The proposed mechanism was adapted to the QoS AODV routing protocol, which takes the quality of service requirements of the data flow into account in the route discovery process. TAC-AODV estimates the idle time of the physical medium based on the frames listened. The incoming traffic is admitted according to the offered load as well as the intra-flow interference, calculated based on the numbe...

  10. Design of mechanical coxa joints based on three-degree-of-freedom spherical parallel manipulators

    Li, Yanbiao; Ji, Shiming; Wang, Zhongfei; Jin, Mingsheng; Liu, Yi; Jin, Zhenlin

    2013-01-01

    We addressed the issue of the design of mechanical coxa joints based on three-degree-of-freedom spherical parallel manipulators using the parameter statistics optimum method based on index atlases. The coxa joints have the advantages of high payload, high accuracy, and good technological efficiency. The first step of the design and prototyping used in this paper develops the direct and inverse displacement equations from the layout feature of the mechanical coxa joints. Then, the shapes of a constant-orientation workspace of the mechanical coxa joints are described, and the effects of the design parameters on the workspace volume are studied quantitatively. The next step deals with the graphical representation of the atlases that illustrates the relationship between performance evaluation index and design parameters based on the kinematics and torque analysis of the mechanical coxa joints. Finally, the geometric parameters of the coxa joints are obtained by the parameter statistics optimum method based on the index atlases. Considering assembly conditions, the design scheme of the mechanical coxa joints is developed, which provides a theoretical basis for the application of the mechanical coxa joints.

  11. Dynamic Symmetric Key Mobile Commerce Scheme Based on Self-Verified Mechanism

    Jiachen Yang

    2014-01-01

    Full Text Available In terms of the security and efficiency of mobile e-commerce, the authors summarized the advantages and disadvantages of several related schemes, especially the self-verified mobile payment scheme based on the elliptic curve cryptosystem (ECC and then proposed a new type of dynamic symmetric key mobile commerce scheme based on self-verified mechanism. The authors analyzed the basic algorithm based on self-verified mechanisms and detailed the complete transaction process of the proposed scheme. The authors analyzed the payment scheme based on the security and high efficiency index. The analysis shows that the proposed scheme not only meets the high efficiency of mobile electronic payment premise, but also takes the security into account. The user confirmation mechanism at the end of the proposed scheme further strengthens the security of the proposed scheme. In brief, the proposed scheme is more efficient and practical than most of the existing schemes.

  12. Charge transport model in nanodielectric composites based on quantum tunneling mechanism and dual-level traps

    Li, Guochang; Chen, George, E-mail: gc@ecs.soton.ac.uk, E-mail: sli@mail.xjtu.edu.cn [State Key Laboratory of Electrical Insulation and Power Equipment, Xi' an Jiaotong University, Xi' an 710049 (China); School of Electronic and Computer Science, University of Southampton, Southampton SO17 1BJ (United Kingdom); Li, Shengtao, E-mail: gc@ecs.soton.ac.uk, E-mail: sli@mail.xjtu.edu.cn [State Key Laboratory of Electrical Insulation and Power Equipment, Xi' an Jiaotong University, Xi' an 710049 (China)

    2016-08-08

    Charge transport properties in nanodielectrics present different tendencies for different loading concentrations. The exact mechanisms that are responsible for charge transport in nanodielectrics are not detailed, especially for high loading concentration. A charge transport model in nanodielectrics has been proposed based on quantum tunneling mechanism and dual-level traps. In the model, the thermally assisted hopping (TAH) process for the shallow traps and the tunnelling process for the deep traps are considered. For different loading concentrations, the dominant charge transport mechanisms are different. The quantum tunneling mechanism plays a major role in determining the charge conduction in nanodielectrics with high loading concentrations. While for low loading concentrations, the thermal hopping mechanism will dominate the charge conduction process. The model can explain the observed conductivity property in nanodielectrics with different loading concentrations.

  13. Investigating the dental toolkit of primates based on food mechanical properties: Feeding action does matter.

    Thiery, Ghislain; Guy, Franck; Lazzari, Vincent

    2017-06-01

    Although conveying an indisputable morphological and behavioral signal, traditional dietary categories such as frugivorous or folivorous tend to group a wide range of food mechanical properties together. Because food/tooth interactions are mostly mechanical, it seems relevant to investigate the dental morphology of primates based on mechanical categories. However, existing mechanical categories classify food by its properties but cannot be used as factors to classify primate dietary habits. This comes from the fact that one primate species might be adapted to a wide range of food mechanical properties. To tackle this issue, what follows is an original framework based on action-related categories. The proposal here is to classify extant primates based on the range of food mechanical properties they can process through one given action. The resulting categories can be used as factors to investigate the dental tools available to primates. Furthermore, cracking, grinding, and shearing categories assigned depending on the hardness and the toughness of food are shown to be supported by morphological data (3D relative enamel thickness) and topographic data (relief index, occlusal complexity, and Dirichlet normal energy). Inferring food mechanical properties from dental morphology is especially relevant for the study of extinct primates, which are mainly documented by dental remains. Hence, we use action-related categories to investigate the molar morphology of an extinct colobine monkey Mesopithecus pentelicus from the Miocene of Pikermi, Greece. Action-related categories show contrasting results compared with classical categories and give us new insights into the dietary adaptations of this extinct primate. Finally, we provide some possible directions for future research aiming to test action-related categories. In particular, we suggest acquiring more data on mechanically challenging fallback foods and advocate the use of other food mechanical properties such as

  14. Mechanical Systems based on Dry Friction Force used for Building Isolation against Seismic Actions

    Fanel Dorel Şcheaua

    2017-11-01

    Full Text Available Today there are multiple solutions intended to avoid the earthquake damaging effects on building structures. There are methods based on the use of special mechanical systems attached directly to the structure's resistance frames, by means of which an improved building behavior is achieved during the earthquake. The systems used work on the principle of structure base isolation based on the dry friction force (Coulomb friction. Some constructive types of these isolation systems patterns are described in this paper

  15. Thermo-mechanical modeling of the obduction process based on the Oman ophiolite case

    Duretz , Thibault; Agard , Philippe; Yamato , Philippe; Ducassou , Céline; Burov , Evgenii ,; Gerya , T. V.

    2016-01-01

    International audience; Obduction emplaces regional-scale fragments of oceanic lithosphere (ophiolites) over continental lithosphere margins of much lower density. For this reason, the mechanisms responsible for obduction remain enigmatic in the framework of plate tectonics. We present two-dimensional (2D) thermo-mechanical models of obduction and investigate possible dynamics and physical controls of this process. Model geometry and boundary conditions are based on available geological and g...

  16. Thermo-mechanical constitutive modeling of unsaturated clays based on the critical state concepts

    Tourchi, Saeed; Hamidi, Amir

    2015-01-01

    A thermo-mechanical constitutive model for unsaturated clays is constructed based on the existing model for saturated clays originally proposed by the authors. The saturated clays model was formulated in the framework of critical state soil mechanics and modified Cam-clay model. The existing model has been generalized to simulate the experimentally observed behavior of unsaturated clays by introducing Bishop's stress and suction as independent stress parameters and modifying the hardening rul...

  17. Humidity Responsive Photonic Sensor based on a Carboxymethyl Cellulose Mechanical Actuator

    Hartings, Matthew; Douglass, Kevin O.; Neice, Claire; Ahmed, Zeeshan

    2017-01-01

    We describe an intuitive and simple method for exploiting humidity-driven volume changes in carboxymethyl cellulose (CMC) to fabricate a humidity responsive actuator on a glass fiber substrate. We optimize this platform to generate a photonic-based humidity sensor where CMC coated on a fiber optic containing a fiber Bragg grating (FBG) actuates a mechanical strain in response to humidity changes. The humidity-driven mechanical deformation of the FBG results in a large lin...

  18. A Mobile IPv6 based Distributed Mobility Management Mechanism of Mobile Internet

    Yan, Shi; Jiayin, Cheng; Shanzhi, Chen

    A flatter architecture is one of the trends of mobile Internet. Traditional centralized mobility management mechanism faces the challenges such as scalability and UE reachability. A MIPv6 based distributed mobility management mechanism is proposed in this paper. Some important network entities and signaling procedures are defined. UE reachability is also considered in this paper through extension to DNS servers. Simulation results show that the proposed approach can overcome the scalability problem of the centralized scheme.

  19. Mechanical properties of permeable materials with an organized structure on the base of continuous metal fibers

    Karpinos, D.M.; Rutkovskij, A.E.; Zorin, V.A.; Ivanchuk, A.A.

    1979-01-01

    The mechanical properties were studied for permeable fibrous materials with an organized structure on the base of continuous metal fibers (from Kh18N9T steel) subjected to preliminary reprocessing volumetric net half-finished products. The effect of geometrical parameters of the net half-finished products and of their orientation in packing are shown to affect the mechanical properties within a wide range of porosities

  20. Shear design and assessment of reinforced and prestressed concrete beams based on a mechanical model

    Marí Bernat, Antonio Ricardo; Bairán García, Jesús Miguel; Cladera Bohigas, Antoni; Oller Ibars, Eva

    2016-01-01

    Safe and economical design and assessment of reinforced (RC) and prestressed concrete (PC) beams requires the availability of accurate but simple formulations which adequately capture the structural response. In this paper, a mechanical model for the prediction of the shear-flexural strength of PC and RC members with rectangular, I, or T sections, with and without shear reinforcement, is presented. The model is based on the principles of concrete mechanics and on assumptions supported by the ...

  1. [Present status and trend of heart fluid mechanics research based on medical image analysis].

    Gan, Jianhong; Yin, Lixue; Xie, Shenghua; Li, Wenhua; Lu, Jing; Luo, Anguo

    2014-06-01

    With introduction of current main methods for heart fluid mechanics researches, we studied the characteristics and weakness for three primary analysis methods based on magnetic resonance imaging, color Doppler ultrasound and grayscale ultrasound image, respectively. It is pointed out that particle image velocity (PIV), speckle tracking and block match have the same nature, and three algorithms all adopt block correlation. The further analysis shows that, with the development of information technology and sensor, the research for cardiac function and fluid mechanics will focus on energy transfer process of heart fluid, characteristics of Chamber wall related to blood fluid and Fluid-structure interaction in the future heart fluid mechanics fields.

  2. Silicon Nano fabrication by Atomic Force Microscopy-Based Mechanical Processing

    Miyake, Sh.; Wang, M.; Kim, J.

    2014-01-01

    This paper reviews silicon nano fabrication processes using atomic force microscopy (AFM). In particular, it summarizes recent results obtained in our research group regarding AFM-based silicon nano fabrication through mechanochemical local oxidation by diamond tip sliding, as well as mechanical, electrical, and electromechanical processing using an electrically conductive diamond tip. Microscopic three-dimensional manufacturing mainly relies on etching, deposition, and lithography. Therefore, a special emphasis was placed on nano mechanical processes, mechanochemical reaction by potassium hydroxide solution etching, and mechanical and electrical approaches. Several important surface characterization techniques consisting of scanning tunneling microscopy and related techniques, such as scanning probe microscopy and AFM, were also discussed.

  3. Mechanical characterisation of agarose-based chromatography resins for biopharmaceutical manufacture.

    Nweke, Mauryn C; McCartney, R Graham; Bracewell, Daniel G

    2017-12-29

    Mechanical characterisation of agarose-based resins is an important factor in ensuring robust chromatographic performance in the manufacture of biopharmaceuticals. Pressure-flow profiles are most commonly used to characterise these properties. There are a number of drawbacks with this method, including the potential need for several re-packs to achieve the desired packing quality, the impact of wall effects on experimental set up and the quantities of chromatography media and buffers required. To address these issues, we have developed a dynamic mechanical analysis (DMA) technique that characterises the mechanical properties of resins based on the viscoelasticity of a 1ml sample of slurry. This technique was conducted on seven resins with varying degrees of mechanical robustness and the results were compared to pressure-flow test results on the same resins. Results show a strong correlation between the two techniques. The most mechanically robust resin (Capto Q) had a critical velocity 3.3 times higher than the weakest (Sepharose CL-4B), whilst the DMA technique showed Capto Q to have a slurry deformation rate 8.3 times lower than Sepharose CL-4B. To ascertain whether polymer structure is indicative of mechanical strength, scanning electron microscopy images were also used to study the structural properties of each resin. Results indicate that DMA can be used as a small volume, complementary technique for the mechanical characterisation of chromatography media. Copyright © 2017 The Author(s). Published by Elsevier B.V. All rights reserved.

  4. Constraint-plane-based synthesis and topology variation of a class of metamorphic parallel mechanisms

    Gan, Dongming; Dias, Jorge; Seneviratne, Lakmal; Dai, Jian S.

    2014-01-01

    This paper investigates various topologies and mobility of a class of metamorphic parallel mechanisms synthesized with reconfigurable rTPS limbs. Based on the reconfigurable Hooke (rT) joint, the rTPS limb has two phases which result in parallel mechanisms having ability of mobility change. While in one phase the limb has no constraint to the platform, in the other it constrains the spherical joint center to lie on a plane which is used to demonstrate different topologies of the nrTPS metamorphic parallel mechanisms by investigating various relations (parallel or intersecting) among the n constraint planes (n = 2,3,..,6). Geometric constraint equations of the platform rotation matrix and translation vector are set up based on the point-plane constraint, which reveals mobility and redundant geometric conditions of the mechanism topologies. By altering the limbs into the non-constraint phase without constraint plane, new mechanism phases are deduced with mobility change based on each mechanism topology.

  5. Molecular medicine of fragile X syndrome: based on known molecular mechanisms.

    Luo, Shi-Yu; Wu, Ling-Qian; Duan, Ran-Hui

    2016-02-01

    Extensive research on fragile X mental retardation gene knockout mice and mutant Drosophila models has largely expanded our knowledge on mechanism-based treatment of fragile X syndrome (FXS). In light of these findings, several clinical trials are now underway for therapeutic translation to humans. Electronic literature searches were conducted using the PubMed database and ClinicalTrials.gov. The search terms included "fragile X syndrome", "FXS and medication", "FXS and therapeutics" and "FXS and treatment". Based on the publications identified in this search, we reviewed the neuroanatomical abnormalities in FXS patients and the potential pathogenic mechanisms to monitor the progress of FXS research, from basic studies to clinical trials. The pathological mechanisms of FXS were categorized on the basis of neuroanatomy, synaptic structure, synaptic transmission and fragile X mental retardation protein (FMRP) loss of function. The neuroanatomical abnormalities in FXS were described to motivate extensive research into the region-specific pathologies in the brain responsible for FXS behavioural manifestations. Mechanism-directed molecular medicines were classified according to their target pathological mechanisms, and the most recent progress in clinical trials was discussed. Current mechanism-based studies and clinical trials have greatly contributed to the development of FXS pharmacological therapeutics. Research examining the extent to which these treatments provided a rescue effect or FMRP compensation for the developmental impairments in FXS patients may help to improve the efficacy of treatments.

  6. Study of network resource allocation based on market and game theoretic mechanism

    Liu, Yingmei; Wang, Hongwei; Wang, Gang

    2004-04-01

    We work on the network resource allocation issue concerning network management system function based on market-oriented mechanism. The scheme is to model the telecommunication network resources as trading goods in which the various network components could be owned by different competitive, real-world entities. This is a multidisciplinary framework concentrating on the similarity between resource allocation in network environment and the market mechanism in economic theory. By taking an economic (market-based and game theoretic) approach in routing of communication network, we study the dynamic behavior under game-theoretic framework in allocating network resources. Based on the prior work of Gibney and Jennings, we apply concepts of utility and fitness to the market mechanism with an intention to close the gap between experiment environment and real world situation.

  7. Linking project-based mechanisms with domestic greenhouse gas emissions trading schemes

    Bygrave, S.; Bosi, M.

    2004-01-01

    Although there are a number of possible links between emission trading and project-based mechanisms, the focus of this paper is on linking domestic GHG emission trading schemes with: (1) domestic; and, (2) international (JI and CDM) GHG reduction project activities. The objective is to examine some of the challenges in linking DETs and project-based mechanisms, as well as some possible solutions to address these challenges. The link between JI / CDM and intergovernmental international emissions trading (i.e. Article 17 of the Kyoto Protocol) is defined by the Kyoto Protocol, and therefore is not covered in this paper. The paper is written in the context of: (a) countries adhering to the Kyoto Protocol and elaborating their strategies to meet their GHG emission commitments, including through the use of the emissions trading and project-based mechanisms. For example, the European Union (EU) will be commencing a GHG Emissions Trading Scheme in January 2005, and recently, the Council of ministers and the European Parliament agreed on a text for an EU Linking Directive allowing the use of JI and CDM emission units in the EU Emission Trading Scheme (EU-ETS); and (b) all countries (and/or regions within countries) with GHG emission obligations that may choose to use domestic emissions trading and project-based mechanisms to meet their GHG commitments. The paper includes the following elements: (1) an overview of the different flexibility mechanisms (i.e. GHG emissions trading and PBMs), including a brief description and comparisons between the mechanisms (Section 3); (2) an exploration of the issues that emerge when project-based mechanisms link with domestic emissions trading schemes, as well as possible solutions to address some of the challenges raised (Section 4); (3) a case study examining the EU-ETS and the EU Linking Directive on project-based mechanisms, in particular on how the EU is addressing in a practical context relevant linking issues (Section 5); (4) a

  8. Heterocyclic Schiff bases as non toxic antioxidants: Solvent effect, structure activity relationship and mechanism of action

    Shanty, Angamaly Antony; Mohanan, Puzhavoorparambil Velayudhan

    2018-03-01

    Phenolic heterocyclic imine based Schiff bases from Thiophene-2-carboxaldehyde and Pyrrole-2-carboxaldehyde were synthesized and characterized as novel antioxidants. The solvent effects of these Schiff bases were determined and compared with standard antioxidants, BHA employing DPPH assay and ABTS assay. Fixed reaction time and Steady state measurement were used for study. IC50 and EC50 were calculated. Structure-activity relationship revealed that the electron donating group in the phenolic ring increases the activity where as the electron withdrawing moiety decreases the activity. The Schiff base derivatives showed antioxidant property by two different pathways namely SPLET and HAT mechanisms in DPPH assay. While in ABTS method, the reaction between ABTS radical and Schiff bases involves electron transfer followed by proton transfer (ET-PT) mechanism. The cytotoxicity of these compounds has been evaluated by MTT assay. The results showed that all these compounds are non toxic in nature.

  9. Mechanisms and regulation of DNA replication initiation in eukaryotes.

    Parker, Matthew W; Botchan, Michael R; Berger, James M

    2017-04-01

    Cellular DNA replication is initiated through the action of multiprotein complexes that recognize replication start sites in the chromosome (termed origins) and facilitate duplex DNA melting within these regions. In a typical cell cycle, initiation occurs only once per origin and each round of replication is tightly coupled to cell division. To avoid aberrant origin firing and re-replication, eukaryotes tightly regulate two events in the initiation process: loading of the replicative helicase, MCM2-7, onto chromatin by the origin recognition complex (ORC), and subsequent activation of the helicase by its incorporation into a complex known as the CMG. Recent work has begun to reveal the details of an orchestrated and sequential exchange of initiation factors on DNA that give rise to a replication-competent complex, the replisome. Here, we review the molecular mechanisms that underpin eukaryotic DNA replication initiation - from selecting replication start sites to replicative helicase loading and activation - and describe how these events are often distinctly regulated across different eukaryotic model organisms.

  10. Application of mechanical alloying to synthesis of intermetallic phases based alloys

    Dymek, S.

    2001-01-01

    Mechanical alloying is the process of synthesis of powder materials during milling in high energetic mills, usually ball mills. The central event in mechanical alloying is the ball-powder-ball collision. Powder particles are trapped between the colliding balls during milling and undergo deformation and/or fracture. Fractured parts are cold welded. The continued fracture and cold welding results in a uniform size and chemical composition of powder particles. The main applications of mechanical alloying are: processing of ODS alloys, syntheses of intermetallic phases, synthesis of nonequilibrium structures (amorphous alloys, extended solid solutions, nanocrystalline, quasi crystals) and magnetic materials. The present paper deals with application of mechanical alloying to synthesis Ni A l base intermetallic phases as well as phases from the Nb-Al binary system. The alloy were processed from elemental powders. The course of milling was monitored by scanning electron microscopy and X-ray diffraction. After milling, the collected powders were sieved by 45 μm grid and hot pressed (Nb alloys and NiAl) or hot extruded (NiAl). The resulting material was fully dense and exhibited fine grain (< 1 μm) and uniform distribution of oxide dispersoid. The consolidated material was compression and creep tested. The mechanical properties of mechanically alloys were superior to properties of their cast counterparts both in the room and elevated temperatures. Higher strength of mechanically alloyed materials results from their fine grains and from the presence of dispersoid. At elevated temperatures, the Nb-Al alloys have higher compression strength than NiAl-based alloys processed at the same conditions. The minimum creep rates of mechanically alloyed Nb alloys are an order of magnitude lower than analogously processed NiAl-base alloys. (author)

  11. Mechanical properties correlation to processing parameters for advanced alumina based refractories

    Dimitrijević Marija M.

    2012-01-01

    Full Text Available Alumina based refractories are usually used in metallurgical furnaces and their thermal shock resistance is of great importance. In order to improve thermal shock resistance and mechanical properties of alumina based refractories short ceramic fibers were added to the material. SEM technique was used to compare the microstructure of specimens and the observed images gave the porosity and morphological characteristics of pores in the specimens. Standard compression test was used to determine the modulus of elasticity and compression strength. Results obtained from thermal shock testing and mechanical properties measurements were used to establish regression models that correlated specimen properties to process parameters.

  12. Synthesis and SAR studies of 5-(pyridin-4-yl)-1,3,4-thiadiazol-2-amine derivatives as potent inhibitors of Bloom helicase

    Rosenthal, Andrew S; Dexheimer, Thomas S; Gileadi, Opher

    2013-01-01

    complementary strands of duplex DNA as well as atypical DNA structures such as Holliday junctions. Mutations of the BLM gene can result in Bloom syndrome, an autosomal recessive disorder associated with cancer predisposition. BLM-deficient cells exhibit increased sensitivity to DNA damaging agents indicating...... and related analogs, which possess potent BLM inhibition and exhibit selectivity over related helicases. Moreover, these compounds demonstrated cellular activity by inducing sister chromatid exchanges, a hallmark of Bloom syndrome....

  13. RECQ5 helicase associates with the C-terminal repeat domain of RNA polymerase II during productive elongation phase of transcription

    Kanagaraj, R.; Huehn, D.; Mackellar, A.; Menigatti, M.; Zheng, L.; Urban, Václav; Shevelev, Igor; Greenleaf, A.L.; Janščák, Pavel

    2010-01-01

    Roč. 38, č. 22 (2010), s. 8131-8140 ISSN 0305-1048 R&D Projects: GA ČR GA204/09/0565 Grant - others:SNSF(CH) 3100A0-116008; NIH(US) GM040505 Institutional research plan: CEZ:AV0Z50520514 Keywords : RECQ5 DNA helicase * transcription * genome stability Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 7.836, year: 2010

  14. Cloning and expression of NS3 helicase fragment of hepatitis C virus and the study of its immunoreactivity in HCV infected patients

    Mahrou Sadri

    2015-02-01

    Full Text Available Objective(s: Hepatitis C is a major cause of liver failure worldwide. Current therapies applied for this disease are not fully effective and produce side effects in most cases. Non-structural protein 3 helicase (NS3 of HCV is one of the key enzymes in viral replication and infection. Therefore, this region is a promising target to design new drugs and therapies against HCV infection. The aim of this study was cloning and expression of HCV NS3 helicase fragment in Escherichia coli BL21 (DE3 using pET102/D-TOPO expression vector and studying immunoreactivity of the expressed antigen in Iranian infected with hepatitis C. Materials and Methods: The viral RNA was extracted from the serum of HCV infected patient. The NS3 helicase region was amplified by RT-PCR. The PCR product was directionally cloned into the expression vector pET102/D-TOPO and transformed into the BL21 strain of E. coli (DE3. The transformed bacteria were then induced by adding 1mM isopropyl-β-D-thiogalactopyranoside (IPTG into the culture medium to enhance the protein expression. SDS-PAGE and western blotting were carried out to identify the protein under investigation, and finally purified recombinant fusion protein was used as the antigen for ELISA method. Results: Theinsertion of theDNA fragment of the NS3 regioninto the expression vectorwas further confirmed by PCR and sequencing. SDS-PAGE analysis showed the successful expression of the recombinant protein of interest. Furthermore, immunoreactivity of fusion NS3 helicase was confirmed by ELISA and western blotting. Conclusion: It seems that this recombinant protein could be a useful source of antigen for future studies on HCV diagnosis and therapy.

  15. On the hydro-mechanical behaviour of MX80 bentonite-based materials

    Yu-Jun Cui

    2017-06-01

    Full Text Available Bentonite-based materials have been considered in many countries as engineered barrier/backfilling materials in deep geological disposal of high-level radioactive waste. During the long period of waste storage, these materials will play an essential role in ensuring the integrity of the storage system that consists of the waste canisters, the engineered barrier/backfill, the retaining structures as well as the geological barrier. Thus, it is essential to well understand the hydro-mechanical behaviours of these bentonite-based materials. This review paper presents the recent advances of knowledge on MX80 bentonite-based materials, in terms of water retention properties, hydraulic behaviour and mechanical behaviour. Emphasis is put on the effect of technological voids and the role of the dry density of bentonite. The swelling anisotropy is also discussed based on the results from swelling tests with measurements of both axial and radial swelling pressures on a sand-bentonite mixture compacted at different densities. Microstructure observation was used to help the interpretation of macroscopic hydro-mechanical behaviour. Also, the evolution of soil microstructure thus the soil density over time is discussed based on the results from mock-up tests. This evolution is essential for understanding the long-term hydro-mechanical behaviour of the engineered barrier/backfill.

  16. A novel vibration assisted polishing device based on the flexural mechanism driven by the piezoelectric actuators

    Guilian Wang

    2018-01-01

    Full Text Available The vibration assisted polishing has widely application fields because of higher machining frequency and better polishing quality, especially the polishing with the non-resonant mode that is regarded as a kind of promising polishing method. This paper reports a novel vibration assisted polishing device, consisting of the flexible hinge mechanism driven by the piezoelectric actuators, which is suitable for polishing planes or curve surfaces with slow curvature. Firstly, the generation methods of vibration trajectory are investigated for the same frequency and different frequency signals’ inputs, respectively, and then the types of elliptic and Lissajous’s vibration trajectories are generated respectively. Secondly, a flexural mechanism consisting of the right circular flexible hinges and the leaf springs is developed to produce two-dimensional vibration trajectory. Statics and dynamics investigating of this flexible mechanism are finished in detail. The analytical models about input and output compliances of the flexural mechanism are established according to the matrix-based compliance modeling, and the dynamic model of the flexural mechanism based on the Euler-Lagrange equation is also presented. The finite element model of the flexural mechanism was established to carry out the numerical simulation in order to testify the rationality of device design. Finally, the polishing experiment is carried out to prove the effectiveness of the vibration device. The experimental results show that this novel vibration assisted polishing device developed in this study can remove more effectively the cutting marks left by last process and obviously reduce the workpiece surface roughness.

  17. Comparison of the mechanical properties between carbon nanotube and nanocrystalline cellulose polypropylene based nano-composites

    Huang, Jun; Rodrigue, Denis

    2015-01-01

    Highlights: • SWCNT and NCC can effectively improve the mechanical properties of nano-composites. • SWCNT is more effective than NCC to increase modulus and strength. • Longer NCC is more effective to improve the mechanical properties of nano-composites. • It is more economic to use NCC than SWCNT to improve mechanical properties. - Abstract: Using beam and tetrahedron elements to simulate nanocrystalline cellulose (NCC), single wall carbon nanotube (SWCNT) and polypropylene (PP), finite element method (FEM) is used to predict the mechanical properties of nano-composites. The bending, shear and torsion behaviors of nano-composites are especially investigated due to the limited amount of information in the present literature. First, mixed method (MM) and FEM are used to compare the bending stiffness of NCC/PP and SWCNT/PP composites. Second, based on mechanics of materials, the shear moduli of both types of nano-composites are obtained. Finally, fixing the number of fibers and for different volume contents, four NCC lengths are used to determine the mechanical properties of the composites. The bending and shearing performances are also compared between NCC and SWCNT based composites. In all cases, the elastic–plastic analyses are carried out and the stress or strain distributions for specific regions are also investigated. From all the results obtained, an economic analysis shows that NCC is more interesting than SWCNT to reinforce PP

  18. A morphing approach to couple state-based peridynamics with classical continuum mechanics

    Han, Fei

    2016-01-04

    A local/nonlocal coupling technique called the morphing method is developed to couple classical continuum mechanics with state-based peridynamics. State-based peridynamics, which enables the description of cracks that appear and propagate spontaneously, is applied to the key domain of a structure, where damage and fracture are considered to have non-negligible effects. In the rest of the structure, classical continuum mechanics is used to reduce computational costs and to simultaneously satisfy solution accuracy and boundary conditions. Both models are glued by the proposed morphing method in the transition region. The morphing method creates a balance between the stiffness tensors of classical continuum mechanics and the weighted coefficients of state-based peridynamics through the equivalent energy density of both models. Linearization of state-based peridynamics is derived by Taylor approximations based on vector operations. The discrete formulation of coupled models is also described. Two-dimensional numerical examples illustrate the validity and accuracy of the proposed technique. It is shown that the morphing method, originally developed for bond-based peridynamics, can be successfully extended to state-based peridynamics through the original developments presented here.

  19. A morphing approach to couple state-based peridynamics with classical continuum mechanics

    Han, Fei; Lubineau, Gilles; Azdoud, Yan; Askari, Abe

    2016-01-01

    A local/nonlocal coupling technique called the morphing method is developed to couple classical continuum mechanics with state-based peridynamics. State-based peridynamics, which enables the description of cracks that appear and propagate spontaneously, is applied to the key domain of a structure, where damage and fracture are considered to have non-negligible effects. In the rest of the structure, classical continuum mechanics is used to reduce computational costs and to simultaneously satisfy solution accuracy and boundary conditions. Both models are glued by the proposed morphing method in the transition region. The morphing method creates a balance between the stiffness tensors of classical continuum mechanics and the weighted coefficients of state-based peridynamics through the equivalent energy density of both models. Linearization of state-based peridynamics is derived by Taylor approximations based on vector operations. The discrete formulation of coupled models is also described. Two-dimensional numerical examples illustrate the validity and accuracy of the proposed technique. It is shown that the morphing method, originally developed for bond-based peridynamics, can be successfully extended to state-based peridynamics through the original developments presented here.

  20. Origami-based tunable truss structures for non-volatile mechanical memory operation.

    Yasuda, Hiromi; Tachi, Tomohiro; Lee, Mia; Yang, Jinkyu

    2017-10-17

    Origami has recently received significant interest from the scientific community as a method for designing building blocks to construct metamaterials. However, the primary focus has been placed on their kinematic applications by leveraging the compactness and auxeticity of planar origami platforms. Here, we present volumetric origami cells-specifically triangulated cylindrical origami (TCO)-with tunable stability and stiffness, and demonstrate their feasibility as non-volatile mechanical memory storage devices. We show that a pair of TCO cells can develop a double-well potential to store bit information. What makes this origami-based approach more appealing is the realization of two-bit mechanical memory, in which two pairs of TCO cells are interconnected and one pair acts as a control for the other pair. By assembling TCO-based truss structures, we experimentally verify the tunable nature of the TCO units and demonstrate the operation of purely mechanical one- and two-bit memory storage prototypes.Origami is a popular method to design building blocks for mechanical metamaterials. Here, the authors assemble a volumetric origami-based structure, predict its axial and rotational movements during folding, and demonstrate the operation of mechanical one- and two-bit memory storage.

  1. Cytotoxicity and mechanical behavior of chitin-bentonite clay based polyurethane bio-nanocomposites.

    Zia, Khalid Mahmood; Zuber, Mohammad; Barikani, Mehdi; Hussain, Rizwan; Jamil, Tahir; Anjum, Sohail

    2011-12-01

    Chitin based polyurethane bio-nanocomposites (PUBNC) were prepared using chitin, Delite HPS bentonite nanoclay enriched in montmorillonite (MMT), 4,4'-diphenylmethane diisocyanate (MDI) and polycaprolactone polyol CAPA 231 (3000 g/mol(-1)). The prepolymers having different concentration of Delite HPS bentonite nanoclay were extended with 2 moles of chitin. The structures of the resulted polymers were determined by FT-IR technique. The effect of nanoclay contents on mechanical properties and in vitro biocompatibility was investigated. The mechanical properties of the synthesized materials were improved with increase in the Delite HPS bentonite nanoclay contents. Optimum mechanical properties were obtained from the PU bio-nanocomposite samples having 4% Delite HPS bentonite nanoclay. The results revealed that the final PU bio-nanocomposite having 2% Delite HPS bentonite nanoclay contents is ideal contenders for surgical threads with on going investigations into their in vitro biocompatibility, non-toxicity, and mechanical properties. Copyright © 2011 Elsevier B.V. All rights reserved.

  2. Protein structure refinement using a quantum mechanics-based chemical shielding predictor

    Bratholm, Lars Andersen; Jensen, Jan Halborg

    2017-01-01

    The accurate prediction of protein chemical shifts using a quantum mechanics (QM)-based method has been the subject of intense research for more than 20 years but so far empirical methods for chemical shift prediction have proven more accurate. In this paper we show that a QM-based predictor...... of a protein backbone and CB chemical shifts (ProCS15, PeerJ, 2016, 3, e1344) is of comparable accuracy to empirical chemical shift predictors after chemical shift-based structural refinement that removes small structural errors. We present a method by which quantum chemistry based predictions of isotropic...

  3. Characterization of Staphylococcus aureus Primosomal DnaD Protein: Highly Conserved C-Terminal Region Is Crucial for ssDNA and PriA Helicase Binding but Not for DnaA Protein-Binding and Self-Tetramerization.

    Yen-Hua Huang

    Full Text Available The role of DnaD in the recruitment of replicative helicase has been identified. However, knowledge of the DNA, PriA, and DnaA binding mechanism of this protein for the DnaA- and PriA-directed replication primosome assemblies is limited. We characterized the DNA-binding properties of DnaD from Staphylococcus aureus (SaDnaD and analyzed its interactions with SaPriA and SaDnaA. The gel filtration chromatography analysis of purified SaDnaD and its deletion mutant proteins (SaDnaD1-195, SaDnaD1-200 and SaDnaD1-204 showed a stable tetramer in solution. This finding indicates that the C-terminal region aa 196-228 is not crucial for SaDnaD oligomerization. SaDnaD forms distinct complexes with ssDNA of different lengths. In fluorescence titrations, SaDnaD bound to ssDNA with a binding-site size of approximately 32 nt. A stable complex of SaDnaD1-195, SaDnaD1-200, and SaDnaD1-204 with ssDNA dT40 was undetectable, indicating that the C-terminal region of SaDnaD (particularly aa 205-228 is crucial for ssDNA binding. The SPR results revealed that SaDnaD1-195 can interact with SaDnaA but not with SaPriA, which may indicate that DnaD has different binding sites for PriA and DnaA. Both SaDnaD and SaDnaDY176A mutant proteins, but not SaDnaD1-195, can significantly stimulate the ATPase activity of SaPriA. Hence, the stimulation effect mainly resulted from direct contact within the protein-protein interaction, not via the DNA-protein interaction. Kinetic studies revealed that the SaDnaD-SaPriA interaction increases the Vmax of the SaPriA ATPase fivefold without significantly affecting the Km. These results indicate that the conserved C-terminal region is crucial for ssDNA and PriA helicase binding, but not for DnaA protein-binding and self-tetramerization.

  4. Effect of brief heat-curing on microstructure and mechanical properties in fresh cement based mortars

    Ballester, P.; Hidalgo, A.; Marmol, I.; Morales, J.; Sanchez, L.

    2009-01-01

    The effect of temperature on fresh mortar and cement paste was evaluated by simulating the curing conditions of external buildings plastering applied under extremely hot weather. The specimens were heated at controlled temperatures in the 40-80 o C range by exposure to IR radiation over short periods. The effect of soaking for a short time was also examined. The results of compressive strength tests, scanning electron microscopy, infrared spectroscopy and mercury porosimetry helped to characterize the mechanical and physico-chemical properties of the studied sample. Early age behaviour (28 days) in neat cement was barely affected by the temperature. By contrast, exposure to high temperatures caused significant microstructural changes in the mortar. However, successive soaking over short periods was found to reactivate the mechanism of curing and restore the expected mechanical properties. Based on the results, application of cement based mortar at high temperatures is effective when followed by a short, specific soaking process.

  5. From evidence based medicine to mechanism based medicine : Reviewing the role of pharmacogenetics

    Wilffert, Bob; Swen, Jesse; Mulder, Hans; Touw, Daan; Maitland-Van der Zee, Anke-Hilse; Deneer, Vera

    Aim of the review The translation of evidence based medicine to a specific patient presents a considerable challenge. We present by means of the examples nortriptyline, tramadol, clopidogrel, coumarins, abacavir and antipsychotics the discrepancy between available pharmacogenetic information and its

  6. From evidence based medicine to mechanism based medicine. Reviewing the role of pharmacogenetics

    Wilffert, Bob; Swen, Jesse; Mulder, Hans; Touw, Daan; Maitland-Van der Zee, Anke-Hilse; Deneer, Vera

    Aim of the review The translation of evidence based medicine to a specific patient presents a considerable challenge. We present by means of the examples nortriptyline, tramadol, clopidogrel, coumarins, abacavir and antipsychotics the discrepancy between available pharmacogenetic information and its

  7. New Drugs for Anemia Treatment Based on a New Understanding of the Mechanisms of Stress Erythropoiesis

    2015-11-01

    Award Number: W81XWH-12-1-0449 TITLE: New Drugs for Anemia Treatment Based on a New Understanding of the Mechanisms of Stress Erythropoiesis...COVERED 1Sep2012 - 31Aug2015 4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER New Drugs for Anemia Treatment Based on a New Understanding of the...cell formation in "Nan" (neonatal anemia ) mice, raising the level of red cells to almost normal. It also causes an increase in the numbers of splenic

  8. Architectural, Mechanical, Electrical, Plumbing, and Site Documents Associated with Sustainability Base, N232

    Grymes, Rosalind A.; Martin, Rodney Alexander; Dianati, Soheila

    2016-01-01

    These files contain more precise and accurate representations of the architectural, mechanical, electrical, plumbing, and site information pertaining to Sustainability Base, aka Collaborative Support Facility, aka N232. These supersede the 'bid' drawings released in STI 8112 previously. They are useful for NASA researchers and collaborators in modeling the performance characteristics of the facility. Otherwise, they do not contain new data.

  9. Representation of Coordination Mechanisms in IMS Learning Design to Support Group-based Learning

    Miao, Yongwu; Burgos, Daniel; Griffiths, David; Koper, Rob

    2007-01-01

    Miao, Y., Burgos, D., Griffiths, D., & Koper, R. (2008). Representation of Coordination Mechanisms in IMS Learning Design to Support Group-based Learning. In L. Lockyer, S. Bennet, S. Agostinho & B. Harper (Eds.), Handbook of Research on Learning Design and Learning Objects: Issues, Applications and

  10. Understanding the antimicrobial mechanism of TiO2-based nanocomposite films in a pathogenic bacterium

    Kubacka, A.; Suarez Diez, M.; Rojo, D.; Bargiela, R.; Ciordia, S.; Zapico, I.; Albar, J.P.; Barbas, C.; Martins Dos Santos, V.A.P.; Fernández-García, M.; Ferrer, M.

    2014-01-01

    Titania (TiO2)-based nanocomposites subjected to light excitation are remarkably effective in eliciting microbial death. However, the mechanism by which these materials induce microbial death and the effects that they have on microbes are poorly understood. Here, we assess the low dose

  11. Investigation of peptide based surface functionalization for copper ions detection using an ultrasensitive mechanical microresonator

    Cagliani, Alberto; Fischer, Lee MacKenzie; Rasmussen, Jakob Lyager

    2011-01-01

    In the framework of developing a portable label-free sensor for multi arrayed detection of heavy metals in drinking water, we present a mechanical resonator-based copper ions sensor, which uses a recently synthesized peptide Cysteine–Glycine–Glycine–Histidine (CGGH) and the l-Cysteine (Cys) peptide...

  12. Articulated, Performance-Based Instruction Objectives Guide for Automotive Mechanics. Final Document. Revised.

    Henderson, William Edward, Jr.

    Developed during a project designed to provide continuous, performance-based vocational training at the secondary and postsecondary levels, this instructional guide is intended to help teachers implement a laterally and vertically articulated secondary level automotive mechanics program. Introductory materials include descriptions of Automotive…

  13. A New Inverted Torsion Pendulum-Based Mechanical Spectrometer to Study Soft Matter

    Lei M.L.

    2016-03-01

    Full Text Available A new multifunctional mechanical spectrometer is developed based on an inverted torsion pendulum showing high precision in a wide frequency range to study soft matter. Apart from measuring the internal friction of solids it can also be used to study viscoelasticity. In this report we describe basic principles of the novel instrument.

  14. Location-based restoration mechanism for multi-domain GMPLS networks

    Manolova, Anna Vasileva; Calle, Eusibi; Ruepp, Sarah Renée

    2009-01-01

    In this paper we propose and evaluate the efficiency of a location-based restoration mechanism in a dynamic multi-domain GMPLS network. We focus on inter-domain link failures and utilize the correlation between the actual position of a failed link along the path with the applied restoration...

  15. Loeb's and streamer-based mechanism for negative corona current pulses

    Vagnerova, L.; Skalny, J.D.; Cermak, M.

    1998-01-01

    The negative point-to-plane corona discharge in electronegative gaseous mixtures is studied experimentally and the basic mechanisms controlling the corona phenomena are discussed. The typical shapes of the current pulse waveforms observed in experiments with the nitrogen-freon mixtures are explained in terms of the theory by Loeb and of the positive-streamer-based model. (J.U.)

  16. Mechanical, electrical and microstructural properties of cement-based materials in conditions of current flow

    Susanto, A.; Koleva, D.A.; Copuroglu, O.; Van Beek, C.; Van Breugel, K.

    2012-01-01

    Corrosion in reinforced concrete structures is not only induced by the penetration of aggressive substances (e.g. chlorides and/or CO2) but also influenced by stray currents. Further, the degradation mechanisms in reinforced cement-based systems due to the combined effect of stray current and

  17. Mechanism of Process-Induced Salt-to-Free Base Transformation of Pharmaceutical Products

    Bruun Hansen, Thomas; Qu, Haiyan

    2014-01-01

    pH-solubility profiles of a model drug in salt form was established and the mechanism of salt-to-free base form transformation was investigated by increasing pH of the system. Wet massing experiments along with suspension experiments were used to investigate the effects of excipients on the stabi...

  18. Design issues of a back-pressure-based congestion control mechanism

    Malhotra, R.; Mandjes, M.R.H.; Scheinhardt, W.R.W.; Berg, van den J.L.

    2008-01-01

    Congestion control in packet-based networks is often realized by feedback protocols -- in this paper we assess the performance under a back-pressure mechanism that has been proposed and standardized for Ethernet metropolitan networks. Relying on our earlier results for feedback fluid queues, we

  19. Strategies for adding adaptive learning mechanisms to rule-based diagnostic expert systems

    Stclair, D. C.; Sabharwal, C. L.; Bond, W. E.; Hacke, Keith

    1988-01-01

    Rule-based diagnostic expert systems can be used to perform many of the diagnostic chores necessary in today's complex space systems. These expert systems typically take a set of symptoms as input and produce diagnostic advice as output. The primary objective of such expert systems is to provide accurate and comprehensive advice which can be used to help return the space system in question to nominal operation. The development and maintenance of diagnostic expert systems is time and labor intensive since the services of both knowledge engineer(s) and domain expert(s) are required. The use of adaptive learning mechanisms to increment evaluate and refine rules promises to reduce both time and labor costs associated with such systems. This paper describes the basic adaptive learning mechanisms of strengthening, weakening, generalization, discrimination, and discovery. Next basic strategies are discussed for adding these learning mechanisms to rule-based diagnostic expert systems. These strategies support the incremental evaluation and refinement of rules in the knowledge base by comparing the set of advice given by the expert system (A) with the correct diagnosis (C). Techniques are described for selecting those rules in the in the knowledge base which should participate in adaptive learning. The strategies presented may be used with a wide variety of learning algorithms. Further, these strategies are applicable to a large number of rule-based diagnostic expert systems. They may be used to provide either immediate or deferred updating of the knowledge base.

  20. The RNA helicase DDX1 is involved in restricted HIV-1 Rev function in human astrocytes

    Fang Jianhua; Acheampong, Edward; Dave, Rajnish; Wang Fengxiang; Mukhtar, Muhammad; Pomerantz, Roger J.

    2005-01-01

    cytoplasmic, as input of exogenous DDX1 significantly altered both Rev sub-cellular localization from cytoplasmic to nuclear predominance and concomitantly increased HIV-1 viral production in these human astrocytes. We conclude that altered DDX1 expression in human astrocytes is, at least in part, responsible for the unfavorable cellular microenvironment for Rev function in these CNS-based cells. Thus, these data suggest a molecular mechanism(s) for restricted replication in astrocytes as a potential low-level site of residual HIV-1 in vivo

  1. Optimization of rhombic drive mechanism used in beta-type Stirling engine based on dimensionless analysis

    Cheng, Chin-Hsiang; Yang, Hang-Suin

    2014-01-01

    In the present study, optimization of rhombic drive mechanism used in a beta-type Stirling engine is performed based on a dimensionless theoretical model toward maximization of shaft work output. Displacements of the piston and the displacer with the rhombic drive mechanism and variations of volumes and pressure in the chambers of the engine are firstly expressed in dimensionless form. Secondly, Schmidt analysis is incorporated with Senft's shaft work theory to build a dimensionless thermodynamic model, which is employed to yield the dimensionless shaft work. The dimensionless model is verified with experimental data. It is found that the relative error between the experimental and the theoretical data in dimensionless shaft work is lower than 5.2%. This model is also employed to investigate the effects of the influential geometric parameters on the shaft work, and the optimization of these parameters is attempted. Eventually, design charts that help design the optimal geometry of the rhombic drive mechanism are presented in this report. - Highlights: • Specifically dealing with optimization of rhombic-drive mechanism used in Stirling engine based on dimensionless model. • Propose design charts that help determine the optimal geometric parameters of the rhombic drive mechanism. • Complete study of influential factors affecting the shaft work output

  2. Soft tissue deformation modelling through neural dynamics-based reaction-diffusion mechanics.

    Zhang, Jinao; Zhong, Yongmin; Gu, Chengfan

    2018-05-30

    Soft tissue deformation modelling forms the basis of development of surgical simulation, surgical planning and robotic-assisted minimally invasive surgery. This paper presents a new methodology for modelling of soft tissue deformation based on reaction-diffusion mechanics via neural dynamics. The potential energy stored in soft tissues due to a mechanical load to deform tissues away from their rest state is treated as the equivalent transmembrane potential energy, and it is distributed in the tissue masses in the manner of reaction-diffusion propagation of nonlinear electrical waves. The reaction-diffusion propagation of mechanical potential energy and nonrigid mechanics of motion are combined to model soft tissue deformation and its dynamics, both of which are further formulated as the dynamics of cellular neural networks to achieve real-time computational performance. The proposed methodology is implemented with a haptic device for interactive soft tissue deformation with force feedback. Experimental results demonstrate that the proposed methodology exhibits nonlinear force-displacement relationship for nonlinear soft tissue deformation. Homogeneous, anisotropic and heterogeneous soft tissue material properties can be modelled through the inherent physical properties of mass points. Graphical abstract Soft tissue deformation modelling with haptic feedback via neural dynamics-based reaction-diffusion mechanics.

  3. Influence of geometry on mechanical properties of bio-inspired silica-based hierarchical materials

    Dimas, Leon S; Buehler, Markus J

    2012-01-01

    Diatoms, bone, nacre and deep-sea sponges are mineralized natural structures found abundantly in nature. They exhibit mechanical properties on par with advanced engineering materials, yet their fundamental building blocks are brittle and weak. An intriguing characteristic of these structures is their heterogeneous distribution of mechanical properties. Specifically, diatoms exhibit nanoscale porosity in specific geometrical configurations to create regions with distinct stress strain responses, notably based on a single and simple building block, silica. The study reported here, using models derived from first principles based full atomistic studies with the ReaxFF reactive force field, focuses on the mechanics and deformation mechanisms of silica-based nanocomposites inspired by mineralized structures. We examine single edged notched tensile specimens and analyze stress and strain fields under varied sample size in order to gain fundamental insights into the deformation mechanisms of structures with distinct ordered arrangements of soft and stiff phases. We find that hierarchical arrangements of silica nanostructures markedly change the stress and strain transfer in the samples. The combined action of strain transfer in the deformable phase, and stress transfer in the strong phase, acts synergistically to reduce the intensity of stress concentrations around a crack tip, and renders the resulting composites less sensitive to the presence of flaws, for certain geometrical configurations it even leads to stable crack propagation. A systematic study allows us to identify composite structures with superior fracture mechanical properties relative to their constituents, akin to many natural biomineralized materials that turn the weaknesses of building blocks into a strength of the overall system. (paper)

  4. Kinetics and mechanism of thermal degradation of pentose- and hexose-based carbohydrate polymers.

    Akbar, Jamshed; Iqbal, Mohammad S; Massey, Shazma; Masih, Rashid

    2012-10-15

    This work aims at study of thermal degradation kinetics and mechanism of pentose- and hexose-based carbohydrate polymers isolated from Plantago ovata (PO), Salvia aegyptiaca (SA) and Ocimum basilicum (OB). The analysis was performed by isoconversional method. The materials exhibited mainly two-stage degradation. The weight loss at ambient-115°C characterized by low activation energy corresponds to loss of moisture. The kinetic triplets consisting of E, A and g(α) model of the materials were determined. The major degradation stage represents a loss of high boiling volatile components. This stage is exothermic in nature. Above 340°C complete degradation takes place leaving a residue of 10-15%. The master plots of g(α) function clearly differentiated the degradation mechanism of hexose-based OB and SA polymers and pentose-based PO polymer. The pentose-based carbohydrate polymer showed D(4) type and the hexose-based polymers showed A(4) type degradation mechanism. Copyright © 2012 Elsevier Ltd. All rights reserved.

  5. A Multistage Control Mechanism for Group-Based Machine-Type Communications in an LTE System

    Wen-Chien Hung

    2013-01-01

    Full Text Available When machine-type communication (MTC devices perform the long-term evolution (LTE attach procedure without bit rate limitations, they may produce congestion in the core network. To prevent this congestion, the LTE standard suggests using group-based policing to regulate the maximum bit rate of all traffic generated by a group of MTC devices. However, previous studies on the access point name-aggregate maximum bit rate based on group-based policing are relatively limited. This study proposes a multistage control (MSC mechanism to process the operations of maximum bit rate allocation based on resource-use information. For performance evaluation, this study uses a Markov chain with to analyze MTC application in a 3GPP network. Traffic flow simulations in an LTE system indicate that the MSC mechanism is an effective bandwidth allocation method in an LTE system with MTC devices. Experimental results show that the MSC mechanism achieves a throughput 22.5% higher than that of the LTE standard model using the group-based policing, and it achieves a lower delay time and greater long-term fairness as well.

  6. Low Complexity Signed Response Based Sybil Attack Detection Mechanism in Wireless Sensor Networks

    M. Saud Khan

    2016-01-01

    Full Text Available Security is always a major concern in wireless sensor networks (WSNs. Identity based attacks such as spoofing and sybil not only compromise the network but also slow down its performance. This paper proposes a low complexity sybil attack detection scheme, that is, based on signed response (SRES authentication mechanism developed for Global System for Mobile (GSM communications. A probabilistic model is presented which analyzes the proposed authentication mechanism for its probability of sybil attack. The paper also presents a simulation based comparative analysis of the existing sybil attack schemes with respect to the proposed scheme. It is observed that the proposed sybil detection scheme exhibits lesser computational cost and power consumption as compared to the existing schemes for the same sybil detection performance.

  7. MEMS-based platforms for mechanical manipulation and characterization of cells

    Pan, Peng; Wang, Wenhui; Ru, Changhai; Sun, Yu; Liu, Xinyu

    2017-12-01

    Mechanical manipulation and characterization of single cells are important experimental techniques in biological and medical research. Because of the microscale sizes and highly fragile structures of cells, conventional cell manipulation and characterization techniques are not accurate and/or efficient enough or even cannot meet the more and more demanding needs in different types of cell-based studies. To this end, novel microelectromechanical systems (MEMS)-based technologies have been developed to improve the accuracy, efficiency, and consistency of various cell manipulation and characterization tasks, and enable new types of cell research. This article summarizes existing MEMS-based platforms developed for cell mechanical manipulation and characterization, highlights their specific design considerations making them suitable for their designated tasks, and discuss their advantages and limitations. In closing, an outlook into future trends is also provided.

  8. Greenhouse gas emissions trading and project-based mechanisms. Proceedings - CATEP

    NONE

    2004-01-01

    Greenhouse gas emissions trading and project-based mechanisms for greenhouse gas reduction are emerging market-based instruments for climate change policy. This book presents a selection of papers from an international workshop co-sponsored by the OECD and Concerted Action on Tradeable Emissions Permits (CATEP), to discuss key research and policy issues relating to the design and implementation of these instruments. The papers cover the experience of developing and transition countries with greenhouse gas emissions trading and project-based mechanisms. In addition, the papers examine the use of tradeable permits in policy mixes and harmonisation of emissions trading schemes, as well as transition issues relating to greenhouse gas emissions trading markets.

  9. Development of a web-based learning medium on mechanism of labour for nursing students.

    Gerdprasert, Sailom; Pruksacheva, Tassanee; Panijpan, Bhinyo; Ruenwongsa, Pintip

    2010-07-01

    This study aimed to develop a web-based learning media on the process and mechanism of labour for the third-year university nursing and midwifery students. This media was developed based on integrating principles of the mechanism of labour with the 5Es inquiry cycle and interactive features of information technology. In this study, the web-based learning unit was used to supplement the conventional lecture as in the traditional teaching. Students' achievements were assessed by using the pre- and post-test on factual knowledge and semi-structured interviews on attitude to the unit. Supplementation with this learning unit made learning significantly more effective than the traditional lecture by itself. The students also showed positive attitude toward the learning unit. Copyright 2009 Elsevier Ltd. All rights reserved.

  10. Intrusion recognition for optic fiber vibration sensor based on the selective attention mechanism

    Xu, Haiyan; Xie, Yingjuan; Li, Min; Zhang, Zhuo; Zhang, Xuewu

    2017-11-01

    Distributed fiber-optic vibration sensors receive extensive investigation and play a significant role in the sensor panorama. A fiber optic perimeter detection system based on all-fiber interferometric sensor is proposed, through the back-end analysis, processing and intelligent identification, which can distinguish effects of different intrusion activities. In this paper, an intrusion recognition based on the auditory selective attention mechanism is proposed. Firstly, considering the time-frequency of vibration, the spectrogram is calculated. Secondly, imitating the selective attention mechanism, the color, direction and brightness map of the spectrogram is computed. Based on these maps, the feature matrix is formed after normalization. The system could recognize the intrusion activities occurred along the perimeter sensors. Experiment results show that the proposed method for the perimeter is able to differentiate intrusion signals from ambient noises. What's more, the recognition rate of the system is improved while deduced the false alarm rate, the approach is proved by large practical experiment and project.

  11. Trajectory Generation and Stability Analysis for Reconfigurable Klann Mechanism Based Walking Robot

    Jaichandar Kulandaidaasan Sheba

    2016-06-01

    Full Text Available Reconfigurable legged robots based on one degree of freedom are highly desired because they are effective on rough and irregular terrains and they provide mobility in such terrain with simple control schemes. It is necessary that reconfigurable legged robots should maintain stability during rest and motion, with a minimum number of legs while maintaining their full range of walking patterns resulting from different gait configuration. In this paper we present a method to generate input trajectory for reconfigurable quadruped robots based on Klann mechanism to properly synchronize movement. Six useful gait cycles based on this reconfigurable Klann mechanism for quadruped robots has been clearly shown here. The platform stability for these six useful gait cycles are validated through simulated results which clearly shows the capabilities of reconfigurable design.

  12. An Empirical Study of the Transmission Power Setting for Bluetooth-Based Indoor Localization Mechanisms.

    Castillo-Cara, Manuel; Lovón-Melgarejo, Jesús; Bravo-Rocca, Gusseppe; Orozco-Barbosa, Luis; García-Varea, Ismael

    2017-06-07

    Nowadays, there is a great interest in developing accurate wireless indoor localization mechanisms enabling the implementation of many consumer-oriented services. Among the many proposals, wireless indoor localization mechanisms based on the Received Signal Strength Indication (RSSI) are being widely explored. Most studies have focused on the evaluation of the capabilities of different mobile device brands and wireless network technologies. Furthermore, different parameters and algorithms have been proposed as a means of improving the accuracy of wireless-based localization mechanisms. In this paper, we focus on the tuning of the RSSI fingerprint to be used in the implementation of a Bluetooth Low Energy 4.0 (BLE4.0) Bluetooth localization mechanism. Following a holistic approach, we start by assessing the capabilities of two Bluetooth sensor/receiver devices. We then evaluate the relevance of the RSSI fingerprint reported by each BLE4.0 beacon operating at various transmission power levels using feature selection techniques. Based on our findings, we use two classification algorithms in order to improve the setting of the transmission power levels of each of the BLE4.0 beacons. Our main findings show that our proposal can greatly improve the localization accuracy by setting a custom transmission power level for each BLE4.0 beacon.

  13. An Empirical Study of the Transmission Power Setting for Bluetooth-Based Indoor Localization Mechanisms

    Manuel Castillo-Cara

    2017-06-01

    Full Text Available Nowadays, there is a great interest in developing accurate wireless indoor localization mechanisms enabling the implementation of many consumer-oriented services. Among the many proposals, wireless indoor localization mechanisms based on the Received Signal Strength Indication (RSSI are being widely explored. Most studies have focused on the evaluation of the capabilities of different mobile device brands and wireless network technologies. Furthermore, different parameters and algorithms have been proposed as a means of improving the accuracy of wireless-based localization mechanisms. In this paper, we focus on the tuning of the RSSI fingerprint to be used in the implementation of a Bluetooth Low Energy 4.0 (BLE4.0 Bluetooth localization mechanism. Following a holistic approach, we start by assessing the capabilities of two Bluetooth sensor/receiver devices. We then evaluate the relevance of the RSSI fingerprint reported by each BLE4.0 beacon operating at various transmission power levels using feature selection techniques. Based on our findings, we use two classification algorithms in order to improve the setting of the transmission power levels of each of the BLE4.0 beacons. Our main findings show that our proposal can greatly improve the localization accuracy by setting a custom transmission power level for each BLE4.0 beacon.

  14. A Gleason-Type Theorem for Any Dimension Based on a Gambling Formulation of Quantum Mechanics

    Benavoli, Alessio; Facchini, Alessandro; Zaffalon, Marco

    2017-07-01

    Based on a gambling formulation of quantum mechanics, we derive a Gleason-type theorem that holds for any dimension n of a quantum system, and in particular for n=2. The theorem states that the only logically consistent probability assignments are exactly the ones that are definable as the trace of the product of a projector and a density matrix operator. In addition, we detail the reason why dispersion-free probabilities are actually not valid, or rational, probabilities for quantum mechanics, and hence should be excluded from consideration.

  15. Dodecylamine functionalization of carbon nanotubes to improve dispersion, thermal and mechanical properties of polyethylene based nanocomposites

    Ferreira, F. V.; Franceschi, W.; Menezes, B. R. C.; Brito, F. S.; Lozano, K.; Coutinho, A. R.; Cividanes, L. S.; Thim, G. P.

    2017-07-01

    This study presents the effect of dodecylamine (DDA) functionalization of carbon nanotubes (CNTs) on the thermo-physical and mechanical properties of high-density polyethylene (HDPE) based composites. Here, we showed that the functionalization with DDA improved the dispersion of the CNTs as well as the interfacial adhesion with the HDPE matrix via non-covalent interactions. The better dispersion and interaction of CNT in the HDPE matrix as a function of the surface chemistry was correlated with the improved thermo-physical and mechanical properties.

  16. Prediction of Sliding Friction Coefficient Based on a Novel Hybrid Molecular-Mechanical Model.

    Zhang, Xiaogang; Zhang, Yali; Wang, Jianmei; Sheng, Chenxing; Li, Zhixiong

    2018-08-01

    Sliding friction is a complex phenomenon which arises from the mechanical and molecular interactions of asperities when examined in a microscale. To reveal and further understand the effects of micro scaled mechanical and molecular components of friction coefficient on overall frictional behavior, a hybrid molecular-mechanical model is developed to investigate the effects of main factors, including different loads and surface roughness values, on the sliding friction coefficient in a boundary lubrication condition. Numerical modelling was conducted using a deterministic contact model and based on the molecular-mechanical theory of friction. In the contact model, with given external loads and surface topographies, the pressure distribution, real contact area, and elastic/plastic deformation of each single asperity contact were calculated. Then asperity friction coefficient was predicted by the sum of mechanical and molecular components of friction coefficient. The mechanical component was mainly determined by the contact width and elastic/plastic deformation, and the molecular component was estimated as a function of the contact area and interfacial shear stress. Numerical results were compared with experimental results and a good agreement was obtained. The model was then used to predict friction coefficients in different operating and surface conditions. Numerical results explain why applied load has a minimum effect on the friction coefficients. They also provide insight into the effect of surface roughness on the mechanical and molecular components of friction coefficients. It is revealed that the mechanical component dominates the friction coefficient when the surface roughness is large (Rq > 0.2 μm), while the friction coefficient is mainly determined by the molecular component when the surface is relatively smooth (Rq < 0.2 μm). Furthermore, optimal roughness values for minimizing the friction coefficient are recommended.

  17. Real-time electricity pricing mechanism in China based on system dynamics

    He, Yongxiu; Zhang, Jixiang

    2015-01-01

    Highlights: • The system dynamics is used to research the real-time electricity pricing mechanism. • Four kinds of the real-time electricity pricing models are carried out and simulated. • It analysed the electricity price, the user satisfaction and the social benefits under the different models. • Market pricing is the trend of the real-time electricity pricing mechanism. • Initial development path of the real-time price mechanism for China is designed between 2015 and 2030. - Abstract: As an important means of demand-side response, the reasonable formulation of the electricity price mechanism will have an important impact on the balance between the supply and demand of electric power. With the introduction of Chinese intelligence apparatus and the rapid development of smart grids, real-time electricity pricing, as the frontier electricity pricing mechanism in the smart grid, will have great significance on the promotion of energy conservation and the improvement of the total social surplus. From the perspective of system dynamics, this paper studies different real-time electricity pricing mechanisms based on load structure, cost structure and bidding and analyses the situation of user satisfaction and the total social surplus under different pricing mechanisms. Finally, through the comparative analysis of examples under different real-time pricing scenarios, this paper aims to explore and design the future dynamic real-time electricity pricing mechanism in China, predicts the dynamic real-time pricing level and provides a reference for real-time electricity price promotion in the future

  18. Inference of Cell Mechanics in Heterogeneous Epithelial Tissue Based on Multivariate Clone Shape Quantification

    Tsuboi, Alice; Umetsu, Daiki; Kuranaga, Erina; Fujimoto, Koichi

    2017-01-01

    Cell populations in multicellular organisms show genetic and non-genetic heterogeneity, even in undifferentiated tissues of multipotent cells during development and tumorigenesis. The heterogeneity causes difference of mechanical properties, such as, cell bond tension or adhesion, at the cell–cell interface, which determine the shape of clonal population boundaries via cell sorting or mixing. The boundary shape could alter the degree of cell–cell contacts and thus influence the physiological consequences of sorting or mixing at the boundary (e.g., tumor suppression or progression), suggesting that the cell mechanics could help clarify the physiology of heterogeneous tissues. While precise inference of mechanical tension loaded at each cell–cell contacts has been extensively developed, there has been little progress on how to distinguish the population-boundary geometry and identify the cause of geometry in heterogeneous tissues. We developed a pipeline by combining multivariate analysis of clone shape with tissue mechanical simulations. We examined clones with four different genotypes within Drosophila wing imaginal discs: wild-type, tartan (trn) overexpression, hibris (hbs) overexpression, and Eph RNAi. Although the clones were previously known to exhibit smoothed or convoluted morphologies, their mechanical properties were unknown. By applying a multivariate analysis to multiple criteria used to quantify the clone shapes based on individual cell shapes, we found the optimal criteria to distinguish not only among the four genotypes, but also non-genetic heterogeneity from genetic one. The efficient segregation of clone shape enabled us to quantitatively compare experimental data with tissue mechanical simulations. As a result, we identified the mechanical basis contributed to clone shape of distinct genotypes. The present pipeline will promote the understanding of the functions of mechanical interactions in heterogeneous tissue in a non-invasive manner. PMID

  19. High temperature deformation mechanisms of L12-containing Co-based superalloys

    Titus, Michael Shaw

    Ni-based superalloys have been used as the structural material of choice for high temperature applications in gas turbine engines since the 1940s, but their operating temperature is becoming limited by their melting temperature (Tm =1300degrees C). Despite decades of research, no viable alternatives to Ni-based superalloys have been discovered and developed. However, in 2006, a ternary gamma' phase was discovered in the Co-Al-W system that enabled a new class of Co-based superalloys to be developed. These new Co-based superalloys possess a gamma-gamma' microstructure that is nearly identical to Ni-based superalloys, which enables these superalloys to achieve extraordinary high temperature mechanical properties. Furthermore, Co-based alloys possess the added benefit of exhibiting a melting temperature of at least 100degrees C higher than commercial Ni-based superalloys. Superalloys used as the structural materials in high pressure turbine blades must withstand large thermomechanical stresses imparted from the rotating disk and hot, corrosive gases present. These stresses induce time-dependent plastic deformation, which is commonly known as creep, and new superalloys must possess adequate creep resistance over a broad range of temperature in order to be used as the structural materials for high pressure turbine blades. For these reasons, this research focuses on quantifying high temperature creep properties of new gamma'-containing Co-based superalloys and identifying the high temperature creep deformation mechanisms. The high temperature creep properties of new Co- and CoNi-based alloys were found to be comparable to Ni-based superalloys with respect to minimum creep rates and creep-rupture lives at 900degrees C up to the solvus temperature of the gamma' phase. Co-based alloys exhibited a propensity for extended superlattice stacking fault formation in the gamma' precipitates resulting from dislocation shearing events. When Ni was added to the Co-based compositions

  20. Non-Watson–Crick interactions between PNA and DNA inhibit the ATPase activity of bacteriophage T4 Dda helicase

    Tackett, Alan J.; Corey, David R.; Raney, Kevin D.

    2002-01-01

    Peptide nucleic acid (PNA) is a DNA mimic in which the nucleobases are linked by an N-(2-aminoethyl) glycine backbone. Here we report that PNA can interact with single-stranded DNA (ssDNA) in a non-sequence-specific fashion. We observed that a 15mer PNA inhibited the ssDNA-stimulated ATPase activity of a bacteriophage T4 helicase, Dda. Surprisingly, when a fluorescein-labeled 15mer PNA was used in binding studies no interaction was observed between PNA and Dda. However, fluorescence polarization did reveal non-sequence-specific interactions between PNA and ssDNA. Thus, the inhibition of ATPase activity of Dda appears to result from depletion of the available ssDNA due to non-Watson–Crick binding of PNA to ssDNA. Inhibition of the ssDNA-stimulated ATPase activity was observed for several PNAs of varying length and sequence. To study the basis for this phenomenon, we examined self-aggregation by PNAs. The 15mer PNA readily self-aggregates to the point of precipitation. Since PNAs are hydrophobic, they aggregate more than DNA or RNA, making the study of this phenomenon essential for understanding the properties of PNA. Non-sequence-specific interactions between PNA and ssDNA were observed at moderate concentrations of PNA, suggesting that such interactions should be considered for antisense and antigene applications. PMID:11842106