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Sample records for integrase a4 helix

  1. An unusual helix turn helix motif in the catalytic core of HIV-1 integrase binds viral DNA and LEDGF.

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    Hayate Merad

    Full Text Available BACKGROUND: Integrase (IN of the type 1 human immunodeficiency virus (HIV-1 catalyzes the integration of viral DNA into host cellular DNA. We identified a bi-helix motif (residues 149-186 in the crystal structure of the catalytic core (CC of the IN-Phe185Lys variant that consists of the alpha(4 and alpha(5 helices connected by a 3 to 5-residue turn. The motif is embedded in a large array of interactions that stabilize the monomer and the dimer. PRINCIPAL FINDINGS: We describe the conformational and binding properties of the corresponding synthetic peptide. This displays features of the protein motif structure thanks to the mutual intramolecular interactions of the alpha(4 and alpha(5 helices that maintain the fold. The main properties are the binding to: 1- the processing-attachment site at the LTR (long terminal repeat ends of virus DNA with a K(d (dissociation constant in the sub-micromolar range; 2- the whole IN enzyme; and 3- the IN binding domain (IBD but not the IBD-Asp366Asn variant of LEDGF (lens epidermal derived growth factor lacking the essential Asp366 residue. In our motif, in contrast to the conventional HTH (helix-turn-helix, it is the N terminal helix (alpha(4 which has the role of DNA recognition helix, while the C terminal helix (alpha(5 would rather contribute to the motif stabilization by interactions with the alpha(4 helix. CONCLUSION: The motif, termed HTHi (i, for inverted emerges as a central piece of the IN structure and function. It could therefore represent an attractive target in the search for inhibitors working at the DNA-IN, IN-IN and IN-LEDGF interfaces.

  2. The HIV-1 Integrase α4-Helix Involved in LTR-DNA Recognition Is also a Highly Antigenic Peptide Element

    Science.gov (United States)

    Azzi, Sandy; Parissi, Vincent; Maroun, Richard G.; Eid, Pierre; Mauffret, Olivier; Fermandjian, Serge

    2010-01-01

    Monoclonal antibodies (MAbas) constitute remarkable tools to analyze the relationship between the structure and the function of a protein. By immunizing a mouse with a 29mer peptide (K159) formed by residues 147 to 175 of the HIV-1 integrase (IN), we obtained a monoclonal antibody (MAba4) recognizing an epitope lying in the N-terminal portion of K159 (residues 147–166 of IN). The boundaries of the epitope were determined in ELISA assays using peptide truncation and amino acid substitutions. The epitope in K159 or as a free peptide (pep-a4) was mostly a random coil in solution, while in the CCD (catalytic core domain) crystal, the homologous segment displayed an amphipathic helix structure (α4-helix) at the protein surface. Despite this conformational difference, a strong antigenic crossreactivity was observed between pep-a4 and the protein segment, as well as K156, a stabilized analogue of pep-a4 constrained into helix by seven helicogenic mutations, most of them involving hydrophobic residues. We concluded that the epitope is freely accessible to the antibody inside the protein and that its recognition by the antibody is not influenced by the conformation of its backbone and the chemistry of amino acids submitted to helicogenic mutations. In contrast, the AA →Glu mutations of the hydrophilic residues Gln148, Lys156 and Lys159, known for their interactions with LTRs (long terminal repeats) and inhibitors (5 CITEP, for instance), significantly impaired the binding of K156 to the antibody. Moreover, we found that in competition ELISAs, the processed and unprocessed LTR oligonucleotides interfered with the binding of MAba4 to IN and K156, confirming that the IN α4-helix uses common residues to interact with the DNA target and the MAba4 antibody. This also explains why, in our standard in vitro concerted integration assays, MAba4 strongly impaired the IN enzymatic activity. PMID:21209864

  3. Reversible Dimerization of Acid-Denatured ACBP Controlled by Helix A4

    DEFF Research Database (Denmark)

    Fieber, Wolfgang; Kragelund, Birthe Brandt; Meldal, Morten Peter

    2005-01-01

    denaturing conditions at pH 2.3, helix conformations are still populated in 24% of the ensemble of molecules. The structure of HA4 at atomic resolution was assessed using nuclear magnetic resonance (NMR) spectroscopy. Long-range NOEs between remote residues at opposite peptide ends suggested the formation...... of dimers and revealed a cooperative stabilization of helix A4 in this process. This emphasizes its special role in the structure formation in the denatured state of ACBP. No dimers are formed in the presence of guanidine hydrochloride, which underlines the fundamental difference between the nature...

  4. [Resistance to integrase inhibitors].

    Science.gov (United States)

    Garrido, Carolina; de Mendoza, Carmen; Soriano, Vicente

    2008-11-01

    Integrase inhibitors are the most recently approved family of antiretroviral agents for the treatment of HIV infection. As with other antiretroviral agents, under pharmacological pressure, the virus selects resistance mutations if viral suppression is incomplete. Mutations are selected in the integrase gene, specifically in positions proximal to the catalytic center. Because clinical experience with these drugs is scarce, information on resistance is limited. Virologic failure with raltegravir is associated with selection of primary mutations such as N155H (40%) and distinct changes in position Q148 (28%). Less frequently, Y143R (6.6%) and E92Q are selected. The most frequently observed mutations in failure with elvitegravir are E92Q, E138K, Q148R/K/H and N155H, and less frequently S147G and T66A/I/K. The most common resistance pattern seems to be E138K + E147G + Q148R. There is a high grade of cross resistance between raltegravir and elvitegravir, making sequencing between these two drugs impossible.

  5. Integrase and integration: biochemical activities of HIV-1 integrase

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    Deprez Eric

    2008-12-01

    Full Text Available Abstract Integration of retroviral DNA is an obligatory step of retrovirus replication because proviral DNA is the template for productive infection. Integrase, a retroviral enzyme, catalyses integration. The process of integration can be divided into two sequential reactions. The first one, named 3'-processing, corresponds to a specific endonucleolytic reaction which prepares the viral DNA extremities to be competent for the subsequent covalent insertion, named strand transfer, into the host cell genome by a trans-esterification reaction. Recently, a novel specific activity of the full length integrase was reported, in vitro, by our group for two retroviral integrases (HIV-1 and PFV-1. This activity of internal cleavage occurs at a specific palindromic sequence mimicking the LTR-LTR junction described into the 2-LTR circles which are peculiar viral DNA forms found during viral infection. Moreover, recent studies demonstrated the existence of a weak palindromic consensus found at the integration sites. Taken together, these data underline the propensity of retroviral integrases for binding symmetrical sequences and give perspectives for targeting specific sequences used for gene therapy.

  6. Pharmacology of HIV integrase inhibitors.

    Science.gov (United States)

    Adams, Jessica L; Greener, Benjamin N; Kashuba, Angela D M

    2012-09-01

    The purpose of this study is to review recent and relevant pharmacology data for three HIV integrase inhibitors: raltegravir (marketed), dolutegravir, and elvitegravir (both in phase III drug development). Data from January 2011 to April 2012 were evaluated. These data better characterized integrase inhibitor pharmacokinetics, assessed dosing regimens, and investigated previously undescribed drug-drug interactions. Due to formulation challenges, raltegravir inter-patient and intra-patient pharmacokinetic variability is high. Twice-daily 400  mg dosing has been shown to be clinically superior to 800  mg once-daily dosing. A pediatric formulation of raltegravir with less variable pharmacokinetics and greater bioavailability was US Food and Drug Administration (US FDA)-approved in December 2011. Cobicistat-boosted elvitegravir, and the second-generation integrase inhibitor dolutegravir, have lower pharmacokinetic variability and are dosed once daily. Dolutegravir drug interactions are similar to raltegravir, whereas boosted elvitegravir participates in additional CYP3A-mediated interactions. Raltegravir's potent antiretroviral activity has resulted in widespread use in both treatment-naïve and experienced patients. Dolutegravir and cobicistat-boosted elvitegravir have some pharmacokinetic advantages. Pharmacokinetic data in special populations (pregnancy, pediatrics) to optimize dosing are still required.

  7. Prediction of reorganization free energies for biological electron transfer: a comparative study of Ru-modified cytochromes and a 4-helix bundle protein.

    Science.gov (United States)

    Tipmanee, Varomyalin; Oberhofer, Harald; Park, Mina; Kim, Kwang S; Blumberger, Jochen

    2010-12-01

    The acceleration of electron transfer (ET) rates in redox proteins relative to aqueous solutes can be attributed to the protein's ability to reduce the nuclear response or reorganization upon ET, while maintaining sufficiently high electronic coupling. Quantitative predictions of reorganization free energy remain a challenge, both experimentally and computationally. Using density functional calculations and molecular dynamics simulation with an electronically polarizable force field, we report reorganization free energies for intraprotein ET in four heme-containing ET proteins that differ in their protein fold, hydrophilicity, and solvent accessibility of the electron-accepting group. The reorganization free energies for ET from the heme cofactors of cytochrome c and b(5) to solvent exposed Ru-complexes docked to histidine residues at the surface of these proteins fall within a narrow range of 1.2-1.3 eV. Reorganization free energy is significantly lowered in a designed 4-helix bundle protein where both redox active cofactors are protected from the solvent. For all ET reactions investigated, the major components of reorganization are the solvent and the protein, with the solvent contributing close to or more than 50% of the total. In three out of four proteins, the protein reorganization free energy can be viewed as a collective effect including many residues, each of which contributing a small fraction. These results have important implications for the design of artificial electron transport proteins. They suggest that reorganization free energy may in general not be effectively controlled by single point mutations, but to a large extent by the degree of solvent exposure of the ionizable cofactors.

  8. HIV-2 Integrase Variation in Integrase Inhibitor-Naïve Adults in Senegal, West Africa

    Science.gov (United States)

    Gottlieb, Geoffrey S.; Smith, Robert A.; Dia Badiane, Ndeye Mery; Ba, Selly; Hawes, Stephen E.; Toure, Macoumba; Starling, Alison K.; Traore, Fatou; Sall, Fatima; Cherne, Stephen L.; Stern, Joshua; Wong, Kim G.; Lu, Paul; Kim, Moon; Raugi, Dana N.; Lam, Airin; Mullins, James I.; Kiviat, Nancy B.

    2011-01-01

    Background Antiretroviral therapy for HIV-2 infection is hampered by intrinsic resistance to many of the drugs used to treat HIV-1. Limited studies suggest that the integrase inhibitors (INIs) raltegravir and elvitegravir have potent activity against HIV-2 in culture and in infected patients. There is a paucity of data on genotypic variation in HIV-2 integrase that might confer intrinsic or transmitted INI resistance. Methods We PCR amplified and analyzed 122 HIV-2 integrase consensus sequences from 39 HIV-2–infected, INI-naive adults in Senegal, West Africa. We assessed genetic variation and canonical mutations known to confer INI-resistance in HIV-1. Results No amino acid-altering mutations were detected at sites known to be pivotal for INI resistance in HIV-1 (integrase positions 143, 148 and 155). Polymorphisms at several other HIV-1 INI resistance-associated sites were detected at positions 72, 95, 125, 154, 165, 201, 203, and 263 of the HIV-2 integrase protein. Conclusion Emerging genotypic and phenotypic data suggest that HIV-2 is susceptible to the new class of HIV integrase inhibitors. We hypothesize that intrinsic HIV-2 integrase variation at “secondary” HIV-1 INI-resistance sites may affect the genetic barrier to HIV-2 INI resistance. Further studies will be needed to assess INI efficacy as part of combination antiretroviral therapy in HIV-2–infected patients. PMID:21765953

  9. HIV-2 integrase variation in integrase inhibitor-naïve adults in Senegal, West Africa.

    Directory of Open Access Journals (Sweden)

    Geoffrey S Gottlieb

    Full Text Available Antiretroviral therapy for HIV-2 infection is hampered by intrinsic resistance to many of the drugs used to treat HIV-1. Limited studies suggest that the integrase inhibitors (INIs raltegravir and elvitegravir have potent activity against HIV-2 in culture and in infected patients. There is a paucity of data on genotypic variation in HIV-2 integrase that might confer intrinsic or transmitted INI resistance.We PCR amplified and analyzed 122 HIV-2 integrase consensus sequences from 39 HIV-2-infected, INI-naive adults in Senegal, West Africa. We assessed genetic variation and canonical mutations known to confer INI-resistance in HIV-1.No amino acid-altering mutations were detected at sites known to be pivotal for INI resistance in HIV-1 (integrase positions 143, 148 and 155. Polymorphisms at several other HIV-1 INI resistance-associated sites were detected at positions 72, 95, 125, 154, 165, 201, 203, and 263 of the HIV-2 integrase protein.Emerging genotypic and phenotypic data suggest that HIV-2 is susceptible to the new class of HIV integrase inhibitors. We hypothesize that intrinsic HIV-2 integrase variation at "secondary" HIV-1 INI-resistance sites may affect the genetic barrier to HIV-2 INI resistance. Further studies will be needed to assess INI efficacy as part of combination antiretroviral therapy in HIV-2-infected patients.

  10. RT-173: Helix, 2017 Helix Technical Report

    Science.gov (United States)

    2018-01-16

    genders across Helix dataset...few exceptions. Each individual, again, has several characteristics, including gender , whether they are a systems engineer or a...Experience Less than 9 years Equal to or greater than 9 years Equal to or greater than 9 years Positions Title’s - 0 years titled as

  11. Integrase of Mason-Pfizer monkey virus

    Czech Academy of Sciences Publication Activity Database

    Snášel, Jan; Krejčík, Zdeněk; Jenčová, Věra; Rosenberg, Ivan; Ruml, Tomáš; Alexandratos, J.; Gustchina, A.; Pichová, Iva

    2005-01-01

    Roč. 272, č. 1 (2005), s. 203-216 ISSN 1742-464X R&D Projects: GA AV ČR(CZ) IAA4055304 Institutional research plan: CEZ:AV0Z4055905 Keywords : integrase * Mason-Pfizer monkey virus * HIV -1 Subject RIV: CE - Biochemistry

  12. Comparison of Newly Assembled Full Length HIV-1 Integrase With Prototype Foamy Virus Integrase: Structure-Function Prospective.

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    Dayer, Mohammad Reza

    2016-05-01

    Drug design against human immunodeficiency virus type 1 (HIV-1) integrase through its mechanistic study is of great interest in the area in biological research. The main obstacle in this area is the absence of the full-length crystal structure for HIV-1 integrase to be used as a model. A complete structure, similar to HIV-1 of a prototype foamy virus integrase in complex with DNA, including all conservative residues, is available and has been extensively used in recent investigations. The aim of this study was to determine whether the above model is precisely representative of HIV-1 integrase. This would critically determine the success of any designed drug using the model in deactivation of integrase and AIDS treatment. Primarily, a new structure for HIV-1 was constructed, using a crystal structure of prototype foamy virus as the starting structure. The constructed structure of HIV-1 integrase was simultaneously simulated with a prototype foamy virus integrase on a separate occasion. Our results indicate that the HIV-1 system behaves differently from the prototype foamy virus in terms of folding, hydration, hydrophobicity of binding site and stability. Based on our findings, we can conclude that HIV-1 integrase is vastly different from the prototype foamy virus integrase and does not resemble it, and the modeling output of the prototype foamy virus simulations could not be simply generalized to HIV-1 integrase. Therefore, our HIV-1 model seems to be more representative and more useful for future research.

  13. Helix mimetics: Recent developments.

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    Wilson, Andrew J

    2015-10-01

    The development of protein-protein interaction (PPIs) inhibitors represents a challenging goal in chemical biology and drug discovery. PPIs are problematic targets because they involve large surfaces with less well defined features and recognition motifs that are less amenable to conventional experimental and computational ligand discovery methodologies. α-Helix mediated PPIs represent a sub group with a clearly defined interface and thus may be more amenable to the development of generic ligand discovery methods. Indeed, this is borne out in numerous studies using peptides covalently constrained into a helical conformation resulting in improvement of myriad biophysical and cellular properties. It is however desirable to have small molecule alternatives: a helix mimetic (proteomimetic) is a generic small molecule scaffold that projects functional groups in a similar spatial orientation so as to mimic the presentation of key amino acid side chains from the helix that mediates the PPI. The first true example of a helix mimetic was described over a decade ago however this approach has not yet been elaborated to the extent that it receives similar levels of attention to constrained peptides. This review explores recent significant developments in the area of small molecule α-helix mimetics and provides a critical overview of success stories, potential limitations of the approach, and areas for future development. Copyright © 2015 Elsevier Ltd. All rights reserved.

  14. Raltegravir: first in class HIV integrase inhibitor

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    Zelalem Temesgen

    2008-06-01

    Full Text Available Zelalem Temesgen1, Dawd S Siraj21Mayo Clinic, Rochester, MN, USA; 2East Carolina University Greenville, NC, USAAbstract: On October 16, 2007, the US Food and Drug Administration (FDA approved raltegravir for treatment of human immunodeficiency virus (HIV-1 infection in combination with other antiretroviral agents in treatment-experienced adult patients who have evidence of viral replication and HIV-1 strains resistant to multiple antiretroviral agents. Raltegravir is first in a novel class of antiretroviral drugs known as integrase inhibitors. It has demonstrated potent anti HIV activity in both antiretroviral treatment-naïve and experienced patients. The most common adverse events reported with raltegravir during phase 2 and 3 clinical trials were diarrhea, nausea, and headache. Laboratory abnormalities include mild elevations in liver transaminases and creatine phosphokinase.Keywords: raltegravir, HIV, antiretroviral agents, integrase inhibitors

  15. Cloning and Biochemical Characterization of HIV Integrase

    Science.gov (United States)

    1989-12-15

    biochemical assays for the interaction of the HIV Intebration protein, IN, with a specific DNA target, the viral LTRs. The gene encoding IN has been subcloned ...FILE COPY CO CONTRACT NO.: DAMD7-88-C-8126 r TITLE: CLONING AND BIOCHEMICAL CHARACTERIZATION OF HIV INTEGRASE I- SPRINCIPAL INVESTIGATOR: ELLEN...ELEMENT No. NO. 392- NO. IACCESSION NO. 1 . TITLE (indude Security Classification)630A I315D49C08 (U) Cloning and Biochemical Characterization of the HIV

  16. HIV-1 integrase inhibitors as new components of antiviral therapy

    Science.gov (United States)

    Prikazchikova, T. A.; Sycheva, A. M.; Agapkina, Y. Y.; Aleksandrov, D. A.; Gottikh, M. B.

    2008-05-01

    Structural and functional features of HIV-1 integrase are considered and the state of the art in the quest for effective inhibitors of this enzyme is reported. The major classes of integrase inhibitors with known mechanisms of action as well as their in vitro and in vivo inhibitory activities are presented.

  17. HIV-1 integrase inhibitors as new components of antiviral therapy

    Energy Technology Data Exchange (ETDEWEB)

    Prikazchikova, T A; Aleksandrov, D A; Gottikh, M B [A. N. Belozersky Institute of Physico-Chemical Biology, M. V. Lomonosov Moscow State University, Moscow (Russian Federation); Sycheva, A M; Agapkina, Y Y [Department of Chemistry, M.V. Lomonosov Moscow State University, Moscow (Russian Federation)

    2008-05-31

    Structural and functional features of HIV-1 integrase are considered and the state of the art in the quest for effective inhibitors of this enzyme is reported. The major classes of integrase inhibitors with known mechanisms of action as well as their in vitro and in vivo inhibitory activities are presented.

  18. Primary resistance to integrase strand-transfer inhibitors in Europe

    DEFF Research Database (Denmark)

    Casadellà, M; van Ham, P M; Noguera-Julian, M

    2015-01-01

    OBJECTIVES: The objective of this study was to define the natural genotypic variation of the HIV-1 integrase gene across Europe for epidemiological surveillance of integrase strand-transfer inhibitor (InSTI) resistance. METHODS: This was a multicentre, cross-sectional study within the European...... SPREAD HIV resistance surveillance programme. A representative set of 300 samples was selected from 1950 naive HIV-positive subjects newly diagnosed in 2006-07. The prevalence of InSTI resistance was evaluated using quality-controlled baseline population sequencing of integrase. Signature raltegravir......, elvitegravir and dolutegravir resistance mutations were defined according to the IAS-USA 2014 list. In addition, all integrase substitutions relative to HXB2 were identified, including those with a Stanford HIVdb score ≥10 to at least one InSTI. To rule out circulation of minority InSTI-resistant HIV, 65...

  19. On the helix equation

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    Taouil Hajer

    2012-08-01

    Full Text Available This paper is devoted to the helices processes, i.e. the solutions H : ℝ × Ω → ℝd, (t, ω ↦ H(t, ω of the helix equation egin{eqnarray} H(0,o=0; quad H(s+t,o= H(s,Phi(t,o +H(t,oonumber end{eqnarray} H ( 0 ,ω = 0 ;   H ( s + t,ω = H ( s, Φ ( t,ω + H ( t,ω where Φ : ℝ × Ω → Ω, (t, ω ↦ Φ(t, ω is a dynamical system on a measurable space (Ω, ℱ. More precisely, we investigate dominated solutions and non differentiable solutions of the helix equation. For the last case, the Wiener helix plays a fundamental role. Moreover, some relations with the cocycle equation defined by Φ, are investigated. Ce papier est consacré aux hélices, c’est-à-dire les solutions H : ℝ × Ω → ℝd, (t, ω ↦ H(t, ω de l’équation fonctionnelle egin{eqnarray} H(0,o=0; quad H(s+t,o= H(s,Phi(t,o +H(t,o onumber end{eqnarray} H ( 0 ,ω = 0 ;   H ( s + t,ω = H ( s, Φ ( t,ω + H ( t,ω où Φ : ℝ × Ω → Ω, (t, ω ↦ Φ(t, ω est un système dynamique défini sur un espace mesurable (Ω, ℱ. Plus présisément, nous déterminons d’abord les hélices dominées puis nous caractérisons les hélices non différentiables. Dans ce dernier cas, l’hélice de Wiener joue un rôle important. Nous précisons aussi quelques relations des hélices avec les cocycles définis par Φ.

  20. Complementation between HIV integrase proteins mutated in different domains

    NARCIS (Netherlands)

    D.C. van Gent (Dik); C. Vink (Cornelis); A.A. Groeneger; R.H. Plassterk

    1993-01-01

    textabstractHIV integrase (IN) cleaves two nucleotides off the 3' end of viral DNA and integrates viral DNA into target DNA. Previously, three functional domains in the HIV IN protein have been identified: (i) the central catalytic domain, (ii) the C-terminal DNA binding domain,

  1. An Algorithm for Protein Helix Assignment Using Helix Geometry.

    Science.gov (United States)

    Cao, Chen; Xu, Shutan; Wang, Lincong

    2015-01-01

    Helices are one of the most common and were among the earliest recognized secondary structure elements in proteins. The assignment of helices in a protein underlies the analysis of its structure and function. Though the mathematical expression for a helical curve is simple, no previous assignment programs have used a genuine helical curve as a model for helix assignment. In this paper we present a two-step assignment algorithm. The first step searches for a series of bona fide helical curves each one best fits the coordinates of four successive backbone Cα atoms. The second step uses the best fit helical curves as input to make helix assignment. The application to the protein structures in the PDB (protein data bank) proves that the algorithm is able to assign accurately not only regular α-helix but also 310 and π helices as well as their left-handed versions. One salient feature of the algorithm is that the assigned helices are structurally more uniform than those by the previous programs. The structural uniformity should be useful for protein structure classification and prediction while the accurate assignment of a helix to a particular type underlies structure-function relationship in proteins.

  2. HIV-1 group O integrase displays lower enzymatic efficiency and higher susceptibility to raltegravir than HIV-1 group M subtype B integrase.

    Science.gov (United States)

    Depatureaux, Agnès; Quashie, Peter K; Mesplède, Thibault; Han, Yingshan; Koubi, Hannah; Plantier, Jean-Christophe; Oliveira, Maureen; Moisi, Daniela; Brenner, Bluma; Wainberg, Mark A

    2014-12-01

    HIV-1 group O (HIV-O) is a rare HIV-1 variant characterized by a high number of polymorphisms, especially in the integrase coding region. As HIV-O integrase enzymes have not previously been studied, our aim was to assess the impact of HIV-O integrase polymorphisms on enzyme function and susceptibility to integrase inhibitors. Accordingly, we cloned and purified integrase proteins from each of HIV-1 group O clades A and B, an HIV-O divergent strain, and HIV-1 group M (HIV-M, subtype B), used as a reference. To assess enzymatic function of HIV-O integrase, we carried out strand transfer and 3' processing assays with various concentrations of substrate (DNA target and long terminal repeats [LTR], respectively) and characterized these enzymes for susceptibility to integrase strand transfer inhibitors (INSTIs) in cell-free assays and in tissue culture, in the absence or presence of various concentrations of several INSTIs. The inhibition constant (Ki) and 50% effective concentration (EC50) values were calculated for HIV-O integrases and HIV-O viruses, respectively, and compared with those of HIV-M. The results showed that HIV-O integrase displayed lower activity in strand transfer assays than did HIV-M enzyme, whereas 3' processing activities were similar to those of HIV-M. HIV-O integrases were more susceptible to raltegravir (RAL) in competitive inhibition assays and in tissue culture than were HIV-M enzymes and viruses, respectively. Molecular modeling suggests that two key polymorphic residues that are close to the integrase catalytic site, 74I and 153A, may play a role in these differences. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  3. Comparative Clinical Pharmacokinetics and Pharmacodynamics of HIV-1 Integrase Strand Transfer Inhibitors.

    Science.gov (United States)

    Podany, Anthony T; Scarsi, Kimberly K; Fletcher, Courtney V

    2017-01-01

    Dolutegravir (DTG), elvitegravir (EVG) and raltegravir (RAL) are members of the latest class of antiretrovirals (ARVs) that have become available to treat human immunodeficiency virus (HIV) infection: integrase strand transfer inhibitors (INSTIs). INSTIs are potent inhibitors of the HIV integrase enzyme, with protein binding-adjusted concentration inhibiting viral replication by 90/95 % [IC 90/95 ] values in the low nanogram per millilitre range, and they retain antiviral activity against strains of HIV with acquired resistance to other classes of ARVs. Each of the INSTIs has unique pharmacokinetic/pharmacodynamic properties, influencing its role in clinical use in specific subsets of patients. RAL and DTG have minimal drug-drug interaction profiles, as their metabolism has minimal cytochrome P450 (CYP) involvement. Conversely, EVG metabolism occurs primarily via CYP3A4 and requires pharmacokinetic boosting to achieve systemic exposures amenable to once-daily dosing. EVG and DTG have the added benefit of availability of fixed-dose combination tablets, allowing for convenient and simplified ARV regimens. RAL is the only INSTI to be listed as a preferred agent in the current US perinatal treatment guidelines. All three INSTIs are recommended regimens for treatment-naïve individuals in the US adult and adolescent HIV treatment guidelines. This review summarizes and compares the pharmacokinetics and pharmacodynamics of the INSTIs, and describes specific pharmacokinetic considerations for special patient conditions: hepatic impairment, renal dysfunction, pregnancy and co-infections.

  4. Prophage Integrase Typing Is a Useful Indicator of Genomic Diversity in Salmonella enterica

    Science.gov (United States)

    Colavecchio, Anna; D’Souza, Yasmin; Tompkins, Elizabeth; Jeukens, Julie; Freschi, Luca; Emond-Rheault, Jean-Guillaume; Kukavica-Ibrulj, Irena; Boyle, Brian; Bekal, Sadjia; Tamber, Sandeep; Levesque, Roger C.; Goodridge, Lawrence D.

    2017-01-01

    Salmonella enterica is a bacterial species that is a major cause of illness in humans and food-producing animals. S. enterica exhibits considerable inter-serovar diversity, as evidenced by the large number of host adapted serovars that have been identified. The development of methods to assess genome diversity in S. enterica will help to further define the limits of diversity in this foodborne pathogen. Thus, we evaluated a PCR assay, which targets prophage integrase genes, as a rapid method to investigate S. enterica genome diversity. To evaluate the PCR prophage integrase assay, 49 isolates of S. enterica were selected, including 19 clinical isolates from clonal serovars (Enteritidis and Heidelberg) that commonly cause human illness, and 30 isolates from food-associated Salmonella serovars that rarely cause human illness. The number of integrase genes identified by the PCR assay was compared to the number of integrase genes within intact prophages identified by whole genome sequencing and phage finding program PHASTER. The PCR assay identified a total of 147 prophage integrase genes within the 49 S. enterica genomes (79 integrase genes in the food-associated Salmonella isolates, 50 integrase genes in S. Enteritidis, and 18 integrase genes in S. Heidelberg). In comparison, whole genome sequencing and PHASTER identified a total of 75 prophage integrase genes within 102 intact prophages in the 49 S. enterica genomes (44 integrase genes in the food-associated Salmonella isolates, 21 integrase genes in S. Enteritidis, and 9 integrase genes in S. Heidelberg). Collectively, both the PCR assay and PHASTER identified the presence of a large diversity of prophage integrase genes in the food-associated isolates compared to the clinical isolates, thus indicating a high degree of diversity in the food-associated isolates, and confirming the clonal nature of S. Enteritidis and S. Heidelberg. Moreover, PHASTER revealed a diversity of 29 different types of prophages and 23

  5. Developing a Dynamic Pharmacophore Model for HIV-1 Integrase

    International Nuclear Information System (INIS)

    Carlson, Heather A.; Masukawa, Keven M.; Rubins, Kathleen; Bushman, Frederic; Jorgensen, William L.; Lins, Roberto; Briggs, James; Mccammon, Andy

    2000-01-01

    We present the first receptor-based pharmacophore model for HIV-1 integrase. The development of ''dynamic'' pharmacophore models is a new method that accounts for the inherent flexibility of the active site and aims to reduce the entropic penalties associated with binding a ligand. Furthermore, this new drug discovery method overcomes the limitation of an incomplete crystal structure of the target protein. A molecular dynamics (MD) simulation describes the flexibility of the uncomplexed protein. Many conformational models of the protein are saved from the MD simulations and used in a series of multi-unit search for interacting conformers (MUSIC) simulations. MUSIC is a multiple-copy minimization method, available in the BOSS program; it is used to determine binding regions for probe molecules containing functional groups that complement the active site. All protein conformations from the MD are overlaid, and conserved binding regions for the probe molecules are identified. Those conserved binding regions define the dynamic pharmacophore model. Here, the dynamic model is compared to known inhibitors of the integrase as well as a three-point, ligand-based pharmacophore model from the literature. Also, a ''static'' pharmacophore model was determined in the standard fashion, using a single crystal structure. Inhibitors thought to bind in the active site of HIV-1 integrase fit the dynamic model but not the static model. Finally, we have identified a set of compounds from the Available Chemicals Directory that fit the dynamic pharmacophore model, and experimental testing of the compounds has confirmed several new inhibitors

  6. Implementation and application of helix-helix distance and crossing angle restraint potentials.

    Science.gov (United States)

    Lee, Jinhyuk; Im, Wonpil

    2007-02-01

    Based on the definition of helix-helix distance and crossing angle introduced by Chothia et al. (J Mol Biol 1981, 145, 215), we have developed the restraint potentials by which the distance and crossing angle of two selected helices can be maintained around target values during molecular dynamics simulations. A series of assessments show that calculated restraint forces are numerically accurate. Since the restraint forces are only exerted on atoms which define the helical principal axes, each helix can rotate along its helical axis, depending on the helix-helix intermolecular interactions. Such a restraint potential enables us to characterize the helix-helix interactions at atomic details by sampling their conformational space around specific distance and crossing angle with (restraint) force-dependent fluctuations. Its efficacy is illustrated by calculating the potential of mean force as a function of helix-helix distance between two transmembrane helical peptides in an implicit membrane model.

  7. Multiple helix ecosystems for sustainable competitiveness

    CERN Document Server

    Ferreira, João; Farinha, Luís; Fernandes, Nuno

    2016-01-01

    This book discusses the main issues, challenges, opportunities, and trends involving the interactions between academia, industry, government and society. Specifically, it aims to explore how these interactions enhance the ways in which companies deliver products and services in order to achieve sustainable competitiveness in the marketplace. Sustainable competitiveness has been widely discussed by academics and practitioners, considering the importance of protecting the environment while sustaining the economic goals of organizations. The Quintuple Helix innovation model is a framework for facilitating knowledge, innovation and sustainable competitive advantage. It embeds the Triple and the Quadruple Helix models by adding a fifth helix, the “natural environment.” The Triple Helix model focuses on the university-industry-government triad, while the Quadruple adds civil society (the media- and culture-driven public) as a fourth helix. The Quintuple Helix model facilitates research, public policy, and pract...

  8. Inhibition of HIV-1 Integrase gene expression by 10-23 DNAzyme

    Indian Academy of Sciences (India)

    We have designed three novel DNAzymes, DIN54, DIN116, and DIN152, against HIV-1 Integrase gene using Mfold software and evaluated them for target site cleavage activity on the in vitro transcribed mRNA. All DNAzymes were tested for its inhibition of expression of HIV Integrase protein in the transiently transfected cell ...

  9. Kevlar: Transitioning Helix from Research to Practice

    Science.gov (United States)

    2015-04-01

    KEVLAR : TRANSITIONING HELIX FROM RESEARCH TO PRACTICE UNIVERSITY OF VIRGINIA APRIL 2015 FINAL TECHNICAL REPORT...DATES COVERED (From - To) FEB 2013 – NOV 2014 4. TITLE AND SUBTITLE KEVLAR : TRANSITIONING HELIX FROM RESEARCH TO PRACTICE 5a. CONTRACT NUMBER...possible exploitation. Our technology, called Kevlar , includes key security technologies are protective transformations and targeted recovery. The

  10. Kevlar: Transitioning Helix for Research to Practice

    Science.gov (United States)

    2016-03-01

    KEVLAR : TRANSITIONING HELIX FROM RESEARCH TO PRACTICE UNIVERSITY OF VIRGINIA MARCH 2016 FINAL TECHNICAL REPORT...REPORT 3. DATES COVERED (From - To) FEB 2015 – SEP 2015 4. TITLE AND SUBTITLE KEVLAR : TRANSITIONING HELIX FROM RESEARCH TO PRACTICE 5a. CONTRACT...subject to possible exploitation. Our technology, called Kevlar , includes key security technologies are protective transformations and targeted

  11. Double Helix Nodal Line Superconductor

    Science.gov (United States)

    Sun, Xiao-Qi; Lian, Biao; Zhang, Shou-Cheng

    2017-10-01

    Time-reversal invariant superconductors in three dimensions may contain nodal lines in the Brillouin zone, which behave as Wilson loops of 3D momentum-space Chern-Simons theory of the Berry connection. Here we study the conditions of realizing linked nodal lines (Wilson loops), which yield a topological contribution to the thermal magnetoelectric coefficient that is given by the Chern-Simons action. We find the essential conditions are the existence of torus or higher genus Fermi surfaces and spiral spin textures. We construct such a model with two torus Fermi surfaces, where a generic spin-dependent interaction leads to double-helix-like linked nodal lines as the superconductivity is developed.

  12. Structure-Guided Optimization of HIV Integrase Strand Transfer Inhibitors.

    Science.gov (United States)

    Zhao, Xue Zhi; Smith, Steven J; Maskell, Daniel P; Métifiot, Mathieu; Pye, Valerie E; Fesen, Katherine; Marchand, Christophe; Pommier, Yves; Cherepanov, Peter; Hughes, Stephen H; Burke, Terrence R

    2017-09-14

    Integrase mutations can reduce the effectiveness of the first-generation FDA-approved integrase strand transfer inhibitors (INSTIs), raltegravir (RAL) and elvitegravir (EVG). The second-generation agent, dolutegravir (DTG), has enjoyed considerable clinical success; however, resistance-causing mutations that diminish the efficacy of DTG have appeared. Our current findings support and extend the substrate envelope concept that broadly effective INSTIs can be designed by filling the envelope defined by the DNA substrates. Previously, we explored 1-hydroxy-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamides as an INSTI scaffold, making a limited set of derivatives, and concluded that broadly effective INSTIs can be developed using this scaffold. Herein, we report an extended investigation of 6-substituents as well the first examples of 7-substituted analogues of this scaffold. While 7-substituents are not well-tolerated, we have identified novel substituents at the 6-position that are highly effective, with the best compound (6p) retaining better efficacy against a broad panel of known INSTI resistant mutants than any analogues we have previously described.

  13. Serine integrase chimeras with activity in E. coli and HeLa cells

    Directory of Open Access Journals (Sweden)

    Alfonso P. Farruggio

    2014-09-01

    Full Text Available In recent years, application of serine integrases for genomic engineering has increased in popularity. The factor-independence and unidirectionality of these large serine recombinases makes them well suited for reactions such as site-directed vector integration and cassette exchange in a wide variety of organisms. In order to generate information that might be useful for altering the specificity of serine integrases and to improve their efficiency, we tested a hybridization strategy that has been successful with several small serine recombinases. We created chimeras derived from three characterized members of the serine integrase family, phiC31, phiBT1, and TG1 integrases, by joining their amino- and carboxy-terminal portions. We found that several phiBT1-phiC31 (BC and phiC31-TG1 (CT hybrid integrases are active in E. coli. BC chimeras function on native att-sites and on att-sites that are hybrids between those of the two donor enzymes, while CT chimeras only act on the latter att-sites. A BC hybrid, BC{−1}, was also active in human HeLa cells. Our work is the first to demonstrate chimeric serine integrase activity. This analysis sheds light on integrase structure and function, and establishes a potentially tractable means to probe the specificity of the thousands of putative large serine recombinases that have been revealed by bioinformatics studies.

  14. Clinical use of HIV integrase inhibitors: a systematic review and meta-analysis.

    Directory of Open Access Journals (Sweden)

    Peter Messiaen

    Full Text Available BACKGROUND: Optimal regimen choice of antiretroviral therapy is essential to achieve long-term clinical success. Integrase inhibitors have swiftly been adopted as part of current antiretroviral regimens. The purpose of this study was to review the evidence for integrase inhibitor use in clinical settings. METHODS: MEDLINE and Web-of-Science were screened from April 2006 until November 2012, as were hand-searched scientific meeting proceedings. Multiple reviewers independently screened 1323 citations in duplicate to identify randomized controlled trials, nonrandomized controlled trials and cohort studies on integrase inhibitor use in clinical practice. Independent, duplicate data extraction and quality assessment were conducted. RESULTS: 48 unique studies were included on the use of integrase inhibitors in antiretroviral therapy-naive patients and treatment-experienced patients with either virological failure or switching to integrase inhibitors while virologically suppressed. On the selected studies with comparable outcome measures and indication (n = 16, a meta-analysis was performed based on modified intention-to-treat (mITT, on-treatment (OT and as-treated (AT virological outcome data. In therapy-naive patients, favorable odds ratios (OR for integrase inhibitor-based regimens were observed, (mITT OR 0.71, 95% CI 0.59-0.86. However, integrase inhibitors combined with protease inhibitors only did not result in a significant better virological outcome. Evidence further supported integrase inhibitor use following virological failure (mITT OR 0.27; 95% CI 0.11-0.66, but switching to integrase inhibitors from a high genetic barrier drug during successful treatment was not supported (mITT OR 1.43; 95% CI 0.89-2.31. Integrase inhibitor-based regimens result in similar immunological responses compared to other regimens. A low genetic barrier to drug-resistance development was observed for raltegravir and elvitegravir, but not for dolutegravir

  15. Helix-Hopes on Finite Hyperfields

    Directory of Open Access Journals (Sweden)

    Thomas Vougiouklis

    2016-12-01

    Full Text Available Hyperstructure theory can overcome restrictions which ordinary algebraic structures have. A hyperproduct on non-square ordinary matrices can be defined by using the so called helix-hyperoperations. We study the helix-hyperstructures on the representations using ordinary fields. The related theory can be faced by defining the hyperproduct on the set of non square matrices. The main tools of the Hyperstructure Theory are the fundamental relations which connect the largest class of hyperstructures, the Hv-structures, with the corresponding classical ones. We focus on finite dimensional helix-hyperstructures and on small Hv-fields, as well.

  16. Insight into the ERVK integrase – propensity for DNA damage

    Directory of Open Access Journals (Sweden)

    Samantha Bray

    2016-12-01

    Full Text Available Retroviruses create permanently integrated proviruses that exist in the host genome. Retroviral genomes encode for functionally conserved gag, pro, pol and env regions, as well as integrase (IN, which is required for retroviral integration. IN mediates viral genome insertion through 3´ end processing of the viral DNA and the strand transfer reaction. This process requires the formation of a pre-integration complex, comprised of IN, viral DNA and cellular proteins. Viral insertion causes DNA damage, leading to the requirement of host DNA repair mechanisms. Therefore, a failure of DNA repair pathways may result in genomic instability and potentially cause host cell death. Considering the numerous human diseases associated with genomic instability, the endogenous retrovirus-K (ERVK IN should be considered as a putative contributor to DNA damage in human cells. Future research and drug discovery should focus on ERVK IN activity and its role in human conditions, such as neurological disease and cancers.

  17. QSAR study of curcumine derivatives as HIV-1 integrase inhibitors.

    Science.gov (United States)

    Gupta, Pawan; Sharma, Anju; Garg, Prabha; Roy, Nilanjan

    2013-03-01

    A QSAR study was performed on curcumine derivatives as HIV-1 integrase inhibitors using multiple linear regression. The statistically significant model was developed with squared correlation coefficients (r(2)) 0.891 and cross validated r(2) (r(2) cv) 0.825. The developed model revealed that electronic, shape, size, geometry, substitution's information and hydrophilicity were important atomic properties for determining the inhibitory activity of these molecules. The model was also tested successfully for external validation (r(2) pred = 0.849) as well as Tropsha's test for model predictability. Furthermore, the domain analysis was carried out to evaluate the prediction reliability of external set molecules. The model was statistically robust and had good predictive power which can be successfully utilized for screening of new molecules.

  18. Sketching the historical development of pyrimidones as the inhibitors of the HIV integrase

    Czech Academy of Sciences Publication Activity Database

    Patel, Rahul V.; Keum, Y.S.; Park, S.W.

    2015-01-01

    Roč. 97, JUN 5 (2015), s. 649-663 ISSN 0223-5234 Institutional support: RVO:61389030 Keywords : Pyrimidones * Anti- HIV * Integrase inhibitors Subject RIV: CE - Biochemistry Impact factor: 3.902, year: 2015

  19. Teaching helix and problems connected with helix using GeoGebra

    Science.gov (United States)

    Bímová, Daniela

    2017-12-01

    The contribution presents the dynamic applets created in GeoGebra that show the origin and main properties of a helix and it also presents some constructive problems connected with the helix. There are created the step by step algorithms of some constructions in the chosen applets. Three-dimensional applets include illustrative helix samples and spatial animations that help students better see problems concerning the helix spatially. There is mentioned the website in the contribution on which there is situated GeoGebra book dedicated to the topic "Helix" and containing the mentioned applets. The created applets and materials of the GeoGebra book "Helix" help in teaching and studying the course Constructive Geometry determined for the students of the Faculty of Mechanical Engineering of the Technical University of Liberec.

  20. Small molecule inhibitors of the LEDGF site of human immunodeficiency virus integrase identified by fragment screening and structure based design.

    Directory of Open Access Journals (Sweden)

    Thomas S Peat

    Full Text Available A fragment-based screen against human immunodeficiency virus type 1 (HIV integrase led to a number of compounds that bound to the lens epithelium derived growth factor (LEDGF binding site of the integrase catalytic core domain. We determined the crystallographic structures of complexes of the HIV integrase catalytic core domain for 10 of these compounds and quantitated the binding by surface plasmon resonance. We demonstrate that the compounds inhibit the interaction of LEDGF with HIV integrase in a proximity AlphaScreen assay, an assay for the LEDGF enhancement of HIV integrase strand transfer and in a cell based assay. The compounds identified represent a potential framework for the development of a new series of HIV integrase inhibitors that do not bind to the catalytic site of the enzyme.

  1. Transmembrane Helix-helix Association: Relative Stabilities at Low pH†

    Science.gov (United States)

    Valluru, Neelima; Silva, Frances; Dhage, Manmath; Rodriguez, Gustavo; Alloor, Srinivas R.; Renthal, Robert

    2008-01-01

    We have previously studied the unfolding equilibrium of bacterio-opsin in a single phase solvent, using Förster mechanism fluorescence resonance energy transfer (FRET) as a probe, from tryptophan donors to a dansyl acceptor. We observed an apparent unfolding transition in bacterio-opsin perturbed by increasing ethanol concentrations [Nannepaga et al.(2004) Biochemistry 43, 50–59]. We have further investigated this transition and find the unfolding is pH-dependent. We have now measured the apparent pK of acid-induced unfolding of bacterio-opsin in 90% ethanol. When the acceptor is on helix B (Lys 41), the apparent pK for unfolding is 4.75; on the EF connecting loop (Cys 163), 5.15; and on helix G (Cys 222), 5.75. Five-helix proteolytic fragments are less stable. The apparent unfolding pKs are, for residues 72-248 (Cys 163), 5.46; and 1-166 (Lys 41), 7.36. When interpreted in terms of a simple equilibrium model for unfolding, the apparent pKs give relative free energies of unfolding, in the range of −0.54 to −3.5 kcal/mol. The results suggest that the C-terminal helix of bacterio-opsin is less stably folded than the N-terminal helices. We analyzed the pair-wise helix-helix interaction surfaces of bacteriorhodopsin and three other 7-transmembrane helix proteins, based on crystal structures. The results show that the interaction surfaces are smoother and the helix axis separations are closer in the amino-terminal two-thirds of the proteins compared with the carboxyl terminal one-third. However, the F helix is important in stabilizing the folded structure, as shown by the instability of the 1-166 fragment. Considering the high resolution crystal structure of bacteriorhodopsin, there are no obvious helix-helix interactions involving protein side chains which would be destabilized by protonation at the estimated pH of the unfolding transitions. However, a number of helix-bridging water molecules could become protonated, thereby weakening the helix-helix

  2. Transmembrane helix-helix association: relative stabilities at low pH.

    Science.gov (United States)

    Valluru, Neelima; Silva, Frances; Dhage, Manmath; Rodriguez, Gustavo; Alloor, Srinivas R; Renthal, Robert

    2006-04-11

    We have previously studied the unfolding equilibrium of bacterioopsin in a single phase solvent, using Förster mechanism fluorescence resonance energy transfer (FRET) as a probe, from tryptophan donors to a dansyl acceptor. We observed an apparent unfolding transition in bacterioopsin perturbed by increasing ethanol concentrations [Nannepaga et al. (2004) Biochemistry 43, 50-59]. We have further investigated this transition and find that the unfolding is pH-dependent. We have now measured the apparent pK of acid-induced unfolding of bacterioopsin in 90% ethanol. When the acceptor is on helix B (Lys 41), the apparent pK for unfolding is 4.75; on the EF connecting loop (Cys 163), 5.15; and on helix G (Cys 222), 5.75. Five-helix proteolytic fragments are less stable. The apparent unfolding pKs are 5.46 for residues 72-248 (Cys 163) and 7.36 for residues 1-166 (Lys 41). When interpreted in terms of a simple equilibrium model for unfolding, the apparent pKs give relative free energies of unfolding in the range of -0.54 to -3.5 kcal/mol. The results suggest that the C-terminal helix of bacterioopsin is less stably folded than the N-terminal helices. We analyzed the pairwise helix-helix interaction surfaces of bacteriorhodopsin and three other seven-transmembrane-helix proteins on the basis of crystal structures. The results show that the interaction surfaces are smoother and the helix axis separations are closer in the amino-terminal two-thirds of the proteins compared with the carboxyl-terminal one-third. However, the F helix is important in stabilizing the folded structure, as shown by the instability of the 1-166 fragment. Considering the high-resolution crystal structure of bacteriorhodopsin, there are no obvious helix-helix interactions involving protein side chains which would be destabilized by protonation at the estimated pH of the unfolding transitions. However, a number of helix-bridging water molecules could become protonated, thereby weakening the helix-helix

  3. The swimming of a perfect deforming helix

    Science.gov (United States)

    Koens, Lyndon; Zhang, Hang; Mourran, Ahmed; Lauga, Eric

    2017-11-01

    Many bacteria rotate helical flagellar filaments in order to swim. When at rest or rotated counter-clockwise these flagella are left handed helices but they undergo polymorphic transformations to right-handed helices when the motor is reversed. These helical deformations themselves can generate motion, with for example Rhodobacter sphaeroides using the polymorphic transformation of the flagellum to generate rotation, or Spiroplasma propagating a change of helix handedness across its body's length to generate forward motion. Recent experiments reported on an artificial helical microswimmer generating motion without a propagating change in handedness. Made of a temperature sensitive gel, these swimmers moved by changing the dimensions of the helix in a non-reciprocal way. Inspired by these results and helix's ubiquitous presence in the bacterial world, we investigate how a deforming helix moves within a viscous fluid. Maintaining a single handedness along its entire length, we discuss how a perfect deforming helix can create a non-reciprocal swimming stroke, identify its principle directions of motion, and calculate the swimming kinematics asymptotically.

  4. Nanoparticle biofabrication using English ivy (Hedera helix

    Directory of Open Access Journals (Sweden)

    Burris Jason N

    2012-10-01

    Full Text Available Abstract Background English ivy (Hedera helix is well known for its adhesive properties and climbing ability. Essential to its ability to adhere to vertical surfaces is the secretion of a nanocomposite adhesive containing spherical nanoparticles, 60–85 nm in diameter, produced exclusively by root hairs present on adventitious roots. These organic nanoparticles have shown promise in biomedical and cosmetic applications, and represent a safer alternative to metal oxide nanoparticles currently available. Results It was discovered that the maximum adventitious root production was achieved by a 4 h application of 1 mg/ml indole-3 butyric acid (IBA to juvenile English ivy shoot segments cultured in custom vessels. After incubation of the shoots under continuous light at 83 μmol/m2 s at 20°C for 2 weeks, the adventitious roots were harvested from the culture system and it was possible to isolate 90 mg of dry weight nanoparticles per 12 g of roots. The nanoparticle morphology was characterized by atomic force microscopy, and found to be similar to previous studies. Conclusions An enhanced system for the production of English ivy adventitious roots and their nanoparticles by modifying GA7 Magenta boxes and identifying the optimal concentration of IBA for adventitious root growth was developed. This system is the first such platform for growing and harvesting organic nanoparticles from plants, and represents an important step in the development of plant-based nanomanufacturing. It is a significant improvement on the exploitation of plant systems for the formation of metallic nanoparticles, and represents a pathway for the generation of bulk ivy nanoparticles for translation into biomedical applications.

  5. X-ray crystal structure of the N-terminal region of Moloney murine leukemia virus integrase and its implications for viral DNA recognition: N-Terminal Region of M-MuLV Integrase

    Energy Technology Data Exchange (ETDEWEB)

    Guan, Rongjin [Center for Advanced Biotechnology and Medicine, Rutgers, The State University of New Jersey, Piscataway New Jersey 08854; Northeast Structural Genomics Consortium, Rutgers, The State University of New Jersey, Piscataway New Jersey 08854; Department of Molecular Biology and Biochemistry, Rutgers, The State University of New Jersey, Piscataway New Jersey 08854; Aiyer, Sriram [Department of Pharmacology, Robert Wood Johnson Medical School, Rutgers, The State University of New Jersey, Piscataway New Jersey 08854; Cote, Marie L. [Department of Biochemistry, Robert Wood Johnson Medical School, UMDNJ, Piscataway New Jersey 08854; Xiao, Rong [Center for Advanced Biotechnology and Medicine, Rutgers, The State University of New Jersey, Piscataway New Jersey 08854; Northeast Structural Genomics Consortium, Rutgers, The State University of New Jersey, Piscataway New Jersey 08854; Department of Molecular Biology and Biochemistry, Rutgers, The State University of New Jersey, Piscataway New Jersey 08854; Jiang, Mei [Center for Advanced Biotechnology and Medicine, Rutgers, The State University of New Jersey, Piscataway New Jersey 08854; Northeast Structural Genomics Consortium, Rutgers, The State University of New Jersey, Piscataway New Jersey 08854; Department of Molecular Biology and Biochemistry, Rutgers, The State University of New Jersey, Piscataway New Jersey 08854; Acton, Thomas B. [Center for Advanced Biotechnology and Medicine, Rutgers, The State University of New Jersey, Piscataway New Jersey 08854; Northeast Structural Genomics Consortium, Rutgers, The State University of New Jersey, Piscataway New Jersey 08854; Department of Molecular Biology and Biochemistry, Rutgers, The State University of New Jersey, Piscataway New Jersey 08854; Roth, Monica J. [Department of Pharmacology, Robert Wood Johnson Medical School, Rutgers, The State University of New Jersey, Piscataway New Jersey 08854; Montelione, Gaetano T. [Center for Advanced Biotechnology and Medicine, Rutgers, The State University of New Jersey, Piscataway New Jersey 08854; Northeast Structural Genomics Consortium, Rutgers, The State University of New Jersey, Piscataway New Jersey 08854; Department of Molecular Biology and Biochemistry, Rutgers, The State University of New Jersey, Piscataway New Jersey 08854; Department of Biochemistry, Robert Wood Johnson Medical School, UMDNJ, Piscataway New Jersey 08854

    2017-02-03

    The retroviral integrase (IN) carries out the integration of a dsDNA copy of the viral genome into the host DNA, an essential step for viral replication. All IN proteins have three general domains, the N-terminal domain (NTD), the catalytic core domain, and the C-terminal domain. The NTD includes an HHCC zinc finger-like motif, which is conserved in all retroviral IN proteins. Two crystal structures of Moloney murine leukemia virus (M-MuLV) IN N-terminal region (NTR) constructs that both include an N-terminal extension domain (NED, residues 1–44) and an HHCC zinc-finger NTD (residues 45–105), in two crystal forms are reported. The structures of IN NTR constructs encoding residues 1–105 (NTR1–105) and 8–105 (NTR8–105) were determined at 2.7 and 2.15 Å resolution, respectively and belong to different space groups. While both crystal forms have similar protomer structures, NTR1–105 packs as a dimer and NTR8–105 packs as a tetramer in the asymmetric unit. The structure of the NED consists of three anti-parallel β-strands and an α-helix, similar to the NED of prototype foamy virus (PFV) IN. These three β-strands form an extended β-sheet with another β-strand in the HHCC Zn2+ binding domain, which is a unique structural feature for the M-MuLV IN. The HHCC Zn2+ binding domain structure is similar to that in HIV and PFV INs, with variations within the loop regions. Differences between the PFV and MLV IN NEDs localize at regions identified to interact with the PFV LTR and are compared with established biochemical and virological data for M-MuLV. Proteins 2017; 85:647–656.

  6. Nucleic acid amplification of HIV-1 integrase sequence subtypes CRF01_AE and B for development of HIV anti-integrase drug resistance genotyping assay

    Science.gov (United States)

    Adlar, F. R.; Bela, B.

    2017-08-01

    To anticipate the potential use of anti-integrase drugs in Indonesia for treatment of HIV-1 infection, the development of a drug resistance genotyping assay for anti-integrase is crucial in identifying the genetic drug resistance profile of Indonesian HIV-1 strains. This experiment aimed to amplify a target region in the integrase gene of Indonesian HIV-1 subtypes CRF01_AE and B that contain genetic mutations known to confer resistance to anti-integrase drug. Eleven archived plasma samples from individuals living with HIV-1 were obtained from the Virology and Cancer Pathobiology Research Center for Health Service (VCPRC FKUI-RSCM) laboratory. One of the plasma samples contained HIV-1 subtype B, and the remaining plasma samples contained subtype CRF01_AE. The target regions for all samples were amplified through RT-PCR, with an annealing temperature of 55 °C, using the primer pair AE_POL 4086F and AE_POL 5232R that were designed by VCPRC FKUI-RSCM. The results of this experiment show that 18.2% (2/11) of the samples were successfully amplified using the one-step RT-PCR. While the primer pair was effective in amplifying the target region in the integrase gene sequence for subtype B (100%; 1/1), it had a low efficacy (10%, 1/10) for subtype CRF01_AE. In conclusion, the primer pair can be used to amplify the target region in Indonesian HIV-1 strain subtypes CRF01_AE and B. However, optimization of the PCR condition and an increased number of samples would help to determine an accurate representation of the efficacy of the primer pair.

  7. Structural basis for functional tetramerization of lentiviral integrase.

    Directory of Open Access Journals (Sweden)

    Stephen Hare

    2009-07-01

    Full Text Available Experimental evidence suggests that a tetramer of integrase (IN is the protagonist of the concerted strand transfer reaction, whereby both ends of retroviral DNA are inserted into a host cell chromosome. Herein we present two crystal structures containing the N-terminal and the catalytic core domains of maedi-visna virus IN in complex with the IN binding domain of the common lentiviral integration co-factor LEDGF. The structures reveal that the dimer-of-dimers architecture of the IN tetramer is stabilized by swapping N-terminal domains between the inner pair of monomers poised to execute catalytic function. Comparison of four independent IN tetramers in our crystal structures elucidate the basis for the closure of the highly flexible dimer-dimer interface, allowing us to model how a pair of active sites become situated for concerted integration. Using a range of complementary approaches, we demonstrate that the dimer-dimer interface is essential for HIV-1 IN tetramerization, concerted integration in vitro, and virus infectivity. Our structures moreover highlight adaptable changes at the interfaces of individual IN dimers that allow divergent lentiviruses to utilize a highly-conserved, common integration co-factor.

  8. Crystal structure of an active two-domain derivative of Rous sarcoma virus integrase.

    Science.gov (United States)

    Yang, Z N; Mueser, T C; Bushman, F D; Hyde, C C

    2000-02-18

    Integration of retroviral cDNA is a necessary step in viral replication. The virally encoded integrase protein and DNA sequences at the ends of the linear viral cDNA are required for this reaction. Previous studies revealed that truncated forms of Rous sarcoma virus integrase containing two of the three protein domains can carry out integration reactions in vitro. Here, we describe the crystal structure at 2.5 A resolution of a fragment of the integrase of Rous sarcoma virus (residues 49-286) containing both the conserved catalytic domain and a modulatory DNA-binding domain (C domain). The catalytic domains form a symmetric dimer, but the C domains associate asymmetrically with each other and together adopt a canted conformation relative to the catalytic domains. A binding path for the viral cDNA is evident spanning both domain surfaces, allowing modeling of the larger integration complexes that are known to be active in vivo. The modeling suggests that formation of an integrase tetramer (a dimer of dimers) is necessary and sufficient for joining both viral cDNA ends at neighboring sites in the target DNA. The observed asymmetric arrangement of C domains suggests that they could form a rotationally symmetric tetramer that may be important for bridging integrase complexes at each cDNA end.

  9. HIV-1 integrase inhibitor resistance among treatment naïve patients in the West of Scotland.

    Science.gov (United States)

    Bradley-Stewart, A; Urcia, C; MacLean, A; Aitken, C; Gunson, R

    2017-07-01

    Transmitted integrase inhibitor resistance is rare, with only a small number of cases reported world-wide to date. The aim of this study was to assess whether transmitted integrase inhibitor resistance has occurred in Scotland and if so, could there be a case for performing genotypic integrase resistance testing at baseline. The study population consisted of 106 treatment naïve, newly diagnosed, HIV positive patients. The patient samples were collected between October 2015 and March 2016 at the time of HIV diagnosis and prior to initiation of anti-retroviral therapy. The integrase region was amplified and sequenced. We detected integrase inhibitor resistance (T66I/T) at baseline in one patient sample. This is a non-polymorphic mutation seen in patients receiving elvitegravir which confers high-level resistance to elvitegravir and intermediate resistance to raltegravir. A further 10 patients had accessory mutations which have minimal or no effect on susceptibility to integrase inhibitors. Transmitted integrase inhibitor resistance remains rare. The results of the present study do not support performing integrase resistance testing at baseline. Crown Copyright © 2017. Published by Elsevier B.V. All rights reserved.

  10. The Discovery of the Double Helix

    CERN Multimedia

    CERN. Geneva

    2011-01-01

    Professor James D. Watson has kindly agreed to make a presentation on the 1953 finding of the Double Helix at the Cavendish Laboratory by Francis Crick and himself. Being one of the greatest scientific discoveries in human history, little else needs to be added.

  11. The Modular Construction of DNA Double Helix

    Indian Academy of Sciences (India)

    DNA or the left-handed double helix,. Z- DNA. Construction of the Module .... 12. DNA, namely, replication and transcription. In the former case, Figure 3. 8 would represent a DNA single strand-generated by splitting of the mother duplex ...

  12. Helix reactor: great potential for flow chemistry

    NARCIS (Netherlands)

    Geerdink, P.; Runstraat, A. van den; Roelands, C.P.M.; Goetheer, E.L.V.

    2009-01-01

    The Helix reactor is highly suited for precise reaction control based on good hydrodynamics. The hydrodynamics are controlled by the Dean vortices, which create excellent heat transfer properties, approach plug flow and avoid turbulence. The flexibility of this reactor has been demonstrated using a

  13. A method for producing transgenic cells using a multi-integrase system on a human artificial chromosome vector.

    Directory of Open Access Journals (Sweden)

    Shigeyuki Yamaguchi

    Full Text Available The production of cells capable of expressing gene(s of interest is important for a variety of applications in biomedicine and biotechnology, including gene therapy and animal transgenesis. The ability to insert transgenes at a precise location in the genome, using site-specific recombinases such as Cre, FLP, and ΦC31, has major benefits for the efficiency of transgenesis. Recent work on integrases from ΦC31, R4, TP901-1 and Bxb1 phages demonstrated that these recombinases catalyze site-specific recombination in mammalian cells. In the present study, we examined the activities of integrases on site-specific recombination and gene expression in mammalian cells. We designed a human artificial chromosome (HAC vector containing five recombination sites (ΦC31 attP, R4 attP, TP901-1 attP, Bxb1 attP and FRT; multi-integrase HAC vector and de novo mammalian codon-optimized integrases. The multi-integrase HAC vector has several functions, including gene integration in a precise locus and avoiding genomic position effects; therefore, it was used as a platform to investigate integrase activities. Integrases carried out site-specific recombination at frequencies ranging from 39.3-96.8%. Additionally, we observed homogenous gene expression in 77.3-87.5% of colonies obtained using the multi-integrase HAC vector. This vector is also transferable to another cell line, and is capable of accepting genes of interest in this environment. These data suggest that integrases have high DNA recombination efficiencies in mammalian cells. The multi-integrase HAC vector enables us to produce transgene-expressing cells efficiently and create platform cell lines for gene expression.

  14. Ethyl malonate amides: a diketo acid offspring fragment for HIV integrase inhibition.

    Science.gov (United States)

    Serafin, Katarzyna; Mazur, Pawel; Bak, Andrzej; Laine, Elodie; Tchertanov, Luba; Mouscadet, Jean-François; Polanski, Jaroslaw

    2011-08-15

    While searching for new HIV integrase inhibitors we discovered that some ethyl malonate amides (EMA) are active against this enzyme. Surprisingly, the main function can only very rarely be found among the reported drug candidates. We synthesised a series of compounds in order to establish and analyse the structure-activity relationship. The similarity to the important classes of HIV integrase inhibitors as well as the synthetic availability of the different targets including this pharmacophore makes EMA compounds an interesting object of investigations. Copyright © 2011 Elsevier Ltd. All rights reserved.

  15. Triple helix interactions for eco-innovation

    DEFF Research Database (Denmark)

    Hermann, Roberto Rivas; Riisgaard, Henrik; Remmen, Arne

    the role of science parks in promoting eco-innovation. This study uses qualitative data gathered in two units of analysis: Panama Canal Authority and City of Knowledge Science Park. The study examines how Triple Helix interactions have built the regional system of eco-innovation at the Panama Canal....... Overall, the research found that the Panamanian national innovation system facilitates eco-innovation by: providing research and development, building competence and financing of innovation processes. The “green maritime route” is an example of institutional eco-innovation promoted by the Panama Canal...... Authority with insights from consultants, universities and donnor agencies. The proximity of the science park to the canal, has hitherto not yielded with the creation of a “green cluster”, which could be a precedent to promote eco-innovations. These findings suggest that, Triple Helix interactions...

  16. FPGA helix tracking algorithm for PANDA

    Energy Technology Data Exchange (ETDEWEB)

    Liang, Yutie; Galuska, Martin; Gessler, Thomas; Kuehn, Wolfgang; Lange, Jens Soeren; Muenchow, David; Spruck, Bjoern [II. Physikalisches Institut, Giessen University (Germany); Ye, Hua [Institute of High Energy Physics, Beijing (China); Collaboration: PANDA-Collaboration

    2015-07-01

    The PANDA detector is a general-purpose detector for physics with high luminosity cooled antiproton beams, planed to operate at the FAIR facility in Darmstadt, Germany. The central detector includes a silicon Micro Vertex Detector (MVD) and a Straw Tube Tracker (STT). Without any hardware trigger, large amounts of raw data are streaming into the data acquisition system. The data reduction task is performed in the online system by reconstruction algorithms programmed on FPGAs (Field Programmable Gate Arrays) as first level and on a farm of GPUs or PCs as a second level. One important part in the system is the online track reconstruction. In this presentation, an online tracking algorithm for helix tracking reconstruction in the solenoidal field is shown. The tracking algorithm is composed by two parts, a road finding module followed by an iterative helix parameter calculation module. A performance study using C++ and the status of the VHDL implementation are presented.

  17. Mapping of HIV-1 integrase preferences for target site selection with various oligonucleotides

    Czech Academy of Sciences Publication Activity Database

    Snášel, Jan; Rosenberg, Ivan; Pačes, Ondřej; Pichová, Iva

    2009-01-01

    Roč. 488, č. 2 (2009), s. 153-162 ISSN 0003-9861 R&D Projects: GA MŠk 1M0508; GA MŠk(CZ) LC06077 Institutional research plan: CEZ:AV0Z40550506 Keywords : HIV integrase * strand transfer reaction * oligonucleotide substrates Subject RIV: CE - Biochemistry Impact factor: 3.046, year: 2009

  18. A Mos1 transposase in vivo assay to screen new HIV-1 integrase inhibitors.

    Science.gov (United States)

    Cancian, Mariana; Loreto, Elgion L S

    2018-04-01

    The integrase and transposase enzymes of retrovirus and transposons, respectively, share the catalytic DDE domain. In vitro assays showed that inhibitors of HIV-1 integrase generally inhibit the mariner Mos1 transposase. Using a Drosophila strain in which the mobilisation of the mariner element can be quantified by mosaic eyes, we showed that flies maintained in medium containing 210 µM to 4 mM of raltegravir, or 1 or 2 mM of dolutegravir, which are HIV-1 integrase inhibitor used in AIDS treatment, have 23-33% less somatic mobilisation in mosaic eyes when treated with raltegravir and 28-32% when treated with dolutegravir. The gene expression of the mariner transposase gene, estimated by qPCR, is similar among treated and control flies. The results suggest that in vivo assays using Drosophila can be used as a primary screening of inhibitory drugs for transposase and retroviral integrase. The advantages of this assay are that it is easy, quick, cheap and is an in vivo test, meaning that the tested substance has to have been taken in by cells and has arrived at the target site, which is not the case when in vitro assays are applied.

  19. Natural variation of HIV-1 group M integrase: Implications for a new class of antiretroviral inhibitors

    Directory of Open Access Journals (Sweden)

    Gifford Robert J

    2008-08-01

    Full Text Available Abstract HIV-1 integrase is the third enzymatic target of antiretroviral (ARV therapy. However, few data have been published on the distribution of naturally occurring amino acid variation in this enzyme. We therefore characterized the distribution of integrase variants among more than 1,800 published group M HIV-1 isolates from more than 1,500 integrase inhibitor (INI-naïve individuals. Polymorphism rates equal or above 0.5% were found for 34% of the central core domain positions, 42% of the C-terminal domain positions, and 50% of the N-terminal domain positions. Among 727 ARV-naïve individuals in whom the complete pol gene was sequenced, integrase displayed significantly decreased inter- and intra-subtype diversity and a lower Shannon's entropy than protease or RT. All primary INI-resistance mutations with the exception of E157Q – which was present in 1.1% of sequences – were nonpolymorphic. Several accessory INI-resistance mutations including L74M, T97A, V151I, G163R, and S230N were also polymorphic with polymorphism rates ranging between 0.5% to 2.0%.

  20. The Stringent Response Promotes Antibiotic Resistance Dissemination by Regulating Integron Integrase Expression in Biofilms

    Directory of Open Access Journals (Sweden)

    Emilie Strugeon

    2016-08-01

    Full Text Available Class 1 integrons are genetic systems that enable bacteria to capture and express gene cassettes. These integrons, when isolated in clinical contexts, most often carry antibiotic resistance gene cassettes. They play a major role in the dissemination of antibiotic resistance among Gram-negative bacteria. The key element of integrons is the integrase, which allows gene cassettes to be acquired and shuffled. Planktonic culture experiments have shown that integrase expression is regulated by the bacterial SOS response. In natural settings, however, bacteria generally live in biofilms, which are characterized by strong antibiotic resilience and by increased expression of stress-related genes. Here, we report that under biofilm conditions, the stringent response, which is induced upon starvation, (i increases basal integrase and SOS regulon gene expression via induction of the SOS response and (ii exerts biofilm-specific regulation of the integrase via the Lon protease. This indicates that biofilm environments favor integron-mediated acquisition of antibiotic resistance and other adaptive functions encoded by gene cassettes.

  1. Bacteriophage φC31 Integrase Mediated Transgenesis in Xenopus laevis for Protein Expression at Endogenous Levels

    Science.gov (United States)

    Allen, Bryan G.; Weeks, Daniel L.

    Bacteriophage φC31 inserts its genome into that of its host bacterium via the integrase enzyme which catalyzes recombination between a phage attachment site (attP) and a bacterial attachment site (attB). Integrase requires no accessory factors, has a high efficiency of recombination, and does not need perfect sequence fidelity for recognition and recombination between these attachment sites. These imperfect attachment sites, or pseudo-attachment sites, are present in many organisms and have been used to insert transgenes in a variety of species. Here we describe the φC31 integrase approach to make transgenic Xenopus laevis embryos.

  2. Performance of Celera RUO integrase resistance assay across multiple HIV-1 subtypes.

    Science.gov (United States)

    Wallis, Carole L; Viana, Raquel V; Saravanan, Shanmugam; Silva de Jesus, Carlos; Zeh, Clement; Halvas, Elias K; Mellors, John W

    2017-03-01

    HIV-1 sequence variation is a major obstacle to developing molecular based assays for multiple subtypes. This study sought to independently assess performance characteristics of the ViroSeq™ HIV-1 Integrase RUO Genotyping Kit (Celera, US) for samples of multiple different HIV-1 subtypes. 264 samples were tested in the validation, 106 from integrase inhibitor naïve patients' sent for routine HIV-1 drug resistance testing after failing a 1st- or 2nd-line regimen, and 158 samples from an external virology quality assurance program (VQA). For the latter, 53 unique VQA samples were tested in two to five different laboratories to assess assay reproducibility. For all assays, viral RNA was extracted using the ViroSeq extraction module, reverse transcribed, and amplified in a one-step reaction. Four sequencing primers were used to span codons 1-288 of integrase. The Rega subtyping tool was used for subtype assignment. Integrase polymorphisms and mutations were determined as differences from the HXB2 sequence and by the Stanford database, respectively. Sequences obtained from the different laboratories were aligned and sequence homology determined. HIV-1 RNA in the 264 samples ranged from 3.15 to 6.74 log copies/ml. Successful amplification was obtained for 97% of samples (n=256). The 8 samples that failed to amplify were subtype D (n=3), subtype C (n=1), CRF01_AE (n=1), subtype A1 (n=2), and an unassigned subtype (n=1). Of the 256 that successfully amplified samples, 203 (79%) were successfully sequenced with bidirectional coverage. Of the 53 unsuccessful samples, 13 (5%) failed sequencing and 40 (16%) did not have full bidirectional sequence, as a result of failure of sequencing primers: Primer A (n=1); Primer B (n=18); Primer C (n=1); Primer D (n=7) or short sequences (n=16). For the 135 VQA samples (30 unique samples) that were assayed by different laboratories, homology of the sequences obtained ranged from 92.1% to 100%. However, Laboratory 2 detected more mixtures

  3. Discovery of a novel HIV-1 integrase inhibitor from natural compounds through structure based virtual screening and cell imaging.

    Science.gov (United States)

    Gu, Wan-Gang; Zhang, Xuan; Ip, Denis Tsz-Ming; Yang, Liu-Meng; Zheng, Yong-Tang; Wan, David Chi-Cheong

    2014-09-17

    The interaction between HIV-1 integrase and LEDGF/P75 has been validated as a target for anti-HIV drug development. Based on the crystal structure of integrase in complex with LEDGF/P75, a library containing 80 thousand natural compounds was filtered with virtual screening. 11 hits were selected for cell based assays. One compound, 3-(1,3-benzothiazol-2-yl)-8-{[bis(2-hydroxyethyl)amino]methyl}-7-hydroxy-2H-chromen-2-one (D719) inhibited integrase nuclear translocation in cell imaging. The binding mode of D719 was analyzed with molecular simulation. The anti-HIV activity of D719 was assayed by measuring the p24 antigen production in acute infection. The structure characteristics of D719 may provide valuable information for integrase inhibitor design. Copyright © 2014 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  4. Prevalence of SOS-mediated control of integron integrase expression as an adaptive trait of chromosomal and mobile integrons

    Directory of Open Access Journals (Sweden)

    Cambray Guillaume

    2011-04-01

    Full Text Available Abstract Background Integrons are found in hundreds of environmental bacterial species, but are mainly known as the agents responsible for the capture and spread of antibiotic-resistance determinants between Gram-negative pathogens. The SOS response is a regulatory network under control of the repressor protein LexA targeted at addressing DNA damage, thus promoting genetic variation in times of stress. We recently reported a direct link between the SOS response and the expression of integron integrases in Vibrio cholerae and a plasmid-borne class 1 mobile integron. SOS regulation enhances cassette swapping and capture in stressful conditions, while freezing the integron in steady environments. We conducted a systematic study of available integron integrase promoter sequences to analyze the extent of this relationship across the Bacteria domain. Results Our results showed that LexA controls the expression of a large fraction of integron integrases by binding to Escherichia coli-like LexA binding sites. In addition, the results provide experimental validation of LexA control of the integrase gene for another Vibrio chromosomal integron and for a multiresistance plasmid harboring two integrons. There was a significant correlation between lack of LexA control and predicted inactivation of integrase genes, even though experimental evidence also indicates that LexA regulation may be lost to enhance expression of integron cassettes. Conclusions Ancestral-state reconstruction on an integron integrase phylogeny led us to conclude that the ancestral integron was already regulated by LexA. The data also indicated that SOS regulation has been actively preserved in mobile integrons and large chromosomal integrons, suggesting that unregulated integrase activity is selected against. Nonetheless, additional adaptations have probably arisen to cope with unregulated integrase activity. Identifying them may be fundamental in deciphering the uneven distribution of

  5. Designing cooperatively folded abiotic uni- and multimolecular helix bundles

    Science.gov (United States)

    de, Soumen; Chi, Bo; Granier, Thierry; Qi, Ting; Maurizot, Victor; Huc, Ivan

    2018-01-01

    Abiotic foldamers, that is foldamers that have backbones chemically remote from peptidic and nucleotidic skeletons, may give access to shapes and functions different to those of peptides and nucleotides. However, design methodologies towards abiotic tertiary and quaternary structures are yet to be developed. Here we report rationally designed interactional patterns to guide the folding and assembly of abiotic helix bundles. Computational design facilitated the introduction of hydrogen-bonding functionalities at defined locations on the aromatic amide backbones that promote cooperative folding into helix-turn-helix motifs in organic solvents. The hydrogen-bond-directed aggregation of helices not linked by a turn unit produced several thermodynamically and kinetically stable homochiral dimeric and trimeric bundles with structures that are distinct from the designed helix-turn-helix. Relative helix orientation within the bundles may be changed from parallel to tilted on subtle solvent variations. Altogether, these results prefigure the richness and uniqueness of abiotic tertiary structure behaviour.

  6. A Classification of Basic Helix-Loop-Helix Transcription Factors of Soybean

    Directory of Open Access Journals (Sweden)

    Karen A. Hudson

    2015-01-01

    Full Text Available The complete genome sequence of soybean allows an unprecedented opportunity for the discovery of the genes controlling important traits. In particular, the potential functions of regulatory genes are a priority for analysis. The basic helix-loop-helix (bHLH family of transcription factors is known to be involved in controlling a wide range of systems critical for crop adaptation and quality, including photosynthesis, light signalling, pigment biosynthesis, and seed pod development. Using a hidden Markov model search algorithm, 319 genes with basic helix-loop-helix transcription factor domains were identified within the soybean genome sequence. These were classified with respect to their predicted DNA binding potential, intron/exon structure, and the phylogeny of the bHLH domain. Evidence is presented that the vast majority (281 of these 319 soybean bHLH genes are expressed at the mRNA level. Of these soybean bHLH genes, 67% were found to exist in two or more homeologous copies. This dataset provides a framework for future studies on bHLH gene function in soybean. The challenge for future research remains to define functions for the bHLH factors encoded in the soybean genome, which may allow greater flexibility for genetic selection of growth and environmental adaptation in this widely grown crop.

  7. HELIX: The High Energy Light Isotope Experiment

    Science.gov (United States)

    Wakely, Scott

    This is the lead proposal for a new suborbital program, HELIX (High-Energy Light Isotope eXperiment), designed to make measurements of the isotopic composition of light cosmic-ray nuclei from ~200 MeV/nuc to ~10 GeV/nuc. Past measurements of this kind have provided profound insights into the nature and origin of cosmic rays, revealing, for instance, information on acceleration and confinement time scales, and exposing some conspicuous discrepancies between solar and cosmic-ray abundances. The most detailed information currently available comes from the ACE/CRIS mission, but is restricted to energies below a few 100 MeV/nuc. HELIX aims at extending this energy range by over an order of magnitude, where, in most cases, no measurements of any kind exist, and where relativistic time dilation affects the apparent lifetime of radioactive clock nuclei. The HELIX measurements will provide essential information for understanding the propagation history of cosmic rays in the galaxy. This is crucial for properly interpreting several intriguing anomalies reported in recent cosmic-ray measurements, pertaining to the energy spectra of protons, helium, and heavier nuclei, and to the anomalous rise in the positron fraction at higher energy. HELIX employs a high-precision magnet spectrometer to provide measurements which are not achievable by any current or planned instrument. The superconducting magnet originally used for the HEAT payload in five successful high-altitude flights will be combined with state-of-the-art detectors to measure the charge, time-of-flight, magnetic rigidity, and velocity of cosmic-ray particles with high precision. The instrumentation includes plastic scintillators, silicon-strip detectors repurposed from Fermilab's CDF detector, a high-performance gas drift chamber, and a ring-imaging Cherenkov counter employing aerogel radiators and silicon photomultipliers. To reduce cost and technical risk, the HELIX program will be structured in two stages. The first

  8. Loop-to-helix transition in the structure of multidrug regulator AcrR at the entrance of the drug-binding cavity

    Energy Technology Data Exchange (ETDEWEB)

    Manjasetty, Babu A.; Halavaty, Andrei S.; Luan, Chi-Hao; Osipiuk, Jerzy; Mulligan, Rory; Kwon, Keehwan; Anderson, Wayne F.; Joachimiak, Andrzej

    2016-04-01

    Multidrug transcription regulator AcrR from Salmonella enterica subsp. enterica serovar Typhimurium str. LT2 belongs to the tetracycline repressor family, one of the largest groups of bacterial transcription factors. The crystal structure of dimeric AcrR was determined and refined to 1.56 Å resolution. The tertiary and quaternary structures of AcrR are similar to those of its homologs. The multidrug binding site was identified based on structural alignment with homologous proteins and has a di(hydroxyethyl)ether molecule bound. Residues from helices a4 and a7 shape the entry into this binding site. The structure of AcrR reveals that the extended helical conformation of helix a4 is stabilized by the hydrogen bond between Glu67 (helix a4) and Gln130 (helix a7). Based on the structural comparison with the closest homolog structure, the Escherichia coli AcrR, we propose that this hydrogen bond is responsible for control of the loop-to-helix transition within helix a4. This local conformational switch of helix a4 may be a key step in accessing the multidrug binding site and securing ligands at the binding site. Solution smallmolecule binding studies suggest that AcrR binds ligands with their core chemical structure resembling the tetracyclic ring of cholesterol.

  9. Identification of minority resistance mutations in the HIV-1 integrase coding region using next generation sequencing

    DEFF Research Database (Denmark)

    Fonager, Jannik; Larsson, Jonas T; Hussing, Christian

    2015-01-01

    BACKGROUND: The current widely applied standard method to screen for HIV-1 genotypic resistance is based on Sanger population sequencing (Sseq), which does not allow for the identification of minority variants (MVs) below the limit of detection for the Sseq-method in patients receiving integrase...... strand-transfer inhibitors (INSTI). Next generation sequencing (NGS) has facilitated the detection of MVs at a much deeper level than Sseq. OBJECTIVES: Here, we compared Illumina MiSeq and Sseq approaches to evaluate the detection of MVs involved in resistance to the three commonly used INSTI......: raltegravir (RAL), elvitegravir (EVG) and dolutegravir (DTG). STUDY DESIGN: NGS and Sseq were used to analyze RT-PCR products of the HIV-1 integrase coding region from six patients and in serial samples from two patients. NGS sequences were assembled and analyzed using the low frequency variant detection...

  10. phiC31 integrase-mediated site-specific recombination in barley.

    Directory of Open Access Journals (Sweden)

    Eszter Kapusi

    Full Text Available The Streptomyces phage phiC31 integrase was tested for its feasibility in excising transgenes from the barley genome through site-specific recombination. We produced transgenic barley plants expressing an active phiC31 integrase and crossed them with transgenic barley plants carrying a target locus for recombination. The target sequence involves a reporter gene encoding green fluorescent protein (GFP, which is flanked by the attB and attP recognition sites for the phiC31 integrase. This sequence disruptively separates a gusA coding sequence from an upstream rice actin promoter. We succeeded in producing site-specific recombination events in the hybrid progeny of 11 independent barley plants carrying the above target sequence after crossing with plants carrying a phiC31 expression cassette. Some of the hybrids displayed fully executed recombination. Excision of the GFP gene fostered activation of the gusA gene, as visualized in tissue of hybrid plants by histochemical staining. The recombinant loci were detected in progeny of selfed F(1, even in individuals lacking the phiC31 transgene, which provides evidence of stability and generative transmission of the recombination events. In several plants that displayed incomplete recombination, extrachromosomal excision circles were identified. Besides the technical advance achieved in this study, the generated phiC31 integrase-expressing barley plants provide foundational stock material for use in future approaches to barley genetic improvement, such as the production of marker-free transgenic plants or switching transgene activity.

  11. Development of a novel in vitro assay for the evaluation of integron DNA integrase activity

    Directory of Open Access Journals (Sweden)

    Fatemeh Tohidi

    2016-05-01

    Full Text Available Integrons play an important role in multidrug resistance. The integron platform codes for integrase (intI that is required for gene cassette integration through site-specific recombination. The recombination crossover occurs between the G and TT nucleotides in non-palindromic attI and palindromic attC sites. The aim of this study was to establish an efficient in vitro assay for integrase purification and activity detection. To this end, the intI gene was cloned into the pET-22b plasmid. Then, the resulting recombinant plasmid was transformed into Escherichia coli Origami™ strain. The recombinant protein expression was confirmed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE and western blot assays. The recombinant intI protein was purified by nickel–nitrilotriacetic acid (Ni–NTA affinity chromatography, and its activity was measured by a newly introduced assay. Briefly, specific primers for each side of attI and attC were used, thereby, a polymerase chain reaction would be performed, if a fused plasmid containing both attI and attC sites was created upon recombination. SDS-PAGE and western blotting confirmed the presence of a 38-kDa recombinant protein. Optimum conditions were established for the measurement of the integrase activity and a new model assay was conducted to analyse the recombination activity in vitro. Although the electrophoretic mobility shift assay is an efficient and reliable method, the newly introduced assay provided new or enhanced capability to determine the integrase activity, suggesting that there is no need for expensive and advanced equipment.

  12. Efficient site-specific integration in Plasmodium falciparum chromosomes mediated by mycobacteriophage Bxb1 integrase.

    Science.gov (United States)

    Nkrumah, Louis J; Muhle, Rebecca A; Moura, Pedro A; Ghosh, Pallavi; Hatfull, Graham F; Jacobs, William R; Fidock, David A

    2006-08-01

    Here we report an efficient, site-specific system of genetic integration into Plasmodium falciparum malaria parasite chromosomes. This is mediated by mycobacteriophage Bxb1 integrase, which catalyzes recombination between an incoming attP and a chromosomal attB site. We developed P. falciparum lines with the attB site integrated into the glutaredoxin-like cg6 gene. Transfection of these attB(+) lines with a dual-plasmid system, expressing a transgene on an attP-containing plasmid together with a drug resistance gene and the integrase on a separate plasmid, produced recombinant parasites within 2 to 4 weeks that were genetically uniform for single-copy plasmid integration. Integrase-mediated recombination resulted in proper targeting of parasite proteins to intra-erythrocytic compartments, including the apicoplast, a plastid-like organelle. Recombinant attB x attP parasites were genetically stable in the absence of drug and were phenotypically homogeneous. This system can be exploited for rapid genetic integration and complementation analyses at any stage of the P. falciparum life cycle, and it illustrates the utility of Bxb1-based integrative recombination for genetic studies of intracellular eukaryotic organisms.

  13. Mucosal immunization with integrase-defective lentiviral vectors protects against influenza virus challenge in mice.

    Directory of Open Access Journals (Sweden)

    Judith M Fontana

    Full Text Available Recent reports highlight the potential for integrase-defective lentiviral vectors (IDLV to be developed as vaccines due to their ability to elicit cell-mediated and humoral immune responses after intramuscular administration. Differently from their integrase-competent counterpart, whose utility for vaccine development is limited by the potential for insertional mutagenesis, IDLV possess a mutation in their integrase gene that prevents genomic integration. Instead, they are maintained as episomal DNA circles that retain the ability to stably express functional proteins. Despite their favorable profile, it is unknown whether IDLV elicit immune responses after intranasal administration, a route that could be advantageous in the case of infection with a respiratory agent. Using influenza as a model, we constructed IDLV expressing the influenza virus nucleoprotein (IDLV-NP, and tested their ability to generate NP-specific immune responses and protect from challenge in vivo. We found that administration of IDLV-NP elicited NP-specific T cell and antibody responses in BALB/c mice. Importantly, IDLV-NP was protective against homologous and heterosubtypic influenza virus challenge only when given by the intranasal route. This is the first report demonstrating that IDLV can induce protective immunity after intranasal administration, and suggests that IDLV may represent a promising vaccine platform against infectious agents.

  14. Design, Synthesis, and Biological Evaluation of 1,2-Dihydroisoquinolines as HIV-1 Integrase Inhibitors.

    Science.gov (United States)

    Tandon, Vibha; Urvashi; Yadav, Pooja; Sur, Souvik; Abbat, Sheenu; Tiwari, Vinod; Hewer, Raymond; Papathanasopoulos, Maria A; Raja, Rameez; Banerjea, Akhil C; Verma, Akhilesh K; Kukreti, Shrikant; Bharatam, Prasad V

    2015-10-08

    6-Endo-dig-cyclization is an efficient method for the synthesis of 1,2-dihydroisoquinolines. We have synthesized few 1,2-dihydroisoquinolines having different functionality at the C-1, C-3, C-7, and N-2 positions for evaluation against HIV-1 integrase (HIV1-IN) inhibitory activity. A direct nitro-Mannich condensation of o-alkynylaldimines and dual activation of o-alkynyl aldehydes by inexpensive cobalt chloride yielded desired compounds. Out of 24 compounds, 4m and 6c came out as potent integrase inhibitors in in vitro strand transfer (ST) assay, with IC50 value of 0.7 and 0.8 μM, respectively. Molecular docking of these compounds in integrase revealed strong interaction between metal and ligands, which stabilizes the enzyme-inhibitor complex. The ten most active compounds were subjected to antiviral assay. Out of those, 6c reduced the level of p24 viral antigen by 91%, which is comparable to RAL in antiviral assay. Interestingly, these compounds showed similar ST inhibitory activity in G140S mutant, suggesting they can act against resistant strains.

  15. Government and Governance of Regional Triple Helix Interactions

    Science.gov (United States)

    Danson, Mike; Todeva, Emanuela

    2016-01-01

    This conceptual paper contributes to the discussion of the role of regional government and regional Triple Helix constellations driving economic development and growth within regional boundaries. The impact of regionalism and subsidiarity on regional Triple Helix constellations, and the questions of governmentality, governance and institutional…

  16. FPGA helix tracking algorithm for PANDA

    Energy Technology Data Exchange (ETDEWEB)

    Liang, Yutie; Galuska, Martin; Gessler, Thomas; Kuehn, Wolfgang; Lange, Jens Soeren; Muenchow, David [II. Physikalisches Institut, University of Giessen (Germany); Ye, Hua [Institute of High Energy Physics, CAS (China); Collaboration: PANDA-Collaboration

    2016-07-01

    The PANDA detector is a general-purpose detector for physics with high luminosity cooled antiproton beams, planed to operate at the FAIR facility in Darmstadt, Germany. The central detector includes a silicon Micro Vertex Detector (MVD) and a Straw Tube Tracker (STT). Without any hardware trigger, large amounts of raw data are streaming into the data acquisition system. The data reduction task is performed in the online system by reconstruction algorithms programmed on FPGAs (Field Programmable Gate Arrays) as first level and on a farm of GPUs or PCs as a second level. One important part in the system is the online track reconstruction. In this presentation, an online tracking algorithm for helix tracking reconstruction in the solenoidal field is shown. The VHDL-based algorithm is tested with different types of events, at different event rate. Furthermore, a study of T0 extraction from the tracking algorithm is performed. A concept of simultaneous tracking and T0 determination is presented.

  17. FPGA helix tracking algorithm for PANDA

    Energy Technology Data Exchange (ETDEWEB)

    Liang, Yutie; Galuska, Martin; Gessler, Thomas; Hu, Jifeng; Kuehn, Wolfgang; Lange, Jens Soeren; Muenchow, David; Spruck, Bjoern [II. Physikalisches, Giessen University (Germany); Ye, Hua [II. Physikalisches, Giessen University (Germany); Institute of High Energy Physics, Beijing (China); Collaboration: PANDA-Collaboration

    2014-07-01

    The PANDA detector is a general-purpose detector for physics with high luminosity cooled antiproton beams, planed to operate at the FAIR facility in Darmstadt, Germany. The central detector includes a silicon Micro Vertex Detector (MVD) and a Straw Tube Tracker (STT). Without any hardware trigger, large amounts of raw data are streaming into the data acquisition system. The data reduction task is performed in the online system by reconstruction algorithms programmed in VHDL (Very High Speed Integrated Circuit Hardware Description Language) on FPGAs (Field Programmable Gate Arrays) as first level and on a farm of GPUs or PCs as a second level. One important part in the system is the online track reconstruction. In this presentation, an online tracking finding algorithm for helix track reconstruction in the solenoidal field is shown. A performance study using C++ and the status of the VHDL implementation are presented.

  18. Biochemical analysis of the role of G118R-linked dolutegravir drug resistance substitutions in HIV-1 integrase.

    Science.gov (United States)

    Quashie, Peter K; Mesplède, Thibault; Han, Ying-Shan; Veres, Tamar; Osman, Nathan; Hassounah, Said; Sloan, Richard D; Xu, Hong-Tao; Wainberg, Mark A

    2013-12-01

    Drug resistance mutations (DRMs) have been reported for all currently approved anti-HIV drugs, including the latest integrase strand transfer inhibitors (INSTIs). We previously used the new INSTI dolutegravir (DTG) to select a G118R integrase resistance substitution in tissue culture and also showed that secondary substitutions emerged at positions H51Y and E138K. Now, we have characterized the impact of the G118R substitution, alone or in combination with either H51Y or E138K, on 3' processing and integrase strand transfer activity. The results show that G118R primarily impacted the strand transfer step of integration by diminishing the ability of integrase-long terminal repeat (LTR) complexes to bind target DNA. The addition of H51Y and E138K to G118R partially restored strand transfer activity by modulating the formation of integrase-LTR complexes through increasing LTR DNA affinity and total DNA binding, respectively. This unique mechanism, in which one function of HIV integrase partially compensates for the defect in another function, has not been previously reported. The G118R substitution resulted in low-level resistance to DTG, raltegravir (RAL), and elvitegravir (EVG). The addition of either of H51Y or E138K to G118R did not enhance resistance to DTG, RAL, or EVG. Homology modeling provided insight into the mechanism of resistance conferred by G118R as well as the effects of H51Y or E138K on enzyme activity. The G118R substitution therefore represents a potential avenue for resistance to DTG, similar to that previously described for the R263K substitution. For both pathways, secondary substitutions can lead to either diminished integrase activity and/or increased INSTI susceptibility.

  19. Basic helix-loop-helix transcription factors and epidermal cell fate determination in Arabidopsis.

    Science.gov (United States)

    Zhao, Hongtao; Li, Xia; Ma, Ligeng

    2012-12-01

    Cell fate determination is an important process in multicellular organisms. Plant epidermis is a readily-accessible, well-used model for the study of cell fate determination. Our knowledge of cell fate determination is growing steadily due to genetic and molecular analyses of root hairs, trichomes, and stomata, which are derived from the epidermal cells of roots and aerial tissues. Studies have shown that a large number of factors are involved in the establishment of these cell types, especially members of the basic helix-loop-helix (bHLH) superfamily, which is an important family of transcription factors. In this mini-review, we focus on the role of bHLH transcription factors in cell fate determination in Arabidopsis.

  20. NMR-based approach to measure the free energy of transmembrane helix-helix interactions.

    Science.gov (United States)

    Mineev, Konstantin S; Lesovoy, Dmitry M; Usmanova, Dinara R; Goncharuk, Sergey A; Shulepko, Mikhail A; Lyukmanova, Ekaterina N; Kirpichnikov, Mikhail P; Bocharov, Eduard V; Arseniev, Alexander S

    2014-01-01

    Knowledge of the energetic parameters of transmembrane helix-helix interactions is necessary for the establishment of a structure-energy relationship for α-helical membrane domains. A number of techniques have been developed to measure the free energies of dimerization and oligomerization of transmembrane α-helices, and all of these have their advantages and drawbacks. In this study we propose a methodology to determine the magnitudes of the free energy of interactions between transmembrane helices in detergent micelles. The suggested approach employs solution nuclear magnetic resonance (NMR) spectroscopy to determine the population of the oligomeric states of the transmembrane domains and introduces a new formalism to describe the oligomerization equilibrium, which is based on the assumption that both the dimerization of the transmembrane domains and the dissociation of the dimer can occur only upon the collision of detergent micelles. The technique has three major advantages compared with other existing approaches: it may be used to analyze both weak and relatively strong dimerization/oligomerization processes, it works well for the analysis of complex equilibria, e.g. when monomer, dimer and high-order oligomer populations are simultaneously present in the solution, and it can simultaneously yield both structural and energetic characteristics of the helix-helix interaction under study. The proposed methodology was applied to investigate the oligomerization process of transmembrane domains of fibroblast growth factor receptor 3 (FGFR3) and vascular endothelium growth factor receptor 2 (VEGFR2), and allowed the measurement of the free energy of dimerization of both of these objects. In addition the proposed method was able to describe the multi-state oligomerization process of the VEGFR2 transmembrane domain. © 2013 Elsevier B.V. All rights reserved.

  1. Effect of Hedera helix on lung histopathology in chronic asthma.

    Science.gov (United States)

    Hocaoglu, Arzu Babayigit; Karaman, Ozkan; Erge, Duygu Olmez; Erbil, Guven; Yilmaz, Osman; Kivcak, Bijen; Bagriyanik, H Alper; Uzuner, Nevin

    2012-12-01

    Hedera helix is widely used to treat bronchial asthma for many years. However, effects of this herb on lung histopathology is still far from clear. We aimed to determine the effect of oral administration of Hedera helix on lung histopathology in a murine model of chronic asthma.BALB/c mice were divided into four groups; I (Placebo), II (Hedera helix), III (Dexamethasone) and IV (Control). All mice except controls were sensitized and challenged with ovalbumin. Then, mice in group I received saline, group II 100 mg/kg Hedera helix and group III 1 mg/kg dexamethasone via orogastic gavage once daily for one week. Airway histopathology was evaluated by using light and electron microscopy in all groups.Goblet cell numbers and thicknesses of basement membrane were found significantly lower in group II, but there was no statistically significant difference in terms of number of mast cells, thicknesses of epithelium and subepithelial smooth muscle layers between group I and II. When Hedera helix and dexamethasone groups were compared with each other, thickness of epithelium, subepithelial muscle layers, number of mast cells and goblet cells of group III were significantly ameliorated when compared with the group II. Although Hedera helix administration reduced only goblet cell counts and the thicknesses of basement membrane in the asthmatic airways, dexamethasone ameliorated all histopathologic parameters except thickness of basement membrane better than Hedera helix.

  2. Effects of radiation on DNA's double helix

    Science.gov (United States)

    2003-01-01

    The blueprint of life, DNA's double helix is found in the cells of everything from bacteria to astronauts. Exposure to radiation(depicted at right) such as X-rays (upper) or heavy ion particles (lower), can damage DNA and cause dire consequences both to the organism itself and to future generations. One of NASA's main goals is to develop better radiation shielding materials to protect astronauts from destructive radiation in space. This is particularly important for long space missions. NASA has selected researchers to study materials that provide better shielding. This research is managed by NASA's Office of Biological and Physical Research and is supported by the Microgravity Science and Applications Department at NASA's Marshall Center. During International Space Station Expedition Six, the Extravehicular Activity Radiation Monitoring (EVARM) will continue to measure radiation dosage encountered by the eyes, internal organs and skin during specific spacewalks, and relate it to the type of activity, location and other factors. An analysis of this information may be useful in mitigating potential exposure to space walkers in the future. (Illustration by Dr. Frank Cucinotta, NASA/Johnson Space Center, and Prem Saganti, Lockheed Martin)

  3. Generating structured light with phase helix and intensity helix using reflection-enhanced plasmonic metasurface at 2 μm

    Science.gov (United States)

    Zhao, Yifan; Du, Jing; Zhang, Jinrun; Shen, Li; Wang, Jian

    2018-04-01

    Mid-infrared (2-20 μm) light has been attracting great attention in many areas of science and technology. Beyond the extended wavelength range from visible and near-infrared to mid-infrared, shaping spatial structures may add opportunities to grooming applications of mid-infrared photonics. Here, we design and fabricate a reflection-enhanced plasmonic metasurface and demonstrate efficient generation of structured light with the phase helix and intensity helix at 2 μm. This work includes two distinct aspects. First, structured light (phase helix, intensity helix) generation at 2 μm, which is far beyond the ability of conventional spatial light modulators, is enabled by the metasurface with sub-wavelength engineered structures. Second, the self-referenced intensity helix against environmental noise is generated without using a spatially separated light. The demonstrations may open up advanced perspectives to structured light applications at 2 μm, such as phase helix for communications and non-communications (imaging, sensing) and intensity helix for enhanced microscopy and advanced metrology.

  4. The dolutegravir R263K resistance mutation in HIV-1 integrase is incompatible with the emergence of resistance against raltegravir.

    Science.gov (United States)

    Oliveira, Maureen; Mesplède, Thibault; Moïsi, Daniela; Ibanescu, Ruxandra-Ilinca; Brenner, Bluma; Wainberg, Mark A

    2015-11-01

    Although the integrase inhibitor dolutegravir (DTG) has demonstrated greater resilience than other antiretroviral drugs at withstanding the emergence of HIV-1 resistance mutations, such substitutions can develop, albeit rarely, in treatment-experienced integrase inhibitor-naïve individuals. The most common substitution in integrase under those circumstances is R263K whereas another substitution that was selected against DTG in tissue culture was G118R. The objective of this study was to determine the effects of these DTG-specific resistance substitutions on the ability of HIV-1 to become resistant against either of two other integrase inhibitors, raltegravir (RAL) and elvitegravir (EVG). We performed tissue culture selection experiments using DTG-resistant viruses containing integrase substitutions at positions R263K, H51Y/R263K, E138K/R263K, G118R and H51Y/G118R in the presence of increasing concentrations of either RAL or EVG. Changes in integrase sequences were monitored by genotyping. The presence of the R263K substitution delayed the emergence of resistance against RAL whereas the simultaneous presence of either the H51Y or E138K secondary substitutions in combination with R263K somewhat mitigated this inhibitory effect. In contrast, resistance against EVG appeared earlier than in wild-type virus in viruses containing the R263K and E138K/R263K DTG-associated resistance substitutions. The DTG-resistant R263K substitution antagonized the development of HIV-1 resistance against RAL while partially facilitating the occurrence of resistance against EVG.

  5. Cancer-specific binary expression system activated in mice by bacteriophage HK022 Integrase

    DEFF Research Database (Denmark)

    Elias, Amer; Spector, Itay; Sogolovsky-Bard, Ilana

    2016-01-01

    of a cytotoxic gene. In the present study we developed a new cancer specific binary expression system activated by the Integrase (Int) of the lambdoid phage HK022. We demonstrate the validity of this system by the specific expression of a luciferase (luc) reporter in human embryonic kidney 293T (HEK293T) cells...... and in a lung cancer mouse model. Due to the absence viral vectors and of cytotoxicity the Int based binary system offers advantages over previously described counterparts and may therefore be developed into a safer cancer cell killing system....

  6. Ferrocenyl chalcone difluoridoborates inhibit HIV-1 integrase and display low activity towards cancer and endothelial cells.

    Science.gov (United States)

    Monserrat, Jean-Philippe; Al-Safi, Rasha I; Tiwari, Keshri Nath; Quentin, Lionel; Chabot, Guy G; Vessières, Anne; Jaouen, Gérard; Neamati, Nouri; Hillard, Elizabeth A

    2011-10-15

    We report here the discovery of a potent series of HIV-1 integrase (IN) inhibitors based on the ferrocenyl chalcone difluoridoborate structure. Ten new compounds have been synthesized and were generally found to have similar inhibitory activities against the IN 3' processing and strand transfer (ST) processes. IC(50) values were found to be in the low micromolar range, and significantly lower than those found for the non-coordinated ferrocenyl chalcones and other ferrocene molecules. The ferrocenyl chalcone difluoridoborates furthermore exhibited low cytotoxicity against cancer cells and low morphological activity against epithelial cells. Copyright © 2011 Elsevier Ltd. All rights reserved.

  7. Discovery of 2-Pyridinone Aminals: A Prodrug Strategy to Advance a Second Generation of HIV-1 Integrase Strand Transfer Inhibitors

    Energy Technology Data Exchange (ETDEWEB)

    Raheem, Izzat T.; Walji, Abbas M.; Klein, Daniel; Sanders, John M.; Powell, David A.; Abeywickrema, Pravien; Barbe, Guillaume; Bennet, Amrith; Clas, SophieDorothee; Dubost, David; Embrey, Mark; Grobler, Jay; Hafey, Michael J.; Hartingh, Timothy J.; Hazuda, Daria J.; Miller, Michael D.; Moore, Keith P.; Pajkovic, Natasa; Patel, Sangita; Rada, Vanessa; Rearden, Paul; Schreier, John D.; Sisko, John; Steele, Thomas G.; Truchon, Jean-François; Wai, John; Xu, Min; Coleman, Paul J. (Merck)

    2015-10-22

    The search for new molecular constructs that resemble the critical two-metal binding pharmacophore required for HIV integrase strand transfer inhibition represents a vibrant area of research within drug discovery. Here we present the discovery of a new class of HIV integrase strand transfer inhibitors based on the 2-pyridinone core of MK-0536. These efforts led to the identification of two lead compounds with excellent antiviral activity and preclinical pharmacokinetic profiles to support a once-daily human dose prediction. Dose escalating PK studies in dog revealed significant issues with limited oral absorption and required an innovative prodrug strategy to enhance the high-dose plasma exposures of the parent molecules.

  8. Controlling chirality with helix inversion in cholesteric liquid crystals

    NARCIS (Netherlands)

    Katsonis, Nathalie Hélène; Lacaze, E.; Ferrarini, A.

    2012-01-01

    The helical organization of cholesteric liquid crystals is omnipresent in living matter. Achieving control over the structure of the cholesteric helix consequently holds great potential for developing stimuli-responsive materials matching the level of sophistication of biological systems. In

  9. Hedera helix L. and damages in Tlos Ancient City

    Directory of Open Access Journals (Sweden)

    Elinç, Z.K.

    2013-03-01

    Full Text Available There are various plant types in Tlos Ancient City of Fethiye district in the Province of Mugla, a city where different residential ruins of Lycia Civilization starting from Classical Age until Byzantine Period. Tlos is an important city in West-Lycia and is situated right on the control point of Lycia Way. Hedera helix L. is one of the plants living in this area, which attracts the attention as it mostly harms the ancient ruins. One of the most important reasons why Hedera helix L. is growing commonly in this region is the perfect ecological circumstances for the growth of this plant of the location where this ancient city is situated in. Additionally the fact that the ruins of the city are left on their fate, is another perfect circumstance for the Hedera helix L. to grow. Climbing or creeping stems of Hedera helix L. stick easily to the objects it touches and encircle them. Due to this characteristic, the walls of the ancient city are covered by this plant. Nevertheless, Hedera helix L. does not only harm the ancient constructions and natural rocks but also woody plants. The harm caused by dried out or cut Hedera helix L. are more than the harm caused by them when they were untouched. The subject of this study is to prove the shape and level of the harm caused by Hedera helix L. on ancient ruins of Tlos. At the same time, this study will underline the fighting methods against Hedera helix L. by comparing similar studies in other countries. Knowledge collected after this study will offer an insight into the excavation and restoration studies undertaken in all Mediterranean countries.

  10. A genome-wide survey on basic helix-loop-helix transcription factors in giant panda.

    Directory of Open Access Journals (Sweden)

    Chunwang Dang

    Full Text Available The giant panda (Ailuropoda melanoleuca is a critically endangered mammalian species. Studies on functions of regulatory proteins involved in developmental processes would facilitate understanding of specific behavior in giant panda. The basic helix-loop-helix (bHLH proteins play essential roles in a wide range of developmental processes in higher organisms. bHLH family members have been identified in over 20 organisms, including fruit fly, zebrafish, mouse and human. Our present study identified 107 bHLH family members being encoded in giant panda genome. Phylogenetic analyses revealed that they belong to 44 bHLH families with 46, 25, 15, 4, 11 and 3 members in group A, B, C, D, E and F, respectively, while the remaining 3 members were assigned into "orphan". Compared to mouse, the giant panda does not encode seven bHLH proteins namely Beta3a, Mesp2, Sclerax, S-Myc, Hes5 (or Hes6, EBF4 and Orphan 1. These results provide useful background information for future studies on structure and function of bHLH proteins in the regulation of giant panda development.

  11. Expanding the diversity of oenococcal bacteriophages: insights into a novel group based on the integrase sequence.

    Science.gov (United States)

    Jaomanjaka, Fety; Ballestra, Patricia; Dols-lafargue, Marguerite; Le Marrec, Claire

    2013-09-02

    Temperate bacteriophages are a contributor of the genetic diversity in the lactic acid bacterium Oenococcus oeni. We used a classification scheme for oenococcal prophages based on integrase gene polymorphism, to analyze a collection of Oenococcus strains mostly isolated in the area of Bordeaux, which represented the major lineages identified through MLST schemes in the species. Genome sequences of oenococcal prophages were clustered into four integrase groups (A to D) which were related to the chromosomal integration site. The prevalence of each group was determined and we could show that members of the intB- and intC-prophage groups were rare in our panel of strains. Our study focused on the so far uncharacterized members of the intD-group. Various intD viruses could be easily isolated from wine samples, while intD lysogens could be induced to produce phages active against two permissive O. oeni isolates. These data support the role of this prophage group in the biology of O. oeni. Global alignment of three relevant intD-prophages revealed significant conservation and highlighted a number of unique ORFs that may contribute to phage and lysogen fitness. © 2013 Elsevier B.V. All rights reserved.

  12. Lead Screening for HIV-1 Integrase (IN Inhibited by Traditional Chinese Medicine

    Directory of Open Access Journals (Sweden)

    Tzu-Chieh Hung

    2014-01-01

    Full Text Available Human immunodeficiency virus causes the acquired immunodeficiency syndrome (AIDS and becomes a serious world-wide problem because of this disease's rapid propagation and incurability. Integrase strand transfer inhibitors (INSTIs supports HIV have rapid drug resistance for antitreatment. Screening the traditional Chinese medicine (TCM database by simulating molecular docking and molecular dynamics may select molecular compounds to inhibit INSTIs against HIV drug resistance. (S-cathinone and (1S,2S-norpseudoephedrine are selected based on structure and ligand-based drugs are designed and then get higher bioactivity predicted score from SVM than Raltegravir and other TCM compounds. The molecular dynamics are helpful in the analysis and detection of protein-ligand interactions. According to the docking poses, hydrophobic interactions and hydrogen bond variations define the main regions of important amino acids in integrase. In addition to the detection of TCM compound efficacy, we suggest (1S,2S-norpseudoephedrine is better than the others based on the analysis of interaction and the effect on the structural variation.

  13. Effect of raltegravir resistance mutations in HIV-1 integrase on viral fitness.

    Science.gov (United States)

    Hu, Zixin; Kuritzkes, Daniel R

    2010-10-01

    Raltegravir resistance is conferred by mutations at integrase codons 143, 148, and 155 together with associated secondary mutations. The N155H mutants emerge first, and are eventually replaced by Q148H mutants, usually in combination with G140S. These mutations have different effects on susceptibility and replication capacity, but data on the relative fitness of RAL-resistant viruses are limited. To understand the impact of the different RAL resistance pathways on viral fitness, mutations at integrase codons 74, 92, 138, 140, 148, 155, and/or 163 were introduced into HIV-1NL4-3 by site-directed mutagenesis and expressed in recombinant viruses. Relative fitness and drug susceptibility were determined in the absence or presence of RAL. In the absence of drug, RAL-resistant mutants were less fit than wild type, and the Q148H mutant was significantly less fit than the N155H mutant. Fitness was partially restored by the presence of additional RAL resistance mutations at positions G140S and E92Q or E138K, respectively. In the presence of RAL, the N155H mutant remained fitter than the Q148H mutant, but the G140S/Q148H double mutant was fitter than single mutants or the E92Q/N155H double mutant. These findings correspond well with the clinical trials data and help explain the temporal pattern of RAL resistance evolution.

  14. Discovery and optimization of 2-pyridinone aminal integrase strand transfer inhibitors for the treatment of HIV.

    Science.gov (United States)

    Schreier, John D; Embrey, Mark W; Raheem, Izzat T; Barbe, Guillaume; Campeau, Louis-Charles; Dubost, David; McCabe Dunn, Jamie; Grobler, Jay; Hartingh, Timothy J; Hazuda, Daria J; Klein, Daniel; Miller, Michael D; Moore, Keith P; Nguyen, Natalie; Pajkovic, Natasa; Powell, David A; Rada, Vanessa; Sanders, John M; Sisko, John; Steele, Thomas G; Wai, John; Walji, Abbas; Xu, Min; Coleman, Paul J

    2017-05-01

    HIV integrase strand transfer inhibitors (InSTIs) represent an important class of antiviral therapeutics with proven efficacy and excellent tolerability for the treatment of HIV infections. In 2007, Raltegravir became the first marketed strand transfer inhibitor pioneering the way to a first-line therapy for treatment-naïve patients. Challenges with this class of therapeutics remain, including frequency of the dosing regimen and the genetic barrier to resistance. To address these issues, research towards next-generation integrase inhibitors has focused on imparting potency against RAL-resistent mutants and improving pharmacokinetic profiles. Herein, we detail medicinal chemistry efforts on a novel class of 2-pyridinone aminal InSTIs, inpsired by MK-0536, which led to the discovery of important lead molecules for our program. Systematic optimization carried out at the amide and aminal positions on the periphery of the core provided the necessary balance of antiviral activity and physiochemical properties. These efforts led to a novel aminal lead compound with the desired virological profile and preclinical pharmacokinetic profile to support a once-daily human dose prediction. Copyright © 2017 Elsevier Ltd. All rights reserved.

  15. Contribution of HIV minority variants to drug resistance in an integrase strand transfer inhibitor-based therapy

    Czech Academy of Sciences Publication Activity Database

    Weber, Jan; Gibson, R. M.; Meyer, A. M.; Winner, D.; Robertson, D. L.; Miller, M. D.; Quinones-Mateu, M. E.

    2013-01-01

    Roč. 18, Suppl. 1 (2013), A66-A66 ISSN 1359-6535. [International Workshop on HIV & Hepatitis Virus Drug Resistance Curative Strategies. 04.06.2013-08.06.2013, Toronto] Institutional support: RVO:61388963 Keywords : HIV minority variants * integrase inhibitor * replicative fitness Subject RIV: CE - Biochemistry

  16. Novel Bifunctional Quinolonyl Diketo Acid Derivatives as HIV-1 Integrase Inhibitors: Design, Synthesis, Biological Activities and Mechanism of Action

    Science.gov (United States)

    Di Santo, Roberto; Costi, Roberta; Roux, Alessandra; Artico, Marino; Lavecchia, Antonio; Marinelli, Luciana; Novellino, Ettore; Palmisano, Lucia; Andreotti, Mauro; Amici, Roberta; Galluzzo, Clementina Maria; Nencioni, Lucia; Palamara, Anna Teresa; Pommier, Yves; Marchand, Christophe

    2008-01-01

    The virally encoded integrase protein is an essential enzyme in the life cycle of the HIV-1 virus and represents an attractive and validated target in the development of therapeutics against HIV infection. Drugs that selectively inhibit this enzyme, when used in combination with inhibitors of reverse transcriptase and protease, are believed to be highly effective in suppressing the viral replication. Among the HIV-1 integrase inhibitors, the β-diketo acids (DKAs) represent a major lead for anti-HIV-1drug development. In this study, novel bifunctional quinolonyl diketo acid derivatives were designed, synthesized and tested for their inhibitory ability against HIV-1 integrase. The compounds are potent inhibitors of integrase activity. Particularly, derivative 8 is a potent IN inhibitor for both steps of the reaction (3′-processing and strand transfer) and exhibits both high antiviral activity against HIV-1 infected cells and low cytotoxicity. Molecular modeling studies provide a plausible mechanism of action, which is consistent with ligand SARs and enzyme photo-crosslinking experiments. PMID:16539381

  17. Nanobody-based chimeric receptor gene integration in Jurkat cells mediated by PhiC31 integrase

    Energy Technology Data Exchange (ETDEWEB)

    Iri-Sofla, Farnoush Jafari [Department of Medical Biotechnology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran (Iran, Islamic Republic of); Rahbarizadeh, Fatemeh, E-mail: rahbarif@modares.ac.ir [Department of Medical Biotechnology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran (Iran, Islamic Republic of); Ahmadvand, Davoud [Center of Pharmaceutical Nanotechnology and Nanotoxicology, Department of Pharmaceutics and Analytical Chemistry, University of Copenhagen, Universitetsparken 2, DK-2100 Copenhagen O (Denmark); Rasaee, Mohammad J. [Department of Medical Biotechnology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran (Iran, Islamic Republic of)

    2011-11-01

    The crucial role of T lymphocytes in anti-tumor immunity has led to the development of novel strategies that can target and activate T cells against tumor cells. Recombinant DNA technology has been used to generate non-MHC-restricted chimeric antigen receptors (CARs). Here, we constructed a panel of recombinant CAR that harbors the anti-MUC1 nanobody and the signaling and co-signaling moieties (CD3{zeta}/CD28) with different spacer regions derived from human IgG3 with one or two repeats of the hinge sequence or the hinge region of Fc{gamma}RII. The PhiC31 integrase system was employed to investigate if the recombination efficiency could be recruited for high and stable expression of T cell chimeric receptor genes. The effect of nuclear localization signal (NLS) and two different promoters (CMV and CAG) on efficacy of PhiC31 integrase in human T cell lines was evaluated. The presence of integrase in combination with NLS, mediated up to 7.6 and 8.5 fold increases in CAR expression in ZCHN-attB and ZCHHN-attB cassette integrated T cells, respectively. Our results showed that highly efficient and stable transduction of the Jurkat cell line by PhiC31 integrase is a feasible modality for generating anti-cancer chimeric T cells for use in cancer immunotherapy.

  18. Nanobody-based chimeric receptor gene integration in Jurkat cells mediated by PhiC31 integrase

    International Nuclear Information System (INIS)

    Iri-Sofla, Farnoush Jafari; Rahbarizadeh, Fatemeh; Ahmadvand, Davoud; Rasaee, Mohammad J.

    2011-01-01

    The crucial role of T lymphocytes in anti-tumor immunity has led to the development of novel strategies that can target and activate T cells against tumor cells. Recombinant DNA technology has been used to generate non-MHC-restricted chimeric antigen receptors (CARs). Here, we constructed a panel of recombinant CAR that harbors the anti-MUC1 nanobody and the signaling and co-signaling moieties (CD3ζ/CD28) with different spacer regions derived from human IgG3 with one or two repeats of the hinge sequence or the hinge region of FcγRII. The PhiC31 integrase system was employed to investigate if the recombination efficiency could be recruited for high and stable expression of T cell chimeric receptor genes. The effect of nuclear localization signal (NLS) and two different promoters (CMV and CAG) on efficacy of PhiC31 integrase in human T cell lines was evaluated. The presence of integrase in combination with NLS, mediated up to 7.6 and 8.5 fold increases in CAR expression in ZCHN-attB and ZCHHN-attB cassette integrated T cells, respectively. Our results showed that highly efficient and stable transduction of the Jurkat cell line by PhiC31 integrase is a feasible modality for generating anti-cancer chimeric T cells for use in cancer immunotherapy.

  19. Sample Extraction Bsaed on Helix Scattering for Polarimetric SAR Calibratio

    Science.gov (United States)

    Chang, Y.; Yang, J.; Li, P.; Zhao, L.; Shi, L.

    2017-09-01

    Polarimetric calibration (PolCAL) of Synthetic Aperture Radar (SAR) images is a significant preprocessing for further applications. Since the reflection symmetry property of distributed objects can provide stable constraints for PolCAL. It is reasonable to extract these reference samples before calibration. The helix scattering generally appears in complex urban area and disappears for a natural scatterer, making it a good measure to extract distributed objects. In this paper, a novel technique that extracts reflecting symmetry samples is proposed by using helix scattering. The helix scattering information is calculated by Yamaguchi four-component decomposition algorithm. An adaptive threshold selection algorithm based on generalized Gaussian distribution is also utilized to scale the helix scattering components automatically, getting rid of the problem of various numerical range. The extracting results will be taken as PolCAL reference samples and the Quegan method are utilized to calibrate these PolSAR images. A C-band airborne PolSAR data was taken as examples to evaluate its ability in improving calibration precision. Traditional method i.e. extracting samples with span power was also evaluated as contrast experiment. The results showed that the samples extracting method based on helix scattering can improve the Polcal precision preferably.

  20. SAMPLE EXTRACTION BSAED ON HELIX SCATTERING FOR POLARIMETRIC SAR CALIBRATIO

    Directory of Open Access Journals (Sweden)

    Y. Chang

    2017-09-01

    Full Text Available Polarimetric calibration (PolCAL of Synthetic Aperture Radar (SAR images is a significant preprocessing for further applications. Since the reflection symmetry property of distributed objects can provide stable constraints for PolCAL. It is reasonable to extract these reference samples before calibration. The helix scattering generally appears in complex urban area and disappears for a natural scatterer, making it a good measure to extract distributed objects. In this paper, a novel technique that extracts reflecting symmetry samples is proposed by using helix scattering. The helix scattering information is calculated by Yamaguchi four-component decomposition algorithm. An adaptive threshold selection algorithm based on generalized Gaussian distribution is also utilized to scale the helix scattering components automatically, getting rid of the problem of various numerical range. The extracting results will be taken as PolCAL reference samples and the Quegan method are utilized to calibrate these PolSAR images. A C-band airborne PolSAR data was taken as examples to evaluate its ability in improving calibration precision. Traditional method i.e. extracting samples with span power was also evaluated as contrast experiment. The results showed that the samples extracting method based on helix scattering can improve the Polcal precision preferably.

  1. Novel 3′-Processing Integrase Activity Assay by Real-Time PCR for Screening and Identification of HIV-1 Integrase Inhibitors

    Directory of Open Access Journals (Sweden)

    Supachai Sakkhachornphop

    2015-01-01

    Full Text Available The 3′-end processing (3′P of each viral long terminal repeat (LTR during human immunodeficiency virus type-1 (HIV-1 integration is a vital step in the HIV life cycle. Blocking the 3′P using 3′P inhibitor has recently become an attractive strategy for HIV-1 therapeutic intervention. Recently, we have developed a novel real-time PCR based assay for the detection of 3′P activity in vitro. The methodology usually involves biotinylated HIV-1 LTR, HIV-1 integrase (IN, and specific primers and probe. In this novel assay, we designed the HIV-1 LTR substrate based on a sequence with a homology to HIV-1 LTR labeled at its 3′ end with biotin on the sense strand. Two nucleotides at the 3′ end were subsequently removed by IN activity. Only two nucleotides labeled biotin were captured on an avidin-coated tube; therefore, inhibiting the binding of primers and probe results in late signals in the real-time PCR. This novel assay has successfully detected both the 3′P activity of HIV-1 IN and the anti-IN activity by Raltegravir and sodium azide agent. This real-time PCR assay has been shown to be effective and inexpensive for a high-throughput screening of novel IN inhibitors.

  2. Addition of E138K to R263K in HIV integrase increases resistance to dolutegravir, but fails to restore activity of the HIV integrase enzyme and viral replication capacity.

    Science.gov (United States)

    Mesplède, Thibault; Osman, Nathan; Wares, Melissa; Quashie, Peter K; Hassounah, Said; Anstett, Kaitlin; Han, Yingshan; Singhroy, Diane N; Wainberg, Mark A

    2014-10-01

    The results of several clinical trials suggest that the integrase inhibitor dolutegravir may be less prone than other drugs to the emergence of HIV drug resistance mutations in treatment-naive patients. We have shown that the R263K mutation commonly emerged during tissue culture selection studies with dolutegravir and conferred low levels of resistance to this drug while simultaneously diminishing both HIV replication capacity and integrase enzymatic activity. E138K has been identified as a secondary mutation for dolutegravir in selection studies and has also been observed as a secondary mutation in the clinic for the integrase inhibitors raltegravir and elvitegravir. We used biochemical cell-free strand-transfer assays and tissue culture assays to characterize the effects of the E138K/R263K combination of mutations on resistance to dolutegravir, integrase enzyme activity and HIV-1 replication capacity. We show here that the addition of the E138K substitution to R263K increased the resistance of HIV-1 to dolutegravir but failed to restore viral replication capacity, integrase strand-transfer activity and integration within cellular DNA. We also show that the addition of E138K to R263K did not increase the resistance to raltegravir or elvitegravir. The addition of the E138K substitution to R263K was also less detrimental to integrase strand-transfer activity and integration than a different secondary mutation at position H51Y that had also been selected in culture. The E138K substitution failed to restore the defect in viral replication capacity that is associated with R263K, confirming previous selection studies that failed to identify compensatory mutation(s) for the latter primary mutation. This study suggests that the R263K resistance pathway may represent an evolutionary dead end for HIV in treatment-naive individuals who are treated with dolutegravir and will need to be confirmed by the long-term use of dolutegravir in the clinic. © The Author 2014. Published

  3. Site-specific integrase-mediated transgenesis in mice via pronuclear injection

    Science.gov (United States)

    Tasic, Bosiljka; Hippenmeyer, Simon; Wang, Charlene; Gamboa, Matthew; Zong, Hui; Chen-Tsai, Yanru; Luo, Liqun

    2011-01-01

    Microinjection of recombinant DNA into zygotic pronuclei has been widely used for producing transgenic mice. However, with this method, the insertion site, integrity, and copy number of the transgene cannot be controlled. Here, we present an integrase-based approach to produce transgenic mice via pronuclear injection, whereby an intact single-copy transgene can be inserted into predetermined chromosomal loci with high efficiency (up to 40%), and faithfully transmitted through generations. We show that neighboring transgenic elements and bacterial DNA within the transgene cause profound silencing and expression variability of the transgenic marker. Removal of these undesirable elements leads to global high-level marker expression from transgenes driven by a ubiquitous promoter. We also obtained faithful marker expression from a tissue-specific promoter. The technique presented here will greatly facilitate murine transgenesis and precise structure/function dissection of mammalian gene function and regulation in vivo. PMID:21464299

  4. Design, Synthesis and Structure-activity Studies of Rhodanine Derivatives as HIV-1 Integrase Inhibitors

    Directory of Open Access Journals (Sweden)

    Kavya Ramkumar

    2010-06-01

    Full Text Available Raltegravir was the first HIV-1 integrase inhibitor that gained FDA approval for use in the treatment of HIV-1 infection. Because of the emergence of IN inhibitor-resistant viral strains, there is a need to identify innovative second-generation IN inhibitors. Previously, we identified 2-thioxo-4-thiazolidinone (rhodanine-containing compounds as IN inhibitors. Herein, we report the design, synthesis and docking studies of a series of novel rhodanine derivatives as IN inhibitors. All these compounds were further tested against human apurinic/apyrimidinic endonuclease 1 (APE1 to determine their selectivity. Two compounds showed significant cytotoxicity in a panel of human cancer cell lines. Taken together, our results show that rhodanines are a promising class of compounds for developing drugs with antiviral and anticancer properties.

  5. Comparative docking and CoMFA analysis of curcumine derivatives as HIV-1 integrase inhibitors.

    Science.gov (United States)

    Gupta, Pawan; Garg, Prabha; Roy, Nilanjan

    2011-08-01

    The docking studies and comparative molecular field analysis (CoMFA) were performed on highly active molecules of curcumine derivatives against 3' processing activity of HIV-1 integrase (IN) enzyme. The optimum CoMFA model was selected with statistically significant cross-validated r(2) value of 0.815 and non-cross validated r (2) value of 0.99. The common pharmacophore of highly active molecules was used for screening of HIV-1 IN inhibitors. The high contribution of polar interactions in pharmacophore mapping is well supported by docking and CoMFA results. The results of docking, CoMFA, and pharmacophore mapping give structural insights as well as important binding features of curcumine derivatives as HIV-1 IN inhibitors which can provide guidance for the rational design of novel HIV-1 IN inhibitors.

  6. QSAR and docking studies of coumarin derivatives as potent HIV-1 integrase inhibitors

    Directory of Open Access Journals (Sweden)

    V.K. Srivastav

    2017-02-01

    Full Text Available Human immunodeficiency virus integrase (HIV-1IN is an emerging and potential drug target for anti-HIV therapy. It is an enzyme essential for 3′ processing and integration step in the life cycle of HIV. In the present study a series of coumarin derivatives (containing 26 compounds as HIV-1IN inhibitors was subjected to quantitative structure–activity relationship (QSAR analysis. For building the regression models two different variable selection approaches namely, genetic function approximation (GFA and sequential multiple linear regression (SQ-MLR were used and compared to predict the HIV-1IN inhibition activity. Based on prediction, the best validation model for 3′ processing inhibition activity with squared correlation coefficient (r2 = 0.8965, cross validated correlation coefficient (Q2 = 0.8307 and external prediction ability pred_r2 = 0.5400 showed that Henry’s law Constant (HLC, Partition Coefficient (PC and Dipole moment-Z component (D3 were the positive contributors, whereas for integration inhibition activity, parameters r2 = 0.8904, Q2 = 0.8174 and pred_r2 = 0.7159 showed HLC, Logarithm of Partition Coefficient (LogP and Dipole moment-Y component (D2 contributed positively to the activity. The binding mode pattern of the compounds to the binding site of integrase enzyme was confirmed by docking studies. The results of the present study may be useful for designing more potent HIV-1IN inhibitors.

  7. Resistance mutations against dolutegravir in HIV integrase impair the emergence of resistance against reverse transcriptase inhibitors.

    Science.gov (United States)

    Oliveira, Maureen; Mesplède, Thibault; Quashie, Peter K; Moïsi, Daniela; Wainberg, Mark A

    2014-03-27

    Among 1222 antiretroviral-naive patients who received dolutegravir (DTG) as part of first-line therapy, none has developed resistance against this compound after 48-96 weeks of follow-up. Moreover, only four occurrences of virological failure with resistance mutations have been documented in previously drug-experienced patients who received DTG as a first time integrase inhibitor as a component of a second-line regimen. The R263K integrase resistance mutation was observed in two of these individuals who received suboptimal background regimens. We have previously selected mutations at position R263K, G118R, H51Y, and E138K as being associated with low-level resistance to DTG. Now, we sought to investigate the facility with which resistance on the part of R263K-containing viruses might develop. We tested the ability of DTG-resistant viruses containing either the R263K or G118R and/or H51Y mutations to develop further resistance against several reverse transcriptase inhibitors during in-vitro selection experiments. Our results show that DTG-resistant viruses are impaired in their ability to acquire further resistance to each of nevirapine and lamivudine as a consequence of their relative inability to develop resistance mutations associated with these two compounds. Our findings provide an explanation for the fact that no individual has yet progressed to virological failure with resistance mutations associated with dolutegravir in clinical trials in which patients received dolutegravir together with an optimized background regimen.

  8. Antiviral Activity of Bictegravir and Cabotegravir against Integrase Inhibitor-Resistant SIVmac239 and HIV-1.

    Science.gov (United States)

    Hassounah, Said A; Alikhani, Ahmad; Oliveira, Maureen; Bharaj, Simrat; Ibanescu, Ruxandra-Ilinca; Osman, Nathan; Xu, Hong-Tao; Brenner, Bluma G; Mesplède, Thibault; Wainberg, Mark A

    2017-12-01

    Animal models are essential to study novel antiretroviral drugs, resistance-associated mutations (RAMs), and treatment strategies. Bictegravir (BIC) is a novel potent integrase strand transfer inhibitor (INSTI) that has shown promising results against HIV-1 infection in vitro and in vivo and against clinical isolates with resistance against INSTIs. BIC has a higher genetic barrier to the development of resistance than two clinically approved INSTIs, termed raltegravir and elvitegravir. Another clinically approved INSTI, dolutegravir (DTG) also possesses a high genetic barrier to resistance, while a fourth compound, termed cabotegravir (CAB), is currently in late phases of clinical development. Here we report the susceptibilities of simian immunodeficiency virus (SIV) and HIV-1 integrase (IN) mutants containing various RAMs to BIC, CAB, and DTG. BIC potently inhibited SIV and HIV-1 in single cycle infection with 50% effective concentrations (EC 50 s) in the low nM range. In single cycle SIV infections, none of the E92Q, T97A, Y143R, or N155H substitutions had a significant effect on susceptibility to BIC (≤4-fold increase in EC 50 ), whereas G118R and R263K conferred ∼14-fold and ∼6-fold increases in EC 50 , respectively. In both single and multiple rounds of HIV-1 infections, BIC remained active against the Y143R, N155H, R263K, R263K/M50I, and R263K/E138K mutants (≤4-fold increase in EC 50 ). In multiple rounds of infection, the G140S/Q148H combination of substitutions decreased HIV-1 susceptibility to BIC 4.8-fold compared to 16.8- and 7.4-fold for CAB and DTG, respectively. BIC possesses an excellent resistance profile in regard to HIV and SIV and could be useful in nonhuman primate models of HIV infection. Copyright © 2017 American Society for Microbiology.

  9. Simian-tropic HIV as a model to study drug resistance against integrase inhibitors.

    Science.gov (United States)

    Wares, Melissa; Hassounah, Said; Mesplède, Thibault; Sandstrom, Paul A; Wainberg, Mark A

    2015-04-01

    Drug resistance represents a key aspect of human immunodeficiency virus (HIV) treatment failure. It is important to develop nonhuman primate models for studying issues of drug resistance and the persistence and transmission of drug-resistant viruses. However, relatively little work has been conducted using either simian immunodeficiency virus (SIV) or SIV/HIV recombinant viruses for studying resistance against integrase strand transfer inhibitors (INSTIs). Here, we used a T-cell-tropic SIV/HIV recombinant virus in which the capsid and vif regions of HIV-1 were replaced with their SIV counterparts (simian-tropic HIV-1 [stHIV-1](SCA,SVIF)) to study the impact of a number of drug resistance substitutions in the integrase coding region at positions E92Q, G118R, E138K, Y143R, S153Y, N155H, and R263K on drug resistance, viral infectivity, and viral replication capacity. Our results show that each of these substitutions exerted effects that were similar to their effects in HIV-1. Substitutions associated with primary resistance against dolutegravir were more detrimental to stHIV-1(SCA,SVIF) infectiousness than were resistance substitutions associated with raltegravir and elvitegravir, consistent with data that have been reported for HIV-1. These findings support the role of stHIV-1(SCA,SVIF) as a useful model with which to evaluate the role of INSTI resistance substitutions on viral persistence, transmissibility, and pathogenesis in a nonhuman primate model. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  10. In vivo and in vitro characterization of site-specific recombination of a novel serine integrase from the temperate phage EFC-1

    Energy Technology Data Exchange (ETDEWEB)

    Yoon, Bohyun; Kim, Inki; Nam, Ja-Ae [Asan Institute for Life Sciences, Asan Medical Center, College of Medicine, University of Ulsan, 86 Asanbyeoungwon-gil, Songpa-gu, Seoul 138-736 (Korea, Republic of); Chang, Hyo-Ihl [College of Life Sciences & Biotechnology, Korea University, 5-1 Anam-dong, Seongbuk-gu, Seoul 136-701 (Korea, Republic of); Ha, Chang Hoon, E-mail: chhoonha@amc.seoul.kr [Asan Institute for Life Sciences, Asan Medical Center, College of Medicine, University of Ulsan, 86 Asanbyeoungwon-gil, Songpa-gu, Seoul 138-736 (Korea, Republic of)

    2016-04-22

    EFC-1 integrase is a site-specific recombinase that belongs to the large family of serine recombinase. In previously study, we isolated the temperate phage EFC-1, and characterized its genomic sequence. Within its genome, Orf28 was predicted encode a 464 amino acid of a putative integrase gene. In this study, EFC-1 integrase was characterized in vitro and in vivo. In vitro assay was performed using purified His-tag fusion integrase. Also, to identify which serine is involved in the catalytic domain, we used site-directed mutagenesis and analyzed by a recombination assay in vitro. In vivo assay involved PCR and confocal microscopy in HEK293 cells, and determined the minimal lengths of attP and attB sites. According to our results, the EFC-1 integrase-mediated recombination was site-specific and unidirectional system in vitro and in vivo. Serine 21 of EFC-1 integrase plays a major role in the catalytic domain, and minimal sizes of attB and attP was defined 48 and 54 bp. Our finding may help develop a useful tool for gene therapy and gene delivery system. - Highlights: • EFC-1 integrase-mediated recombination was site-specific and unidirectional system. • Serine 21 of EFC-1 integrase plays a major role in the catalytic domain. • The functional minimal sizes of attB and attP was defined 48 and 54 bp.

  11. Tight regulation of the intS gene of the KplE1 prophage: a new paradigm for integrase gene regulation.

    Directory of Open Access Journals (Sweden)

    Gaël Panis

    2010-10-01

    Full Text Available Temperate phages have the ability to maintain their genome in their host, a process called lysogeny. For most, passive replication of the phage genome relies on integration into the host's chromosome and becoming a prophage. Prophages remain silent in the absence of stress and replicate passively within their host genome. However, when stressful conditions occur, a prophage excises itself and resumes the viral cycle. Integration and excision of phage genomes are mediated by regulated site-specific recombination catalyzed by tyrosine and serine recombinases. In the KplE1 prophage, site-specific recombination is mediated by the IntS integrase and the TorI recombination directionality factor (RDF. We previously described a sub-family of temperate phages that is characterized by an unusual organization of the recombination module. Consequently, the attL recombination region overlaps with the integrase promoter, and the integrase and RDF genes do not share a common activated promoter upon lytic induction as in the lambda prophage. In this study, we show that the intS gene is tightly regulated by its own product as well as by the TorI RDF protein. In silico analysis revealed that overlap of the attL region with the integrase promoter is widely encountered in prophages present in prokaryotic genomes, suggesting a general occurrence of negatively autoregulated integrase genes. The prediction that these integrase genes are negatively autoregulated was biologically assessed by studying the regulation of several integrase genes from two different Escherichia coli strains. Our results suggest that the majority of tRNA-associated integrase genes in prokaryotic genomes could be autoregulated and that this might be correlated with the recombination efficiency as in KplE1. The consequences of this unprecedented regulation for excessive recombination are discussed.

  12. Npas4, a novel helix-loop-helix PAS domain protein, is regulated in response to cerebral ischemia

    DEFF Research Database (Denmark)

    Shamloo, Mehrdad; Soriano, Liza; von Schack, David

    2006-01-01

    Basic helix-loop-helix PAS domain proteins form a growing family of transcription factors. These proteins are involved in the process of adaptation to cellular stresses and environmental factors such as a change in oxygen concentration. We describe the identification and characterization of a rec...... lead to a decrease in the 200 kDa form and a simultaneous increase in the 100 kDa immunoreactivity. This could indicate a novel regulatory mechanism for activation and/or deactivation of this protein in response to ischemic brain injury....

  13. An effective HIV-1 integrase inhibitor screening platform: Rationality validation of drug screening, conformational mobility and molecular recognition analysis for PFV integrase complex with viral DNA.

    Science.gov (United States)

    Du, Wenyi; Zuo, Ke; Sun, Xin; Liu, Wei; Yan, Xiao; Liang, Li; Wan, Hua; Chen, Fengzheng; Hu, Jianping

    2017-11-01

    As an important target for the development of novel anti-AIDS drugs, HIV-1 integrase (IN) has been widely concerned. However, the lack of a complete accurate crystal structure of HIV-1 IN greatly blocks the discovery of novel inhibitors. In this work, an effective HIV-1 IN inhibitor screening platform, namely PFV IN, was filtered from all species of INs. Next, the 40.8% similarity with HIV-1 IN, as well as the high efficiency of virtual screening and the good agreement between calculated binding free energies and experimental ones all proved PFV IN is a promising screening platform for HIV-1 IN inhibitors. Then, the molecular recognition mechanism of PFV IN by its substrate viral DNA and six naphthyridine derivatives (NRDs) inhibitors was investigated through molecular docking, molecular dynamics simulations and water-mediated interactions analyses. The functional partition of NRDs IN inhibitors could be divided into hydrophobic and hydrophilic ones, and the Mg 2+ ions, water molecules and conserved DDE motif residues all interacted with the hydrophilic partition, while the bases in viral DNA and residues like Tyr212, Pro214 interacted with the hydrophobic one. Finally, the free energy landscape (FEL) and cluster analyses were performed to explore the molecular motion of PFV IN-DNA system. It is found that the association with NRDs inhibitors would obviously decrease the motion amplitude of PFV IN-DNA, which may be one of the most potential mechanisms of IN inhibitors. This work will provide a theoretical basis for the inhibitor design based on the structure of HIV-1 IN. Copyright © 2017 Elsevier Inc. All rights reserved.

  14. Nucleic acid helix structure determination from NMR proton chemical shifts

    International Nuclear Information System (INIS)

    Werf, Ramon M. van der; Tessari, Marco; Wijmenga, Sybren S.

    2013-01-01

    We present a method for de novo derivation of the three-dimensional helix structure of nucleic acids using non-exchangeable proton chemical shifts as sole source of experimental restraints. The method is called chemical shift de novo structure derivation protocol employing singular value decomposition (CHEOPS) and uses iterative singular value decomposition to optimize the structure in helix parameter space. The correct performance of CHEOPS and its range of application are established via an extensive set of structure derivations using either simulated or experimental chemical shifts as input. The simulated input data are used to assess in a defined manner the effect of errors or limitations in the input data on the derived structures. We find that the RNA helix parameters can be determined with high accuracy. We finally demonstrate via three deposited RNA structures that experimental proton chemical shifts suffice to derive RNA helix structures with high precision and accuracy. CHEOPS provides, subject to further development, new directions for high-resolution NMR structure determination of nucleic acids.

  15. Probabilistic grammatical model for helix‐helix contact site classification

    Science.gov (United States)

    2013-01-01

    Background Hidden Markov Models power many state‐of‐the‐art tools in the field of protein bioinformatics. While excelling in their tasks, these methods of protein analysis do not convey directly information on medium‐ and long‐range residue‐residue interactions. This requires an expressive power of at least context‐free grammars. However, application of more powerful grammar formalisms to protein analysis has been surprisingly limited. Results In this work, we present a probabilistic grammatical framework for problem‐specific protein languages and apply it to classification of transmembrane helix‐helix pairs configurations. The core of the model consists of a probabilistic context‐free grammar, automatically inferred by a genetic algorithm from only a generic set of expert‐based rules and positive training samples. The model was applied to produce sequence based descriptors of four classes of transmembrane helix‐helix contact site configurations. The highest performance of the classifiers reached AUCROC of 0.70. The analysis of grammar parse trees revealed the ability of representing structural features of helix‐helix contact sites. Conclusions We demonstrated that our probabilistic context‐free framework for analysis of protein sequences outperforms the state of the art in the task of helix‐helix contact site classification. However, this is achieved without necessarily requiring modeling long range dependencies between interacting residues. A significant feature of our approach is that grammar rules and parse trees are human‐readable. Thus they could provide biologically meaningful information for molecular biologists. PMID:24350601

  16. The Triple Helix Model and the Knowledge-Based Economy

    NARCIS (Netherlands)

    Leydesdorff, L.; Meyer, M.

    2010-01-01

    The Triple Helix model of university-industry-government relations can be generalized from a neo-institutional model of networks of relations to a neo-evolutionary model of how three selection environments operate upon one another. Two selection mechanisms operating upon each other can mutually

  17. Living Labs as boundary-spanners between Triple Helix actors

    NARCIS (Netherlands)

    van Geenhuizen, M.S.

    2016-01-01

    Living labs are an increasingly popular methodology to enhance innovation. Living labs aim to span boundaries between different organizations, among others Triple helix actors, by acting as a network organization typically in a real-life environment to foster co-creation by user-groups. This paper

  18. Cloning, purification and structure determination of the HIV integrase-binding domain of lens epithelium-derived growth factor.

    Science.gov (United States)

    Hannon, Clare; Cruz-Migoni, Abimael; Platonova, Olga; Owen, Robin L; Nettleship, Joanne E; Miller, Ami; Carr, Stephen B; Harris, Gemma; Rabbitts, Terence H; Phillips, Simon E V

    2018-03-01

    Lens epithelium-derived growth factor (LEDGF)/p75 is the dominant binding partner of HIV-1 integrase in human cells. The crystal structure of the HIV integrase-binding domain (IBD) of LEDGF has been determined in the absence of ligand. IBD was overexpressed in Escherichia coli, purified and crystallized by sitting-drop vapour diffusion. X-ray diffraction data were collected at Diamond Light Source to a resolution of 2.05 Å. The crystals belonged to space group P2 1 , with eight polypeptide chains in the asymmetric unit arranged as an unusual octamer composed of four domain-swapped IBD dimers. IBD exists as a mixture of monomers and dimers in concentrated solutions, but the dimers are unlikely to be biologically relevant.

  19. Site-directed zebrafish transgenesis into single landing sites with the phiC31 integrase system.

    Science.gov (United States)

    Mosimann, Christian; Puller, Ann-Christin; Lawson, Katy L; Tschopp, Patrick; Amsterdam, Adam; Zon, Leonard I

    2013-08-01

    Linear DNA-based and Tol2-mediated transgenesis are powerful tools for the generation of transgenic zebrafish. However, the integration of multiple copies or transgenes at random genomic locations complicates comparative transgene analysis and makes long-term transgene stability unpredictable with variable expression. Targeted, site-directed transgene integration into pre-determined genomic loci can circumvent these issues. The phiC31 integrase catalyzes the unidirectional recombination reaction between heterotypic attP and attB sites and is an efficient platform for site-directed transgenesis. We report the implementation of the phiC31 integrase-mediated attP/attB recombination for site-directed zebrafish transgenics of attB-containing transgene vectors into single genomic attP landing sites. We generated Tol2-based single-insertion attP transgenic lines and established their performance in phiC31 integrase-catalyzed integration of an attB-containing transgene vector. We found stable germline transmission into the next generation of an attB reporter transgene in 34% of all tested animals. We further characterized two functional attP landing site lines and determined their genomic location. Our experiments also demonstrate tissue-specific transgene applications as well as long-term stability of phiC31-mediated transgenes. Our results establish phiC31 integrase-controlled site-directed transgenesis into single, genomic attP sites as space-, time-, and labor-efficient zebrafish transgenesis technique. The described reagents are available for distribution to the zebrafish community. Copyright © 2013 Wiley Periodicals, Inc.

  20. Identification of a hexapeptide inhibitor of the human immunodeficiency virus integrase protein by using a combinatorial chemical library.

    OpenAIRE

    Puras Lutzke, R A; Eppens, N A; Weber, P A; Houghten, R A; Plasterk, R H

    1995-01-01

    Integration of human immunodeficiency virus (HIV) DNA into the human genome requires the virus-encoded integrase (IN) protein, and therefore the IN protein is a suitable target for antiviral strategies. To find a potent HIV IN inhibitor, we screened a "synthetic peptide combinatorial library." We identified a hexapeptide with the sequence HCKFWW that inhibits IN-mediated 3'-processing and integration with an IC50 of 2 microM. The peptide is active on IN proteins from other retroviruses such a...

  1. The close-packed triple helix as a possible new structural motif for collagen

    DEFF Research Database (Denmark)

    Bohr, Jakob; Olsen, Kasper

    2010-01-01

    The one-dimensional problem of selecting the triple helix with the highest volume fraction is solved and hence the condition for a helix to be close-packed is obtained. The close-packed triple helix is shown to have a pitch angle of v CP = 43.3°. Contrary to the conventional notion, we suggest th...

  2. Generating Customized Transgene Landing Sites and Multi-Transgene Arrays in Drosophila Using phiC31 Integrase

    Science.gov (United States)

    Knapp, Jon-Michael; Chung, Phuong; Simpson, Julie H.

    2015-01-01

    Transgenesis in numerous eukaryotes has been facilitated by the use of site-specific integrases to stably insert transgenes at predefined genomic positions (landing sites). However, the utility of integrase-mediated transgenesis in any system is constrained by the limited number and variable expression properties of available landing sites. By exploiting the nonstandard recombination activity exhibited by a phiC31 integrase mutant, we developed a rapid and inexpensive method for isolating landing sites that exhibit desired expression properties. Additionally, we devised a simple technique for constructing arrays of transgenes at a single landing site, thereby extending the utility of previously characterized landing sites. Using the fruit fly Drosophila melanogaster, we demonstrate the feasibility of these approaches by isolating new landing sites optimized to express transgenes in the nervous system and by building fluorescent reporter arrays at several landing sites. Because these strategies require the activity of only a single exogenous protein, we anticipate that they will be portable to species such as nonmodel organisms, in which genetic manipulation is more challenging, expediting the development of genetic resources in these systems. PMID:25680812

  3. Secondary mutations in viruses resistant to HIV-1 integrase inhibitors that restore viral infectivity and replication kinetics.

    Science.gov (United States)

    Nakahara, Koichiro; Wakasa-Morimoto, Chiaki; Kobayashi, Masanori; Miki, Shigeru; Noshi, Takeshi; Seki, Takahiro; Kanamori-Koyama, Mikiko; Kawauchi, Shinobu; Suyama, Akemi; Fujishita, Toshio; Yoshinaga, Tomokazu; Garvey, Edward P; Johns, Brian A; Foster, Scott A; Underwood, Mark R; Sato, Akihiko; Fujiwara, Tamio

    2009-02-01

    Passage of HIV-1 in the presence of integrase inhibitors (INIs) generates resistant viruses that have mutations in the integrase region. Integrase-resistant mutations Q148K and Q148R were identified as primary mutations with the passage of HIV-1 IIIB in the presence of INIs S-1360 or S/GSK-364735, respectively. Secondary amino acid substitutions E138K or G140S were observed when passage with INI was continued. The role of these mutations was investigated with molecular clones. Relative to Q148K alone, Q148K/E138K had 2- and >6-fold increases in resistance to S-1360 and S/GSK-364735, respectively, and the double mutant had slightly better infectivity and replication kinetics. In contrast, Q148K/G140S and Q148R/E138K had nearly equivalent or slightly reduced fold resistance to the INI compared with their respective Q148 primary mutants, and had increases in infectivity and replication kinetics. Recovery of these surrogates of viral fitness coincided with the recovery of integration efficiency of viral DNA into the host cell chromosome for these double mutants. These data show that recovery of viral integration efficiency can be an important factor for the emergence and maintenance of INI-resistant mutations.

  4. Development of a receptor model for efficient in silico screening of HIV-1 integrase inhibitors.

    Science.gov (United States)

    Quevedo, Mario A; Ribone, Sergio R; Briñón, Margarita C; Dehaen, Wim

    2014-07-01

    Integrase (IN) is a key viral enzyme for the replication of the type-1 human immunodeficiency virus (HIV-1), and as such constitutes a relevant therapeutic target for the development of anti-HIV agents. However, the lack of crystallographic data of HIV IN complexed with the corresponding viral DNA has historically hindered the application of modern structure-based drug design techniques to the discovery of new potent IN inhibitors (INIs). Consequently, the development and validation of reliable HIV IN structural models that may be useful for the screening of large databases of chemical compounds is of particular interest. In this study, four HIV-1 IN homology models were evaluated respect to their capability to predict the inhibition potency of a training set comprising 36 previously reported INIs with IC50 values in the low nanomolar to the high micromolar range. Also, 9 inactive structurally related compounds were included in this training set. In addition, a crystallographic structure of the IN-DNA complex corresponding to the prototype foamy virus (PFV) was also evaluated as structural model for the screening of inhibitors. The applicability of high throughput screening techniques, such as blind and ligand-guided exhaustive rigid docking was assessed. The receptor models were also refined by molecular dynamics and clustering techniques to assess protein sidechain flexibility and solvent effect on inhibitor binding. Among the studied models, we conclude that the one derived from the X-ray structure of the PFV integrase exhibited the best performance to rank the potencies of the compounds in the training set, with the predictive power being further improved by explicitly modeling five water molecules within the catalytic side of IN. Also, accounting for protein sidechain flexibility enhanced the prediction of inhibition potencies among the studied compounds. Finally, an interaction fingerprint pattern was established for the fast identification of potent IN

  5. Impact of hydrodynamic injection and phiC31 integrase on tumor latency in a mouse model of MYC-induced hepatocellular carcinoma.

    Directory of Open Access Journals (Sweden)

    Lauren E Woodard

    2010-06-01

    Full Text Available Hydrodynamic injection is an effective method for DNA delivery in mouse liver and is being translated to larger animals for possible clinical use. Similarly, phiC31 integrase has proven effective in mediating long-term gene therapy in mice when delivered by hydrodynamic injection and is being considered for clinical gene therapy applications. However, chromosomal aberrations have been associated with phiC31 integrase expression in tissue culture, leading to questions about safety.To study whether hydrodynamic delivery alone, or in conjunction with delivery of phiC31 integrase for long-term transgene expression, could facilitate tumor formation, we used a transgenic mouse model in which sustained induction of the human C-MYC oncogene in the liver was followed by hydrodynamic injection. Without injection, mice had a median tumor latency of 154 days. With hydrodynamic injection of saline alone, the median tumor latency was significantly reduced, to 105 days. The median tumor latency was similar, 106 days, when a luciferase donor plasmid and backbone plasmid without integrase were administered. In contrast, when active or inactive phiC31 integrase and donor plasmid were supplied to the mouse liver, the median tumor latency was 153 days, similar to mice receiving no injection.Our data suggest that phiC31 integrase does not facilitate tumor formation in this C-MYC transgenic mouse model. However, in groups lacking phiC31 integrase, hydrodynamic injection appeared to contribute to C-MYC-induced hepatocellular carcinoma in adult mice. Although it remains to be seen to what extent these findings may be extrapolated to catheter-mediated hydrodynamic delivery in larger species, they suggest that caution should be used during translation of hydrodynamic injection to clinical applications.

  6. Design and synthesis of DNA four-helix bundles

    International Nuclear Information System (INIS)

    Rangnekar, Abhijit; Gothelf, Kurt V; LaBean, Thomas H

    2011-01-01

    The field of DNA nanotechnology has evolved significantly in the past decade. Researchers have succeeded in synthesizing tile-based structures and using them to form periodic lattices in one, two and three dimensions. Origami-based structures have also been used to create nanoscale structures in two and three dimensions. Design and construction of DNA bundles with fixed circumference has added a new dimension to the field. Here we report the design and synthesis of a DNA four-helix bundle. It was found to be extremely rigid and stable. When several such bundles were assembled using appropriate sticky-ends, they formed micrometre-long filaments. However, when creation of two-dimensional sheet-like arrays of the four-helix bundles was attempted, nanoscale rings were observed instead. The exact reason behind the nanoring formation is yet to be ascertained, but it provides an exciting prospect for making programmable circular nanostructures using DNA.

  7. Crowding effect on helix-coil transition: Beyond entropic stabilization

    International Nuclear Information System (INIS)

    Koutsioubas, A.; Lairez, D.; Combet, S.; Longeville, S.; Fadda, G. C.; Zalczer, G.

    2012-01-01

    We report circular dichroism measurements on the helix-coil transition of poly(L-glutamic acid) in solution with polyethylene glycol (PEG) as a crowding agent. The PEG solutions have been characterized by small angle neutron scattering and are well described by the picture of a network of mesh size ξ, usual for semi-dilute chains in good solvent. We show that the increase of PEG concentration stabilizes the helices and increases the transition temperature. But more unexpectedly, we also notice that the increase of concentration of crowding agent reduces the mean helix extent at the transition, or in other words reduces its cooperativity. This result cannot be taken into account for by an entropic stabilization mechanism. Comparing the mean length of helices at the transition and the mesh size of the PEG network, our results strongly suggest two regimes: helices shorter or longer than the mesh size.

  8. Superconducting Helix loaded cavity for heavy ion acceleration

    International Nuclear Information System (INIS)

    Ramstein, G.; Cauvin, B.; Fouan, J.P.

    1987-01-01

    Half-wave helix loaded structures have been designed to increase the mass range of heavy ions which will be accelerated by the CEN Saclay superconducting accelerator. In this paper we shall list the main characteristics of these resonators; especially, the quality factor and the energy gain per charge measured with a 12 C beam. The important advantages of these new structures are the broadness of the transit time factor curve which allows us to accelerate low velocity beams with a sufficient efficiency, and the simplicity of mechanical design which requires only two helix welds. We shall also give the performances achieved by the series of resonators which will be mounted into two of the six machine cryostats. The new superconducting accelerator will provide the first beams at the end of this year [fr

  9. Design and synthesis of DNA four-helix bundles

    Energy Technology Data Exchange (ETDEWEB)

    Rangnekar, Abhijit; Gothelf, Kurt V [Department of Chemistry, Centre for DNA Nanotechnology (CDNA) and Interdisciplinary Nanoscience Center (iNANO), Aarhus University, DK-8000 Aarhus C (Denmark); LaBean, Thomas H, E-mail: kvg@chem.au.dk, E-mail: thl@cs.duke.edu [Department of Chemistry, Duke University, Durham, NC 27708 (United States)

    2011-06-10

    The field of DNA nanotechnology has evolved significantly in the past decade. Researchers have succeeded in synthesizing tile-based structures and using them to form periodic lattices in one, two and three dimensions. Origami-based structures have also been used to create nanoscale structures in two and three dimensions. Design and construction of DNA bundles with fixed circumference has added a new dimension to the field. Here we report the design and synthesis of a DNA four-helix bundle. It was found to be extremely rigid and stable. When several such bundles were assembled using appropriate sticky-ends, they formed micrometre-long filaments. However, when creation of two-dimensional sheet-like arrays of the four-helix bundles was attempted, nanoscale rings were observed instead. The exact reason behind the nanoring formation is yet to be ascertained, but it provides an exciting prospect for making programmable circular nanostructures using DNA.

  10. Single-Molecule Spectroscopic Investigations of Amphipathic Helix Formation

    Science.gov (United States)

    Cunningham, Joy Ann; Okamoto, Kenji; English, Douglas

    2004-03-01

    We are using single molecule spectroscopy to examine surface-induced conformational states occurring through interaction of a polypeptide with an interface. Specifically, we investigate the folding of amphipathic helices by using single-molecule fluorescence resonance energy transfer to construct peptide conformational distributions in solution and at interfaces. Analysis of the conformational distributions and kinetics of peptides in different environments reveals properties of the free energy surface for helix formation at an interface relative to formation in solution.

  11. Chiral transformation: From single nanowire to double helix

    KAUST Repository

    Wang, Yong

    2011-12-21

    We report a new type of water-soluble ultrathin Au-Ag alloy nanowire (NW), which exhibits unprecedented behavior in a colloidal solution. Upon growth of a thin metal (Pd, Pt, or Au) layer, the NW winds around itself to give a metallic double helix. We propose that the winding originates from the chirality within the as-synthesized Au-Ag NWs, which were induced to untwist upon metal deposition. © 2011 American Chemical Society.

  12. Facilitating Quintuple helix innovation with urban living labs

    OpenAIRE

    Baccarne, Bastiaan; Schuurman, Dimitri; De Marez, Lieven

    2015-01-01

    This paper discusses the Urban Living Lab approach as a way to put the Quintuple Helix model for innovation into practice. In this analysis we focus on the concepts innovation democracy, ‘mode 3’ knowledge production, the innovation ecosystem as a system of societal subsystems and socioecological transition. The empirical analysis is performed by means of a multidimensional case study design, applied on a project-based ad hoc collaborative innovation development process in an ecological doma...

  13. Snail mucus - glandular origin and composition in Helix pomatia.

    Science.gov (United States)

    Greistorfer, Sophie; Klepal, Waltraud; Cyran, Norbert; Gugumuck, Andreas; Rudoll, Livia; Suppan, Johannes; von Byern, Janek

    2017-06-01

    Apart from their well-known culinary use, gastropod species such as Helix, which have a hydrogel-like mucus, are increasingly being exploited for cosmetic, bioengineering and medical applications. However, not only are the origin and composition of these "sticky" secretions far from being fully characterized, the number and morphology of the mucus glands involved is also uncertain. This study aims to characterize in detail the cutaneous glands of the Helix pomatia foot on morphological, histochemical and immunohistochemical levels. Hereby the focus is on the gland position and appearance on the foot sole as well as on the chemical nature of the different gland secretions. At least five different gland types can be distinguished by their microanatomy; three are located on the dorsal side and two on the ventral side of the foot sole. Most glands are reactive for acidic proteins and sugars such as mannose and fucose, indicating the presence of acidic glycosaminoglycans. One dorsal gland type shows high reactivity for acidic proteins only. The isolated mucus includes a certain amount of the elements chlorine, potassium and calcium; evidence for lipids was also confirmed in the isolated mucus. The present results for Helix pomatia show a clear difference in the number of glands compared to the related species Helix aspersa (only four mucus glands); histochemically, the glands of both species similarly produce acidic proteins as well as acidic glycosaminoglycans. While calcium ions are known to play a role in mucus formation, the presence and function of other ions such as potassium still need to be clarified. Copyright © 2017 Elsevier GmbH. All rights reserved.

  14. Cellular uptake of Aib-containing amphipathic helix peptide.

    Science.gov (United States)

    Wada, Shun-ichi; Tsuda, Hirokazu; Okada, Terumi; Urata, Hidehito

    2011-10-01

    Cell-penetrating peptides (CPPs) are useful tools for the delivery of hydrophilic bioactive molecules, such as peptides, proteins, and oligonucleotides, across the cell membrane. To realize the delivery of therapeutic macromolecules by CPPs, the CPPs are required to show resistance to protease and no cytotoxicity. In order to produce potent non-toxic and protease-resistant CPPs with high cellular uptake, we designed an amphipathic helix peptide using α-aminoisobutyric acid (Aib, U) and named it MAP(Aib). In the MAP(Aib) molecule, five Aib residues are aligned on the hydrophobic face of the helix and five lysine (K) residues are aligned on the hydrophilic face. MAP(Aib) showed potent resistance to trypsin and pronase compared with MAP, an amphipathic helix peptide formed by usual amino acids. Fluorescein-labeled MAP(Aib) efficiently traversed the A549 cell membrane, diffusing into the cytoplasm and slightly into the nucleus without exerting any cytotoxicity. In contrast, MAP was poorly taken up by the cell. These results indicate that the incorporation of Aib residues into CPPs markedly improves cellular uptake and MAP(Aib) may be a useful tool for the delivery of hydrophilic macromolecules. Copyright © 2011 Elsevier Ltd. All rights reserved.

  15. HIV integrase variability and genetic barrier in antiretroviral naïve and experienced patients

    Directory of Open Access Journals (Sweden)

    Comolli Giuditta

    2011-03-01

    Full Text Available Abstract Background HIV-1 integrase (IN variability in treatment naïve patients with different HIV-1 subtypes is a major issue. In fact, the effect of previous exposure to antiretrovirals other than IN inhibitors (INI on IN variability has not been satisfactorily defined. In addition, the genetic barrier for specific INI resistance mutations remains to be calculated. Methods IN variability was analyzed and compared with reverse transcriptase (RT and protease (PR variability in 41 treatment naïve and 54 RT inhibitor (RTI and protease inhibitor (PRI experienced patients from subjects infected with subtype B and non-B strains. In addition, four HIV-2 strains were analyzed in parallel. Frequency and distribution of IN mutations were compared between HAART-naïve and RTI/PI-experienced patients; the genetic barrier for 27 amino acid positions related to INI susceptibility was calculated as well. Results Primary mutations associated with resistance to INI were not detected in patients not previously treated with this class of drug. However, some secondary mutations which have been shown to contribute to INI resistance were found. Only limited differences in codon usage distribution between patient groups were found. HIV-2 strains from INI naïve patients showed the presence of both primary and secondary resistance mutations. Conclusion Exposure to antivirals other than INI does not seem to significantly influence the emergence of mutations implicated in INI resistance. HIV-2 strain might have reduced susceptibility to INI.

  16. Structural Basis for the Recognition Between HIV-1 Integrase and Transcriptional Coactivator p75

    Energy Technology Data Exchange (ETDEWEB)

    Cherepanov,P.; Ambrosio, A.; Rahman, S.; Ellenberger, T.; Engelman, A.

    2005-01-01

    Integrase (IN) is an essential retroviral enzyme, and human transcriptional coactivator p75, which is also referred to as lens epithelium-derived growth factor (LEDGF), is the dominant cellular binding partner of HIV-1 IN. Here, we report the crystal structure of the dimeric catalytic core domain of HIV-1 IN complexed to the IN-binding domain of LEDGF. Previously identified LEDGF hotspot residues anchor the protein to both monomers at the IN dimer interface. The principal structural features of IN that are recognized by the host factor are the backbone conformation of residues 168-171 from one monomer and a hydrophobic patch that is primarily comprised of {alpha}-helices 1 and 3 of the second IN monomer. Inspection of diverse retroviral primary and secondary sequence elements helps to explain the apparent lentiviral tropism of the LEDGF-IN interaction. Because the lethal phenotypes of HIV-1 mutant viruses unable to interact with LEDGF indicate that IN function is highly sensitive to perturbations of the structure around the LEDGF-binding site, we propose that small molecule inhibitors of the protein-protein interaction might similarly disrupt HIV-1 replication.

  17. Mapping side chain interactions at protein helix termini.

    Science.gov (United States)

    Newell, Nicholas E

    2015-07-25

    Interactions that involve one or more amino acid side chains near the ends of protein helices stabilize helix termini and shape the geometry of the adjacent loops, making a substantial contribution to overall protein structure. Previous work has identified key helix-terminal motifs, such as Asx/ST N-caps, the capping box, and hydrophobic and electrostatic interactions, but important questions remain, including: 1) What loop backbone geometries are favoured by each motif? 2) To what extent are multi-amino acid motifs likely to represent genuine cooperative interactions? 3) Can new motifs be identified in a large, recent dataset using the latest bioinformatics tools? Three analytical tools are applied here to answer these questions. First, helix-terminal structures are partitioned by loop backbone geometry using a new 3D clustering algorithm. Next, Cascade Detection, a motif detection algorithm recently published by the author, is applied to each cluster to determine which sequence motifs are overrepresented in each geometry. Finally, the results for each motif are presented in a CapMap, a 3D conformational heatmap that displays the distribution of the motif's overrepresentation across loop geometries, enabling the rapid isolation and characterization of the associated side chain interaction. This work identifies a library of geometry-specific side chain interactions that provides a new, detailed picture of loop structure near the helix terminus. Highlights include determinations of the favoured loop geometries for the Asx/ST N-cap motifs, capping boxes, "big" boxes, and other hydrophobic, electrostatic, H-bond, and pi stacking interactions, many of which have not been described before. This work demonstrates that the combination of structural clustering and motif detection in the sequence space can efficiently identify side chain motifs and map them to the loop geometries which they support. Protein designers should find this study useful, because it identifies side

  18. Evaluation of a high-precision gear measuring machine for helix measurement using helix and wedge artifacts

    Science.gov (United States)

    Taguchi, Tetsuya; Kondo, Yohan

    2016-08-01

    High-precision gears are required for advanced motion and power transmission. The reliability of the measured value becomes important as the gear accuracy increases, and the establishment of a traceability system is needed. Therefore, a high-precision gear measuring machine (GMM) with a smaller uncertainty is expected to improve the gear calibration uncertainty. For this purpose, we developed a prototype of a high-precision GMM that adopts a direct drive mechanism and other features. Then, the high measurement capability of the developed GMM was verified using gear artifacts. Recently, some new measurement methods using simple shapes such as spheres and planes have been proposed as standards. We have verified the tooth profile measurement using a sphere artifact and reported the results that the developed GMM had a high capability in tooth profile measurement. Therefore, we attempted to devise a new evaluation method for helix measurement using a wedge artifact (WA) whose plane was treated as the tooth flank, and the high measurement capability of the developed GMM was verified. The results will provide a part of information to fully assess measurement uncertainty as our future work. This paper describes the evaluation results of the developed GMM for helix measurement using both a helix artifact and the WA, and discusses the effectiveness of the WA as a new artifact to evaluate the GMMs.

  19. Design, synthesis and biological evaluation of (E)-3,4-dihydroxystyryl 4-acylaminophenethyl sulfone, sulfoxide derivatives as dual inhibitors of HIV-1 CCR5 and integrase.

    Science.gov (United States)

    Sun, Yixing; Xu, Weisi; Fan, Ningning; Sun, Xuefeng; Ning, Xianling; Ma, Liying; Liu, Junyi; Wang, Xiaowei

    2017-02-01

    Aiming at the limited effectiveness of current clinical therapeutic effect of AIDS, novel series of compounds bearing (E)-3,4-dihydroxystyryl sulfone (or sulfoxide) and anilide fragments were designed and synthesized as dual inhibitors of HIV-1 CCR5/IN. The biological results indicated that several target compounds showed inhibitory activity against HIV-1 Bal (R5) infection in TZM-bl cells. Besides targeting the chemokine receptor on the host cell surface, they also displayed binding affinities with HIV-1 integrase using the surface plasmon resonance (SPR) binding assays. Molecular docking studies have inferred the possible binding mode of target compounds against integrase. These data demonstrate that the structure of (E)-3,4-dihydroxystyryl sulfone and sulfoxide derivatives have the potential to derive potent dual inhibitors of HIV-1 Integrase and CCR5. Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. BF integrase genes of HIV-1 circulating in São Paulo, Brazil, with a recurrent recombination region.

    Directory of Open Access Journals (Sweden)

    Atila Iamarino

    Full Text Available Although some studies have shown diversity in HIV integrase (IN genes, none has focused particularly on the gene evolving in epidemics in the context of recombination. The IN gene in 157 HIV-1 integrase inhibitor-naïve patients from the São Paulo State, Brazil, were sequenced tallying 128 of subtype B (23 of which were found in non-B genomes, 17 of subtype F (8 of which were found in recombinant genomes, 11 integrases were BF recombinants, and 1 from subtype C. Crucially, we found that 4 BF recombinant viruses shared a recurrent recombination breakpoint region between positions 4900 and 4924 (relative to the HXB2 that includes 2 gRNA loops, where the RT may stutter. Since these recombinants had independent phylogenetic origin, we argue that these results suggest a possible recombination hotspot not observed so far in BF CRF in particular, or in any other HIV-1 CRF in general. Additionally, 40% of the drug-naïve and 45% of the drug-treated patients had at least 1 raltegravir (RAL or elvitegravir (EVG resistance-associated amino acid change, but no major resistance mutations were found, in line with other studies. Importantly, V151I was the most common minor resistance mutation among B, F, and BF IN genes. Most codon sites of the IN genes had higher rates of synonymous substitutions (dS indicative of a strong negative selection. Nevertheless, several codon sites mainly in the subtype B were found under positive selection. Consequently, we observed a higher genetic diversity in the B portions of the mosaics, possibly due to the more recent introduction of subtype F on top of an ongoing subtype B epidemics and a fast spread of subtype F alleles among the B population.

  1. Influence of Drug Resistance Mutations on the Activity of HIV-1 Subtypes A and B Integrases: a Comparative Study.

    Science.gov (United States)

    Shadrina, O A; Zatsepin, T S; Agapkina, Yu Yu; Isaguliants, M G; Gottikh, M B

    2015-01-01

    Integration of human immunodeficiency virus (HIV-1) DNA into the genome of an infected cell is one of the key steps in the viral replication cycle. The viral enzyme integrase (IN), which catalyzes the integration, is an attractive target for the development of new antiviral drugs. However, the HIV-1 therapy often results in the IN gene mutations inducing viral resistance to integration inhibitors. To assess the impact of drug resistance mutations on the activity of IN of HIV-1 subtype A strain FSU-A, which is dominant in Russia, variants of the consensus IN of this subtype containing the primary resistance mutations G118R and Q148K and secondary compensatory substitutions E138K and G140S were prepared and characterized. Comparative study of these enzymes with the corresponding mutants of IN of HIV-1 subtype B strains HXB-2 was performed. The mutation Q148K almost equally reduced the activity of integrases of both subtypes. Its negative effect was partially compensated by the secondary mutations E138K and G140S. Primary substitution G118R had different influence on the activity of proteins of the subtypes A and B, and the compensatory effect of the secondary substitution E138K also depended on the viral subtype. Comparison of the mutants resistance to the known strand transfer inhibitors raltegravir and elvitegravir, and a new inhibitor XZ-259 (a dihydro-1H-isoindol derivative), showed that integrases of both subtypes with the Q148K mutation were insensitive to raltegravir and elvitegravir but were effectively inhibited by XZ-259. The substitution G118R slightly reduced the efficiency of IN inhibition by raltegravir and elvitegravir and caused no resistance to XZ_259.

  2. Non-catalytic site HIV-1 integrase inhibitors disrupt core maturation and induce a reverse transcription block in target cells.

    Science.gov (United States)

    Balakrishnan, Mini; Yant, Stephen R; Tsai, Luong; O'Sullivan, Christopher; Bam, Rujuta A; Tsai, Angela; Niedziela-Majka, Anita; Stray, Kirsten M; Sakowicz, Roman; Cihlar, Tomas

    2013-01-01

    HIV-1 integrase (IN) is the target for two classes of antiretrovirals: i) the integrase strand-transfer inhibitors (INSTIs) and ii) the non-catalytic site integrase inhibitors (NCINIs). NCINIs bind at the IN dimer interface and are thought to interfere primarily with viral DNA (vDNA) integration in the target cell by blocking IN-vDNA assembly as well as the IN-LEDGF/p75 interaction. Herein we show that treatment of virus-producing cells, but not of mature virions or target cells, drives NCINI antiviral potency. NCINIs target an essential late-stage event in HIV replication that is insensitive to LEDGF levels in the producer cells. Virus particles produced in the presence of NCINIs displayed normal Gag-Pol processing and endogenous reverse transcriptase activity, but were defective at initiating vDNA synthesis following entry into the target cell. NCINI-resistant virus carrying a T174I mutation in the IN dimer interface was less sensitive to the compound-induced late-stage effects, including the reverse transcription block. Wild-type, but not T174I virus, produced in the presence of NCINIs exhibited striking defects in core morphology and an increased level of IN oligomers that was not observed upon treatment of mature cell-free particles. Collectively, these results reveal that NCINIs act through a novel mechanism that is unrelated to the previously observed inhibition of IN activity or IN-LEDGF interaction, and instead involves the disruption of an IN function during HIV-1 core maturation and assembly.

  3. Mechanism of inhibition of HIV-1 integrase by G-tetrad-forming oligonucleotides in Vitro.

    Science.gov (United States)

    Jing, N; Marchand, C; Liu, J; Mitra, R; Hogan, M E; Pommier, Y

    2000-07-14

    The G-tetrad-forming oligonucleotides and have been identified as potent inhibitors of human immunodeficiency virus type 1 integrase (HIV-1 IN) activity (Rando, R. F., Ojwang, J., Elbaggari, A., Reyes, G. R., Tinder, R., McGrath, M. S., and Hogan, M. E. (1995) J. Biol. Chem. 270, 1754-1760; Mazumder, A., Neamati, N., Ojwang, J. O., Sunder, S., Rando, R. F., and Pommier, Y. (1996) Biochemistry 35, 13762-13771; Jing, N., and Hogan, M. E. (1998) J. Biol. Chem. 273, 34992-34999). To understand the inhibition of HIV-1 IN activity by the G-quartet inhibitors, we have designed the oligonucleotides and, composed of three and four G-quartets with stem lengths of 19 and 24 A, respectively. The fact that increasing the G-quartet stem length from 15 to 24 A kept inhibition of HIV-1 IN activity unchanged suggests that the binding interaction occurs between a GTGT loop domain of the G-quartet inhibitors and a catalytic site of HIV-1 IN, referred to as a face-to-face interaction. Docking the NMR structure of (Jing and Hogan (1998)) into the x-ray structure of the core domain of HIV-1 IN, HIV-1 IN-(51-209) (Maignan, S., Guilloteau, J.-P. , Qing, Z.-L., Clement-Mella, C., and Mikol, V. (1998) J. Mol. Biol. 282, 359-368), was performed using the GRAMM program. The statistical distributions of hydrogen bonding between HIV-1 IN and were obtained from the analyses of 1000 random docking structures. The docking results show a high probability of interaction between the GTGT loop residues of the G-quartet inhibitors and the catalytic site of HIV-1 IN, in agreement with the experimental observation.

  4. Efficient and specific internal cleavage of a retroviral palindromic DNA sequence by tetrameric HIV-1 integrase.

    Directory of Open Access Journals (Sweden)

    Olivier Delelis

    Full Text Available BACKGROUND: HIV-1 integrase (IN catalyses the retroviral integration process, removing two nucleotides from each long terminal repeat and inserting the processed viral DNA into the target DNA. It is widely assumed that the strand transfer step has no sequence specificity. However, recently, it has been reported by several groups that integration sites display a preference for palindromic sequences, suggesting that a symmetry in the target DNA may stabilise the tetrameric organisation of IN in the synaptic complex. METHODOLOGY/PRINCIPAL FINDINGS: We assessed the ability of several palindrome-containing sequences to organise tetrameric IN and investigated the ability of IN to catalyse DNA cleavage at internal positions. Only one palindromic sequence was successfully cleaved by IN. Interestingly, this symmetrical sequence corresponded to the 2-LTR junction of retroviral DNA circles-a palindrome similar but not identical to the consensus sequence found at integration sites. This reaction depended strictly on the cognate retroviral sequence of IN and required a full-length wild-type IN. Furthermore, the oligomeric state of IN responsible for this cleavage differed from that involved in the 3'-processing reaction. Palindromic cleavage strictly required the tetrameric form, whereas 3'-processing was efficiently catalysed by a dimer. CONCLUSIONS/SIGNIFICANCE: Our findings suggest that the restriction-like cleavage of palindromic sequences may be a general physiological activity of retroviral INs and that IN tetramerisation is strongly favoured by DNA symmetry, either at the target site for the concerted integration or when the DNA contains the 2-LTR junction in the case of the palindromic internal cleavage.

  5. Stretched versus compressed exponential kinetics in α-helix folding

    International Nuclear Information System (INIS)

    Hamm, Peter; Helbing, Jan; Bredenbeck, Jens

    2006-01-01

    In a recent paper (J. Bredenbeck, J. Helbing, J.R. Kumita, G.A. Woolley, P. Hamm, α-helix formation in a photoswitchable peptide tracked from picoseconds to microseconds by time resolved IR spectroscopy, Proc. Natl. Acad. Sci USA 102 (2005) 2379), we have investigated the folding of a photo-switchable α-helix with a kinetics that could be fit by a stretched exponential function exp(-(t/τ) β ). The stretching factor β became smaller as the temperature was lowered, a result which has been interpreted in terms of activated diffusion on a rugged energy surface. In the present paper, we discuss under which conditions diffusion problems occur with stretched exponential kinetics (β 1). We show that diffusion problems do have a strong tendency to yield stretched exponential kinetics, yet, that there are conditions (strong perturbation from equilibrium, performing the experiment in the folding direction) under which compressed exponential kinetics would be expected instead. We discuss the kinetics on free energy surfaces predicted by simple initiation-propagation models (zipper models) of α-helix folding, as well as by folding funnel models. We show that our recent experiment has been performed under condition for which models with strong downhill driving force, such as the zipper model, would predict compressed, rather than stretched exponential kinetics, in disagreement with the experimental observation. We therefore propose that the free energy surface along a reaction coordinate that governs the folding kinetics must be relatively flat and has a shape similar to a 1D golf course. We discuss how this conclusion can be unified with the thermodynamically well established zipper model by introducing an additional kinetic reaction coordinate

  6. The self-propulsion of a helix in granular matter

    Science.gov (United States)

    Valdes, Rogelio; Angeles, Veronica; de La Calleja, Elsa; Zenit, Roberto

    2017-11-01

    The effect of the shape of helicoidal on the displacement of magnetic robots in granular media is studied experimentally. We quantify the influences of three main parameters of the shape of the helicoidal swimmers: body diameter, step, and the angle. We compare the experimental measurements with an empirically modified resistive force theory prediction that accounts for the static friction coefficient of the particles of the granular material, leading to good agreement. Comparisons are also made with the granular resistive force theory proposed by Goldman and collaborators. We found an optimal helix angle to produce movement and determined a relationship between the swimmer size and speed.

  7. Some quality parameters of land snail meat - Helix pomatia

    Directory of Open Access Journals (Sweden)

    Tojagić Slobodan N.

    2004-01-01

    Full Text Available Considering the tradition in our regions to collect land snails (Helix pomatia for export, which is "disrupted" by social control resulting in limited possibilities to develop this attractive activity, there is a great interest lately for land snail breeding and fattening at farms. For this reason it is necessary to investigate systematically the possibilities to develop this activity in a longer period and in larger areas. The first investigations, although covering only nutritive and health safety aspects of the edible parts yielded the results presented in this work. Chemical composition, the content of some elements and organochlorine insecticides were followed as unavoidable in human living and environment.

  8. Toxic effects of Cadmium on the garden snail (Helix aspersa)

    Energy Technology Data Exchange (ETDEWEB)

    Russell, L.K. (Northrop Services Inc., Corvallis, OR); DeHaven, J.I.; Botts, R.P.

    1981-05-01

    Spreading treated municipal wastes on agricultural and forest lands is becoming an established method of disposal. However, there is concern about the deleterious effects of toxicants, particularly cadmium, in the sludges. Cadmium concentrations in sewage sludge have been reported as high as 1500 ppM. The work reported here is a part of a larger project to investigate the ecological effects of municipal wastes on forest lands. Snails, Helix aspersa, were chosen to examine the entrance of cadmium into terrestrial food chains. This experiment was designed to determine cadmium accumulation, acute toxicity, and behavioral, reproductive and growth responses with increasing levels of cadmium.

  9. Ab initio theory of helix <-> coil phase transition

    DEFF Research Database (Denmark)

    Yakubovich, Alexander V.; Solov'yov, Ilia; Solov'yov, Andrey V.

    2008-01-01

    In this paper, we suggest a theoretical method based on the statistical mechanics for treating the alpha-helix random coil transition in alanine polypeptides. We consider this process as a first-order phase transition and develop a theory which is free of model parameters and is based solely...... on fundamental physical principles. It describes essential thermodynamical properties of the system such as heat capacity, the phase transition temperature and others from the analysis of the polypeptide potential energy surface calculated as a function of two dihedral angles, responsible for the polypeptide...

  10. Gold helix photonic metamaterial as broadband circular polarizer.

    Science.gov (United States)

    Gansel, Justyna K; Thiel, Michael; Rill, Michael S; Decker, Manuel; Bade, Klaus; Saile, Volker; von Freymann, Georg; Linden, Stefan; Wegener, Martin

    2009-09-18

    We investigated propagation of light through a uniaxial photonic metamaterial composed of three-dimensional gold helices arranged on a two-dimensional square lattice. These nanostructures are fabricated via an approach based on direct laser writing into a positive-tone photoresist followed by electrochemical deposition of gold. For propagation of light along the helix axis, the structure blocks the circular polarization with the same handedness as the helices, whereas it transmits the other, for a frequency range exceeding one octave. The structure is scalable to other frequency ranges and can be used as a compact broadband circular polarizer.

  11. Low-coupling impedance double-helix structure for use in a ferrite kicker magnet

    International Nuclear Information System (INIS)

    Giordano, S.

    1983-01-01

    In a machine such as the CBA, the ejection ferrite kicker magnet has a very large longitudinal and transverse coupling impedance which could destroy the beam. Using a double-helix structure that surrounds the beam, the beam-induced fields are confined within the helix and, therefore, decoupled from the kicker; but at the same time the helix is transparent to the external fields of the kicker. At first, this may seem paradoxical that the helix is opaque to the fields generated inside the structure by the beam and simultaneously transparent to the external fields generated by the kicker

  12. Clear cell hidradenocarcinoma of the ear helix: report of primary ear helix adnexal carcinoma with regional lymph node metastasis.

    Science.gov (United States)

    Bae, Tae Hui; Kang, Shin Hyuk; Kim, Han Koo; Kim, Woo Seob; Kim, Mi Kyung

    2014-07-01

    Clear cell hidradenocarcinoma is a rare tumor of eccrine sweat gland origin that has a predilection for the head and neck. It has an indolent growth pattern and a higher incidence of regional and distant metastases. Metastasizing adnexal carcinomas are rare; thus, currently there is no uniform treatment guideline. We report a case of an 89-year-old female patient with clear cell hidradenocarcinoma manifesting in the right ear helix that metastasized to the right parotid gland who was treated by wide local excision and radiation therapy.

  13. Phenoloxidase activity of Helix aspersa maxima (garden snail, gastropod) hemocyanin.

    Science.gov (United States)

    Raynova, Yuliana; Doumanova, Lyuba; Idakieva, Krassimira Nikolova

    2013-12-01

    The oxygen-transporting protein, hemocyanin (Hc), of the garden snail Helix aspersa maxima (HaH) was isolated and kinetically characterized. Kinetic parameters of the reaction of catalytic oxidation of catechol to quinone, catalyzed by native HaH were determined: the V max value amounted to 22 nmol min(-1) mg(-1), k cat to 1.1 min(-1). Data were compared to those reported for other molluscan Hcs and phenoloxidases (POs). The o-diphenoloxidase activity of the native HaH is about five times higher than the activity determined for the Hcs of the terrestrial snail Helix pomatia and of the marine snail Rapana thomasiana (k cat values of 0.22 and 0.25 min(-1), respectively). The K m values obtained for molluscan Hcs from different species are comparable to those for true POs, but the low catalytic efficiency of Hcs is probably related to inaccessibility of the active sites to potential substrates. Upon treatment of HaH with subtilisin DY, the enzyme activity against substrate catechol was considerably increased. The relatively high proteolytically induced o-diPO activity of HaH allowed using it for preparation of a biosensor for detection of catechol.

  14. α-helix formation rate of oligopeptides at subzero temperatures

    Science.gov (United States)

    Qin, Zhi-jie; Shimizu, Akio; Li, Jinsong; Ikeguchi, Masamichi; Shinjo, Masaji; Kihara, Hiroshi

    2014-01-01

    In 1999, Clarke et al. ((1999) Proc. Natl. Acad. Sci. USA 96, 7232–7237) reported that the nucleation rate of α-helix of oligopeptide AK16 is as slow as 60 ms. In the present study, we measured the nucleation rate of oligopeptide, C17 (DLTDDIMCVKKILDKVG, corresponding to α-helical region of 84th to 100th amino acids of bovine α-lactalbumin) using the same method as Clarke et al. We found only initial bursts of the increase of α-helices at temperatures higher than −50°C in the presence of 70% methanol. The result with AK16 was the same as Clarke et al. reported. We also found that the folding rate of polyglutamic acid is too fast to be detected by the stopped-flow apparatus at 4°C. These results demonstrate that the α-helix formation rates in C17, AK16 and polyglutamic acid are shorter than the dead time of the stopped-flow apparatus (6 ms). PMID:27493493

  15. LEDGIN-mediated Inhibition of Integrase-LEDGF/p75 Interaction Reduces Reactivation of Residual Latent HIV.

    Science.gov (United States)

    Vranckx, Lenard S; Demeulemeester, Jonas; Saleh, Suha; Boll, Annegret; Vansant, Gerlinde; Schrijvers, Rik; Weydert, Caroline; Battivelli, Emilie; Verdin, Eric; Cereseto, Anna; Christ, Frauke; Gijsbers, Rik; Debyser, Zeger

    2016-06-01

    Persistence of latent, replication-competent Human Immunodeficiency Virus type 1 (HIV-1) provirus is the main impediment towards a cure for HIV/AIDS (Acquired Immune Deficiency Syndrome). Therefore, different therapeutic strategies to eliminate the viral reservoirs are currently being explored. We here propose a novel strategy to reduce the replicating HIV reservoir during primary HIV infection by means of drug-induced retargeting of HIV integration. A novel class of integration inhibitors, referred to as LEDGINs, inhibit the interaction between HIV integrase and the LEDGF/p75 host cofactor, the main determinant of lentiviral integration site selection. We show for the first time that LEDGF/p75 depletion hampers HIV-1 reactivation in cell culture. Next we demonstrate that LEDGINs relocate and retarget HIV integration resulting in a HIV reservoir that is refractory to reactivation by different latency-reversing agents. Taken together, these results support the potential of integrase inhibitors that modulate integration site targeting to reduce the likeliness of viral rebound. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

  16. Comparative molecular genetic analysis of simian and human HIV-1 integrase interactor INI1/SMARCB1/SNF5.

    Science.gov (United States)

    Pyeon, Dohun; Price, Lenore; Park, In-Woo

    2015-12-01

    Human integrase interactor 1 (INI1/SMARCB1/SNF5) is a chromatin-remodeling molecule that binds to HIV-1 integrase and enhances proviral DNA integration. INI1 is also known as a tumor suppressor gene and has been found to be mutated in several aggressive tumors such as rhabdoid and lymphoid tumors. To study the function of simian INI1, we screened and cloned simian INI1 cDNA from B lymphoma cells of rhesus monkeys using RT-PCR. Sequence analysis showed 23 single nucleotide differences compared to the human ortholog, which, however, did not result in amino acid changes, and the amino acid sequence is therefore 100% conserved between human and simian INI1. Two alternatively spliced isoforms, INI1a and INI1b, were also found in simian INI1. These two isoforms did not show any functional difference in HIV-1 proviral DNA integration and nuclear localization, suggesting that the specificity of simian INI1 would not be a factor preventing HIV-1 infection of a simian host. Nevertheless, INI1b is expressed only in established cancer cell lines such as Jurkat and COS-7 cells, and not in primary cells, suggesting that INIlb could be an indicator of cell transformation.

  17. A basic helix-loop-helix transcription factor DvIVS determines flower color intensity in cyanic dahlia cultivars.

    Science.gov (United States)

    Ohno, Sho; Deguchi, Ayumi; Hosokawa, Munetaka; Tatsuzawa, Fumi; Doi, Motoaki

    2013-08-01

    The study was aimed to identify the factors that regulate the intensity of flower color in cyanic dahlia (Dahlia variabilis), using fifteen cultivars with different color intensities in their petals. The cultivars were classified into three groups based on their flavonoid composition: ivory white cultivars with flavones; purple and pink cultivars with flavones and anthocyanins; and red cultivars with flavones, anthocyanins, and chalcones. Among the purple, pink, and ivory white cultivars, an inverse relationship was detected between lightness, which was used as an indicator for color intensity and anthocyanin content. A positive correlation was detected between anthocyanin contents and the expression of some structural genes in the anthocyanin synthesis pathway that are regulated by DvIVS, a basic helix-loop-helix transcription factor. A positive correlation between anthocyanin content and expression of DvIVS was also found. The promoter region of DvIVS was classified into three types, with cultivars carrying Type 1 promoter exhibited deep coloring, those carrying Type 2 and/or Type 3 exhibited pale coloring, and those carrying Type 1 and Type 2 and/or Type 3 exhibited medium coloring. The transcripts of the genes from these promoters encoded full-length predicted proteins. These results suggested that the genotype of the promoter region in DvIVS is one of the key factors determining the flower color intensity.

  18. Phylogeny, Functional Annotation, and Protein Interaction Network Analyses of the Xenopus tropicalis Basic Helix-Loop-Helix Transcription Factors

    Directory of Open Access Journals (Sweden)

    Wuyi Liu

    2013-01-01

    Full Text Available The previous survey identified 70 basic helix-loop-helix (bHLH proteins, but it was proved to be incomplete, and the functional information and regulatory networks of frog bHLH transcription factors were not fully known. Therefore, we conducted an updated genome-wide survey in the Xenopus tropicalis genome project databases and identified 105 bHLH sequences. Among the retrieved 105 sequences, phylogenetic analyses revealed that 103 bHLH proteins belonged to 43 families or subfamilies with 46, 26, 11, 3, 15, and 4 members in the corresponding supergroups. Next, gene ontology (GO enrichment analyses showed 65 significant GO annotations of biological processes and molecular functions and KEGG pathways counted in frequency. To explore the functional pathways, regulatory gene networks, and/or related gene groups coding for Xenopus tropicalis bHLH proteins, the identified bHLH genes were put into the databases KOBAS and STRING to get the signaling information of pathways and protein interaction networks according to available public databases and known protein interactions. From the genome annotation and pathway analysis using KOBAS, we identified 16 pathways in the Xenopus tropicalis genome. From the STRING interaction analysis, 68 hub proteins were identified, and many hub proteins created a tight network or a functional module within the protein families.

  19. Observation of helix associations for insertion of a retinal molecule and distortions of helix structures in bacteriorhodopsin

    Science.gov (United States)

    Urano, Ryo; Okamoto, Yuko

    2015-12-01

    We applied a newly proposed prediction method for membrane protein structures to bacteriorhodopsin that has distorted transmembrane helices in the native structure. This method uses an implicit membrane model, which restricts sampling space during folding in a membrane region, and includes helix bending. Replica-exchange simulations were performed with seven transmembrane helices only without a retinal molecule. Obtained structures were classified into clusters of similar structures, which correspond to local-minimum free energy states. The two lowest free energy states corresponded to a native-like structure with the correct empty space for retinal and a structure with this empty space filled with a helix. Previous experiments of bacteriorhodopsin suggested that association of transmembrane helices enables them to make a room for insertion of a retinal. Our results are consistent with these results. Moreover, distortions of helices in the native-like structures were successfully reproduced. In the distortions, whereas the locations of kinks for all helices were similar to those of Protein Data Bank's data, the amount of bends was more similar for helices away from the retinal than for those close to the retinal in the native structure. This suggests a hypothesis that the amino-acid sequence specifies the location of kinks in transmembrane helices and that the amount of distortions depends on the interactions with the surrounding molecules such as neighboring helices, lipids, and retinal.

  20. Basic helix-loop-helix transcription factor TCF21 is a downstream target of the male sex determining gene SRY.

    Directory of Open Access Journals (Sweden)

    Ramji K Bhandari

    Full Text Available The cascade of molecular events involved in mammalian sex determination has been shown to involve the SRY gene, but specific downstream events have eluded researchers for decades. The current study identifies one of the first direct downstream targets of the male sex determining factor SRY as the basic-helix-loop-helix (bHLH transcription factor TCF21. SRY was found to bind to the Tcf21 promoter and activate gene expression. Mutagenesis of SRY/SOX9 response elements in the Tcf21 promoter eliminated the actions of SRY. SRY was found to directly associate with the Tcf21 promoter SRY/SOX9 response elements in vivo during fetal rat testis development. TCF21 was found to promote an in vitro sex reversal of embryonic ovarian cells to induce precursor Sertoli cell differentiation. TCF21 and SRY had similar effects on the in vitro sex reversal gonadal cell transcriptomes. Therefore, SRY acts directly on the Tcf21 promoter to in part initiate a cascade of events associated with Sertoli cell differentiation and embryonic testis development.

  1. Structure of bacteriophage [phi]29 head fibers has a supercoiled triple repeating helix-turn-helix motif

    Energy Technology Data Exchange (ETDEWEB)

    Xiang, Ye; Rossmann, Michael G. (Purdue)

    2011-12-22

    The tailed bacteriophage {phi}29 capsid is decorated with 55 fibers attached to quasi-3-fold symmetry positions. Each fiber is a homotrimer of gene product 8.5 (gp8.5) and consists of two major structural parts, a pseudohexagonal base and a protruding fibrous portion that is about 110 {angstrom} in length. The crystal structure of the C-terminal fibrous portion (residues 112-280) has been determined to a resolution of 1.6 {angstrom}. The structure is about 150 {angstrom} long and shows three distinct structural domains designated as head, neck, and stem. The stem region is a unique three-stranded helix-turn-helix supercoil that has not previously been described. When fitted into a cryoelectron microscope reconstruction of the virus, the head structure corresponded to a disconnected density at the distal end of the fiber and the neck structure was located in weak density connecting it to the fiber. Thin section studies of Bacillus subtilis cells infected with fibered or fiberless {phi}29 suggest that the fibers might enhance the attachment of the virions onto the host cell wall.

  2. Evaluation of sequence variability in HIV-1 gp41 C-peptide helix-grafted proteins.

    Science.gov (United States)

    Tennyson, Rachel L; Walker, Susanne N; Ikeda, Terumasa; Harris, Reuben S; McNaughton, Brian R

    2017-08-01

    Many therapeutically-relevant protein-protein interactions (PPIs) have been reported that feature a helix and helix-binding cleft at the interface. Given this, different approaches to disrupting such PPIs have been developed. While short peptides (<15 amino acids) typically do not fold into a stable helix, researchers have reported chemical approaches to constraining helix structure. However, these approaches rely on laborious, and often expensive, chemical synthesis and purification. Our premise is that protein-based solutions that stabilize a therapeutically-relevant helix offer a number of advantages. In contrast to chemically constrained helical peptides, or minimal/miniature proteins, which must be synthesized (at great expense and labor), a protein can be expressed in a cellular system (like all current protein therapeutics). If selected properly, the protein scaffold can stabilize the therapeutically-relevant helix. We recently reported a protein engineering strategy, which we call "helix-grafted display", and applied it to the challenge of suppressing HIV entry. We have reported helix-grafted display proteins that inhibit formation of an intramolecular PPI involving HIV gp41 C-peptide helix, and HIV gp41 N-peptide trimer, which contain C-peptide helix-binding clefts. Here, we used yeast display to screen a library of grafted C-peptide helices for N-peptide trimer recognition. Using 'hits' from yeast display library screening, we evaluated the effect helix mutations have on structure, expression, stability, function (target recognition), and suppression of HIV entry. Copyright © 2017. Published by Elsevier Ltd.

  3. Integrase Defective Lentiviral Vector as a Vaccine Platform for Delivering Influenza Antigens

    Directory of Open Access Journals (Sweden)

    Alessandra Gallinaro

    2018-02-01

    Full Text Available Viral vectors represent an attractive technology for vaccine delivery. We exploited the integrase defective lentiviral vector (IDLV as a platform for delivering relevant antigens within the context of the ADITEC collaborative research program. In particular, Influenza virus hemagglutinin (HA and nucleoprotein (NP were delivered by IDLVs while H1N1 A/California/7/2009 subunit vaccine (HAp with or without adjuvant was used to compare the immune response in a murine model of immunization. In order to maximize the antibody response against HA, both IDLVs were also pseudotyped with HA (IDLV-HA/HA and IDLV-NP/HA, respectively. Groups of CB6F1 mice were immunized intramuscularly with a single dose of IDLV-NP/HA, IDLV-HA/HA, HAp alone, or with HAp together with the systemic adjuvant MF59. Six months after the vaccine prime all groups were boosted with HAp alone. Cellular and antibody responses to influenza antigens were measured at different time points after the immunizations. Mice immunized with HA-pseudotyped IDLVs showed similar levels of anti-H1N1 IgG over time, evaluated by ELISA, which were comparable to those induced by HAp + MF59 vaccination, but significantly higher than those induced by HAp alone. The boost with HAp alone induced an increase of antibodies in all groups, and the responses were maintained at higher levels up to 18 weeks post-boost. The antibody response was functional and persistent overtime, capable of neutralizing virus infectivity, as evaluated by hemagglutination inhibition and microneutralization assays. Moreover, since neuraminidase (NA-expressing plasmid was included during IDLV preparation, immunization with IDLV-NP/HA and IDLV-HA/HA also induced functional anti-NA antibodies, evaluated by enzyme-linked lectin assay. IFNγ-ELISPOT showed evidence of HA-specific response in IDLV-HA/HA immunized animals and persistent NP-specific CD8+ T cell response in IDLV-NP/HA immunized mice. Taken together our results indicate

  4. Emergent drug resistance with integrase strand transfer inhibitor-based regimens.

    Science.gov (United States)

    Lepik, Katherine J; Harrigan, P Richard; Yip, Benita; Wang, Lu; Robbins, Marjorie A; Zhang, Wendy W; Toy, Junine; Akagi, Linda; Lima, Viviane D; Guillemi, Silvia; Montaner, Julio S G; Barrios, Rolando

    2017-06-19

    To estimate the incidence of and risk factors for emergent resistance to integrase strand transfer inhibitor (INSTI) and nucleoside(-tide) reverse transcriptase inhibitors (NRTI) in HIV-1-infected adults receiving an INSTI and two NRTIs. Retrospective cohort study. Persons aged at least 19 years were included if they received their first prescription for raltegravir, elvitegravir or dolutegravir in British Columbia, Canada in 2012-2014 and were followed to 31 December 2015. Emergent resistance was defined as new mutations conferring intermediate-high level NRTI or INSTI resistance (score ≥30, Stanford HIV Drug Resistance Algorithm v.7.0.1). First-year resistance rates and 95% confidence intervals (95% CI) were estimated for 'any' (INSTI or NRTI) resistance using Poisson regression. The relationship between any emergent resistance and explanatory variables was modeled by Cox proportional hazards. There were 270 raltegravir, 323 elvitegravir and 392 dolutegravir-treated persons who were predominantly male (77%), antiretroviral therapy (ART)-experienced (81%), with low prevalence of preexisting drug resistance (16%). INSTI and NRTI resistance emerged in both ART-experienced and ART-naive persons (including dolutegravir-treated ART-naive), with no statistically significant differences in 'any' resistance rates (95% CI) between INSTIs: raltegravir 3.80 (1.90, 7.60), elvitegravir 2.37 (1.06, 5.27) and dolutegravir 1.48 (0.62, 3.55)/100 person-years. The strongest factors associated with emergent resistance were CD4 less than 200 cells/μl, adjusted hazard ratio (95% CI) 10.46 (4.67, 23.41) and less than 80% adherence to the INSTI regimen hazard ratio 2.52 (1.11, 5.71). Incident drug resistance rates were low with 'real-world' use of INSTI-based regimens. However, incomplete ART adherence and low CD4 cell count were associated with increased resistance rates regardless of which INSTI was prescribed. Provide adherence support and monitor for drug resistance.

  5. Anti-HIV-1 integrase compounds from Dioscorea bulbifera and molecular docking study.

    Science.gov (United States)

    Chaniad, Prapaporn; Wattanapiromsakul, Chatchai; Pianwanit, Somsak; Tewtrakul, Supinya

    2016-01-01

    Dioscorea bulbifera L. (Dioscoreaceae) has been used in a traditional Thai longevity medicine preparation. Isolation of inhibitors from natural products is a potential source for continuous development of new HIV-1 integrase (IN) inhibitors. The objective of this study is to isolate the compounds and evaluate their anti-HIV-1 IN activity, as well as to predict the potential interactions of the compounds with an IN. The ethyl acetate and water fractions (1-100 μg/mL) of Dioscorea bulbifera bulbils were isolated and tested for their anti-HIV-1 IN activity using the multiplate integration assay (MIA). The interactions of the active compounds with IN were investigated using a molecular docking method. The ethyl acetate and water fractions of Dioscorea bulbifera bulbils afforded seven compounds. Among these, allantoin (1), 2,4,3',5'-tetrahydroxybibenzyl (2), and 5,7,4'-trihydroxy-2-styrylchromone (5) were isolated for the first time from this plant. Myricetin (4) exhibited the most potent activity with an IC50 value of 3.15 μM, followed by 2,4,6,7-tetrahydroxy-9,10-dihydrophenanthrene (3, IC50 value= 14.20 μM), quercetin-3-O-β-D-glucopyranoside (6, IC50 value = 19.39 μM) and quercetin-3-O-β-D-galactopyranoside (7, IC50 value = 21.80 μM). Potential interactions of the active compounds (3, 4, 6, and 7) with the IN active site were additionally investigated. Compound 4 showed the best binding affinity to IN and formed strong interactions with various amino acid residues. These compounds interacted with Asp64, Thr66, His67, Glu92, Asp116, Gln148, Glu152, Asn155, and Lys159, which are involved in both the 3'-processing and strand transfer reactions of IN. In particular, galloyl, catechol, and sugar moieties were successful inhibitors for HIV-1 IN.

  6. The HIV-1 integrase-LEDGF allosteric inhibitor MUT-A: resistance profile, impairment of virus maturation and infectivity but without influence on RNA packaging or virus immunoreactivity

    NARCIS (Netherlands)

    Amadori, Céline; Ubeles van der Velden, Yme; Bonnard, Damien; Orlov, Igor; van Bel, Nikki; Le Rouzic, Erwann; Miralles, Laia; Brias, Julie; Chevreuil, Francis; Spehner, Daniele; Chasset, Sophie; Ledoussal, Benoit; Mayr, Luzia; Moreau, François; García, Felipe; Gatell, José; Zamborlini, Alessia; Emiliani, Stéphane; Ruff, Marc; Klaholz, Bruno P.; Moog, Christiane; Berkhout, Ben; Plana, Montserrat; Benarous, Richard

    2017-01-01

    HIV-1 Integrase (IN) interacts with the cellular co-factor LEDGF/p75 and tethers the HIV preintegration complex to the host genome enabling integration. Recently a new class of IN inhibitors was described, the IN-LEDGF allosteric inhibitors (INLAIs). Designed to interfere with the IN-LEDGF

  7. Genome-wide identification of basic helix-loop-helix and NF-1 motifs underlying GR binding sites in male rat hippocampus

    DEFF Research Database (Denmark)

    Pooley, John R.; Flynn, Ben P.; Grøntved, Lars

    2017-01-01

    in hippocampal GR function. Our findings imply a dosedependent and context-independent action of GRs in the hippocampus. Alterations in the expression or activity of NF-1/basic helix-loop-helix factors may play an as yet undetermined role in glucocorticoid-related disease susceptibility and outcome by altering......Glucocorticoids regulate hippocampal function in part by modulating gene expression through the glucocorticoid receptor (GR). GR binding is highly cell type specific, directed to accessible chromatin regions established during tissue differentiation. Distinct classes of GR binding sites...... linked to structural and organizational roles, an absence of major tethering partners for GRs, and little or no evidence for binding at negative glucocorticoid response elements. A basic helix-loop-helix motif closely resembling a NeuroD1 or Olig2 binding site was found underlying a subset of GR binding...

  8. Control of Recombination Directionality by the Listeria Phage A118 Protein Gp44 and the Coiled-Coil Motif of Its Serine Integrase.

    Science.gov (United States)

    Mandali, Sridhar; Gupta, Kushol; Dawson, Anthony R; Van Duyne, Gregory D; Johnson, Reid C

    2017-06-01

    The serine integrase of phage A118 catalyzes integrative recombination between attP on the phage and a specific attB locus on the chromosome of Listeria monocytogenes , but it is unable to promote excisive recombination between the hybrid attL and attR sites found on the integrated prophage without assistance by a recombination directionality factor (RDF). We have identified and characterized the phage-encoded RDF Gp44, which activates the A118 integrase for excision and inhibits integration. Gp44 binds to the C-terminal DNA binding domain of integrase, and we have localized the primary binding site to be within the mobile coiled-coil (CC) motif but distinct from the distal tip of the CC that is required for recombination. This interaction is sufficient to inhibit integration, but a second interaction involving the N-terminal end of Gp44 is also required to activate excision. We provide evidence that these two contacts modulate the trajectory of the CC motifs as they extend out from the integrase core in a manner dependent upon the identities of the four att sites. Our results support a model whereby Gp44 shapes the Int-bound complexes to control which att sites can synapse and recombine. IMPORTANCE Serine integrases mediate directional recombination between bacteriophage and bacterial chromosomes. These highly regulated site-specific recombination reactions are integral to the life cycle of temperate phage and, in the case of Listeria monocytogenes lysogenized by A118 family phage, are an essential virulence determinant. Serine integrases are also utilized as tools for genetic engineering and synthetic biology because of their exquisite unidirectional control of the DNA exchange reaction. Here, we identify and characterize the recombination directionality factor (RDF) that activates excision and inhibits integration reactions by the phage A118 integrase. We provide evidence that the A118 RDF binds to and modulates the trajectory of the long coiled-coil motif that

  9. Comparative Study on the Adaptation and Growth Dynamics of the Helix pomatia and Helix aspersa Muller Terrestrial Snails Under Different Feeding Regimes

    Directory of Open Access Journals (Sweden)

    Adrian Toader-Williams

    2010-05-01

    Full Text Available We used Helix pomatia and Helix aspersa species and measure their growth as the snails were approaching the hibernation season. Helix pomatia 2yo shown a decrease in weight while being raised in enclosed parcels of 4sqm the younger Helix pomatia 1yo as well as Helix aspersa Muller demonstrated the ability to adapt relatively fast to the same conditions. We established 5 experimental lots in a Helix pomatia farm, GPS coordinates N46.606040 E23.599950. Control lot contained Taraxacum officinales, Sonchus oleraceus, Equisetum arvense and Atriplex hortensis, wild flora found within the farm. The other lots contained the same plants as the control lot plus different combinations of imported plants from other areals. The H. pomatia 2yo weight decreased in the control lot by a mean of -3.86% while H. aspersa 1yo marked an increase of +16.89% in the same lot during the same period. The lot containing lupinus polyphyllus delivered snails with weight gain of +24.66% for H. pomatia 2yo and an increase of only +1.98% for H. aspersa 1yo. As a contrast, H. pomatia 2yo gained only +7.72% while H. aspersa 1yo gained +28.89%, in the lot containing Lavanda officinalis, Foeniculum vulgare and Hyssopus officinalis among the other plants.

  10. Double helix vortex breakdown in a turbulent swirling annular jet flow

    NARCIS (Netherlands)

    Vanierschot, M.; Perçin, M.; van Oudheusden, B.W.

    2018-01-01

    In this paper, we report on the structure and dynamics of double helix vortex breakdown in a turbulent annular swirling jet. Double helix breakdown has been reported previously for the laminar flow regime, but this structure has rarely been observed in turbulent flow. The flow field is

  11. Inactivation of colicin Y by intramembrane helix–helix interaction with its immunity protein

    Czech Academy of Sciences Publication Activity Database

    Šmajs, D.; Doležalová, M.; Macek, Pavel; Žídek, L.

    2008-01-01

    Roč. 275, č. 21 (2008), s. 5325-5331 ISSN 1742-464X Institutional research plan: CEZ:AV0Z50200510 Keywords : colicin immunity * colicin y * helix-helix interaction Subject RIV: CE - Biochemistry Impact factor: 3.139, year: 2008

  12. When proteome meets genome: the alpha helix and the beta strand ...

    Indian Academy of Sciences (India)

    The avoidance of the -helix and the -strand by splice junctions suggests the existence of a selection pressure against their disruption, perhaps underscoring the investment made by nature in building these intricate secondary structures. A corollary is that the helix and the strand are the smallest integral architectural units ...

  13. Assembly of Liposomes Controlled by Triple Helix Formation

    DEFF Research Database (Denmark)

    Vogel, Stefan; Jakobsen, Ulla

    2013-01-01

    Attachment of DNA to the surface of different solid nanoparticles (e.g. gold- and silica nanoparticles) is well established and a number of DNA-modified solid nanoparticle systems have been applied to thermal denaturation analysis of oligonucleotides. We report herein the non...... sequences (G or C-rich) to explore the applicability of the method for different triple helical assembly modes. We demonstrate advantages and limitations of the approach and proof the reversible and reproducible formation of liposome aggregates during thermal denaturation cycles. Nanoparticle tracking......-covalent immobilization of oligonucleotides on the surface of soft nanoparticles (e.g. liposomes) and the subsequent controlled assembly by DNA triple helix formation. The non-covalent approach avoids tedious surface chemistry and necessary purification procedures and can simplify and extend the available methodology...

  14. Associative learning phenomena in the snail (Helix aspersa): conditioned inhibition.

    Science.gov (United States)

    Acebes, Félix; Solar, Patricia; Moris, Joaquín; Loy, Ignacio

    2012-03-01

    Two experiments using garden snails (Helix aspersa) showed conditioned inhibition using both retardation and summation tests. Conditioned inhibition is a procedure by which a stimulus becomes a predictor of the absence of a relevant event--the unconditioned stimulus (US). Typically, conditioned inhibition consists of pairings between an initially neutral conditioned stimulus, CS(2), and an effective excitatory conditioned stimulus, CS(1), in the absence of the US. Retardation and summation tests are required in order to confirm that CS(2) has acquired inhibitory properties. Conditioned inhibition has previously been found in invertebrates; however, these demonstrations did not use the retardation and summation tests required for an unambiguous demonstration of inhibition, allowing for alternative explanations. The implications of our results for the fields of comparative cognition and invertebrate physiological models of learning are discussed.

  15. Significantly enhanced mechanical properties in AlN helix

    Science.gov (United States)

    Zhang, Xinghong; Zhao, Chaoliang; Yao, Tai; Zhou, Shanbao; Han, Jiecai; Li, Jiajie; Gao, Tangling; Wang, Xianjie; Zheng, Kun; Song, Bo

    2017-07-01

    To safely and reliably use aluminum nitride (AlN) helices in the fabrication of novel micro/nanodevices, it is very important to know their mechanical properties. Herein, we investigate the mechanical properties of individual AlN helices using an in situ tensile-bending test. Tensile tests reveal that an AlN helix has an average ε of ∼4.7 ± 0.8% elastic deformation before a typical brittle fracture occurs. The bending test shows a two-step mechanical feature—linear-elastic followed by an elastic-plastic process—with an average ε bent of ∼54.5 ± 0.6%. Our results provide direct cognition about the mechanical properties of AlN helices and their benefit to the design of AlN-based flexible micro/nanodevices.

  16. Beluga (Delphinapterus leucas Novel Bubble Helix Play Behavior

    Directory of Open Access Journals (Sweden)

    Brittany L. Jones

    2014-05-01

    Full Text Available Cetaceans demonstrate considerable ingenuity in their play with bubbles. Both wild and captive cetaceans have been reported to manipulate self-produced bubbles (Delfour & Aulagnier, 1997; Gewalt, 1989; Kuczaj, Makecha, Trone, Paulos, & Ramos, 2006; Kuczaj & Walker, 2006; McCowan, Marino, Vance, Walke, & Reiss, 2000; Pace, 2000; Paulos, Trone, & Kuczaj, 2010; Tizzi, Castellano, & Pace, 2000. The spread of unique and novel play behaviors across a group may involve social learning as well as trial and error learning (Kuczaj et al., 2006; Kuczaj, Yeater, & Highfill, 2012; McCowan et al., 2000; Pace, 2000. We report on a form of bubble play in belugas (Delphinapterus leucas that has not been previously reported. Four belugas at the Shedd Aquarium were videotaped producing bubble helices, smooth, long “helical tubes” that were created by the animal producing a pressure vortex that caused bubbles to merge and elongate based on pressure variation in the vortex (Marten, Shariff, Psarakos, & White, 1996. These observations revealed that belugas create novel bubble play behaviors that are transmitted among members of the group through social learning. When a beluga engaged in bubble helix play following the play of another beluga, it often acted on the bubble in the same manner as the most recent player, consistent with the notion that the second beluga was mimicking the behavior of the first beluga. Kimalu, a calf, was more likely to both observe and interact with Miki, his older brother, during bubble helix play bouts than with Naya (no relation, or Mauyak, his mother. Dolphin calves have also been found to be more likely to imitate the play behaviors of older more competent peers (Kuczaj et al., 2006; Kuczaj et al., 2012. Consistent with previous analyses of cetacean play (Kuczaj et al., 2006, belugas also varied the complexity of the play behavior in order to keep the game stimulating.

  17. Preparation and evaluation of appertized from snail Helix aspersa M

    Directory of Open Access Journals (Sweden)

    Nelson Loyola López

    2015-01-01

    Full Text Available This study includes the development and evaluation of snails (Helix aspersa M. appertized, collected at a heliciculture breeding center, located in Los Niches sector, Curico, Maule region, South-central of Chile. The test was conducted at the Laboratory of Sciences of the Catholic University of Maule, Nuestra Señora del Carmen Campus, Curico. The main objective of this work was to study the influence of appertized on sensory attributes and commercial durability of snail Helix aspersa M. Additionally, some specific objectives were proposed as follow: to provide this mollusc with a commercial alternative for it consume, to evaluate its organoleptic characteristics and guarantee the product from both the microbiological and nutritional points of view. Three media cover were used (T0: water + NaCl 2%; T1: Water + NaCl 2% + citric acid 0.5% + kilol and T2: extra virgin olive oil + spices + tocopherol. The product was assessed at two different times, after 30 and 90 days of storage. Two sensory evaluations were conducted to measure various organoleptic attributes and acceptability of the appertized by 14 trained panelists. Amino acid, vitamins, cholesterol, acidity, heavy metals, phosphorus and organochlorines analysis were performed. The presence of both total and fecal contaminant microorganisms was determined. Attributes such as color, flavor, aroma, texture and overall acceptability were also measured. Preserves made by T0 and T1 treatments were equally accepted by the panelists. However, preserve from treatment T2 was rejected because of the detection in them of a very dark color, odor and mealy texture. Positive results regarding the content of amino acids, vitamin C and low cholesterol, as well as the absence of pathogenic microorganisms were obtained for the three treatments.

  18. Volumetric Physics of Polypeptide Coil-Helix Transitions.

    Science.gov (United States)

    Krobath, Heinrich; Chen, Tao; Chan, Hue Sun

    2016-11-15

    Volumetric properties of proteins bear directly on their biological functions in hyperbaric environments and are useful in general as a biophysical probe. To gain insight into conformation-dependent protein volume, we developed an implicit-solvent atomic chain model that transparently embodies two physical origins of volume: (1) a fundamental geometric term capturing the van der Waals volume of the protein and the particulate, finite-size nature of the water molecules, modeled together by the volume encased by the protein's molecular surface, and (2) a physicochemical term for other solvation effects, accounted for by empirical proportionality relationships between experimental partial molar volumes and solvent-accessible surface areas of model compounds. We tested this construct by Langevin dynamics simulations of a 16-residue polyalanine. The simulated trajectories indicate an average volume decrease of 1.73 ± 0.1 Å 3 /residue for coil-helix transition, ∼80% of which is caused by a decrease in geometric void/cavity volume, and a robust positive activation volume for helical hydrogen bond formation originating from the transient void created by an approaching donor-acceptor pair and nearby atoms. These findings are consistent with prior experiments with alanine-rich peptides and offer an atomistic analysis of the observed overall volume changes. The results suggest, in general, that hydrostatic pressure likely stabilizes helical conformations of short peptides but slows the process of helix formation. In contrast, hydrostatic pressure is more likely to destabilize natural globular proteins because of the void volume entrapped in their folded structures. The conceptual framework of our model thus affords a coherent physical rationalization for experiments.

  19. Connecting two proteins using a fusion alpha helix stabilized by a chemical cross linker

    Science.gov (United States)

    Jeong, Woo Hyeon; Lee, Haerim; Song, Dong Hyun; Eom, Jae-Hoon; Kim, Sun Chang; Lee, Hee-Seung; Lee, Hayyoung; Lee, Jie-Oh

    2016-03-01

    Building a sophisticated protein nano-assembly requires a method for linking protein components in a predictable and stable structure. Most of the cross linkers available have flexible spacers. Because of this, the linked hybrids have significant structural flexibility and the relative structure between their two components is largely unpredictable. Here we describe a method of connecting two proteins via a `fusion α helix' formed by joining two pre-existing helices into a single extended helix. Because simple ligation of two helices does not guarantee the formation of a continuous helix, we used EY-CBS, a synthetic cross linker that has been shown to react selectively with cysteines in α-helices, to stabilize the connecting helix. Formation and stabilization of the fusion helix was confirmed by determining the crystal structures of the fusion proteins with and without bound EY-CBS. Our method should be widely applicable for linking protein building blocks to generate predictable structures.

  20. The hepatitis B virus ribonuclease H is sensitive to inhibitors of the human immunodeficiency virus ribonuclease H and integrase enzymes.

    Directory of Open Access Journals (Sweden)

    John E Tavis

    2013-01-01

    Full Text Available Nucleos(tide analog therapy blocks DNA synthesis by the hepatitis B virus (HBV reverse transcriptase and can control the infection, but treatment is life-long and has high costs and unpredictable long-term side effects. The profound suppression of HBV by the nucleos(tide analogs and their ability to cure some patients indicates that they can push HBV to the brink of extinction. Consequently, more patients could be cured by suppressing HBV replication further using a new drug in combination with the nucleos(tide analogs. The HBV ribonuclease H (RNAseH is a logical drug target because it is the second of only two viral enzymes that are essential for viral replication, but it has not been exploited, primarily because it is very difficult to produce active enzyme. To address this difficulty, we expressed HBV genotype D and H RNAseHs in E. coli and enriched the enzymes by nickel-affinity chromatography. HBV RNAseH activity in the enriched lysates was characterized in preparation for drug screening. Twenty-one candidate HBV RNAseH inhibitors were identified using chemical structure-activity analyses based on inhibitors of the HIV RNAseH and integrase. Twelve anti-RNAseH and anti-integrase compounds inhibited the HBV RNAseH at 10 µM, the best compounds had low micromolar IC(50 values against the RNAseH, and one compound inhibited HBV replication in tissue culture at 10 µM. Recombinant HBV genotype D RNAseH was more sensitive to inhibition than genotype H. This study demonstrates that recombinant HBV RNAseH suitable for low-throughput antiviral drug screening has been produced. The high percentage of compounds developed against the HIV RNAseH and integrase that were active against the HBV RNAseH indicates that the extensive drug design efforts against these HIV enzymes can guide anti-HBV RNAseH drug discovery. Finally, differential inhibition of HBV genotype D and H RNAseHs indicates that viral genetic variability will be a factor during drug

  1. A smallest 6 kda metalloprotease, mini-matrilysin, in living world: a revolutionary conserved zinc-dependent proteolytic domain- helix-loop-helix catalytic zinc binding domain (ZBD

    Directory of Open Access Journals (Sweden)

    Yu Wei-Hsuan

    2012-05-01

    Full Text Available Abstract Background The Aim of this study is to study the minimum zinc dependent metalloprotease catalytic folding motif, helix B Met loop-helix C, with proteolytic catalytic activities in metzincin super family. The metzincin super family share a catalytic domain consisting of a twisted five-stranded β sheet and three long α helices (A, B and C. The catalytic zinc is at the bottom of the cleft and is ligated by three His residues in the consensus sequence motif, HEXXHXXGXXH, which is located in helix B and part of the adjacent Met turn region. An interesting question is - what is the minimum portion of the enzyme that still possesses catalytic and inhibitor recognition?” Methods We have expressed a 60-residue truncated form of matrilysin which retains only the helix B-Met turn-helix C region and deletes helix A and the five-stranded β sheet which form the upper portion of the active cleft. This is only 1/4 of the full catalytic domain. The E. coli derived 6 kDa MMP-7 ZBD fragments were purified and refolded. The proteolytic activities were analyzed by Mca-Pro-Leu-Gly-Leu-Dpa-Ala-Arg-NH2 peptide assay and CM-transferrin zymography analysis. SC44463, BB94 and Phosphoramidon were computationally docked into the 3day structure of the human MMP7 ZBD and TAD and thermolysin using the docking program GOLD. Results This minimal 6 kDa matrilysin has been refolded and shown to have proteolytic activity in the Mca-Pro-Leu-Gly-Leu-Dpa-Ala-Arg-NH2 peptide assay. Triton X-100 and heparin are important factors in the refolding environment for this mini-enzyme matrilysin. This minienzyme has the proteolytic activity towards peptide substrate, but the hexamer and octamer of the mini MMP-7 complex demonstrates the CM-transferrin proteolytic activities in zymographic analysis. Peptide digestion is inhibited by SC44463, specific MMP7 inhibitors, but not phosphorimadon. Interestingly, the mini MMP-7 can be processed by autolysis and producing ~ 6

  2. A smallest 6 kda metalloprotease, mini-matrilysin, in living world: a revolutionary conserved zinc-dependent proteolytic domain- helix-loop-helix catalytic zinc binding domain (ZBD).

    Science.gov (United States)

    Yu, Wei-Hsuan; Huang, Po-Tsang; Lou, Kuo-Long; Yu, Shuan-Su C; Lin, Chen

    2012-05-29

    The Aim of this study is to study the minimum zinc dependent metalloprotease catalytic folding motif, helix B Met loop-helix C, with proteolytic catalytic activities in metzincin super family. The metzincin super family share a catalytic domain consisting of a twisted five-stranded β sheet and three long α helices (A, B and C). The catalytic zinc is at the bottom of the cleft and is ligated by three His residues in the consensus sequence motif, HEXXHXXGXXH, which is located in helix B and part of the adjacent Met turn region. An interesting question is - what is the minimum portion of the enzyme that still possesses catalytic and inhibitor recognition?" We have expressed a 60-residue truncated form of matrilysin which retains only the helix B-Met turn-helix C region and deletes helix A and the five-stranded β sheet which form the upper portion of the active cleft. This is only 1/4 of the full catalytic domain. The E. coli derived 6 kDa MMP-7 ZBD fragments were purified and refolded. The proteolytic activities were analyzed by Mca-Pro-Leu-Gly-Leu-Dpa-Ala-Arg-NH2 peptide assay and CM-transferrin zymography analysis. SC44463, BB94 and Phosphoramidon were computationally docked into the 3 day structure of the human MMP7 ZBD and TAD and thermolysin using the docking program GOLD. This minimal 6 kDa matrilysin has been refolded and shown to have proteolytic activity in the Mca-Pro-Leu-Gly-Leu-Dpa-Ala-Arg-NH2 peptide assay. Triton X-100 and heparin are important factors in the refolding environment for this mini-enzyme matrilysin. This minienzyme has the proteolytic activity towards peptide substrate, but the hexamer and octamer of the mini MMP-7 complex demonstrates the CM-transferrin proteolytic activities in zymographic analysis. Peptide digestion is inhibited by SC44463, specific MMP7 inhibitors, but not phosphorimadon. Interestingly, the mini MMP-7 can be processed by autolysis and producing ~ 6 ~ 7 kDa fragments. Thus, many of the functions of the enzyme are

  3. Pharmacovirological Impact of an Integrase Inhibitor on Human Immunodeficiency Virus Type 1 cDNA Species In Vivo ▿

    Science.gov (United States)

    Goffinet, Christine; Allespach, Ina; Oberbremer, Lena; Golden, Pamela L.; Foster, Scott A.; Johns, Brian A.; Weatherhead, Jason G.; Novick, Steven J.; Chiswell, Karen E.; Garvey, Edward P.; Keppler, Oliver T.

    2009-01-01

    Clinical trials of the first approved integrase inhibitor (INI), raltegravir, have demonstrated a drop in the human immunodeficiency virus type 1 (HIV-1) RNA loads of infected patients that was unexpectedly more rapid than that with a potent reverse transcriptase inhibitor, and apparently dose independent. These clinical outcomes are not understood. In tissue culture, although their inhibition of integration is well documented, the effects of INIs on levels of unintegrated HIV-1 cDNAs have been variable. Furthermore, there has been no report to date on an INI's effect on these episomal species in vivo. Here, we show that prophylactic treatment of transgenic rats with the strand transfer INI GSK501015 reduced levels of viral integrants in the spleen by up to 99.7%. Episomal two-long-terminal-repeat (LTR) circles accumulated up to sevenfold in this secondary lymphoid organ, and this inversely correlated with the impact on the proviral burden. Contrasting raltegravir's dose-ranging study with HIV patients, titration of GSK501015 in HIV-infected animals demonstrated dependence of the INI's antiviral effect on its serum concentration. Furthermore, the in vivo 50% effective concentration calculated from these data best matched GSK501015's in vitro potency when serum protein binding was accounted for. Collectively, this study demonstrates a titratable, antipodal impact of an INI on integrated and episomal HIV-1 cDNAs in vivo. Based on these findings and known biological characteristics of viral episomes, we discuss how integrase inhibition may result in additional indirect antiviral effects that contribute to more rapid HIV-1 decay in HIV/AIDS patients. PMID:19458008

  4. Structural dynamics of native and V260E mutant C-terminal domain of HIV-1 integrase

    Science.gov (United States)

    Sangeetha, Balasubramanian; Muthukumaran, Rajagopalan; Amutha, Ramaswamy

    2015-04-01

    The C-terminal domain (CTD) of HIV-1 integrase is a five stranded β-barrel resembling an SH3 fold. Mutational studies on isolated CTD and full-length IN have reported V260E mutant as either homo-dimerization defective or affecting the stability and folding of CTD. In this study, molecular dynamics simulation techniques were used to unveil the effect of V260E mutation on isolated CTD monomer and dimer. Both monomeric and dimeric forms of wild type and V260E mutant are highly stable during the simulated period. However, the stabilizing π-stacking interaction between Trp243 and Trp243' at the dimer interface is highly disturbed in CTD-V260E (>6 Å apart). The loss in entropy for dimerization is -30 and -25 kcal/mol for CTD-wt and CTD-V260E respectively signifying a weak hydrophobic interaction and its perturbation in CTD-V260E. The mutant Glu260 exhibits strong attraction/repulsion with all the basic/acidic residues of CTD. In addition to this, the dynamics of CTD-wild type and V260E monomers at 498 K was analyzed to elucidate the effect of V260E mutation on CTD folding. Increase in SASA and reduction in the number of contacts in CTD-V260E during simulation highlights the instability caused by the mutation. In general, V260E mutation affects both multimerization and protein folding with a pronounced effect on protein folding rather than multimerization. This study emphasizes the importance of the hydrophobic nature and SH3 fold of CTD in proper functioning of HIV integrase and perturbing this nature would be a rational approach toward designing more selective and potent allosteric anti-HIV inhibitors.

  5. Prevalence of integrase inhibitor resistance mutations in Austrian patients recently diagnosed with HIV from 2008 to 2013.

    Science.gov (United States)

    Zoufaly, A; Kraft, C; Schmidbauer, C; Puchhammer-Stoeckl, E

    2017-04-01

    Treatment guidelines often do not advocate testing for integrase inhibitor resistance associated mutations (IRAM) before initiation of first line ART given the extremely low prevalence of mutations found in older surveillance studies. We aimed to describe the prevalence of IRAM in Austrian patients recently diagnosed with HIV in the 5 years following introduction of integrase inhibitors and to analyse trends and factors associated with their detection. Samples of antiretroviral treatment (ART) naïve patients recently diagnosed with HIV in Austria between 2008 and 2013 were analysed for the existence of IRAM and drug penalty scores were calculated to estimate response to drugs. Demographic and virological data were extracted from a database. Descriptive and comparative statistics were used. A total of 303 samples were analysed. 78 % were male and mean age was 38 years. Overall prevalence of IRAM was 2.3 %. Six percent had at least potentially low-level resistance to raltegravir or elvitegravir, versus 1 % for dolutegravir. One primary mutation was observed (F121Y) in a patient sample from 2012 leading to 5-10-fold reduced susceptibility to raltegravir and elvitegravir. Two patients carried the accessory mutations E138K and G140A, respectively, where both lie on the Q148 pathway. No temporal trend of IRAM prevalence was observed (p = 0.16). Primary IRAM are still rarely found despite the increasing use of INSTI in Austria, but there is a potential for reduced susceptibility to these drugs in selected patients. Routine resistance testing seems prudent to avoid the consequences including accumulation of further mutations and therapeutic failure.

  6. Double helix vortex breakdown in a turbulent swirling annular jet flow

    Science.gov (United States)

    Vanierschot, M.; Percin, M.; van Oudheusden, B. W.

    2018-03-01

    In this paper, we report on the structure and dynamics of double helix vortex breakdown in a turbulent annular swirling jet. Double helix breakdown has been reported previously for the laminar flow regime, but this structure has rarely been observed in turbulent flow. The flow field is investigated experimentally by means of time-resolved tomographic particle image velocimetry. Notwithstanding the axisymmetric nature of the time-averaged flow, analysis of the instantaneous three-dimensional (3D) vortical structures shows the existence of a vortex core along the central axis which breaks up into a double helix downstream. The winding sense of this double helix is opposite to the swirl direction (m =-2 ) and it is wrapped around a central vortex breakdown bubble. This structure is quite different from double helix breakdown found in laminar flows where the helix is formed in the wake of the bubble and not upstream. The double helix precesses around the central axis of the jet with a precessing frequency corresponding to a Strouhal number of 0.27.

  7. Differential effects of the G118R, H51Y, and E138K resistance substitutions in different subtypes of HIV integrase.

    Science.gov (United States)

    Quashie, Peter K; Oliviera, Maureen; Veres, Tamar; Osman, Nathan; Han, Ying-Shan; Hassounah, Said; Lie, Yolanda; Huang, Wei; Mesplède, Thibault; Wainberg, Mark A

    2015-03-01

    Dolutegravir (DTG) is the latest antiretroviral (ARV) approved for the treatment of human immunodeficiency virus (HIV) infection. The G118R substitution, previously identified with MK-2048 and raltegravir, may represent the initial substitution in a dolutegravir resistance pathway. We have found that subtype C integrase proteins have a low enzymatic cost associated with the G118R substitution, mostly at the strand transfer step of integration, compared to either subtype B or recombinant CRF02_AG proteins. Subtype B and circulating recombinant form AG (CRF02_AG) clonal viruses encoding G118R-bearing integrases were severely restricted in their viral replication capacity, and G118R/E138K-bearing viruses had various levels of resistance to dolutegravir, raltegravir, and elvitegravir. In cell-free experiments, the impacts of the H51Y and E138K substitutions on resistance and enzyme efficiency, when present with G118R, were highly dependent on viral subtype. Sequence alignment and homology modeling showed that the subtype-specific effects of these mutations were likely due to differential amino acid residue networks in the different integrase proteins, caused by polymorphic residues, which significantly affect native protein activity, structure, or function and are important for drug-mediated inhibition of enzyme activity. This preemptive study will aid in the interpretation of resistance patterns in dolutegravir-treated patients. Recognized drug resistance mutations have never been reported for naive patients treated with dolutegravir. Additionally, in integrase inhibitor-experienced patients, only R263K and other previously known integrase resistance substitutions have been reported. Here we suggest that alternate resistance pathways may develop in non-B HIV-1 subtypes and explain how "minor" polymorphisms and substitutions in HIV integrase that are associated with these subtypes can influence resistance against dolutegravir. This work also highlights the importance

  8. Solvent-Directed Switch of a Left-Handed 10/12-Helix into a Right-Handed 12/10-Helix in Mixed β-Peptides.

    Science.gov (United States)

    Thodupunuri, Prashanth; Katukuri, Sirisha; Ramakrishna, Kallaganti V S; Sharma, Gangavaram V M; Kunwar, Ajit C; Sarma, Akella V S; Hofmann, Hans-Jörg

    2017-02-17

    Present study describes the synthesis and conformational analysis of β-peptides from C-linked carbo-β-amino acids [β-Caa (l) ] with a d-lyxo furanoside side chain and β-hGly in 1:1 alternation. NMR and CD investigations on peptides with an (S)-β-Caa (l) monomer at the N-terminus revealed a right-handed 10/12-mixed helix. An unprecedented solvent-directed "switch" both in helical pattern and handedness was observed when the sequence begins with a β-hGly residue instead of a (S)-β-Caa (l) constituent. NMR studies on these peptides in chloroform indicated a left-handed 10/12-helix, while the CD spectrum in methanol inferred a right-handed secondary structure. The NMR data for these peptides in CD 3 OH showed the presence of a right-handed 12/10-helix. NMR investigations in acetonitrile indicated the coexistence of both helix types. Quantum chemical studies predicted a small energy difference of 0.3 kcal/mol between the two helix types, which may explain the possibility of solvent influence. Examples for a solvent-directed switch of both the H-bonding pattern and the handedness of foldamer helices are rare so far. A comparable solvent effect was not found in the corresponding peptides with (R)-β-Caa (l) residues, where right-handed 12/10-helices are predominating.

  9. RENNSH: a novel α-helix identification approach for intermediate resolution electron density maps.

    Science.gov (United States)

    Ma, Lingyu; Reisert, Marco; Burkhardt, Hans

    2012-01-01

    Accurate identification of protein secondary structures is beneficial to understand three-dimensional structures of biological macromolecules. In this paper, a novel refined classification framework is proposed, which treats alpha-helix identification as a machine learning problem by representing each voxel in the density map with its Spherical Harmonic Descriptors (SHD). An energy function is defined to provide statistical analysis of its identification performance, which can be applied to all the α-helix identification approaches. Comparing with other existing α-helix identification methods for intermediate resolution electron density maps, the experimental results demonstrate that our approach gives the best identification accuracy and is more robust to the noise.

  10. Thermal helix-coil transition in UV irradiated collagen from rat tail tendon.

    Science.gov (United States)

    Sionkowska, A; Kamińska, A

    1999-05-01

    The thermal helix-coil transition in UV irradiated collagen solution, collagen film and pieces of rat tail tendon (RTT) were compared. Their thermal stability's were determined by differential scanning calorimeter (DSC) and by viscometric measurements. The denaturation temperatures of collagen solution, film and pieces of RTT were different. The helix-coil transition occur near 40 degrees C in collagen solution, near 112 degrees C in collagen film, and near 101 degrees C in pieces of RTT. After UV irradiation the thermal helix-coil transition of collagen samples were changed. These changes depend on the degree of hydratation.

  11. Defining the structural requirements for a helix in 23 S ribosomal RNA that confers erythromycin resistance

    DEFF Research Database (Denmark)

    Douthwaite, S; Powers, T; Lee, J Y

    1989-01-01

    The helix spanning nucleotides 1198 to 1247 (helix 1200-1250) in Escherichia coli 23 S ribosomal RNA (rRNA) is functionally important in protein synthesis, and deletions in this region confer erythromycin resistance. In order to define the structural requirements for resistance, we have dissected....... However, removal of either these or non-conserved nucleotides from helix 1200-1250 measurably reduces the efficiency of 23 S RNA in forming functional ribosomes. We have used chemical probing and a modified primer extension method to investigate erythromycin binding to wild-type and resistant ribosomes...

  12. Characterization of a crosslinked nucleic acid - helix destabilizing protein complex

    Energy Technology Data Exchange (ETDEWEB)

    Karpel, R.L.; Levin, V.Y.; Haley, B.E.

    1986-05-01

    They have enzymatically synthesized /sup 3/H- and /sup 32/P-poly(A,8N/sub 3/A) from 8-N/sub 3/ADP and radiolabeled ADP, and have used this polynucleotide to photoaffinity label T4 gene 32 protein, as well as several other helix-destabilizing proteins (HDPs). Irradiation of /sup 32/P-/sup 3/H-poly(A,N/sub 3/A) mixtures for short durations produces a covalent complex, seen as a high molecular weight, radioactive band on SDS-polyacrylamide gels. Preliminary experiments on other HDPs, from prokaryotic, eukaryotic and animal viral sources, show analogous results. Several successful control experiments indicate that this system is suitable for binding site localization on /sup 32/P. Single-stranded nucleic acids competitively inhibit photolabeling. The effect of NaCl on photolabeling parallels the salt-dependence of /sup 32/P-poly(A,N/sub 3/A) binding. Photolabeling reaches a plateau after approx.1 min, and the formation of the high molecular weight complex parallels the reduction of free /sup 32/P on SDS gels. Staph. nuclease digestion of crosslinked complexes produces a diffuse, radioactive band on SDS gels, migrating just behind free /sup 32/P. When these digested complexes are subjected to reverse-phase HPLC on a C3 Ultrapore column, the nucleic acid /sup 32/P-label is seen to coelute with protein. They are currently employing RP-HPLC methods to locate the label on tryptic peptides of nuclease-digested complexes.

  13. Portrait of a discovery. Watson, Crick, and the double helix.

    Science.gov (United States)

    de Chadarevian, Soraya

    2003-03-01

    This essay examines an iconic image of twentieth-century science: Antony Barrington Brown's photograph of James Watson, Francis Crick, and the double-helical model of DNA. The detailed reconstruction of the production, reception, and uses of the photograph reveals the central role of the image in making the discovery it portrays. Taken in May 1953, two full months after the scientists built the model, to accompany a report on the structure in Time magazine, the photograph (like the report) was never published. It came into circulation only fifteen years later, as an illustration in Watson's best-selling book The Double Helix. While the image served as a historical document and advertisement for the book, only the book provided the description that made the image as well as the people and the model it represented famous. The history of the image provides insights into the retrospective construction of the discovery, which has since been celebrated as the origin of a new science of life.

  14. Calorimetric Study of Helix aspersa Maxima Hemocyanin Isoforms

    Directory of Open Access Journals (Sweden)

    Svetla Todinova

    2018-01-01

    Full Text Available The thermal unfolding of hemocyanin isoforms, β-HaH and αD+N-HaH, isolated from the hemolymph of garden snails Helix aspersa maxima, was studied by means of differential scanning calorimetry (DSC. One transition, with an apparent transition temperature (Tm at 79.88°C, was detected in the thermogram of β-HaH in 20 mM HEPES buffer, containing 0.1 M NaCl, 5 mM CaCl2, and 5 mM MgCl2, pH 7.0, at scan rate of 1.0°C min−1. By means of successive annealing procedure, two individual transitions were identified in the thermogram of αD+N-HaH. Denaturation of both hemocyanins was found to be an irreversible process. The scan-rate dependence of the calorimetric profiles indicated that the thermal unfolding of investigated hemocyanins was kinetically controlled. The thermal denaturation of the isoforms β-HaH and αD+N-HaH was described by the two-state irreversible model, and parameters of the Arrhenius equation were calculated.

  15. Double helix boron-10 powder thermal neutron detector

    Science.gov (United States)

    Wang, Zhehui; Morris, Christopher L.; Bacon, Jeffrey D.

    2015-06-02

    A double-helix Boron-10 powder detector having intrinsic thermal neutron detection efficiency comparable to 36'' long, 2-in diameter, 2-bar Helium-3 detectors, and which can be used to replace such detectors for use in portal monitoring, is described. An embodiment of the detector includes a metallic plate coated with Boron-10 powder for generating alpha and Lithium-7 particles responsive to neutrons impinging thereon supported by insulators affixed to at least two opposing edges; a grounded first wire wound in a helical manner around two opposing insulators; and a second wire having a smaller diameter than that of the first wire, wound in a helical manner around the same insulators and spaced apart from the first wire, the second wire being positively biased. A gas, disposed within a gas-tight container enclosing the plate, insulators and wires, and capable of stopping alpha and Lithium-7 particles and generating electrons produces a signal on the second wire which is detected and subsequently related to the number of neutrons impinging on the plate.

  16. Helix Nebula: sunshine and clouds on the CERN computing horizon

    CERN Multimedia

    Joannah Caborn Wengler

    2012-01-01

    23 petabytes is how much data CERN recorded during 2011, and this number will rise in 2012. In order to respond to the challenge, the IT department is upping its game, amongst other things by participating in the Helix Nebula project, a public-private partnership to create a European cloud-computing platform, as announced in a recent CERN press release.   “We’re not replacing the Grid,” clarifies Bob Jones, responsible for CERN openlab who is also responsible for EC-funded projects in IT, “but looking at three complementary ways of increasing CERN’s computing capacity, so that as demand goes up we can continue to satisfy our users.” “First we are upgrading the electrical and cooling infrastructure of the computer centre in order to increase the availability of critical IT services needed for the Laboratory. This will also provide more floor space in the area called The Barn, allowing for more servers to fit in.”...

  17. RFID Tag Helix Antenna Sensors for Wireless Drug Dosage Monitoring

    Science.gov (United States)

    Huang, Haiyu; Zhao, Peisen; Chen, Pai-Yen; Ren, Yong; Liu, Xuewu; Ferrari, Mauro; Hu, Ye; Akinwande, Deji

    2014-01-01

    Miniaturized helix antennas are integrated with drug reservoirs to function as RFID wireless tag sensors for real-time drug dosage monitoring. The general design procedure of this type of biomedical antenna sensors is proposed based on electromagnetic theory and finite element simulation. A cost effective fabrication process is utilized to encapsulate the antenna sensor within a biocompatible package layer using PDMS material, and at the same time form a drug storage or drug delivery unit inside the sensor. The in vitro experiment on two prototypes of antenna sensor-drug reservoir assembly have shown the ability to monitor the drug dosage by tracking antenna resonant frequency shift from 2.4–2.5-GHz ISM band with realized sensitivity of 1.27 \\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{upgreek} \\usepackage{mathrsfs} \\setlength{\\oddsidemargin}{-69pt} \\begin{document} }{}$\\mu~{\\rm l}/{\\rm MHz}$\\end{document} for transdermal drug delivery monitoring and 2.76-\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{upgreek} \\usepackage{mathrsfs} \\setlength{\\oddsidemargin}{-69pt} \\begin{document} }{}$\\mu~{\\rm l}/{\\rm MHz}$\\end{document} sensitivity for implanted drug delivery monitoring. PMID:27170865

  18. Helix Electrohydrodynamic Printing of Highly Aligned Serpentine Micro/Nanofibers

    Directory of Open Access Journals (Sweden)

    Yongqing Duan

    2017-09-01

    Full Text Available Micro/nano serpentine structures have widespread applications in flexible/stretchable electronics; however, challenges still exist for low-cost, high-efficiency and controllable manufacturing. Helix electrohydrodynamic printing (HE-printing has been proposed here to realize controllable direct-writing of large area, highly aligned serpentine micro/nanofibers by introducing the rope coiling effect into printing process. By manipulating the flying trajectory and solidification degree of the micro/nano jet, the solidified micro/nanofiber flying in a stabilized helical manner and versatile serpentine structures deposited on a moving collector have been achieved. Systematic experiments and theoretical analysis were conducted to study the transformation behavior and the size changing rules for various deposited microstructures, and highly aligned serpentine microfibers were directly written by controlling the applied voltage, nozzle-to-collector distance and collector velocity. Furthermore, a hyper-stretchable piezoelectric device that can detect stretching, bending and pressure has been successfully fabricated using the printed serpentine micro/nanofibers, demonstrating the potential of HE-printing in stretchable electronics manufacturing.

  19. SREBP-2, a second basic-helix-loop-helix-leucine zipper protein that stimulates transcription by binding to a sterol regulatory element.

    OpenAIRE

    Hua, X; Yokoyama, C; Wu, J; Briggs, M R; Brown, M S; Goldstein, J L; Wang, X

    1993-01-01

    We report the cDNA cloning of SREBP-2, the second member of a family of basic-helix-loop-helix-leucine zipper (bHLH-Zip) transcription factors that recognize sterol regulatory element 1 (SRE-1). SRE-1, a conditional enhancer in the promoters for the low density lipoprotein receptor and 3-hydroxy-3-methylglutaryl-coenzyme A synthase genes, increases transcription in the absence of sterols and is inactivated when sterols accumulate. Human SREBP-2 contains 1141 amino acids and is 47% identical t...

  20. Peptides derived from nucleoside beta-amino acids form an unusual 8-helix.

    Science.gov (United States)

    Threlfall, Richard; Davies, Andrew; Howarth, Nicola M; Fisher, Julie; Cosstick, Richard

    2008-02-07

    Peptides of varying length (dimers to octamers) were prepared from nucleoside beta-amino acids and conformational studies, based on NOE observations, show that the beta-peptides form an unusual 8-helix.

  1. Application of the Morita-Baylis-Hillman reaction in the synthesis of 3-[(N-cycloalkylbenzamido)methyl]-2-quinolones as potential HIV-1 integrase inhibitors.

    Science.gov (United States)

    Sekgota, Khethobole C; Majumder, Swarup; Isaacs, Michelle; Mnkandhla, Dumisani; Hoppe, Heinrich C; Khanye, Setshaba D; Kriel, Frederik H; Coates, Judy; Kaye, Perry T

    2017-12-01

    A practicable six-step synthetic pathway has been developed to access a library of novel 3-[(N-cycloalkylbenzamido)methyl]-2-quinolones using Morita-Baylis-Hillman methodology. These compounds and their 3-[(N-cycloalkylamino)methyl]-2-quinolone precursors have been screened as potential HIV-1 integrase (IN) inhibitors. A concomitant survey of their activity against HIV-1 protease and reverse-transcriptase reveals selective inhibition of HIV-1 IN. Copyright © 2017 Elsevier Inc. All rights reserved.

  2. Construction of a stepwise gene integration system by transient expression of actinophage R4 integrase in cyanobacterium Synechocystis sp. PCC 6803.

    Science.gov (United States)

    Miura, Takamasa; Nishizawa, Akito; Nishizawa, Tomoyasu; Asayama, Munehiko; Takahashi, Hideo; Shirai, Makoto

    2014-08-01

    The integrase of actinophage R4, which belongs to the large serine-recombinase family, catalyzes site-specific recombination between two distinct attachment site sequences of the phage (attP) and actinomycete Streptomyces parvulus 2297 chromosome (attB). We previously reported that R4 integrase (Sre) catalyzed site-specific recombination both in vivo and in vitro. In the present study, a Sre-based system was developed for the stepwise site-specific integration of multiple genes into the chromosome of cyanobacterium Synechocystis sp. PCC 6803 (hereafter PCC 6803). A transgene-integrated plasmid with two attP sites and a non-replicative sre-containing plasmid were co-introduced into attB-inserted PCC 6803 cells. The transiently expressed Sre catalyzed highly efficient site-specific integration between one of the two attP sites on the integration plasmid and the attB site on the chromosome of PCC 6803. A second transgene-integrated plasmid with an attB site was integrated into the residual attP site on the chromosome by repeating site-specific recombination. The transformation frequencies (%) of the first and second integrations were approximately 5.1 × 10(-5) and 8.2 × 10(-5), respectively. Furthermore, the expression of two transgenes was detected. This study is the first to apply the multiple gene site-specific integration system based on R4 integrase to cyanobacteria.

  3. D77, one benzoic acid derivative, functions as a novel anti-HIV-1 inhibitor targeting the interaction between integrase and cellular LEDGF/p75

    International Nuclear Information System (INIS)

    Du Li; Zhao Yaxue; Chen, Jing; Yang Liumeng; Zheng Yongtang; Tang Yun; Shen Xu; Jiang Hualiang

    2008-01-01

    Integration of viral-DNA into host chromosome mediated by the viral protein HIV-1 integrase (IN) is an essential step in the HIV-1 life cycle. In this process, Lens epithelium-derived growth factor (LEDGF/p75) is discovered to function as a cellular co-factor for integration. Since LEDGF/p75 plays an important role in HIV integration, disruption of the LEDGF/p75 interaction with IN has provided a special interest for anti-HIV agent discovery. In this work, we reported that a benzoic acid derivative, 4-[(5-bromo-4-{[2,4-dioxo-3-(2-oxo-2-phenylethyl) -1,3-thiazolidin-5-ylidene]methyl}-2-ethoxyphenoxy)methyl]benzoic acid (D77) could potently inhibit the IN-LEDGF/p75 interaction and affect the HIV-1 IN nuclear distribution thus exhibiting antiretroviral activity. Molecular docking with site-directed mutagenesis analysis and surface plasmon resonance (SPR) binding assays has clarified possible binding mode of D77 against HIV-1 integrase. As the firstly discovered small molecular compound targeting HIV-1 integrase interaction with LEDGF/p75, D77 might supply useful structural information for further anti-HIV agent discovery

  4. Molecular dynamics simulation studies of the wild type and E92Q/N155H mutant of Elvitegravir-resistance HIV-1 integrase

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Qi [Shanghai Jiao Tong Univ., Shanghai (China). State Key Lab. of Microbial Metabolism and College of Life Science and Biotechnology; Cheng, Xiaolin [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Center for Molecular Biophysics; Univ. of Tennessee, Knoxville, TN (United States). Dept. of Biochemistry and Cellular and Molecular Biology; Wei, Dongqing [Shanghai Jiao Tong Univ., Shanghai (China). State Key Lab. of Microbial Metabolism and College of Life Science and Biotechnology; Xu, Qin [Shanghai Jiao Tong Univ., Shanghai (China). State Key Lab. of Microbial Metabolism and College of Life Science and Biotechnology

    2014-11-06

    Although Elvitegravir (EVG) is a newly developed antiretrovirals drug to treat the acquired immunodeficiency syndrome (AIDS), drug resistance has already been found in clinic, such as E92Q/N155H and Q148H/G140S. Several structural investigations have already been reported to reveal the molecular mechanism of the drug resistance. As full length crystal structure for HIV-1 integrase is still unsolved, we use in this paper the crystal structure of the full length prototype foamy virus (PFV) in complex with virus DNA and inhibitor Elvitegravir as a template to construct the wild type and E92Q/N155H mutant system of HIV-1 integrase. Molecular dynamic simulations was used to revel the binding mode and the drug resistance of the EVG ligand in E92Q/N155H. Several important interactions were discovered between the mutated residues and the residues in the active site of the E92Q/N155H double mutant pattern, and cross correlation and clustering methods were used for detailed analysis. The results from the MD simulation studies will be used to guide the experimental efforts of developing novel inhibitors against drug-resistant HIV integrase mutants.

  5. Molecular dynamics simulation studies of the wild type and E92Q/N155H mutant of Elvitegravir-resistance HIV-1 integrase.

    Science.gov (United States)

    Chen, Qi; Cheng, Xiaolin; Wei, Dongqing; Xu, Qin

    2015-03-01

    Although Elvitegravir (EVG) is a newly developed antiretrovirals drug to treat the acquired immunodeficiency syndrome (AIDS), drug resistance has already been found in clinic, such as E92Q/N155H and Q148H/G140S. Several structural investigations have already been reported to reveal the molecular mechanism of the drug resistance. As full length crystal structure for HIV-1 integrase is still unsolved, we herein use the crystal structure of the full length prototype foamy virus (PFV) in complex with virus DNA and inhibitor Elvitegravir as a template to construct the wild type and E92Q/N155H mutant system of HIV-1 integrase. Molecular dynamic simulations was used to revel the binding mode and the drug resistance of the EVG ligand in E92Q/N155H. Several important interactions were discovered between the mutated residues and the residues in the active site of the E92Q/N155H double mutant pattern, and cross correlation and clustering methods were used for detailed analysis. The results from the MD simulation studies will be used to guide the experimental efforts of developing novel inhibitors against drug-resistant HIV integrase mutants.

  6. Mutations in the SH1 helix alter the thermal properties of myosin II.

    Science.gov (United States)

    Shibata, Kotomi; Koyama, Tsubasa; Inde, Shohei; Iwai, Sosuke; Chaen, Shigeru

    2017-01-01

    The myosin II SH1 helix is a joint that links the converter subdomain to the rest of the myosin motor domain and possibly plays a key role in the arrangement of the converter/lever arm. Several point mutations within the SH1 helix in human myosin IIs have been shown to cause diseases. To reveal whether these SH1 helix mutations affect not only motile activities but also thermal properties of myosin II, here we introduced the E683K or R686C point mutation into the SH1 helix in Dictyostelium myosin II. Thermal inactivation as well as thermal aggregation rates of these mutant proteins demonstrated that these mutations decreased the thermal stability of myosin II. Temperature dependence of sliding velocities of actin filaments showed that these mutations also reduced the activation energy of a rate-limiting process involved in actin movement. Given that these mutations are likely to alter coupling between the subdomains, and thus their thermal fluctuations, we propose that the SH1 helix is a key structural element that determines the flexibility and thermal properties of the myosin motor. These characteristics of the SH1 helix may contribute to the pathogenesis of the human diseases caused by mutations within this structural element.

  7. Emergence of the persistent spin helix in semiconductor quantum wells

    International Nuclear Information System (INIS)

    Koralek, Jake; Weber, Chris; Orenstein, Joe; Bernevig, Andrei; Zhang, Shoucheng; Mack, Shawn; Awschalom, David

    2008-01-01

    According to Noether's theorem, for every symmetry in nature there is a corresponding conservation law. For example, invariance with respect to spatial translation corresponds to conservation of momentum. In another well-known example, invariance with respect to rotation of the electron's spin, or SU(2) symmetry, leads to conservation of spin polarization. For electrons in a solid, this symmetry is ordinarily broken by spin-orbit (SO) coupling, allowing spin angular momentum to flow to orbital angular momentum. However, it has recently been predicted that SU(2) can be recovered in a two-dimensional electron gas (2DEG), despite the presence of SO coupling. The corresponding conserved quantities include the amplitude and phase of a helical spin density wave termed the 'persistent spin helix' (PSH) .2 SU(2) is restored, in principle, when the strength of two dominant SO interactions, the Rashba (alpha) and linear Dresselhaus (beta 1), are equal. This symmetry is predicted to be robust against all forms of spin-independent scattering, including electron-electron interactions, but is broken by the cubic Dresselhaus term (beta 3) and spin-dependent scattering. When these terms are negligible, the distance over which spin information can propagate is predicted to diverge as alpha approaches beta 1. Here we observe experimentally the emergence of the PSH in GaAs quantum wells (QW's) by independently tuning alpha and beta 1. Using transient spin-grating spectroscopy (TSG), we find a spin-lifetime enhancement of two orders of magnitude near the symmetry point. Excellent quantitative agreement with theory across a wide range of sample parameters allows us to obtain an absolute measure of all relevant SO terms, identifying beta 3 as the main SU(2) violating term in our samples. The tunable suppression of spin-relaxation demonstrated in this work is well-suited for application to spintronics

  8. Emergence of the Persistent Spin Helix in Semiconductor Quantum Wells

    International Nuclear Information System (INIS)

    Koralek, Jake

    2011-01-01

    According to Noether's theorem, for every symmetry in nature there is a corresponding conservation law. For example, invariance with respect to spatial translation corresponds to conservation of momentum. In another well-known example, invariance with respect to rotation of the electron's spin, or SU(2) symmetry, leads to conservation of spin polarization. For electrons in a solid, this symmetry is ordinarily broken by spin-orbit (SO) coupling, allowing spin angular momentum to flow to orbital angular momentum. However, it has recently been predicted that SU(2) can be recovered in a two-dimensional electron gas (2DEG), despite the presence of SO coupling. The corresponding conserved quantities include the amplitude and phase of a helical spin density wave termed the 'persistent spin helix' (PSH). SU(2) is restored, in principle, when the strength of two dominant SO interactions, the Rashba (α) and linear Dresselhaus (β 1 ), are equal. This symmetry is predicted to be robust against all forms of spin-independent scattering, including electron-electron interactions, but is broken by the cubic Dresselhaus term (β 3 ) and spin-dependent scattering. When these terms are negligible, the distance over which spin information can propagate is predicted to diverge as α → β 1 . Here we observe experimentally the emergence of the PSH in GaAs quantum wells (QW's) by independently tuning α and β 1 . Using transient spin-grating spectroscopy (TSG), we find a spin-lifetime enhancement of two orders of magnitude near the symmetry point. Excellent quantitative agreement with theory across a wide range of sample parameters allows us to obtain an absolute measure of all relevant SO terms, identifying β 3 as the main SU(2) violating term in our samples. The tunable suppression of spin-relaxation demonstrated in this work is well-suited for application to spintronics.

  9. Defect-facilitated buckling in supercoiled double-helix DNA

    Science.gov (United States)

    Brahmachari, Sumitabha; Dittmore, Andrew; Takagi, Yasuharu; Neuman, Keir C.; Marko, John F.

    2018-02-01

    We present a statistical-mechanical model for stretched twisted double-helix DNA, where thermal fluctuations are treated explicitly from a Hamiltonian without using any scaling hypotheses. Our model applied to defect-free supercoiled DNA describes the coexistence of multiple plectoneme domains in long DNA molecules at physiological salt concentrations (≈0.1 M Na+) and stretching forces (≈1 pN ) . We find a higher (lower) number of domains at lower (higher) ionic strengths and stretching forces, in accord with experimental observations. We use our model to study the effect of an immobile point defect on the DNA contour that allows a localized kink. The degree of the kink is controlled by the defect size, such that a larger defect further reduces the bending energy of the defect-facilitated kinked end loop. We find that a defect can spatially pin a plectoneme domain via nucleation of a kinked end loop, in accord with experiments and simulations. Our model explains previously reported magnetic tweezer experiments [A. Dittmore et al., Phys. Rev. Lett. 119, 147801 (2017), 10.1103/PhysRevLett.119.147801] showing two buckling signatures: buckling and "rebuckling" in supercoiled DNA with a base-unpaired region. Comparing with experiments, we find that under 1 pN force, a kinked end loop nucleated at a base-mismatched site reduces the bending energy by ≈0.7 kBT per unpaired base. Our model predicts the coexistence of three states at the buckling and rebuckling transitions, which warrants new experiments.

  10. Surfactant in the gas mantle of the snail Helix aspersa.

    Science.gov (United States)

    Daniels, C B; Wood, P G; Loptako, O V; Codd, J R; Johnston, S D; Orgeig, S

    1999-01-01

    Surfactant occurs in cyclically inflating and deflating, gas-holding structures of vertebrates to reduce the surface tension of the inner fluid lining, thereby preventing collapse and decreasing the work of inflation. Here we determined the presence of surfactant in material lavaged from the airspace in the gas mantle of the pulmonate snail Helix aspersa. Surfactant is characterized by the presence of disaturated phospholipid (DSP), especially disaturated phosphatidylcholine (PC), lavaged from the airspace, by the presence of lamellated osmiophilic bodies (LBs) in the airspaces and epithelial tissue, and by the ability of the lavage to reduce surface tension of fluid in a surface balance. Lavage had a DSP/phospholipid (PL) ratio of 0.085, compared to 0.011 in membranes, with the major PL being PC (45.3%). Cholesterol, the primary fluidizer for pulmonary surfactant, was similar in lavage and in lipids extracted from cell homogenates (cholesterol/PL: 0.04 and 0. 03, respectively). LBs were found in the tissues and airspaces. The surface activity of the lavage material is defined as the ability to reduce surface tension under compression to values much lower than that of water. In addition, surface-active lipids will vary surface tension, increasing it upon inspiration as the surface area expands. By these criteria, the surface activity of lavaged material was poor and most similar to that shown by pulmonary lavage of fish and toads. Snail surfactant displays structures, a biochemical PL profile, and biophysical properties similar to surfactant obtained from primitive fish, teleost swim bladders, the lung of the Dipnoan Neoceratodus forsteri, and the amphibian Bufo marinus. However, the cholesterol/PL and cholesterol/DSP ratios are more similar to the amphibian B. marinus than to the fish, and this similarity may indicate a crucial physicochemical relationship for these lipids.

  11. Molecular modeling study on the allosteric inhibition mechanism of HIV-1 integrase by LEDGF/p75 binding site inhibitors.

    Directory of Open Access Journals (Sweden)

    Weiwei Xue

    Full Text Available HIV-1 integrase (IN is essential for the integration of viral DNA into the host genome and an attractive therapeutic target for developing antiretroviral inhibitors. LEDGINs are a class of allosteric inhibitors targeting LEDGF/p75 binding site of HIV-1 IN. Yet, the detailed binding mode and allosteric inhibition mechanism of LEDGINs to HIV-1 IN is only partially understood, which hinders the structure-based design of more potent anti-HIV agents. A molecular modeling study combining molecular docking, molecular dynamics simulation, and binding free energy calculation were performed to investigate the interaction details of HIV-1 IN catalytic core domain (CCD with two recently discovered LEDGINs BI-1001 and CX14442, as well as the LEDGF/p75 protein. Simulation results demonstrated the hydrophobic domain of BI-1001 and CX14442 engages one subunit of HIV-1 IN CCD dimer through hydrophobic interactions, and the hydrophilic group forms hydrogen bonds with HIV-1 IN CCD residues from other subunit. CX14442 has a larger tert-butyl group than the methyl of BI-1001, and forms better interactions with the highly hydrophobic binding pocket of HIV-1 IN CCD dimer interface, which can explain the stronger affinity of CX14442 than BI-1001. Analysis of the binding mode of LEDGF/p75 with HIV-1 IN CCD reveals that the LEDGF/p75 integrase binding domain residues Ile365, Asp366, Phe406 and Val408 have significant contributions to the binding of the LEDGF/p75 to HIV1-IN. Remarkably, we found that binding of BI-1001 and CX14442 to HIV-1 IN CCD induced the structural rearrangements of the 140 s loop and oration displacements of the side chains of the three conserved catalytic residues Asp64, Asp116, and Glu152 located at the active site. These results we obtained will be valuable not only for understanding the allosteric inhibition mechanism of LEDGINs but also for the rational design of allosteric inhibitors of HIV-1 IN targeting LEDGF/p75 binding site.

  12. The C-Terminal RpoN Domain of sigma54 Forms an unpredictedHelix-Turn-Helix Motif Similar to domains of sigma70

    Energy Technology Data Exchange (ETDEWEB)

    Doucleff, Michaeleen; Malak, Lawrence T.; Pelton, Jeffrey G.; Wemmer, David E.

    2005-11-01

    The ''{delta}'' subunit of prokaryotic RNA-polymerase allows gene-specific transcription initiation. Two {sigma} families have been identified, {sigma}{sup 70} and {sigma}{sup 54}, which use distinct mechanisms to initiate transcription and share no detectable sequence homology. Although the {sigma}{sup 70}-type factors have been well characterized structurally by x-ray crystallography, no high-resolution structural information is available for the {sigma}{sup 54}-type factors. Here we present the NMR derived structure of the C-terminal domain of {sigma}{sup 54} from Aquifex aeolicus. This domain (Thr323 to Gly389), which contains the highly conserved RpoN box sequence, consists of a poorly structured N-terminal tail followed by a three-helix bundle, which is surprisingly similar to domains of the {sigma}{sup 70}-type proteins. Residues of the RpoN box, which have previously been shown to be critical for DNA binding, form the second helix of an unpredicted helix-turn-helix motif. This structure's homology with other DNA binding proteins, combined with previous biochemical data, suggest how the C-terminal domain of {sigma}{sup 54} binds to DNA.

  13. CFD analysis and flow model reduction for surfactant production in helix reactor = CFD analiza i redukcija modela strujanja za proizvodnju surfaktanta u helix reaktoru

    NARCIS (Netherlands)

    Nikačević, N.M.; Thielen, L.; Twerda, A.; Hof, P.M.J. van den

    2015-01-01

    Flow pattern analysis in a spiral Helix reactor is conducted, for the application in commercial surfactant production. Step change response curves (SCR) were obtained from numerical tracer experiments by three-dimensional computational fluid dynamics (CFD) simulations. Non-reactive flow is

  14. Evolutionary repair of HIV type 1 gp41 with a kink in the N-terminal helix leads to restoration of the six-helix bundle structure

    NARCIS (Netherlands)

    Sanders, Rogier W.; Busser, Els; Moore, John P.; Lu, Min; Berkhout, Ben

    2004-01-01

    The HIV-1 envelope glycoprotein complex (Env) can be stabilized by the introduction of a disulfide bond between the gp120 and gp41 subunits. The resulting protein is monomeric, but trimerization can be improved by the introduction of a single helix-breaking residue at the conserved Ile559 site in

  15. Production of transgenic cattle highly expressing human serum albumin in milk by phiC31 integrase-mediated gene delivery.

    Science.gov (United States)

    Luo, Yan; Wang, Yongsheng; Liu, Jun; Lan, Hui; Shao, Minghao; Yu, Yuan; Quan, Fusheng; Zhang, Yong

    2015-10-01

    Transgenic cattle expressing high levels of recombinant human serum albumin (HSA) in their milk may as an alternative source for commercial production. Our objective was to produce transgenic cattle highly expressing HSA in milk by using phiC31 integrase system and somatic cell nuclear transfer (SCNT). The mammary-specific expression plasmid pIACH(-), containing the attB recognition site for phiC31 integrase, were co-transfected with integrase expression plasmid pCMVInt into bovine fetal fibroblast cells (BFFs). PhiC31 integrase-mediated integrations in genome of BFFs were screened by nested inverse PCR. After analysis of sequence of the PCR products, 46.0% (23/50) of the both attB-genome junction sites (attL and attR) were confirmed, and four pseudo attP sites were identified. The integration rates in BF3, BF11, BF19 and BF4 sites were 4.0% (2/50), 6.0% (3/50), 16.0% (8/50) and 20.0% (10/50), respectively. BF3 is located in the bovine chromosome 3 collagen alpha-3 (VI) chain isomer 2 gene, while the other three sites are located in the non-coding region. The transgenic cell lines from BF11, BF19 and BF4 sites were used as donors for SCNT. Two calves from transgenic cells BF19 were born, one died within a few hours after birth, and another calf survived healthy. PCR and Southern blot analysis revealed integration of the transgene in the genome of cloned calves. The nested reverse PCR confirmed that the integration site in cloned calves was identical to the donor cells. The western blotting assessment indicated that recombinant HSA was expressed in the milk of transgenic cattle and the expression level was about 4-8 mg/mL. The present study demonstrated that phiC31 integrase system was an efficient and safety gene delivery tool for producing HSA transgenic cattle. The production of recombinant HSA in the milk of cattle may provide a large-scale and cost-effective resource.

  16. The R263K mutation in HIV integrase that is selected by dolutegravir may actually prevent clinically relevant resistance to this compound.

    Science.gov (United States)

    Wainberg, Mark; Anstett, Kaitlin; Mesplede, Thibault; Quashie, Peter; Han, Yingshan; Oliveira, Maureen

    2014-01-01

    Drug resistance against dolutegravir (DTG) or the nucleosides with which it has been co-administered has never been observed in patients receiving this drug in first-line therapy. In contrast, a R263K mutation that confers low-level resistance (3-4 fold) to DTG has been selected by DTG in culture. Our group has ascribed the absence of resistance to DTG to the high fitness cost exacted by the R263K mutation and an inability of HIV to generate compensatory mutations. We generated recombinant integrase enzymes and viruses containing various combinations of mutations and studied these enzymatically and in culture. We also selected for resistance against raltegravir (RAL) using viruses containing the R263K mutation. The R263K mutation alone conferred an approximate 3-fold level of resistance to DTG and a 40% loss in viral replicative capacity and recombinant integrase activity. Secondary mutations selected at positions H51Y or E138K did not individually affect either enzyme activity or DTG resistance, but the combination of R263K together with H51Y or E138K increased DTG resistance to about 7-fold accompanied by a ≈75% loss in each of viral replication capacity, and both in vitro and in vivo integrase activity. Conversely, combinations of R263K together with multiple resistance mutations for RAL and/or EVG at positions 92,143, 148 and 155 resulted in even further diminished enzymatic activity that may be incompatible with viral survival. Modelling of the 3-dimensional structure of integrase suggests that R263K is located in a region that may not permit further mutagenesis if secondary mutations at H51Y or E138K are also present. Moreover, integrase that contains R263K together with substitutions at positions 92, 143, 148 and 155 may be enzymatically inactive. The use of the R263K-containing virus to select for resistance to RAL led to the appearance of RAL-containing mutations but the loss of R263K. Secondary mutations to R263K following selection with DTG have all

  17. Bicyclic 1-hydroxy-2-oxo-1,2-dihydropyridine-3-carboxamide-containing HIV-1 integrase inhibitors having high antiviral potency against cells harboring raltegravir-resistant integrase mutants.

    Science.gov (United States)

    Zhao, Xue Zhi; Smith, Steven J; Métifiot, Mathieu; Johnson, Barry C; Marchand, Christophe; Pommier, Yves; Hughes, Stephen H; Burke, Terrence R

    2014-02-27

    Integrase (IN) inhibitors are the newest class of antiretroviral agents developed for the treatment of HIV-1 infections. Merck's Raltegravir (RAL) (October 2007) and Gilead's Elvitegravir (EVG) (August 2012), which act as IN strand transfer inhibitors (INSTIs), were the first anti-IN drugs to be approved by the FDA. However, the virus develops resistance to both RAL and EVG, and there is extensive cross-resistance to these two drugs. New "2nd-generation" INSTIs are needed that will have greater efficacy against RAL- and EVG-resistant strains of IN. The FDA has recently approved the first second generation INSTI, GSK's Dolutegravir (DTG) (August 2013). Our current article describes the design, synthesis, and evaluation of a series of 1,8-dihydroxy-2-oxo-1,2-dihydroquinoline-3-carboxamides, 1,4-dihydroxy-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamides, and 1-hydroxy-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carboxamides. This resulted in the identification of noncytotoxic inhibitors that exhibited single digit nanomolar EC50 values against HIV-1 vectors harboring wild-type IN in cell-based assays. Importantly, some of these new inhibitors retain greater antiviral efficacy compared to that of RAL when tested against a panel of IN mutants that included Y143R, N155H, G140S/Q148H, G118R, and E138K/Q148K.

  18. pH responsiveness of fibrous assemblies of repeat-sequence amphipathic α-helix polypeptides.

    Science.gov (United States)

    Takei, Toshiaki; Tsumoto, Kouhei; Okonogi, Atsuhito; Kimura, Akiko; Kojima, Shuichi; Yazaki, Kazumori; Takei, Tsunetomo; Ueda, Takuya; Miura, Kin-ichiro

    2015-05-01

    We reported previously that our designed polypeptide α3 (21 residues), which has three repeats of a seven-amino-acid sequence (LETLAKA)3, forms not only an amphipathic α-helix structure but also long fibrous assemblies in aqueous solution. To address the relationship between the electrical states of the polypeptide and its α-helix and fibrous assembly formation, we characterized mutated polypeptides in which charged amino acid residues of α3 were replaced with Ser. We prepared the following polypeptides: 2Sα3 (LSTLAKA)3, in which all Glu residues were replaced with Ser residues; 6Sα3 (LETLASA)3, in which all Lys residues were replaced with Ser; and 2S6Sα3 (LSTLASA)3; in which all Glu and Lys residues were replaced with Ser. In 0.1M KCl, 2Sα3 formed an α-helix under basic conditions and 6Sα3 formed an α-helix under acid conditions. In 1M KCl, they both formed α-helices under a wide pH range. In addition, 2Sα3 and 6Sα3 formed fibrous assemblies under the same buffer conditions in which they formed α-helices. α-Helix and fibrous assembly formation by these polypeptides was reversible in a pH-dependent manner. In contrast, 2S6Sα3 formed an α-helix under basic conditions in 1M KCl. Taken together, these findings reveal that the charge states of the charged amino acid residues and the charge state of the Leu residue located at the terminus play an important role in α-helix formation. © 2015 The Protein Society.

  19. Integrase-independent HIV-1 infection is augmented under conditions of DNA damage and produces a viral reservoir

    International Nuclear Information System (INIS)

    Ebina, Hirotaka; Kanemura, Yuka; Suzuki, Yasutsugu; Urata, Kozue; Misawa, Naoko; Koyanagi, Yoshio

    2012-01-01

    HIV-1 possesses a viral protein, integrase (IN), which is necessary for its efficient integration in target cells. However, it has been reported that an IN-defective HIV strain is still capable of integration. Here, we assessed the ability of wild type (WT) HIV-1 to establish infection in the presence of IN inhibitors. We observed a low, yet clear infection of inhibitor-incubated cells infected with WT HIV which was identical to cells infected with IN-deficient HIV, D64A. Furthermore, the IN-independent integration could be enhanced by the pretreatment of cells with DNA-damaging agents suggesting that integration is mediated by a DNA repair system. Moreover, significantly faster viral replication kinetics with augmented viral DNA integration was observed after infection in irradiated cells treated with IN inhibitor compared to nonirradiated cells. Altogether, our results suggest that HIV DNA has integration potential in the presence of an IN inhibitor and may serve as a virus reservoir.

  20. Adeno-associated virus Rep-mediated targeting of integrase-defective retroviral vector DNA circles into human chromosome 19

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Shuohao [Graduate School of Systems Life Sciences, Kyushu University, 744 Motooka, Nishi-ku, Fukuoka 819-0395 (Japan); Kawabe, Yoshinori; Ito, Akira [Department of Chemical Engineering, Faculty of Engineering, Kyushu University, 744 Motooka, Nishi-ku, Fukuoka 819-0395 (Japan); Kamihira, Masamichi, E-mail: kamihira@chem-eng.kyushu-u.ac.jp [Graduate School of Systems Life Sciences, Kyushu University, 744 Motooka, Nishi-ku, Fukuoka 819-0395 (Japan); Department of Chemical Engineering, Faculty of Engineering, Kyushu University, 744 Motooka, Nishi-ku, Fukuoka 819-0395 (Japan)

    2012-01-06

    Highlights: Black-Right-Pointing-Pointer Adeno-associated virus (AAV) is capable of targeted integration in human cells. Black-Right-Pointing-Pointer Integrase-defective retroviral vector (IDRV) enables a circular DNA delivery. Black-Right-Pointing-Pointer A targeted integration system of IDRV DNA using the AAV integration mechanism. Black-Right-Pointing-Pointer Targeted IDRV integration ameliorates the safety concerns for retroviral vectors. -- Abstract: Retroviral vectors have been employed in clinical trials for gene therapy owing to their relative large packaging capacity, alterable cell tropism, and chromosomal integration for stable transgene expression. However, uncontrollable integrations of transgenes are likely to cause safety issues, such as insertional mutagenesis. A targeted transgene integration system for retroviral vectors, therefore, is a straightforward way to address the insertional mutagenesis issue. Adeno-associated virus (AAV) is the only known virus capable of targeted integration in human cells. In the presence of AAV Rep proteins, plasmids possessing the p5 integration efficiency element (p5IEE) can be integrated into the AAV integration site (AAVS1) in the human genome. In this report, we describe a system that can target the circular DNA derived from non-integrating retroviral vectors to the AAVS1 site by utilizing the Rep/p5IEE integration mechanism. Our results showed that after G418 selection 30% of collected clones had retroviral DNA targeted at the AAVS1 site.

  1. Resistance Analyses of Integrase Strand Transfer Inhibitors within Phase 3 Clinical Trials of Treatment-Naive Patients

    Directory of Open Access Journals (Sweden)

    Kirsten L. White

    2014-07-01

    Full Text Available The integrase (IN strand transfer inhibitors (INSTIs, raltegravir (RAL, elvitegravir (EVG and dolutegravir (DTG, comprise the newest drug class approved for the treatment of HIV-1 infection, which joins the existing classes of reverse transcriptase, protease and binding/entry inhibitors. The efficacy of first-line regimens has attained remarkably high levels, reaching undetectable viral loads in 90% of patients by Week 48; however, there remain patients who require a change in regimen due to adverse events, virologic failure with emergent resistance or other issues of patient management. Large, randomized clinical trials conducted in antiretroviral treatment-naive individuals are required for drug approval in this population in the US, EU and other countries, with the primary endpoint for virologic success at Week 48. However, there are differences in the definition of virologic failure and the evaluation of drug resistance among the trials. This review focuses on the methodology and tabulation of resistance to INSTIs in phase 3 clinical trials of first-line regimens and discusses case studies of resistance.

  2. Resistance Analyses of Integrase Strand Transfer Inhibitors within Phase 3 Clinical Trials of Treatment-Naive Patients

    Science.gov (United States)

    White, Kirsten L.; Raffi, Francois; Miller, Michael D.

    2014-01-01

    The integrase (IN) strand transfer inhibitors (INSTIs), raltegravir (RAL), elvitegravir (EVG) and dolutegravir (DTG), comprise the newest drug class approved for the treatment of HIV-1 infection, which joins the existing classes of reverse transcriptase, protease and binding/entry inhibitors. The efficacy of first-line regimens has attained remarkably high levels, reaching undetectable viral loads in 90% of patients by Week 48; however, there remain patients who require a change in regimen due to adverse events, virologic failure with emergent resistance or other issues of patient management. Large, randomized clinical trials conducted in antiretroviral treatment-naive individuals are required for drug approval in this population in the US, EU and other countries, with the primary endpoint for virologic success at Week 48. However, there are differences in the definition of virologic failure and the evaluation of drug resistance among the trials. This review focuses on the methodology and tabulation of resistance to INSTIs in phase 3 clinical trials of first-line regimens and discusses case studies of resistance. PMID:25054884

  3. A QSAR study of integrase strand transfer inhibitors based on a large set of pyrimidine, pyrimidone, and pyridopyrazine carboxamide derivatives

    Science.gov (United States)

    de Campos, Luana Janaína; de Melo, Eduardo Borges

    2017-08-01

    In the present study, 199 compounds derived from pyrimidine, pyrimidone and pyridopyrazine carboxamides with inhibitory activity against HIV-1 integrase were modeled. Subsequently, a multivariate QSAR study was conducted with 54 molecules employed by Ordered Predictors Selection (OPS) and Partial Least Squares (PLS) for the selection of variables and model construction, respectively. Topological, electrotopological, geometric, and molecular descriptors were used. The selected real model was robust and free from chance correlation; in addition, it demonstrated favorable internal and external statistical quality. Once statistically validated, the training model was used to predict the activity of a second data set (n = 145). The root mean square deviation (RMSD) between observed and predicted values was 0.698. Although it is a value outside of the standards, only 15 (10.34%) of the samples exhibited higher residual values than 1 log unit, a result considered acceptable. Results of Williams and Euclidean applicability domains relative to the prediction showed that the predictions did not occur by extrapolation and that the model is representative of the chemical space of test compounds.

  4. DNA binding specificities of Escherichia coli Cas1–Cas2 integrase drive its recruitment at the CRISPR locus

    Science.gov (United States)

    Moch, Clara; Fromant, Michel; Blanquet, Sylvain

    2017-01-01

    Abstract Prokaryotic adaptive immunity relies on the capture of fragments of invader DNA (protospacers) followed by their recombination at a dedicated acceptor DNA locus. This integrative mechanism, called adaptation, needs both Cas1 and Cas2 proteins. Here, we studied in vitro the binding of an Escherichia coli Cas1–Cas2 complex to various protospacer and acceptor DNA molecules. We show that, to form a long-lived ternary complex containing Cas1–Cas2, the acceptor DNA must carry a CRISPR locus, and the protospacer must not contain 3΄-single-stranded overhangs longer than 5 bases. In addition, the acceptor DNA must be supercoiled. Formation of the ternary complex is synergistic, in such that the binding of Cas1–Cas2 to acceptor DNA is reinforced in the presence of a protospacer. Mutagenesis analysis at the CRISPR locus indicates that the presence in the acceptor plasmid of the palindromic motif found in CRISPR repeats drives stable ternary complex formation. Most of the mutations in this motif are deleterious even if they do not prevent cruciform structure formation. The leader sequence of the CRISPR locus is fully dispensable. These DNA binding specificities of the Cas1–Cas2 integrase are likely to play a major role in the recruitment of this enzyme at the CRISPR locus. PMID:28034956

  5. Integrase-independent HIV-1 infection is augmented under conditions of DNA damage and produces a viral reservoir

    Energy Technology Data Exchange (ETDEWEB)

    Ebina, Hirotaka, E-mail: hebina@virus.kyoto-u.ac.jp; Kanemura, Yuka; Suzuki, Yasutsugu; Urata, Kozue; Misawa, Naoko; Koyanagi, Yoshio

    2012-05-25

    HIV-1 possesses a viral protein, integrase (IN), which is necessary for its efficient integration in target cells. However, it has been reported that an IN-defective HIV strain is still capable of integration. Here, we assessed the ability of wild type (WT) HIV-1 to establish infection in the presence of IN inhibitors. We observed a low, yet clear infection of inhibitor-incubated cells infected with WT HIV which was identical to cells infected with IN-deficient HIV, D64A. Furthermore, the IN-independent integration could be enhanced by the pretreatment of cells with DNA-damaging agents suggesting that integration is mediated by a DNA repair system. Moreover, significantly faster viral replication kinetics with augmented viral DNA integration was observed after infection in irradiated cells treated with IN inhibitor compared to nonirradiated cells. Altogether, our results suggest that HIV DNA has integration potential in the presence of an IN inhibitor and may serve as a virus reservoir.

  6. Identification of novel mutations responsible for resistance to MK-2048, a second-generation HIV-1 integrase inhibitor.

    Science.gov (United States)

    Bar-Magen, Tamara; Sloan, Richard D; Donahue, Daniel A; Kuhl, Björn D; Zabeida, Alexandra; Xu, Hongtao; Oliveira, Maureen; Hazuda, Daria J; Wainberg, Mark A

    2010-09-01

    MK-2048 represents a prototype second-generation integrase strand transfer inhibitor (INSTI) developed with the goal of retaining activity against viruses containing mutations associated with resistance to first-generation INSTIs, raltegravir (RAL) and elvitegravir (EVG). Here, we report the identification of mutations (G118R and E138K) which confer resistance to MK-2048 and not to RAL or EVG. These mutations were selected in vitro and confirmed by site-specific mutagenesis. G118R, which appeared first in cell culture, conferred low levels of resistance to MK-2048. G118R also reduced viral replication capacity to approximately 1% that of the isogenic wild-type (wt) virus. The subsequent selection of E138K partially restored replication capacity to approximately 13% of wt levels and increased resistance to MK-2048 to approximately 8-fold. Viruses containing G118R and E138K remained largely susceptible to both RAL and EVG, suggesting a unique interaction between this second-generation INSTI and the enzyme may be defined by these residues as a potential basis for the increased intrinsic affinity and longer "off" rate of MK-2048. In silico structural analysis suggests that the introduction of a positively charged arginine at position 118, near the catalytic amino acid 116, might decrease Mg(2+) binding, compromising enzyme function and thus leading to the significant reduction in both integration and viral replication capacity observed with these mutations.

  7. Identification of Novel Mutations Responsible for Resistance to MK-2048, a Second-Generation HIV-1 Integrase Inhibitor ▿

    Science.gov (United States)

    Bar-Magen, Tamara; Sloan, Richard D.; Donahue, Daniel A.; Kuhl, Björn D.; Zabeida, Alexandra; Xu, Hongtao; Oliveira, Maureen; Hazuda, Daria J.; Wainberg, Mark A.

    2010-01-01

    MK-2048 represents a prototype second-generation integrase strand transfer inhibitor (INSTI) developed with the goal of retaining activity against viruses containing mutations associated with resistance to first-generation INSTIs, raltegravir (RAL) and elvitegravir (EVG). Here, we report the identification of mutations (G118R and E138K) which confer resistance to MK-2048 and not to RAL or EVG. These mutations were selected in vitro and confirmed by site-specific mutagenesis. G118R, which appeared first in cell culture, conferred low levels of resistance to MK-2048. G118R also reduced viral replication capacity to approximately 1% that of the isogenic wild-type (wt) virus. The subsequent selection of E138K partially restored replication capacity to ≈13% of wt levels and increased resistance to MK-2048 to ≈8-fold. Viruses containing G118R and E138K remained largely susceptible to both RAL and EVG, suggesting a unique interaction between this second-generation INSTI and the enzyme may be defined by these residues as a potential basis for the increased intrinsic affinity and longer “off” rate of MK-2048. In silico structural analysis suggests that the introduction of a positively charged arginine at position 118, near the catalytic amino acid 116, might decrease Mg2+ binding, compromising enzyme function and thus leading to the significant reduction in both integration and viral replication capacity observed with these mutations. PMID:20610719

  8. Design, synthesis, and evaluation of an alpha-helix mimetic library targeting protein-protein interactions.

    Science.gov (United States)

    Shaginian, Alex; Whitby, Landon R; Hong, Sukwon; Hwang, Inkyu; Farooqi, Bilal; Searcey, Mark; Chen, Jiandong; Vogt, Peter K; Boger, Dale L

    2009-04-22

    The design and solution-phase synthesis of an alpha-helix mimetic library as an integral component of a small-molecule library targeting protein-protein interactions are described. The iterative design, synthesis, and evaluation of the candidate alpha-helix mimetic was initiated from a precedented triaryl template and refined by screening the designs for inhibition of MDM2/p53 binding. Upon identifying a chemically and biologically satisfactory design and consistent with the screening capabilities of academic collaborators, the corresponding complete library was assembled as 400 mixtures of 20 compounds (20 x 20 x 20-mix), where the added subunits are designed to mimic all possible permutations of the naturally occurring i, i + 4, i + 7 amino acid side chains of an alpha-helix. The library (8000 compounds) was prepared using a solution-phase synthetic protocol enlisting acid/base liquid-liquid extractions for purification on a scale that insures its long-term availability for screening campaigns. Screening of the library for inhibition of MDM2/p53 binding not only identified the lead alpha-helix mimetic upon which the library was based, but also suggests that a digestion of the initial screening results that accompany the use of such a comprehensive library can provide insights into the nature of the interaction (e.g., an alpha-helix mediated protein-protein interaction) and define the key residues and their characteristics responsible for recognition.

  9. Structure, stability, and thermodynamics of a short intermolecular purine-purine-pyrimidine triple helix

    International Nuclear Information System (INIS)

    Pilch, D.S.; Shafer, R.H.; Levenson, C.

    1991-01-01

    The authors have investigated the structure and physical chemistry of the d(C 3 T 4 C 3 )·2[d(G 3 A 4 G 3 )] triple helix by polyacrylamide gel electrophoresis (PAGE), 1 H NMR, and ultraviolet (UV) absorption spectroscopy. The triplex was stabilized with MgCl 2 at neutral pH. PAGE studies verify the stoichiometry of the strands comprising the triplex and indicate that the orientation of the third strand in purine-purine-pyrimidine (pur-pur-pyr) triplexes is antiparallel with respect to the purine strand of the underlying duplex. Imino proton NMR spectra provide evidence for the existence of new purine-purine (pur·pur) hydrogen bonds, in addition to those of the Watson-Crick (W-C) base pairs, in the triplex structure. These new hydrogen bonds are likely to correspond to the interaction between third-strand guanine NH1 imino protons and the N7 atoms of guanine residues on the puring strand of the underlying duplex. Thermal denaturation of the triplex proceeds to single strands in one step, under the conditions used in this study. Binding of the third strand appears to enhance the thermal stability of the duplex by 1-3 C, depending on the DNA concentration. This marked enhancement in stability, coupled with the lack of an acidic pH requirement, suggests that pur-pur-pyr triplexes are appealing choices for use in applications involving oligonucleotide targeting of duplex DNA in vitro and in vivo

  10. Buried lysine, but not arginine, titrates and alters transmembrane helix tilt.

    Science.gov (United States)

    Gleason, Nicholas J; Vostrikov, Vitaly V; Greathouse, Denise V; Koeppe, Roger E

    2013-01-29

    The ionization states of individual amino acid residues of membrane proteins are difficult to decipher or assign directly in the lipid-bilayer membrane environment. We address this issue for lysines and arginines in designed transmembrane helices. For lysines (but not arginines) at two locations within dioleoyl-phosphatidylcholine bilayer membranes, we measure pK(a) values below 7.0. We find that buried charged lysine, in fashion similar to arginine, will modulate helix orientation to maximize its own access to the aqueous interface or, if occluded by aromatic rings, may cause a transmembrane helix to exit the lipid bilayer. Interestingly, the influence of neutral lysine (vis-à-vis leucine) upon helix orientation also depends upon its aqueous access. Our results suggest that changes in the ionization states of particular residues will regulate membrane protein function and furthermore illustrate the subtle complexity of ionization behavior with respect to the detailed lipid and protein environment.

  11. Golden Helix Institute of Biomedical Research: Interdisciplinary research and educational activities in pharmacogenomics and personalized medicine

    Science.gov (United States)

    Mitropoulos, Konstantinos; Innocenti, Federico; van Schaik, Ron H.; Lezhava, Alexander; Tzimas, Giannis; Kollia, Panagoula; Macek, Milan; Fortina, Paolo; Patrinos, George P.

    2013-01-01

    The Golden Helix Institute of Biomedical Research is an international non-profit scientific organization with interdisciplinary research and educational activities in the field of genome medicine in Europe, Asia and Latin America. These activities are supervised by an international scientific advisory council, consisting of world leaders in the field of genomics and translational medicine. Research activities include the regional coordination of the Pharmacogenomics for Every Nation Initiative in Europe, in an effort to integrate pharmacogenomics in developing countries, the development of several National/Ethnic Genetic databases and related web services and the critical assessment of the impact of genetics and genomic medicine to society in various countries. Also, educational activities include the organization of the Golden Helix Symposia®, which are high profile scientific research symposia in the field of personalized medicine, and the Golden Helix Pharmacogenomics Days, an international educational activity focused on pharmacogenomics, as part of its international pharmacogenomics education and outreach efforts. PMID:22379996

  12. Progesterone modulation of transmembrane helix-helix interactions between the α-subunit of Na/K-ATPase and phospholipid N-methyltransferase in the oocyte plasma membrane

    Directory of Open Access Journals (Sweden)

    Askari Amir

    2010-05-01

    Full Text Available Abstract Background Progesterone binding to the surface of the amphibian oocyte initiates the meiotic divisions. Our previous studies with Rana pipiens oocytes indicate that progesterone binds to a plasma membrane site within the external loop between the M1 and M2 helices of the α-subunit of Na/K-ATPase, triggering a cascade of lipid second messengers and the release of the block at meiotic prophase. We have characterized this site, using a low affinity ouabain binding isoform of the α1-subunit. Results Preparations of isolated plasma membranes from Rana oocytes demonstrate that physiological levels of progesterone (or the non-metabolizable progestin R5020 successively activate phosphatidylethanolamine-N-methyltransferase (PE-NMT and sphingomyelin synthase within seconds. Inhibition of PE-NMT blocks the progesterone induction of meiosis in intact oocytes, whereas its initial product, phosphatidylmonomethylethanolamine (PME, can itself initiate meiosis in the presence of the inhibitor. Published X-ray crystallographic data on Na/K-ATPase, computer-generated 3D projections, heptad repeat analysis and hydrophobic cluster analysis of the transmembrane helices predict that hydrophobic residues L, V, V, I, F and Y of helix M2 of the α1-subunit interact with F, L, G, L, L and F, respectively, of helix M3 of PE-NMT. Conclusion We propose that progesterone binding to the first external loop of the α1-subunit facilitates specific helix-helix interactions between integral membrane proteins to up-regulate PE-NMT, and, that successive interactions between two or more integral plasma membrane proteins induce the signaling cascades which result in completion of the meiotic divisions.

  13. The Quadruple Helix Model Enhancing Innovative Performance Of Indonesian Creative Industry

    Directory of Open Access Journals (Sweden)

    Sri Wahyu Lelly Hana Setyanti

    2017-11-01

    Full Text Available The creative industry in Indonesia has contributed positively to the national economic growth. Creative industry grows from the creativity and innovation performance of the business actors. The challenge of creative industry is how to completely understand the creative and innovative processes in business management. Therefore it requires an approach that combines the synergy between academicians entrepreneurs government and society in a quadruple helix model. The objective of this research is to develop a creativity model through a quadruple helix model in improving innovation performance of the creative industry.

  14. Neurotoxic and Lipidic peroxidation effect of metal dust and Cadmium on Helix aspersa

    Directory of Open Access Journals (Sweden)

    GRARA Nedjoud

    2017-04-01

    Full Text Available In this study we were interested in the evaluation of the impact of the metal dust collected on the level of the iron and steel complex of EL-Hadjar and the Cadmium which is regarded as the most toxic pollutant, most widespread in the environment of the zones to strong human activities and their effects on organizations bioaccumulator and bio indicator of pollution Helix aspersa. With regard to the bio markers we highlighted a reduction in the AChE activity on the level of the head. In addition, the exposure of Helix aspersa to metal dust and Cadmium induces a lipidic peroxidation with release of (MDA.

  15. Gate-controlled switching between persistent and inverse persistent spin helix states

    International Nuclear Information System (INIS)

    Yoshizumi, K.; Sasaki, A.; Kohda, M.; Nitta, J.

    2016-01-01

    We demonstrate gate-controlled switching between persistent spin helix (PSH) state and inverse PSH state, which are detected by quantum interference effect on magneto-conductance. These special symmetric spin states showing weak localization effect give rise to a long spin coherence when the strength of Rashba spin-orbit interaction (SOI) is close to that of Dresselhaus SOI. Furthermore, in the middle of two persistent spin helix states, where the Rashba SOI can be negligible, the bulk Dresselhaus SOI parameter in a modulation doped InGaAs/InAlAs quantum well is determined.

  16. Gate-controlled switching between persistent and inverse persistent spin helix states

    Energy Technology Data Exchange (ETDEWEB)

    Yoshizumi, K.; Sasaki, A.; Kohda, M.; Nitta, J. [Department of Materials Science, Tohoku University, Sendai 980-8579 (Japan)

    2016-03-28

    We demonstrate gate-controlled switching between persistent spin helix (PSH) state and inverse PSH state, which are detected by quantum interference effect on magneto-conductance. These special symmetric spin states showing weak localization effect give rise to a long spin coherence when the strength of Rashba spin-orbit interaction (SOI) is close to that of Dresselhaus SOI. Furthermore, in the middle of two persistent spin helix states, where the Rashba SOI can be negligible, the bulk Dresselhaus SOI parameter in a modulation doped InGaAs/InAlAs quantum well is determined.

  17. The signaling helix: a common functional theme in diverse signaling proteins

    Directory of Open Access Journals (Sweden)

    Aravind L

    2006-09-01

    Full Text Available Abstract Background The mechanism by which the signals are transmitted between receptor and effector domains in multi-domain signaling proteins is poorly understood. Results Using sensitive sequence analysis methods we identify a conserved helical segment of around 40 residues in a wide range of signaling proteins, including numerous sensor histidine kinases such as Sln1p, and receptor guanylyl cyclases such as the atrial natriuretic peptide receptor and nitric oxide receptors. We term this helical segment the signaling (S-helix and present evidence that it forms a novel parallel coiled-coil element, distinct from previously known helical segments in signaling proteins, such as the Dimerization-Histidine phosphotransfer module of histidine kinases, the intra-cellular domains of the chemotaxis receptors, inter-GAF domain helical linkers and the α-helical HAMP module. Analysis of domain architectures allowed us to reconstruct the domain-neighborhood graph for the S-helix, which showed that the S-helix almost always occurs between two signaling domains. Several striking patterns in the domain neighborhood of the S-helix also became evident from the graph. It most often separates diverse N-terminal sensory domains from various C-terminal catalytic signaling domains such as histidine kinases, cNMP cyclase, PP2C phosphatases, NtrC-like AAA+ ATPases and diguanylate cyclases. It might also occur between two sensory domains such as PAS domains and occasionally between a DNA-binding HTH domain and a sensory domain. The sequence conservation pattern of the S-helix revealed the presence of a unique constellation of polar residues in the dimer-interface positions within the central heptad of the coiled-coil formed by the S-helix. Conclusion Combining these observations with previously reported mutagenesis studies on different S-helix-containing proteins we suggest that it functions as a switch that prevents constitutive activation of linked downstream

  18. Mutational analyses of the core domain of Avian Leukemia and Sarcoma Viruses integrase: critical residues for concerted integration and multimerization

    International Nuclear Information System (INIS)

    Moreau, Karen; Faure, Claudine; Violot, Sebastien; Gouet, Patrice; Verdier, Gerard; Ronfort, Corinne

    2004-01-01

    During replicative cycle of retroviruses, the reverse-transcribed viral DNA is integrated into the cell DNA by the viral integrase (IN) enzyme. The central core domain of IN contains the catalytic site of the enzyme and is involved in binding viral ends and cell DNA as well as dimerization. We previously performed single amino acid substitutions in the core domain of an Avian Leukemia and Sarcoma Virus (ALSV) IN [Arch. Virol. 147 (2002) 1761]. Here, we modeled the resulting IN mutants and analyzed the ability of these mutants to mediate concerted DNA integration in an in vitro assay, and to form dimers by protein-protein cross-linking and size exclusion chromatography. The N197C mutation resulted in the inability of the mutant to perform concerted integration that was concomitant with a loss of IN dimerization. Surprisingly, mutations Q102G and A106V at the dimer interface resulted in mutants with higher efficiencies than the wild-type IN in performing two-ended concerted integration of viral DNA ends. The G139D and A195V mutants had a trend to perform one-ended DNA integration of viral ends instead of two-ended integration. More drastically, the I88L and L135G mutants preferentially mediated nonconcerted DNA integration although the proteins form dimers. Therefore, these mutations may alter the formation of IN complexes of higher molecular size than a dimer that would be required for concerted integration. This study points to the important role of core domain residues in the concerted integration of viral DNA ends as well as in the oligomerization of the enzyme

  19. PH4 of petunia is an R2R3-MYB protein that activates vacuolar acidification through interactions with Basic-Helix-Loop-Helix transcription factors of the anthocyanin pathway.

    NARCIS (Netherlands)

    Quattrocchio, F.M.; Verweij, C.W.; Spelt, C.E.; Mol, J.N.M.; Koes, R.E.

    2007-01-01

    The Petunia hybrids genes ANTHOCYANIN1 (AN1) and AN2 encode transcription factors with a basic-helix-loop-helix (BHLH) and a MYB domain, respectively, that are required for anthocyanin synthesis and acidification of the vacuole in petal cells. Mutation of PH4 results in a bluer flower color,

  20. Structure-activity relationship of pyrrolyl diketo acid derivatives as dual inhibitors of HIV-1 integrase and reverse transcriptase ribonuclease H domain.

    Science.gov (United States)

    Cuzzucoli Crucitti, Giuliana; Métifiot, Mathieu; Pescatori, Luca; Messore, Antonella; Madia, Valentina Noemi; Pupo, Giovanni; Saccoliti, Francesco; Scipione, Luigi; Tortorella, Silvano; Esposito, Francesca; Corona, Angela; Cadeddu, Marta; Marchand, Christophe; Pommier, Yves; Tramontano, Enzo; Costi, Roberta; Di Santo, Roberto

    2015-02-26

    The development of HIV-1 dual inhibitors is a highly innovative approach aimed at reducing drug toxic side effects as well as therapeutic costs. HIV-1 integrase (IN) and reverse transcriptase-associated ribonuclease H (RNase H) are both selective targets for HIV-1 chemotherapy, and the identification of dual IN/RNase H inhibitors is an attractive strategy for new drug development. We newly synthesized pyrrolyl derivatives that exhibited good potency against IN and a moderate inhibition of the RNase H function of RT, confirming the possibility of developing dual HIV-1 IN/RNase H inhibitors and obtaining new information for the further development of more effective dual HIV-1 inhibitors.

  1. Equilibrium crossing exhibited by an ethynylhelicene (M)-nonamer during random-coil-to-double-helix thermal transition in solution.

    Science.gov (United States)

    Miyagawa, Masamichi; Yagi, Atsushi; Shigeno, Masanori; Yamaguchi, Masahiko

    2014-11-28

    The structural change between the random-coil and the double-helix of an ethynylhelicene (M)-nonamer during heating crosses equilibrium. This is a phenomenon where a chemical reaction crosses equilibrium and returns to equilibrium. It is due to an accelerated rate of formation of the double-helix by self-catalysis and an equilibrium shift.

  2. Rotational symmetry and the transformation of innovation systems in a Triple Helix of university-industry-government relations

    NARCIS (Netherlands)

    Ivanova, I.A.; Leydesdorff, L.

    2014-01-01

    Using a mathematical model, we show that a Triple Helix (TH) system contains self-interaction, and therefore self-organization of innovations can be expected in waves, whereas a Double Helix (DH) remains determined by its linear constituents. (The mathematical model is fully elaborated in the

  3. Equilibrium shift in solution: molecular shape recognition and precipitation of a synthetic double helix using helicene-grafted silica nanoparticles.

    Science.gov (United States)

    Miyagawa, Masamichi; Ichinose, Wataru; Yamaguchi, Masahiko

    2014-01-27

    Chiral silica nanoparticles (70 nm) grafted with (P)-helicene recognized the molecular shape of double helix and random coil (P)-ethynylhelicene oligomers in solution. A mixture of the (P)-nanoparticles and double helix precipitated much faster than a mixture of the (P)-nanoparticles and random coil, and the precipitate contained only the double helix. The mixture of the (P)-nanoparticles and (P)-ethynylhelicene pentamer reversibly dispersed in trifluoromethylbenzene upon heating at 70 °C and precipitated upon cooling at 25 °C. When a 10:90 equilibrium mixture of the double helix and random coil in solution was treated with the (P)-nanoparticles, the double helix was precipitated in 53% yield and was accompanied by equilibrium shift. Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  4. Restrictions on TWT Helix Voltage Ripple for Acceptable Notch Filter Performance

    Energy Technology Data Exchange (ETDEWEB)

    Hyslop, B.

    1984-12-01

    An ac ripple on the helix voltage of the 1-2 GHz TWT's creates FM sidebands that cause amplitude and phase modulation of the microwave TWT output signal. A limit of 16 volts peak-to-peak is required for acceptable superconducting notch filter performance.

  5. Estudio Histoquímico y ultraestructural del sistema nervioso de helix aspersa

    OpenAIRE

    Coveñas Rodríguez, Rafael

    1982-01-01

    El objetivo de nuestro trabajo, ha sido estudiar el sistema nervioso central de Helix aspersa a nivel ultraestructural e histoquímico. Bajo microscopía electrónica describiremos las observaciones realizadas en la cápsula que envuelve a los elementos nervi

  6. Evidence for genetic control of adult weight plasticity in the snail Helix aspersa

    DEFF Research Database (Denmark)

    Ros, Mathieu; Sorensen, Daniel; Waagepetersen, Rasmus Plenge

    2004-01-01

    of adult weight in the snail Helix aspersa. Several models of heterogeneous variance are fitted using a Bayesin, MCMC approach. Exploratory analyses using posterior predictive model checking and model comparisons based on the deviance information criterion favor a model postulating a genetically structured...... is illustrated numerically using estimates of parameters derived from the snail data set....

  7. Alpha-helix <-> random coil phase transition: analysis of ab initio theory predictions

    DEFF Research Database (Denmark)

    Solov'yov, Ilia; Yakubovich, Alexander V.; Solov'yov, Andrey V.

    2008-01-01

    In the present paper we present results of calculations obtained with the use of the theoretical method described in our preceding paper [Eur. Phys. J. D, DOI 10.1140/epjd/e2007-00328-9] and perform detail analysis of -helix random coil transition in alanine polypeptides of different length. We...

  8. Extension of Helix 12 in Munc18-1 Induces Vesicle Priming

    DEFF Research Database (Denmark)

    Munch, Anders S; Kedar, Girish H; van Weering, Jan R T

    2016-01-01

    Munc18-1 is essential for vesicle fusion and participates in the docking of large dense-core vesicles to the plasma membrane. Recent structural data suggest that conformational changes in the 12th helix of the Munc18-1 domain 3a within the Munc18-1:syntaxin complex result in an additional interac...

  9. A triple Helix Approach to the Future Innovation Flagship of Europe:

    DEFF Research Database (Denmark)

    Jofre, Sergio; Andersen, Per Dannemand

    2009-01-01

    , are transforming the profile of the triple helix relationships. This transformation is bringing the American and the Japanese innovation system to an unprecedented level of commonality and the EU to a yet uncertain stage of transition characterized by the conflict between national and supranational priorities...

  10. Open Innovation, Triple Helix and Regional Innovation Systems: Exploring CATAPULT Centres in the UK

    Science.gov (United States)

    Kerry, Christopher; Danson, Michael

    2016-01-01

    Through the lens of UK CATAPULT Centres this conceptual paper presents an examination of the links between open innovation, the Triple Helix model and regional innovation systems. Highlighting the importance of boundary-spanning intermediaries, the combined role of these concepts is explored in detail. A conceptual model is then proposed which…

  11. Interference of Helix pomatia extracts on the determination of methandriol in veterinary residue control analysis

    OpenAIRE

    LEYSSENS, Luc; VAN PUYMBROECK, Mark; RAUS, Jef

    1998-01-01

    Aqueous solutions containing methandriol (MAD) were incubated with Succus helix pomatia (SHP) or beta-glucuronidase from Escherichia coli (EC). SHP, used for enzymatic hydrolysis of urinary steroid conjugates for residue analysis of anabolic agents, caused transformation of MAD into methyltestosterone. No conversion occurred when bacterial beta-glucuronidase from E. coli was used.

  12. A note on the pollen representation of ivy (Hedera helix L.)

    NARCIS (Netherlands)

    Bottema, S

    2001-01-01

    Pollen productivity and dispersal of ivy (Hedera helix) is discussed on the basis of a simple surface sample study in a situation that has been followed for about 30 yr. The pollen production of a special ivy plant was estimated for the flowering season of 1998. Hedera pollen dispersal was followed

  13. Triple helix networks matching knowledge demand and supply in seven Dutch horticulture Greenport regions

    NARCIS (Netherlands)

    Geerling-Eiff, Florentien A.; Hoes, Anne-Charlotte; Dijkshoorn-Dekker, Marijke

    2017-01-01

    This paper investigates the triple helix (industry, knowledge workers and governments) cooperation on knowledge co-production and valorisation for innovation, which took place in seven horticultural regions in the Netherlands. It thus provides more empirical insight into the functioning of this form

  14. Microsecond and nanosecond polyproline II helix formation in aqueous nanodrops measured by mass spectrometry.

    Science.gov (United States)

    Mortensen, Daniel N; Williams, Evan R

    2016-10-06

    The 1.5 μs and <400 ns time constants for the formation of polyproline II helix structures in 21 and 16 residue peptides, respectively, are measured using rapid mixing from theta-glass emitters coupled with mass spectrometry. Results from these studies should serve as useful benchmarks for comparison with computational simulation results.

  15. Enhancing the innovative capacity of small firms through triple helix interactions : challenges and opportunities

    NARCIS (Netherlands)

    Ranga, Liana Marina; Miedema, Joost; Jorna, Rene

    2008-01-01

    This paper presents the results of a recent exploratory study aiming to enhance the innovative capacity of small firms in the Northern Netherlands, a region lagging behind the rest of the country in terms of economic growth and innovative capacity. The triple helix perspective is adopted to examine

  16. Influence of season, temperature, and photoperiod on growth of the land snail Helix aperta

    NARCIS (Netherlands)

    Benbellil-Tafoughalt, S.; Koene, J.M.

    2015-01-01

    Growth strategies are often plastic and influenced by environmental conditions. Terrestrial gastropods are particularly affected by seasonal and climatic variables, and growth rate and size at maturity are key traits in their life history. Therefore, we investigated juvenile growth of Helix aperta

  17. Glycine Perturbs Local and Global Conformational Flexibility of a Transmembrane Helix

    DEFF Research Database (Denmark)

    Högel, Philipp; Götz, Alexander; Kuhne, Felix

    2018-01-01

    the measured exchange kinetics and reveal, at atomic resolution, a severe packing defect at glycine that enhances local hydration. Furthermore, glycine alters H-bond occupancies and triggers a redistribution of α-helical and 310-helical H-bonds. These effects facilitate local helix bending at the glycine site...

  18. pH-jump induced α-helix folding of poly-L-glutamic acid

    Energy Technology Data Exchange (ETDEWEB)

    Donten, Mateusz L. [Institute of Physical Chemistry, University of Zurich, Winterthurerstrasse 190, CH-8057 Zürich (Switzerland); Hamm, Peter, E-mail: phamm@pci.uzh.ch [Institute of Physical Chemistry, University of Zurich, Winterthurerstrasse 190, CH-8057 Zürich (Switzerland)

    2013-08-30

    Highlights: ► pH-jump as truly biomimetic tool to initiate non-equilibrium dynamics of biomolecules. ► Design criteria to widen the applicability of pH-jumps are developed. ► Folding of poly-L-Glu in dependence of starting pH, pH jump size and helix length. ► Length dependence provides strong evidence for a nucleation–propagation scenario. - Abstract: pH jumps are a truly biomimetic technique to initiate non-equilibrium dynamics of biomolecules. In this work, the pH jump induced α-helix folding of poly-L-glutamic acid is investigated upon proton release from o-nitrobenzaldehyde. The aim of this work is twofold: On the one hand, design criteria of pH jump experiments are discussed, on the other hand, the folding mechanism of poly-L-glutamic acid is clarified by probing the IR response of the amide I band. Its folding kinetics is studied in dependence of the starting pD, the size of the pD jump and the length of the helix. While no dependence on the first two parameters could be detected, the folding time varies from 0.6 μs to 1.8 μs for helix lengths of 20 residue to 440 residue, respectively. It converges to a long-length limit at about 50 residue, a result which is attributed to a nucleation–propagation mechanism.

  19. Large deformation of helix F during the photoreaction cycle of Pharaonis halorhodopsin in complex with azide.

    Science.gov (United States)

    Nakanishi, Taichi; Kanada, Soun; Murakami, Midori; Ihara, Kunio; Kouyama, Tsutomu

    2013-01-22

    Halorhodopsin from Natronomonas pharaonis (pHR), a retinylidene protein that functions as a light-driven chloride ion pump, is converted into a proton pump in the presence of azide ion. To clarify this conversion, we investigated light-induced structural changes in pHR using a C2 crystal that was prepared in the presence of Cl(-) and subsequently soaked in a solution containing azide ion. When the pHR-azide complex was illuminated at pH 9, a profound outward movement (∼4 Å) of the cytoplasmic half of helix F was observed in a subunit with the EF loop facing an open space. This movement created a long water channel between the retinal Schiff base and the cytoplasmic surface, along which a proton could be transported. Meanwhile, the middle moiety of helix C moved inward, leading to shrinkage of the primary anion-binding site (site I), and the azide molecule in site I was expelled out to the extracellular medium. The results suggest that the cytoplasmic half of helix F and the middle moiety of helix C act as different types of valves for active proton transport. Copyright © 2013 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  20. Efficient Fatigue Analysis of Helix Elements in Umbilicals and Flexible Risers: Theory and Applications

    Directory of Open Access Journals (Sweden)

    Geir Skeie

    2012-01-01

    Full Text Available Fatigue analysis of structural components such as helix tensile armors and steel tubes is a critical design issue for dynamic umbilicals and flexible pipes. The basis for assessment of fatigue damage of such elements is the long-term stress cycle distribution at critical locations on the helix elements caused by long-term environmental loading on the system. The long-term stress cycle distribution will hence require global dynamic time domain analysis followed by a detailed cross-sectional analysis in a large number of irregular sea states. An overall computational consistent and efficient fatigue analysis scheme is outlined with due regard of the cross-sectional analysis technique required for fatigue stress calculation with particular attention to the helix elements. The global cross-section is exposed to pure bending, tensile, torsion, and pressure loading. The state of the different cross-section elements is based on the global response. Special emphasis is placed on assessment of friction stresses caused by the stick-slip behavior of helix elements in bending that are of special importance for fatigue life assessments. The described cross-sectional analysis techniques are based on an extensive literature survey and are hence considered to represent industry consensus. The performance of the described calculation scheme is illustrated by case studies.

  1. The triple-helix model of smart cities: a neo-evolutionary perspective

    NARCIS (Netherlands)

    Leydesdorff, L.; Deakin, M.

    2011-01-01

    This paper sets out to demonstrate how the triple-helix model enables us to study the knowledge base of an urban economy in terms of its civil society's support for the evolution of the city as a key component of an innovation system. It argues that cities can be considered as densities in networks

  2. Democracy and environment as references for quadruple and quintuple helix innovation systems

    Science.gov (United States)

    Carayannis, Elias G.; Campbell, David F. J.; Orr, Barron J.

    2015-04-01

    The perspective of democracy and the ecological context define key references for knowledge production and innovation in innovation systems. Particularly under conditions of environmental change where enhancing the potential for adaptation is critical, this requires a closer look at ecological responsibility and sensitivity in the different innovation models and governance regimes. The "Quintuple Helix" innovation model is an approach that stresses the necessary socio-ecological transition of society and economy by adding an environment helix to an innovation system already made up of three (university-industry-government) or four (civil society relations) helices in a way that supports adaptation by incorporating global warming as both a challenge to and a driver of innovation. There is the proposition that knowledge production and innovation co-evolve with democracy (Carayannis and Campbell, 2014). In the Triple Helix model (Etzkowitz and Leydesdorff, 2000) the existence of a democracy does not appear to be necessary for knowledge production and innovation. However, the Quadruple Helix (Carayannis and Campbell, 2009, 2010 and 2014) is defined and represented by additional key attributes and components: "media-based and culture-based public", "civil society" and "arts, artistic research and arts-based innovation" (Bast, Carayannis and Campbell, 2015). Implications of this are that the fourth helix in the Quadruple Helix innovation systems brings in and represents the perspective of "dimension of democracy" or the "context of democracy" for knowledge in general and knowledge production and innovation in more particular. Within theories of democracy there is a competition between narrow and broader concepts of democracy (Campbell, 2013). This is particularly true when democracy is to be understood to transcend more substantially the narrow understanding of being primarily based on or being primarily rooted in government institutions (within a Triple Helix

  3. From Family Based to Industrial Based Production: Local Economic Development Initiatives and the HELIX Model

    Directory of Open Access Journals (Sweden)

    Bartjan W Pennink

    2013-01-01

    Full Text Available To build a strong local economy, good practice tells us that each community should undertake a collaborative, strategically planned process to understand and then act upon its own strengths, weaknesses, opportunities and threats. From this perspective we start with the local communities but how is this related to the perspective from the Helix model in which three actors are explicitly introduced: the Government, the Industry and the Universities? The purpose of local economic development (LED is to build up the economic capacity of a local area to improve its economic future and the quality of life for all. To support  the Local Economic Development in remote areas,   a program  has been developed based on the LED frame work of the world bank. This approach and  the experiences over  the past years with this program are  described in the first part.  In the second part of the paper, We analyse work done with that program with the help of the social capital concept and the triple helix model.  In all cases it is important to pay attention to who is taken the initiative after the first move (and it is not always the governance as actor and for the triple helix we suggest  that the concepts of (national Government, Industry and University need a translation to Local Governance Agency, Cooperation or other ways of cooperation of local communities and Local Universities. Although a push from outside might help  a local region in development the endogenous factors are  also needed. Keywords: Triple Helix model, Local Economic Development, Local Actors, Double Triangle within the Helix Model

  4. Study of Structure-active Relationship for Inhibitors of HIV-1 Integrase LEDGF/p75 Interaction by Machine Learning Methods.

    Science.gov (United States)

    Li, Yang; Wu, Yanbin; Yan, Aixia

    2017-07-01

    HIV-1 integrase (IN) is a promising target for anti-AIDS therapy, and LEDGF/p75 is proved to enhance the HIV-1 integrase strand transfer activity in vitro. Blocking the interaction between IN and LEDGF/p75 is an effective way to inhibit HIV replication infection. In this work, 274 LEDGF/p75-IN inhibitors were collected as the dataset. Support Vector Machine (SVM), Decision Tree (DT), Function Tree (FT) and Random Forest (RF) were applied to build several computational models for predicting whether a compound is an active or weakly active LEDGF/p75-IN inhibitor. Each compound is represented by MACCS fingerprints and CORINA Symphony descriptors. The prediction accuracies for the test sets of all the models are over 70 %. The best model Model 3B built by FT obtained a prediction accuracy and a Matthews Correlation Coefficient (MCC) of 81.08 % and 0.62 on test set, respectively. We found that the hydrogen bond and hydrophobic interactions are important for the bioactivity of an inhibitor. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. The HIV-1 integrase mutations Y143C/R are an alternative pathway for resistance to Raltegravir and impact the enzyme functions.

    Directory of Open Access Journals (Sweden)

    Sandrine Reigadas

    Full Text Available Resistance to HIV-1 integrase (IN inhibitor raltegravir (RAL, is encoded by mutations in the IN region of the pol gene. The emergence of the N155H mutation was replaced by a pattern including the Y143R/C/H mutations in three patients with anti-HIV treatment failure. Cloning analysis of the IN gene showed an independent selection of the mutations at loci 155 and 143. Characterization of the phenotypic evolution showed that the switch from N155H to Y143C/R was linked to an increase in resistance to RAL. Wild-type (WT IN and IN with mutations Y143C or Y143R were assayed in vitro in 3'end-processing, strand transfer and concerted integration assays. Activities of mutants were moderately impaired for 3'end-processing and severely affected for strand transfer. Concerted integration assay demonstrated a decrease in mutant activities using an uncleaved substrate. With 3'end-processing assay, IC(50 were 0.4 microM, 0.9 microM (FC = 2.25 and 1.2 microM (FC = 3 for WT, IN Y143C and IN Y143R, respectively. An FC of 2 was observed only for IN Y143R in the strand transfer assay. In concerted integration, integrases were less sensitive to RAL than in ST or 3'P but mutants were more resistant to RAL than WT.

  6. Mode of inhibition of HIV-1 Integrase by a C-terminal domain-specific monoclonal antibody*

    Directory of Open Access Journals (Sweden)

    Merkel George

    2006-06-01

    Full Text Available Abstract Background To further our understanding of the structure and function of HIV-1 integrase (IN we developed and characterized a library of monoclonal antibodies (mAbs directed against this protein. One of these antibodies, mAb33, which is specific for the C-terminal domain, was found to inhibit HIV-1 IN processing activity in vitro; a corresponding Fv fragment was able to inhibit HIV-1 integration in vivo. Our subsequent studies, using heteronuclear nuclear magnetic resonance spectroscopy, identified six solvent accessible residues on the surface of the C-terminal domain that were immobilized upon binding of the antibody, which were proposed to comprise the epitope. Here we test this hypothesis by measuring the affinity of mAb33 to HIV-1 proteins that contain Ala substitutions in each of these positions. To gain additional insight into the mode of inhibition we also measured the DNA binding capacity and enzymatic activities of the Ala substituted proteins. Results We found that Ala substitution of any one of five of the putative epitope residues, F223, R224, Y226, I267, and I268, caused a decrease in the affinity of the mAb33 for HIV-1 IN, confirming the prediction from NMR data. Although IN derivatives with Ala substitutions in or near the mAb33 epitope exhibited decreased enzymatic activity, none of the epitope substitutions compromised DNA binding to full length HIV-1 IN, as measured by surface plasmon resonance spectroscopy. Two of these derivatives, IN (I276A and IN (I267A/I268A, exhibited both increased DNA binding affinity and uncharacteristic dissociation kinetics; these proteins also exhibited non-specific nuclease activity. Results from these investigations are discussed in the context of current models for how the C-terminal domain interacts with substrate DNA. Conclusion It is unlikely that inhibition of HIV-1 IN activity by mAb33 is caused by direct interaction with residues that are essential for substrate binding. Rather

  7. Targeted transgene insertion into the CHO cell genome using Cre recombinase-incorporating integrase-defective retroviral vectors.

    Science.gov (United States)

    Kawabe, Yoshinori; Shimomura, Takuya; Huang, Shuohao; Imanishi, Suguru; Ito, Akira; Kamihira, Masamichi

    2016-07-01

    Retroviral vectors have served as efficient gene delivery tools in various biotechnology fields. However, viral DNA is randomly inserted into the genome, which can cause problems, such as insertional mutagenesis and gene silencing. Previously, we reported a site-specific gene integration system, in which a transgene is integrated into a predetermined chromosomal locus of Chinese hamster ovary (CHO) cells using integrase-defective retroviral vectors (IDRVs) and Cre recombinase. In this system, a Cre expression plasmid is transfected into founder cells before retroviral transduction. In practical applications of site-specific gene modification such as for hard-to-transfect cells or for in vivo gene delivery, both the transgene and the Cre protein into retroviral virions should be encapsulate. Here, we generated novel hybrid IDRVs in which viral genome and enzymatically active Cre can be delivered (Cre-IDRVs). Cre-IDRVs encoding marker genes, neomycin resistance and enhanced green fluorescent protein (EGFP), flanked by wild-type and mutated loxP sites were produced using an expression plasmid for a chimeric protein of Cre and retroviral gag-pol. After analyzing the incorporation of the Cre protein into retroviral virions by Western blotting, the Cre-IDRV was infected into founder CHO cells, in which marker genes (hygromycin resistance and red fluorescent protein) flanked with corresponding loxP sites are introduced into the genome. G418-resistant colonies expressing GFP appeared and the site-specific integration of the transgene into the expected chromosomal site was confirmed by PCR and sequencing of amplicons. Moreover, when Cre-IDRV carried a gene expression unit for a recombinant antibody, the recombinant cells in which the antibody expression cassette was integrated in a site-specific manner were generated and the cells produced the recombinant antibody. This method may provide a promising tool to perform site-specific gene modification according to Cre

  8. Development of elvitegravir resistance and linkage of integrase inhibitor mutations with protease and reverse transcriptase resistance mutations.

    Directory of Open Access Journals (Sweden)

    Mark A Winters

    Full Text Available Failure of antiretroviral regimens containing elvitegravir (EVG and raltegravir (RAL can result in the appearance of integrase inhibitor (INI drug-resistance mutations (DRMs. While several INI DRMs have been identified, the evolution of EVG DRMs and the linkage of these DRMs with protease inhibitor (PI and reverse transcriptase inhibitor (RTI DRMs have not been studied at the clonal level. We examined the development of INI DRMs in 10 patients failing EVG-containing regimens over time, and the linkage of INI DRMs with PI and RTI DRMs in these patients plus 6 RAL-treated patients. A one-step RT-nested PCR protocol was used to generate a 2.7 kB amplicon that included the PR, RT, and IN coding region, and standard cloning and sequencing techniques were used to determine DRMs in 1,277 clones (mean 21 clones per time point. Results showed all patients had multiple PI, NRTI, and/or NNRTI DRMs at baseline, but no primary INI DRM. EVG-treated patients developed from 2 to 6 strains with different primary INI DRMs as early as 2 weeks after initiation of treatment, predominantly as single mutations. The prevalence of these strains fluctuated and new strains, and/or strains with new combinations of INI DRMs, developed over time. Final failure samples (weeks 14 to 48 typically showed a dominant strain with multiple mutations or N155H alone. Single N155H or multiple mutations were also observed in RAL-treated patients at virologic failure. All patient strains showed evidence of INI DRM co-located with single or multiple PI and/or RTI DRMs on the same viral strand. Our study shows that EVG treatment can select for a number of distinct INI-resistant strains whose prevalence fluctuates over time. Continued appearance of new INI DRMs after initial INI failure suggests a potent, highly dynamic selection of INI resistant strains that is unaffected by co-location with PI and RTI DRMs.

  9. Reovirus FAST Proteins Drive Pore Formation and Syncytiogenesis Using a Novel Helix-Loop-Helix Fusion-Inducing Lipid Packing Sensor

    Science.gov (United States)

    Sarker, Muzaddid; de Antueno, Roberto; Langelaan, David N.; Parmar, Hiren B.; Shin, Kyungsoo; Rainey, Jan K.; Duncan, Roy

    2015-01-01

    Pore formation is the most energy-demanding step during virus-induced membrane fusion, where high curvature of the fusion pore rim increases the spacing between lipid headgroups, exposing the hydrophobic interior of the membrane to water. How protein fusogens breach this thermodynamic barrier to pore formation is unclear. We identified a novel fusion-inducing lipid packing sensor (FLiPS) in the cytosolic endodomain of the baboon reovirus p15 fusion-associated small transmembrane (FAST) protein that is essential for pore formation during cell-cell fusion and syncytiogenesis. NMR spectroscopy and mutational studies indicate the dependence of this FLiPS on a hydrophobic helix-loop-helix structure. Biochemical and biophysical assays reveal the p15 FLiPS preferentially partitions into membranes with high positive curvature, and this partitioning is impeded by bis-ANS, a small molecule that inserts into hydrophobic defects in membranes. Most notably, the p15 FLiPS can be functionally replaced by heterologous amphipathic lipid packing sensors (ALPS) but not by other membrane-interactive amphipathic helices. Furthermore, a previously unrecognized amphipathic helix in the cytosolic domain of the reptilian reovirus p14 FAST protein can functionally replace the p15 FLiPS, and is itself replaceable by a heterologous ALPS motif. Anchored near the cytoplasmic leaflet by the FAST protein transmembrane domain, the FLiPS is perfectly positioned to insert into hydrophobic defects that begin to appear in the highly curved rim of nascent fusion pores, thereby lowering the energy barrier to stable pore formation. PMID:26061049

  10. Reovirus FAST Proteins Drive Pore Formation and Syncytiogenesis Using a Novel Helix-Loop-Helix Fusion-Inducing Lipid Packing Sensor.

    Directory of Open Access Journals (Sweden)

    Jolene Read

    2015-06-01

    Full Text Available Pore formation is the most energy-demanding step during virus-induced membrane fusion, where high curvature of the fusion pore rim increases the spacing between lipid headgroups, exposing the hydrophobic interior of the membrane to water. How protein fusogens breach this thermodynamic barrier to pore formation is unclear. We identified a novel fusion-inducing lipid packing sensor (FLiPS in the cytosolic endodomain of the baboon reovirus p15 fusion-associated small transmembrane (FAST protein that is essential for pore formation during cell-cell fusion and syncytiogenesis. NMR spectroscopy and mutational studies indicate the dependence of this FLiPS on a hydrophobic helix-loop-helix structure. Biochemical and biophysical assays reveal the p15 FLiPS preferentially partitions into membranes with high positive curvature, and this partitioning is impeded by bis-ANS, a small molecule that inserts into hydrophobic defects in membranes. Most notably, the p15 FLiPS can be functionally replaced by heterologous amphipathic lipid packing sensors (ALPS but not by other membrane-interactive amphipathic helices. Furthermore, a previously unrecognized amphipathic helix in the cytosolic domain of the reptilian reovirus p14 FAST protein can functionally replace the p15 FLiPS, and is itself replaceable by a heterologous ALPS motif. Anchored near the cytoplasmic leaflet by the FAST protein transmembrane domain, the FLiPS is perfectly positioned to insert into hydrophobic defects that begin to appear in the highly curved rim of nascent fusion pores, thereby lowering the energy barrier to stable pore formation.

  11. Docking and Molecular Dynamics Calculations of Some Previously Studied and newly Designed Ligands to Catalytic Core Domain of HIV-1 Integrase and an Investigation to Effects of Conformational Changes of Protein on Docking Results

    Directory of Open Access Journals (Sweden)

    Selami Ercan

    2016-10-01

    Full Text Available Nowadays, AIDS still remains as a worldwide pandemic and continues to cause many death which arise from HIV-1 virus. For nearly 35 years, drugs that target various steps of virus life cycle have been developed. HIV-1 integrase is the one of these steps which is essential for virus life cycle. Computer aided drug design is being used in many drug design studies as also used in development of the first HIV-1 integrase inhibitor Raltegravir. In this study 3 ligands which are used as HIV-1 integrase inhibitors and 4 newly designed ligands were docked to catalytic core domain of HIV-1 integrase. Each of ligands docked to three different conformations of protein. Prepared complexes (21 item were carried out by 50 ns MD simulations and results were analyzed. Finally, the binding free energies of ligands were calculated. Hereunder, it was determined that designed ligands L01 and L03 gave favorable results. The questions about the ligands which have low docking scores in a conformation of protein could give better scores in another conformation of protein and if the MD simulations carry the different oriented and different localized ligands in same position at the end of simulation were answered.

  12. Trace metallic elements in Helix aspersa terrestrial snails of a semiarid ecosystem; Elementos metalicos traza en caracoles terrestres Helix aspersa de un ecosistema semiarido

    Energy Technology Data Exchange (ETDEWEB)

    Gaso P, M.I.; Segovia, N.; Zarazua, G.; Montes, F. [Instituto Nacional de Investigaciones Nucleares, A.P. 18-1027, 11801 Mexico D.F. (Mexico); Morton, O.; Armienta, M.A.; Hernandez, E. [IGF-UNAM, Ciudad Universitaria, 04510 Mexico D.F. (Mexico)

    2001-07-01

    The concentration of some major elements and traces in soil samples and of Helix aspersa eatable terrestrial snails were analysed at the Radioactive Wastes Storage Center (CADER) and in other reference sites. The methodology includes the use of an atomic absorption spectrophotometer, an X-ray fluorescence equipment and an Icp-mass spectroscope. The concentrations of some toxic elements (Ba, Cd, Cr, Ni, Pb and V) in the soft tissue of the snails were greater than the toxic levels reported in the literature for such trace elements. The snails compared with another wild eatable foods present transfer coefficients soil-snail high relatively. (Author)

  13. Tunable single photonic defect-mode in cholesteric liquid crystals with laser-induced local modifications of helix

    International Nuclear Information System (INIS)

    Yoshida, Hiroyuki; Lee, Chee Heng; Fujii, Akihiko; Ozaki, Masanori

    2006-01-01

    The authors demonstrate a tunable single photonic defect-mode in a single cholesteric liquid crystal material based on a structural defect introduced by local modification of the helix. An unpolymerized region of cholesteric liquid crystal acting as the defect was left between two polymerized regions via a two-photon excitation laser-lithography process. Upon polymerization, the cholesteric liquid crystal helix elongated and became thermally stable, and a single photonic defect mode was exhibited due to the contrast in the helix pitch at the defect. The defect mode showed tunability upon heating, and a 36 nm redshift was seen over a temperature range of 30 deg. C

  14. Analysis of eco-innovation with triple helix approach: case-study of biofloc catfish farming in Yogyakarta

    Science.gov (United States)

    Purwadi, D.; Nurlaily, I.

    2018-03-01

    Concerning environmental into focus of innovation process will expand the number of actor involved. Eco-innovation and triple helix are often frameworks applied to analyse how environmental concern are integrated in innovation process and how different stakeholder groups are having inter relation. Case study from biofloc catfish farming in Yogyakarta is presented to demonstrate a possible approach for researching the success of triple helix frameworks. This case is considered on basic of the result of a survey among farmers, academician and government. The paper concludes the creating of full triple helix encounters problem in practice. It also includes suggestion for further research on fisheries development.

  15. RelB and RelE of Escherichia coli Form a Tight Complex That Represses Transcription via The Ribbon-Helix-Helix Motif in RelB

    DEFF Research Database (Denmark)

    Overgaard, Martin; Borch, Jonas; Gerdes, Kenn

    2009-01-01

    RelB, the Ribbon-Helix-Helix (RHH) repressor encoded by the relBE toxin-antitoxin locus of Escherichia coli, forms a tight complex with RelE and thereby counteracts the mRNA cleavage activity of RelE. In addition, RelB dimers repress the strong relBE promoter and this repression by RelB is enhanced...... by RelE - that is - RelE functions as a transcriptional co-repressor. RelB is a Lon protease substrate and Lon is required both for activation of relBE transcription and for activation of the mRNA cleavage activity of RelE. Here we characterize the molecular interactions important for transcriptional...... motif recognizes four 6 bp repeats within the bipartite binding site. The spacing between each half-site was found to be essential for cooperative interactions between adjacently bound RelB dimers stabilized by the co-repressor RelE. Kinetic and stoichiometric measurements of the interaction between Rel...

  16. Agrobacterium tumefaciens VirC2 enhances T-DNA transfer and virulence through its C-terminal ribbon-helix-helix DNA-binding fold.

    Science.gov (United States)

    Lu, Jun; den Dulk-Ras, Amke; Hooykaas, Paul J J; Glover, J N Mark

    2009-06-16

    Agrobacterium tumefaciens VirC2 stimulates processing of single-stranded T-DNA that is translocated into plants to induce tumor formation, but how VirC2 functions is unclear. Here, we report the 1.7-A X-ray crystal structure of its trypsin-resistant C-terminal domain, VirC2(82-202), which reveals a form of the ribbon-helix-helix (RHH) DNA-binding fold contained within a single polypeptide chain. DNA-binding assays and mutagenesis indicate that VirC2 uses this RHH fold to bind double-stranded DNA but not single-stranded DNA. Mutations that severely affect VirC2 DNA binding are highly deleterious for both T-DNA transfer into yeast and the virulence of A. tumefaciens in different plants including Nicotiana glauca and Kalanchoe daigremontiana. These data suggest that VirC2 enhances T-DNA transfer and virulence through DNA binding with its RHH fold. The RHH fold of VirC2 is the first crystal structure representing a group of predicted RHH proteins that facilitate endonucleolytic processing of DNA for horizontal gene transfer.

  17. Sequence and function of basic helix-loop-helix proteins required for stomatal development in Arabidopsis are deeply conserved in land plants.

    Science.gov (United States)

    MacAlister, Cora A; Bergmann, Dominique C

    2011-01-01

    Stomata are a broadly conserved feature of land plants with a crucial role regulating transpiration and gas exchange between the plant and atmosphere. Stereotyped cell divisions within a specialized cell lineage of the epidermis generate stomata and define the pattern of their distribution. The behavior of the stomatal lineage varies in its detail among different plant groups, but general features include asymmetric cell divisions and an immediate precursor (the guard mother cell [GMC]) that divides symmetrically to form the pair of cells that will differentiate into the guard cells. In Arabidopsis, the closely related basic helix-loop-helix (bHLH) subgroup Ia transcription factors SPEECHLESS, MUTE, and FAMA promote asymmetric divisions, the acquisition of GMC identity and guard cell differentiation, respectively. Genome sequence data indicate that these key positive regulators of stomatal development are broadly conserved among land plants. While orthologies can be established among individual family members within the angiosperms, more distantly related groups contain subgroup Ia bHLHs of unclear affinity. We demonstrate group Ia members from the moss Physcomitrella patens can partially complement MUTE and FAMA and recapitulate gain of function phenotypes of group Ia genes in multiple steps in the stomatal lineage in Arabidopsis. Our data are consistent with a mechanism whereby a multifunctional transcription factor underwent duplication followed by specialization to provide the three (now nonoverlapping) functions of the angiosperm stomatal bHLHs. © 2011 Wiley Periodicals, Inc.

  18. An unusual helix-turn-helix protease inhibitory motif in a novel trypsin inhibitor from seeds of Veronica (Veronica hederifolia L.).

    Science.gov (United States)

    Conners, Rebecca; Konarev, Alexander V; Forsyth, Jane; Lovegrove, Alison; Marsh, Justin; Joseph-Horne, Timothy; Shewry, Peter; Brady, R Leo

    2007-09-21

    The storage tissues of many plants contain protease inhibitors that are believed to play an important role in defending the plant from invasion by pests and pathogens. These proteinaceous inhibitor molecules belong to a number of structurally distinct families. We describe here the isolation, purification, initial inhibitory properties, and three-dimensional structure of a novel trypsin inhibitor from seeds of Veronica hederifolia (VhTI). The VhTI peptide inhibits trypsin with a submicromolar apparent K(i) and is expected to be specific for trypsin-like serine proteases. VhTI differs dramatically in structure from all previously described families of trypsin inhibitors, consisting of a helix-turn-helix motif, with the two alpha helices tightly associated by two disulfide bonds. Unusually, the crystallized complex is in the form of a stabilized acyl-enzyme intermediate with the scissile bond of the VhTI inhibitor cleaved and the resulting N-terminal portion of the inhibitor remaining attached to the trypsin catalytic serine 195 by an ester bond. A synthetic, truncated version of the VhTI peptide has also been produced and co-crystallized with trypsin but, surprisingly, is seen to be uncleaved and consequently forms a noncovalent complex with trypsin. The VhTI peptide shows that effective enzyme inhibitors can be constructed from simple helical motifs and provides a new scaffold on which to base the design of novel serine protease inhibitors.

  19. Two basic-helix-loop-helix genes (MYC-146 and GL3) from Arabidopsis can activate anthocyanin biosynthesis in a white-flowered Matthiola incana mutant.

    Science.gov (United States)

    Ramsay, Nicola A; Walker, Amanda R; Mooney, Mark; Gray, John C

    2003-06-01

    Basic helix-loop-helix (bHLH) proteins, similar to mammalian Myc transcription factors, regulate the anthocyanin biosynthetic pathway in both monocots and dicots. Two Arabidopsis bHLH genes, GLABRA3 (GL3) and MYC-146, encode proteins that are similar throughout the predicted amino acid sequence to R and DELILA, which regulate anthocyanin production in maize and snapdragon, respectively. Northern blot analysis indicates that MYC-146 is most highly expressed in flower buds and flowers. Expression of a MYC-146 cDNA from the CaMV 35S promoter was unable to complement the anthocyanin deficiency in a ttg1 mutant of Arabidopsis and resulted in no obvious phenotypic change in Columbia plants. However, transient expression of GL3 and MYC-146 upon microprojectile bombardment of petals of a white-flowered mutant of Matthiola incana was able to complement anthocyanin deficiency. The lack of anthocyanin-deficient Arabidopsis mutants mapping to the locations of GL3 and MYC-146 suggests that the two bHLH proteins may be partially redundant and overlap in function.

  20. Alternative generation of well-aligned uniform lying helix texture in a cholesteric liquid crystal cell

    Directory of Open Access Journals (Sweden)

    Chia-Hua Yu

    2017-10-01

    Full Text Available This work demonstrates a simple approach for obtaining a well-aligned uniform lying helix (ULH texture and a tri-bistable feature at ambient temperature in a typical 90°-twisted cell filled with a short-pitch cholesteric liquid crystal. This ULH texture is obtained at room temperature from initially field-induced helix-free homeotropic state by gradually decreasing the applied voltage. Depending on the way and rate of reducing the voltage, three stable states (i.e., Grandjean planar, focal conic, and ULH are generated and switching between any two of them is realized. Moreover, the electrical operation of the cell in the ULH state enables the tunability in phase retardation via the deformation of the ULH. The observations made in this work may be useful for applications such as tunable phase modulators and energy-efficient photonic devices.

  1. Helix Nebula and CERN: A Symbiotic approach to exploiting commercial clouds

    CERN Document Server

    Barreiro Megino, Fernando Harald; Kucharczyk, Katarzyna; Medrano Llamas, Ramón; van der Ster, Daniel

    2014-01-01

    The recent paradigm shift toward cloud computing in IT, and general interest in "Big Data" in particular, have demonstrated that the computing requirements of HEP are no longer globally unique. Indeed, the CERN IT department and LHC experiments have already made significant R&D investments in delivering and exploiting cloud computing resources. While a number of technical evaluations of interesting commercial offerings from global IT enterprises have been performed by various physics labs, further technical, security, sociological, and legal issues need to be address before their large-scale adoption by the research community can be envisaged. Helix Nebula - the Science Cloud is an initiative that explores these questions by joining the forces of three European research institutes (CERN, ESA and EMBL) with leading European commercial IT enterprises. The goals of Helix Nebula are to establish a cloud platform federating multiple commercial cloud providers, along with new business models, which can sustain ...

  2. Helix Nebula and CERN: A Symbiotic approach to exploiting commercial clouds

    CERN Multimedia

    Barreiro Megino, Fernando Harald; Kucharczyk, Katarzyna; Medrano Llamas, Ramón; van der Ster, Daniel

    2013-01-01

    The recent paradigm shift toward cloud computing in IT, and general interest in "Big Data" in particular, have demonstrated that the computing requirements of HEP are no longer globally unique. Indeed, the CERN IT department and LHC experiments have already made significant R&D; investments in delivering and exploiting cloud computing resources. While a number of technical evaluations of interesting commercial offerings from global IT enterprises have been performed by various physics labs, further technical, security, sociological, and legal issues need to be address before their large-scale adoption by the research community can be envisaged. Helix Nebula - the Science Cloud is an initiative that explores these questions by joining the forces of three European research institutes (CERN, ESA and EMBL) with leading European commercial IT enterprises. The goals of Helix Nebula are to establish a cloud platform federating multiple commercial cloud providers, along with new business models, which can sustain...

  3. The Effect of a Helix-Coil Transition on the Extension Elasticity

    Science.gov (United States)

    Buhot, Arnaud; Halperin, Avi

    2000-03-01

    The secondary structure of a polymer affects its deformation behavior in accordance with the Le Chatelier principle. An important example of such secondary structure is the alpha helix encountered in polypeptides. Similar structure was recently proposed for PEO in aqueous media. Our discussion concerns the coupling of the cooperative helix-coil transition and the extension elasticity. In particular, we analyze the extension of a long single chain by use of optical tweezers or AFM. We consider chains that exist in the coil-state when unperturbed. The transition nevertheless occurs because the extension favors the low entropy helical state. As a result, the corresponding force law exhibits a plateau. The analysis of this situation involves two ingredients: (I) the stretching free energy penalty for a rod-coil mutiblock copolymer (II) the entropy associated with the possible placements of the rod and coil blocks.

  4. Structure of a novel winged-helix like domain from human NFRKB protein.

    Directory of Open Access Journals (Sweden)

    Abhinav Kumar

    Full Text Available The human nuclear factor related to kappa-B-binding protein (NFRKB is a 1299-residue protein that is a component of the metazoan INO80 complex involved in chromatin remodeling, transcription regulation, DNA replication and DNA repair. Although full length NFRKB is predicted to be around 65% disordered, comparative sequence analysis identified several potentially structured sections in the N-terminal region of the protein. These regions were targeted for crystallographic studies, and the structure of one of these regions spanning residues 370-495 was determined using the JCSG high-throughput structure determination pipeline. The structure reveals a novel, mostly helical domain reminiscent of the winged-helix fold typically involved in DNA binding. However, further analysis shows that this domain does not bind DNA, suggesting it may belong to a small group of winged-helix domains involved in protein-protein interactions.

  5. Performance of Process Damping in Machining Titanium Alloys at Low Cutting Speed with Different Helix Tools

    International Nuclear Information System (INIS)

    Shaharun, M A; Yusoff, A R; Reza, M S; Jalal, K A

    2012-01-01

    Titanium is a strong, lustrous, corrosion-resistant and transition metal with a silver color to produce strong lightweight alloys for industrial process, automotive, medical instruments and other applications. However, it is very difficult to machine the titanium due to its poor machinability. When machining titanium alloys with the conventional tools, the wear rate of the tool is rapidly accelerate and it is generally difficult to achieve at high cutting speed. In order to get better understanding of machining titanium alloy, the interaction between machining structural system and the cutting process which result in machining instability will be studied. Process damping is a useful phenomenon that can be exploited to improve the limited productivity of low speed machining. In this study, experiments are performed to evaluate the performance of process damping of milling under different tool helix geometries. The results showed that the helix of 42° angle is significantly increase process damping performance in machining titanium alloy.

  6. Mechanical evaluation of quad-helix appliance made of low-nickel stainless steel wire.

    Science.gov (United States)

    dos Santos, Rogério Lacerda; Pithon, Matheus Melo

    2013-01-01

    The objective of this study was to test the hypothesis that there is no difference between stainless steel and low-nickel stainless steel wires as regards mechanical behavior. Force, resilience, and elastic modulus produced by Quad-helix appliances made of 0.032-inch and 0.036-inch wires were evaluated. Sixty Quad-helix appliances were made, thirty for each type of alloy, being fifteen for each wire thickness, 0.032-in and 0.036-in. All the archwires were submitted to mechanical compression test using an EMIC DL-10000 machine simulating activations of 4, 6, 9, and 12 mm. Analysis of variance (ANOVA) with multiple comparisons and Tukey's test were used (p steel alloy had force, resilience, and elastic modulus similar to those made of stainless steel alloy.

  7. Genomics into Healthcare: the 5th Pan Arab Human Genetics Conference and 2013 Golden Helix Symposium.

    Science.gov (United States)

    Fortina, Paolo; Al Khaja, Najib; Al Ali, Mahmoud Taleb; Hamzeh, Abdul Rezzak; Nair, Pratibha; Innocenti, Federico; Patrinos, George P; Kricka, Larry J

    2014-05-01

    The joint 5th Pan Arab Human Genetics conference and 2013 Golden Helix Symposium, "Genomics into Healthcare" was coorganized by the Center for Arab Genomic Studies (http://www.cags.org.ae) in collaboration with the Golden Helix Foundation (http://www.goldenhelix.org) in Dubai, United Arab Emirates from 17 to 19 November, 2013. The meeting was attended by over 900 participants, doctors and biomedical students from over 50 countries and was organized into a series of nine themed sessions that covered cancer genomics and epigenetics, genomic and epigenetic studies, genomics of blood and metabolic disorders, cytogenetic diagnosis and molecular profiling, next-generation sequencing, consanguinity and hereditary diseases, clinical genomics, clinical applications of pharmacogenomics, and genomics in public health. © 2014 WILEY PERIODICALS, INC.

  8. RNA secondary structure prediction based on SHAPE data in helix regions.

    Science.gov (United States)

    Lotfi, Mohadeseh; Zare-Mirakabad, Fatemeh; Montaseri, Soheila

    2015-09-07

    RNA molecules play important and fundamental roles in biological processes. Frequently, the functional form of single-stranded RNA molecules requires a specific tertiary structure. Classically, RNA structure determination has mostly been accomplished by X-Ray crystallography or Nuclear Magnetic Resonance approaches. These experimental methods are time consuming and expensive. In the past two decades, some computational methods and algorithms have been developed for RNA secondary structure prediction. In these algorithms, minimum free energy is known as the best criterion. However, the results of algorithms show that minimum free energy is not a sufficient criterion to predict RNA secondary structure. These algorithms need some additional knowledge about the structure, which has to be added in the methods. Recently, the information obtained from some experimental data, called SHAPE, can greatly improve the consistency between the native and predicted RNA secondary structure. In this paper, we investigate the influence of SHAPE data on four types of RNA substructures, helices, loops, base pairs from the start and end of helices and two base pairs from the start and end of helices. The results show that SHAPE data in helix regions can improve the prediction. We represent a new method to apply SHAPE data in helix regions for finding RNA secondary structure. Finally, we compare the results of the method on a set of RNAs to predict minimum free energy structure based on considering all SHAPE data and only SHAPE data in helix regions as pseudo free energy and without SHAPE data (without any pseudo free energy). The results show that RNA secondary structure prediction based on considering only SHAPE data in helix regions is more successful than not considering SHAPE data and it provides competitive results in comparison with considering all SHAPE data. Copyright © 2015 Elsevier Ltd. All rights reserved.

  9. A rare polyglycine type II-like helix motif in naturally occurring proteins.

    Science.gov (United States)

    Warkentin, Eberhard; Weidenweber, Sina; Schühle, Karola; Demmer, Ulrike; Heider, Johann; Ermler, Ulrich

    2017-11-01

    Common structural elements in proteins such as α-helices or β-sheets are characterized by uniformly repeating, energetically favorable main chain conformations which additionally exhibit a completely saturated hydrogen-bonding network of the main chain NH and CO groups. Although polyproline or polyglycine type II helices (PP II or PG II ) are frequently found in proteins, they are not considered as equivalent secondary structure elements because they do not form a similar self-contained hydrogen-bonding network of the main chain atoms. In this context our finding of an unusual motif of glycine-rich PG II -like helices in the structure of the acetophenone carboxylase core complex is of relevance. These PG II -like helices form hexagonal bundles which appear to fulfill the criterion of a (largely) saturated hydrogen-bonding network of the main-chain groups and therefore may be regarded in this sense as a new secondary structure element. It consists of a central PG II -like helix surrounded by six nearly parallel PG II -like helices in a hexagonal array, plus an additional PG II -like helix extending the array outwards. Very related structural elements have previously been found in synthetic polyglycine fibers. In both cases, all main chain NH and CO groups of the central PG II -helix are saturated by either intra- or intermolecular hydrogen-bonds, resulting in a self-contained hydrogen-bonding network. Similar, but incomplete PG II -helix patterns were also previously identified in a GTP-binding protein and an antifreeze protein. © 2017 Wiley Periodicals, Inc.

  10. Oligomerization state of photosynthetic core complexes is correlated with the dimerization affinity of a transmembrane helix.

    Science.gov (United States)

    Hsin, Jen; LaPointe, Loren M; Kazy, Alla; Chipot, Christophe; Senes, Alessandro; Schulten, Klaus

    2011-09-07

    In the Rhodobacter (Rba.) species of photosynthetic purple bacteria, a single transmembrane α-helix, PufX, is found within the core complex, an essential photosynthetic macromolecular assembly that performs the absorption and the initial processing of light energy. Despite its structural simplicity, many unresolved questions surround PufX, the most important of which is its location within the photosynthetic core complex. One proposed placement of PufX is at the center of a core complex dimer, where two PufX helices associate in the membrane and form a homodimer. Inability for PufX of certain Rba. species to form a homodimer is thought to lead to monomeric core complexes. In the present study, we employ a combination of computational and experimental techniques to test the hypothesized homodimerization of PufX. We carry out a systematic investigation to measure the dimerization affinity of PufX from four Rba. species, Rba. blasticus , Rba. capsulatus , Rba. sphaeroides , and Rba. veldkampii , using a molecular dynamics-based free-energy method, as well as experimental TOXCAT assays. We found that the four PufX helices have substantially different dimerization affinities. Both computational and experimental techniques demonstrate that species with dimeric core complexes have PufX that can potentially form a homodimer, whereas the one species with monomeric core complexes has a PufX with little to no dimerization propensity. Our analysis of the helix-helix interface revealed a number of positions that may be important for PufX dimerization and the formation of a hydrogen-bond network between these GxxxG-containing helices. Our results suggest that the different oligomerization states of core complexes in various Rba. species can be attributed, among other factors, to the different propensity of its PufX helix to homodimerize.

  11. Salt- and pH-Triggered Helix-Coil Transition of Ionic Polypeptides under Physiology Conditions.

    Science.gov (United States)

    Yuan, Jingsong; Zhang, Yi; Sun, Yue; Cai, Zhicheng; Yang, Lijiang; Lu, Hua

    2018-03-26

    Controlling the helix-coil transition of polypeptides under physiological conditions is an attractive way toward smart functional materials. Here we report the synthesis of a series of tertiary amine-functionalized ethylene glycol (EGx)-linked polypeptide electrolytes with their secondary structures tunable under physiological conditions. The resultant polymers, denoted as P(EGxDMA-Glu) (x =1, 2, and 3), show excellent aqueous solubility (> 10 mg/mL) regardless of their charge states. Unlike poly-L-lysine that can form helix only at pH above 10, P(EGxDMA-Glu) undergo a pH-dependent helix-coil switch with their transition points within physiological range (~ pH 5.3-6.5). Meanwhile, P(EGxDMA-Glu) exhibit unusual salt-induced helical conformation presumably owing to the unique properties of EGx linkers. Together, the current work highlights the importance of fine-tuning the linker chemistry in achieving conformation-switchable polypeptides, and represents a facile approach towards stimuli-responsive biopolymers for advanced biological applications.

  12. Left-handed polyproline-II helix revisited: proteins causing proteopathies.

    Science.gov (United States)

    Adzhubei, Alexei A; Anashkina, Anastasia A; Makarov, Alexander A

    2017-09-01

    Left-handed polyproline-II type helix is a regular conformation of polypeptide chain not only of fibrous, but also of folded and natively unfolded proteins and peptides. It is the only class of regular secondary structure substantially represented in non-fibrous proteins and peptides on a par with right-handed alpha-helix and beta-structure. In this study, we have shown that polyproline-II helix is abundant in several peptides and proteins involved in proteopathies, the amyloid-beta peptides, protein tau and prion protein. Polyproline-II helices form two interaction sites in the amyloid-beta peptides, which are pivotal for pathogenesis of Alzheimer's disease (AD). It also with high probability is the structure of the majority of tau phosphorylation sites, important for tau hyperphosphorylation and formation of neurofibrillary tangles, a hallmark of AD. Polyproline-II helices form large parts of the structure of the folded domain of prion protein. They can undergo conversion to beta-structure as a result of relatively small change of one torsional angle of polypeptide chain. We hypothesize that in prions and amyloids, in general polyproline-II helices can serve as structural elements of the normal structure as well as dormant nuclei of structure conversion, and thus play important role in structure changes leading to the formation of fibrils.

  13. Potassium channels in the central nervous system of the snail, Helix pomatia: localization and functional characterization.

    Science.gov (United States)

    Battonyai, I; Krajcs, N; Serfőző, Z; Kiss, T; Elekes, K

    2014-05-30

    The distribution and functional presence of three voltage-dependent potassium channels, Kv2.1, Kv3.4, Kv4.3, respectively, were studied in the central nervous system of the snail Helix pomatia by immunohistochemical and electrophysiological methods. Cell clusters displaying immunoreactivity for the different channels were observed in all parts of the CNS, although their localization and number partly varied. Differences were also found in their intracellular, perikaryonal and axonal localization, as well as in their presence in non-neuronal tissues nearby the CNS, such as the perineurium and the aorta wall. At ultrastructural level Kv4.3 channel immunolabeling was observed in axon profiles containing large 80-100nm granular vesicles. Blotting analyses revealed specific signals for the Kv2.1, Kv3.4 and Kv4.3 channels, confirming the presence of the channels in the Helix CNS. Voltage-clamp recordings proved that outward currents obtained from neurons displaying Kv3.4 or Kv4.3 immunoreactivity contained transient components while Kv2.1 immunoreactive cells were characterized by delayed currents. The distribution of the K(+)-channels containing neurons suggests specific roles in intercellular signaling processes in the Helix CNS, most probably related to well-defined, partly local events. The cellular localization of the K(+)-channels studied supports their involvement in both pre- and postsynaptic events at perikaryonal and axonal levels. Copyright © 2014 IBRO. Published by Elsevier Ltd. All rights reserved.

  14. Involvement of Na,K-pump in SEPYLRFamide-mediated reduction of cholinosensitivity in Helix neurons.

    Science.gov (United States)

    Pivovarov, Arkady S; Foreman, Richard C; Walker, Robert J

    2007-02-01

    SEPYLRFamide acts as an inhibitory modulator of acetylcholine (ACh) receptors in Helix lucorum neurones. Ouabain, a specific inhibitor of Na,K-pump, (0.1 mM, bath application) decreased the ACh-induced inward current (ACh-current) and increased the leak current. Ouabain decreased the modulatory SEPYLRFamide effect on the ACh-current. There was a correlation between the effects of ouabain on the amplitude of the ACh-current and on the modulatory peptide effect. Ouabain and SEPYLRFamide inhibited the activity of Helix aspersa brain Na,K-ATPase. Activation of Na,K-pump by intracellular injection of 3 M Na acetate or 3 M NaCl reduced the modulatory peptide effect on the ACh-current. An inhibitor of Na/Ca-exchange, benzamil (25 muM, bath application), and an inhibitor of Ca(2+)-pump in the endoplasmic reticulum, thapsigargin (TG, applied intracellularly), both prevented the effect of ouabain on SEPYLRFamide-mediated modulatory effect. Another inhibitor of Ca(2+)-pump in the endoplasmic reticulum, cyclopiazonic acid (applied intracellularly), did not prevent the effect of ouabain on SEPYLRFamide-mediated modulatory effect. These results indicate that Na,K-pump is responsible for the SEPYLRFamide-mediated inhibition of ACh receptors in Helix neurons. Na/Ca-exchange and intracellular Ca(2+) released from internal pools containing TG-sensitive Ca(2+)-pump are involved in the Na,K-pump pathway for the SEPYLRFamide-mediated inhibition of ACh receptors.

  15. Structure of the membrane anchor of pestivirus glycoprotein E(rns, a long tilted amphipathic helix.

    Directory of Open Access Journals (Sweden)

    Daniel Aberle

    2014-02-01

    Full Text Available E(rns is an essential virion glycoprotein with RNase activity that suppresses host cellular innate immune responses upon being partially secreted from the infected cells. Its unusual C-terminus plays multiple roles, as the amphiphilic helix acts as a membrane anchor, as a signal peptidase cleavage site, and as a retention/secretion signal. We analyzed the structure and membrane binding properties of this sequence to gain a better understanding of the underlying mechanisms. CD spectroscopy in different setups, as well as Monte Carlo and molecular dynamics simulations confirmed the helical folding and showed that the helix is accommodated in the amphiphilic region of the lipid bilayer with a slight tilt rather than lying parallel to the surface. This model was confirmed by NMR analyses that also identified a central stretch of 15 residues within the helix that is fully shielded from the aqueous layer, which is C-terminally followed by a putative hairpin structure. These findings explain the strong membrane binding of the protein and provide clues to establishing the E(rns membrane contact, processing and secretion.

  16. Structure of the Membrane Anchor of Pestivirus Glycoprotein Erns, a Long Tilted Amphipathic Helix

    Science.gov (United States)

    Aberle, Daniel; Muhle-Goll, Claudia; Bürck, Jochen; Wolf, Moritz; Reißer, Sabine; Luy, Burkhard; Wenzel, Wolfgang; Ulrich, Anne S.; Meyers, Gregor

    2014-01-01

    Erns is an essential virion glycoprotein with RNase activity that suppresses host cellular innate immune responses upon being partially secreted from the infected cells. Its unusual C-terminus plays multiple roles, as the amphiphilic helix acts as a membrane anchor, as a signal peptidase cleavage site, and as a retention/secretion signal. We analyzed the structure and membrane binding properties of this sequence to gain a better understanding of the underlying mechanisms. CD spectroscopy in different setups, as well as Monte Carlo and molecular dynamics simulations confirmed the helical folding and showed that the helix is accommodated in the amphiphilic region of the lipid bilayer with a slight tilt rather than lying parallel to the surface. This model was confirmed by NMR analyses that also identified a central stretch of 15 residues within the helix that is fully shielded from the aqueous layer, which is C-terminally followed by a putative hairpin structure. These findings explain the strong membrane binding of the protein and provide clues to establishing the Erns membrane contact, processing and secretion. PMID:24586172

  17. Structure of the membrane anchor of pestivirus glycoprotein E(rns), a long tilted amphipathic helix.

    Science.gov (United States)

    Aberle, Daniel; Muhle-Goll, Claudia; Bürck, Jochen; Wolf, Moritz; Reißer, Sabine; Luy, Burkhard; Wenzel, Wolfgang; Ulrich, Anne S; Meyers, Gregor

    2014-02-01

    E(rns) is an essential virion glycoprotein with RNase activity that suppresses host cellular innate immune responses upon being partially secreted from the infected cells. Its unusual C-terminus plays multiple roles, as the amphiphilic helix acts as a membrane anchor, as a signal peptidase cleavage site, and as a retention/secretion signal. We analyzed the structure and membrane binding properties of this sequence to gain a better understanding of the underlying mechanisms. CD spectroscopy in different setups, as well as Monte Carlo and molecular dynamics simulations confirmed the helical folding and showed that the helix is accommodated in the amphiphilic region of the lipid bilayer with a slight tilt rather than lying parallel to the surface. This model was confirmed by NMR analyses that also identified a central stretch of 15 residues within the helix that is fully shielded from the aqueous layer, which is C-terminally followed by a putative hairpin structure. These findings explain the strong membrane binding of the protein and provide clues to establishing the E(rns) membrane contact, processing and secretion.

  18. BAROMETRIC PRESSURE and Other Data from ALPHA HELIX From Bering Sea and Others from 19940910 to 19941010 (NODC Accession 9500030)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — The Conductivity, Temperature and Depth (CTD) and other data were collected in Bering Sea and Chukchi Sea. Data was collected from Ship ALPHA HELIX. The data was...

  19. Altered viral fitness and drug susceptibility in HIV-1 carrying mutations that confer resistance to nonnucleoside reverse transcriptase and integrase strand transfer inhibitors.

    Science.gov (United States)

    Hu, Zixin; Kuritzkes, Daniel R

    2014-08-01

    Nonnucleoside reverse transcriptase (RT) inhibitors (NNRTI) and integrase (IN) strand transfer inhibitors (INSTI) are key components of antiretroviral regimens. To explore potential interactions between NNRTI and INSTI resistance mutations, we investigated the combined effects of these mutations on drug susceptibility and fitness of human immunodeficiency virus type 1 (HIV-1). In the absence of drug, single-mutant viruses were less fit than the wild type; viruses carrying multiple mutations were less fit than single-mutant viruses. These findings were explained in part by the observation that mutant viruses carrying NNRTI plus INSTI resistance mutations had reduced amounts of virion-associated RT and/or IN protein. In the presence of efavirenz (EFV), a virus carrying RT-K103N together with IN-G140S and IN-Q148H (here termed IN-G140S/Q148H) mutations was fitter than a virus with a RT-K103N mutation alone. Similarly, in the presence of EFV, the RT-E138K plus IN-G140S/Q148H mutant virus was fitter than one with the RT-E138K mutation alone. No effect of INSTI resistance mutations on the fitness of RT-Y181C mutant viruses was observed. Conversely, RT-E138K and -Y181C mutations improved the fitness of the IN-G140S/Q148H mutant virus in the presence of raltegravir (RAL); the RT-K103N mutation had no effect. The NNRTI resistance mutations had no effect on RAL susceptibility. Likewise, the IN-G140S/Q148H mutations had no effect on EFV or RPV susceptibility. However, both the RT-K103N plus IN-G140S/Q148H and the RT-E138K plus IN-G140S/Q148H mutant viruses had significantly greater fold increases in 50% inhibitory concentration (IC50) of EFV than viruses carrying a single NNRTI mutation. Likewise, the RT-E138K plus IN-G140S/Q148H mutant virus had significantly greater fold increases in RAL IC50 than that of the IN-G140S/Q148H mutant virus. These results suggest that interactions between RT and IN mutations are important for NNRTI and INSTI resistance and viral fitness

  20. Mutations in the catalytic core or the C-terminus of murine leukemia virus (MLV) integrase disrupt virion infectivity and exert diverse effects on reverse transcription

    International Nuclear Information System (INIS)

    Steinrigl, Adolf; Nosek, Dagmara; Ertl, Reinhard; Guenzburg, Walter H.; Salmons, Brian; Klein, Dieter

    2007-01-01

    Understanding of the structures and functions of the retroviral integrase (IN), a key enzyme in the viral replication cycle, is essential for developing antiretroviral treatments and facilitating the development of safer gene therapy vehicles. Thus, four MLV IN-mutants were constructed in the context of a retroviral vector system, harbouring either a substitution in the catalytic centre, deletions in the C-terminus, or combinations of both modifications. IN-mutants were tested for their performance in different stages of the viral replication cycle: RNA-packaging; RT-activity; transient and stable infection efficiency; dynamics of reverse transcription and nuclear entry. All mutant vectors packaged viral RNA with wild-type efficiencies and displayed only slight reductions in RT-activity. Deletion of either the IN C-terminus alone, or in addition to part of the catalytic domain exerted contrasting effects on intracellular viral DNA levels, implying that IN influences reverse transcription in more than one direction

  1. Praevalensen afhud- og slimhindesymptomer blandt gartnere der omgås Ficus benjamina (stuebirk) og Hedera helix (vedbend). Et tvaersnitsstudie

    DEFF Research Database (Denmark)

    Jørs, Erik

    2003-01-01

    Allergic and toxic initiative symptoms from skin, eyes and respiratory tract are well known among gardeners This study reports the prevalence of these symptoms among gardeners working with Ficus Benjamina (Fb) and Hedera helix (Hh).......Allergic and toxic initiative symptoms from skin, eyes and respiratory tract are well known among gardeners This study reports the prevalence of these symptoms among gardeners working with Ficus Benjamina (Fb) and Hedera helix (Hh)....

  2. Removal of bacterial cells, antibiotic resistance genes and integrase genes by on-site hospital wastewater treatment plants: surveillance of treated hospital effluent quality

    KAUST Repository

    Timraz, Kenda Hussain Hassan

    2016-12-15

    This study aims to evaluate the removal efficiency of microbial contaminants, including total cell counts, antibiotic-resistant bacteria (ARB), antibiotic resistance genes (ARGs, e.g. tetO, tetZ, sul1 and sul2) and integrase genes (e.g. intl1 and intl2), by wastewater treatment plants (WWTPs) operated on-site of two hospitals (i.e., SH WWTP and IH WWTP). Both SH and IH WWTPs utilize the conventional activated sludge process but differences in the removal efficiencies were observed. Over the 2 week sampling period, IH WWTP outperformed SH WWTP, and achieved an approximate 0.388 to 2.49-log log removal values (LRVs) for total cell counts compared to the 0.010 to 0.162-log removal in SH WWTP. Although ARB were present in the hospital influent, the treatment process of both hospitals effectively removed ARB from most of the effluent samples. In instances where ARB were recovered in the effluent, none of the viable isolates were identified to be opportunistic pathogenic species based on 16S rRNA gene sequencing. However, sul1 and intl1 genes remained detectable at up to 105 copies per mL or 8 x 10(-1) copies per 16S rRNA gene in the treated effluent, with an LRV of less than 1.2. When the treated effluent is discharged from hospital WWTPs into the public sewer for further treatment as per requirement in many countries, the detected amount of ARGs and integrase genes in the hospital effluent can become a potential source of horizontal gene dissemination in the municipal WWTP. Proper on-site wastewater treatment and surveillance of the effluent quality for emerging contaminants are therefore highly recommended.

  3. Prevalence of primary resistance mutations to integrase inhibitors in treatment-naïve and -experienced patients infected with B and non-B HIV-1 variants.

    Science.gov (United States)

    Gutiérrez, Carolina; Hernández-Novoa, Beatriz; Pérez-Elías, María Jesús; Moreno, Ana María; Holguín, Africa; Dronda, Fernando; Casado, José Luis; Moreno, Santiago

    2013-01-01

    Raltegravir (RAL) constitutes the first available integrase strand transfer inhibitor (INSTI) available in clinical practice. Three independent pathways have been described to confer resistance to RAL. Secondary mutations with little effect on INSTI susceptibility and additional substitutions with an uncertain role have also been described especially in HIV-1 non-B variants. We evaluated the prevalence of primary, secondary, and additional resistance mutations to INSTIs in patients naïve to RAL or elvitegravir (EGV) carrying different HIV-1 variants. A total of 83 patients infected by B HIV-1 subtype (64%) or non-B HIV-1 variants (36%) were evaluated. No primary mutations to RAL or EGV were found in the inte-grase sequences analyzed. Secondary mutations were detected in only 5 patients. Additional mutations were found in both in B and non-B variants. According to the geno2pheno algorithm, some of the secondary mutations detected (L74V, E138K, G163RS, and V151I) have been associated with a reduced estimated susceptibility to RAL and only the E138K mutation has been associated with a decreased estimated susceptibility to EGV. No virological failure was observed after RAL was administrated in 17 patients carrying 1 or more additional substitutions in the absence of primary or secondary mutations. No primary resistance mutations to INSTI were found in treatment-naïve or -experienced patients infected with B or non-B HIV-1 variants. The vast majority had some polymorphic and non-polymorphic substitutions; however response to RAL was excellent in patients who harbored one or more of these mutations. We could not identify any clinical factors associated with the presence of any of these mutations.

  4. Effects of ligands on unfolding of the amyloid β-peptide central helix: mechanistic insights from molecular dynamics simulations.

    Directory of Open Access Journals (Sweden)

    Mika Ito

    Full Text Available Polymerization of the amyloid β-peptide (Aβ, a process which requires that the helical structure of Aβ unfolds beforehand, is suspected to cause neurodegeneration in Alzheimer's disease. According to recent experimental studies, stabilization of the Aβ central helix counteracts Aβ polymerization into toxic assemblies. The effects of two ligands (Dec-DETA and Pep1b, which were designed to bind to and stabilize the Aβ central helix, on unfolding of the Aβ central helix were investigated by molecular dynamics simulations. It was quantitatively demonstrated that the stability of the Aβ central helix is increased by both ligands, and more effectively by Pep1b than by Dec-DETA. In addition, it was shown that Dec-DETA forms parallel conformations with β-strand-like Aβ, whereas Pep1b does not and instead tends to bend unwound Aβ. The molecular dynamics results correlate well with previous experiments for these ligands, which suggest that the simulation method should be useful in predicting the effectiveness of novel ligands in stabilizing the Aβ central helix. Detailed Aβ structural changes upon loss of helicity in the presence of the ligands are also revealed, which gives further insight into which ligand may lead to which path subsequent to unwinding of the Aβ central helix.

  5. The Combination of the R263K and T66I Resistance Substitutions in HIV-1 Integrase Is Incompatible with High-Level Viral Replication and the Development of High-Level Drug Resistance.

    Science.gov (United States)

    Liang, Jiaming; Mesplède, Thibault; Oliveira, Maureen; Anstett, Kaitlin; Wainberg, Mark A

    2015-11-01

    The R263K substitution in integrase has been selected in tissue culture with dolutegravir (DTG) and has been reported for several treatment-experienced individuals receiving DTG as part of salvage therapy. The R263K substitution seems to be incompatible with the presence of common resistance mutations associated with raltegravir (RAL), a different integrase strand transfer inhibitor (INSTI). T66I is a substitution that is common in individuals who have developed resistance against a different INSTI termed elvitegravir (EVG), but it is not known whether these two mutations might be compatible in the context of resistance against DTG or what impact the combination of these substitutions might have on resistance against INSTIs. E138K is a common secondary substitution observed with various primary resistance substitutions in RAL- and EVG-treated individuals. Viral infectivity, replicative capacity, and resistance against INSTIs were measured in cell-based assays. Strand transfer and 3' processing activities were measured biochemically. The combination of the R263K and T66I substitutions decreased HIV-1 infectivity, replicative capacity, and strand transfer activity. The addition of the E138K substitution partially compensated for these deficits and resulted in high levels of resistance against EVG but not against DTG or RAL. These findings suggest that the presence of the T66I substitution will not compromise the activity of DTG and may also help to prevent the additional generation of the R263K mutation. Our observations support the use of DTG in second-line therapy for individuals who experience treatment failure with EVG due to the T66I substitution. The integrase strand transfer inhibitors (INSTIs) elvitegravir and dolutegravir are newly developed inhibitors against human immunodeficiency virus type 1 (HIV-1). HIV drug-resistant mutations in integrase that can arise in individuals treated with elvitegravir commonly include the T66I substitution, whereas R263K is a

  6. Alanine to valine substitutions in the pore helix IIIP1 and linker-helix IIIL45 confer cockroach sodium channel resistance to DDT and pyrethroids.

    Science.gov (United States)

    Chen, Mengli; Du, Yuzhe; Nomura, Yoshiko; Zhu, Guonian; Zhorov, Boris S; Dong, Ke

    2017-05-01

    Pyrethroid insecticides exert toxic effects by prolonging the opening of voltage-gated sodium channels. More than 20 sodium channel mutations from arthropod pests and disease vectors have been confirmed to confer pyrethroid resistance. These mutations have been valuable in elucidating the molecular interaction between pyrethroids and sodium channels, including identification of two pyrethroid receptor sites. Previously, two alanine to valine substitutions, one in the pore helix IIIP1 and the other in the linker-helix connecting S4 and S5 in domain III (IIIL45), were found in Drosophila melanogaster mutants that are resistant to DDT and deltamethrin (a type II pyrethroid with an α-cyano group at the phenylbenzyl alcohol position, which is lacking in type I pyrethroids), but their role in target-site-mediated insecticide resistance has not been functionally confirmed. In this study, we functionally examined the two mutations in cockroach sodium channels expressed in Xenopus laevis oocytes. Both mutations caused depolarizing shifts in the voltage dependence of activation, conferred DDT resistance and also resistance to two Type I pyrethroids by almost abolishing the tail currents induced by Type I pyrethroids. In contrast, neither mutation reduced the amplitude of tail currents induced by the Type II pyrethroids, deltamethrin or cypermethrin. However, both mutations accelerated the decay of Type II pyrethroid-induced tail currents, which normally decay extremely slowly. These results provided new insight into the molecular basis of different actions of Type I and Type II pyrethroids on sodium channels. Computer modeling predicts that both mutations may allosterically affect pyrethroid binding. Copyright © 2016 Elsevier B.V. All rights reserved.

  7. The poplar basic helix-loop-helix transcription factor BEE3 – Like gene affects biomass production by enhancing proliferation of xylem cells in poplar

    Energy Technology Data Exchange (ETDEWEB)

    Noh, Seol Ah, E-mail: s6022029@korea.ac.kr; Choi, Young-Im, E-mail: yichoi99@forest.go.kr; Cho, Jin-Seong, E-mail: jinsung3932@gmail.com; Lee, Hyoshin, E-mail: hslee@forest.go.kr

    2015-06-19

    Brassinosteroids (BRs) play important roles in many aspects of plant growth and development, including regulation of vascular cambium activities and cell elongation. BR-induced BEE3 (brassinosteroid enhanced expression 3) is required for a proper BR response. Here, we identified a poplar (Populus alba × Populus glandulosa) BEE3-like gene, PagBEE3L, encoding a putative basic helix-loop-helix (bHLH)-type transcription factor. Expression of PagBEE3L was induced by brassinolide (BL). Transcripts of PagBEE3L were mainly detected in stems, with the internode having a low level of transcription and the node having a relatively higher level. The function of the PagBEE3L gene was investigated through phenotypic analyses with PagBEE3L-overexpressing (ox) transgenic lines. This work particularly focused on a potential role of PagBEE3L in stem growth and development of polar. The PagBEE3L-ox poplar showed thicker and longer stems than wild-type plants. The xylem cells from the stems of PagBEE3L-ox plants revealed remarkably enhanced proliferation, resulting in an earlier thickening growth than wild-type plants. Therefore, this work suggests that xylem development of poplar is accelerated in PagBEE3L-ox plants and PagBEE3L plays a role in stem growth by increasing the proliferation of xylem cells to promote the initial thickening growth of poplar stems. - Highlights: • We identify the BEE3-like gene form hybrid poplar (Populus alba × Populus glandulosa). • We examine effects of overexpression of PagBEE3L on growth in poplar. • We found that 35S:BEE3L transgenic plants showed more rapid growth than wild-type plants. • BEE3L protein plays an important role in the development of plant stem.

  8. The poplar basic helix-loop-helix transcription factor BEE3 – Like gene affects biomass production by enhancing proliferation of xylem cells in poplar

    International Nuclear Information System (INIS)

    Noh, Seol Ah; Choi, Young-Im; Cho, Jin-Seong; Lee, Hyoshin

    2015-01-01

    Brassinosteroids (BRs) play important roles in many aspects of plant growth and development, including regulation of vascular cambium activities and cell elongation. BR-induced BEE3 (brassinosteroid enhanced expression 3) is required for a proper BR response. Here, we identified a poplar (Populus alba × Populus glandulosa) BEE3-like gene, PagBEE3L, encoding a putative basic helix-loop-helix (bHLH)-type transcription factor. Expression of PagBEE3L was induced by brassinolide (BL). Transcripts of PagBEE3L were mainly detected in stems, with the internode having a low level of transcription and the node having a relatively higher level. The function of the PagBEE3L gene was investigated through phenotypic analyses with PagBEE3L-overexpressing (ox) transgenic lines. This work particularly focused on a potential role of PagBEE3L in stem growth and development of polar. The PagBEE3L-ox poplar showed thicker and longer stems than wild-type plants. The xylem cells from the stems of PagBEE3L-ox plants revealed remarkably enhanced proliferation, resulting in an earlier thickening growth than wild-type plants. Therefore, this work suggests that xylem development of poplar is accelerated in PagBEE3L-ox plants and PagBEE3L plays a role in stem growth by increasing the proliferation of xylem cells to promote the initial thickening growth of poplar stems. - Highlights: • We identify the BEE3-like gene form hybrid poplar (Populus alba × Populus glandulosa). • We examine effects of overexpression of PagBEE3L on growth in poplar. • We found that 35S:BEE3L transgenic plants showed more rapid growth than wild-type plants. • BEE3L protein plays an important role in the development of plant stem

  9. atonal- and achaete-scute-related genes in the annelid Platynereis dumerilii: insights into the evolution of neural basic-Helix-Loop-Helix genes

    Directory of Open Access Journals (Sweden)

    Arendt Detlev

    2008-06-01

    Full Text Available Abstract Background Functional studies in model organisms, such as vertebrates and Drosophila, have shown that basic Helix-loop-Helix (bHLH proteins have important roles in different steps of neurogenesis, from the acquisition of neural fate to the differentiation into specific neural cell types. However, these studies highlighted many differences in the expression and function of orthologous bHLH proteins during neural development between vertebrates and Drosophila. To understand how the functions of neural bHLH genes have evolved among bilaterians, we have performed a detailed study of bHLH genes during nervous system development in the polychaete annelid, Platynereis dumerilii, an organism which is evolutionary distant from both Drosophila and vertebrates. Results We have studied Platynereis orthologs of the most important vertebrate neural bHLH genes, i.e. achaete-scute, neurogenin, atonal, olig, and NeuroD genes, the latter two being genes absent of the Drosophila genome. We observed that all these genes have specific expression patterns during nervous system formation in Platynereis. Our data suggest that in Platynereis, like in vertebrates but unlike Drosophila, (i neurogenin is the main proneural gene for the formation of the trunk central nervous system, (ii achaete-scute and olig genes are involved in neural subtype specification in the central nervous system, in particular in the specification of the serotonergic phenotype. In addition, we found that the Platynereis NeuroD gene has a broad and early neuroectodermal expression, which is completely different from the neuronal expression of vertebrate NeuroD genes. Conclusion Our analysis suggests that the Platynereis bHLH genes have both proneural and neuronal specification functions, in a way more akin to the vertebrate situation than to that of Drosophila. We conclude that these features are ancestral to bilaterians and have been conserved in the vertebrates and annelids lineages, but

  10. A genome-wide identification and analysis of the basic helix-loop-helix transcription factors in the ponerine ant, Harpegnathos saltator

    Directory of Open Access Journals (Sweden)

    Liu Ake

    2012-08-01

    Full Text Available Abstract Background The basic helix-loop-helix (bHLH transcription factors and their homologs form a superfamily that plays essential roles in transcriptional networks of multiple developmental processes. bHLH family members have been identified in over 20 organisms, including fruit fly, zebrafish, human and mouse. Result In this study, we conducted a genome-wide survey for bHLH sequences, and identified 57 bHLH sequences encoded in complete genome sequence of the ponerine ant, Harpegnathos saltator. Phylogenetic analysis of the bHLH domain sequences classified these genes into 38 bHLH families with 23, 14, 10, 1, 8 and 1 members in group A, B, C, D, E and F, respectively. The number of PabHLHs (ponerine ant bHLHs with introns is higher than many other insect species, and they are found to have introns with average lengths only inferior to those of pea aphid. In addition, two H. saltator bHLHs named PaCrp1 and PaSide locate on two separate contigs in the genome. Conclusions A putative full set of PabHLH genes is comparable with other insect species and genes encoding Oligo, MyoRb and Figα were not found in genomes of all insect species of which bHLH family members have been identified. Moreover, in-family phylogenetic analyses indicate that the PabHLH genes are more closely related with Apis mellifera than others. The present study will serve as a solid foundation for further investigations into the structure and function of bHLH proteins in the regulation of H. saltator development.

  11. Triple Helix

    National Research Council Canada - National Science Library

    2008-01-01

    .... In 2008, the FIRST Robotics Competition challenged over 37,000 high-school-aged young people on more than 1,500 teams worldwide to work with 18,000 mentors and 2,000 sponsoring companies to design...

  12. Helix Nebula and CERN: A Symbiotic approach to exploiting commercial clouds

    Science.gov (United States)

    Barreiro Megino, Fernando H.; Jones, Robert; Kucharczyk, Katarzyna; Medrano Llamas, Ramón; van der Ster, Daniel

    2014-06-01

    The recent paradigm shift toward cloud computing in IT, and general interest in "Big Data" in particular, have demonstrated that the computing requirements of HEP are no longer globally unique. Indeed, the CERN IT department and LHC experiments have already made significant R&D investments in delivering and exploiting cloud computing resources. While a number of technical evaluations of interesting commercial offerings from global IT enterprises have been performed by various physics labs, further technical, security, sociological, and legal issues need to be address before their large-scale adoption by the research community can be envisaged. Helix Nebula - the Science Cloud is an initiative that explores these questions by joining the forces of three European research institutes (CERN, ESA and EMBL) with leading European commercial IT enterprises. The goals of Helix Nebula are to establish a cloud platform federating multiple commercial cloud providers, along with new business models, which can sustain the cloud marketplace for years to come. This contribution will summarize the participation of CERN in Helix Nebula. We will explain CERN's flagship use-case and the model used to integrate several cloud providers with an LHC experiment's workload management system. During the first proof of concept, this project contributed over 40.000 CPU-days of Monte Carlo production throughput to the ATLAS experiment with marginal manpower required. CERN's experience, together with that of ESA and EMBL, is providing a great insight into the cloud computing industry and highlighted several challenges that are being tackled in order to ease the export of the scientific workloads to the cloud environments.

  13. Helix Nebula and CERN: A Symbiotic approach to exploiting commercial clouds

    International Nuclear Information System (INIS)

    Megino, Fernando H Barreiro; Jones, Robert; Llamas, Ramón Medrano; Ster, Daniel van der; Kucharczyk, Katarzyna

    2014-01-01

    The recent paradigm shift toward cloud computing in IT, and general interest in 'Big Data' in particular, have demonstrated that the computing requirements of HEP are no longer globally unique. Indeed, the CERN IT department and LHC experiments have already made significant R and D investments in delivering and exploiting cloud computing resources. While a number of technical evaluations of interesting commercial offerings from global IT enterprises have been performed by various physics labs, further technical, security, sociological, and legal issues need to be address before their large-scale adoption by the research community can be envisaged. Helix Nebula – the Science Cloud is an initiative that explores these questions by joining the forces of three European research institutes (CERN, ESA and EMBL) with leading European commercial IT enterprises. The goals of Helix Nebula are to establish a cloud platform federating multiple commercial cloud providers, along with new business models, which can sustain the cloud marketplace for years to come. This contribution will summarize the participation of CERN in Helix Nebula. We will explain CERN's flagship use-case and the model used to integrate several cloud providers with an LHC experiment's workload management system. During the first proof of concept, this project contributed over 40.000 CPU-days of Monte Carlo production throughput to the ATLAS experiment with marginal manpower required. CERN's experience, together with that of ESA and EMBL, is providing a great insight into the cloud computing industry and highlighted several challenges that are being tackled in order to ease the export of the scientific workloads to the cloud environments.

  14. The generalized model of polypeptide chain describing the helix-coil transition in biopolymers

    International Nuclear Information System (INIS)

    Mamasakhlisov, E.S.; Badasyan, A.V.; Tsarukyan, A.V.; Grigoryan, A.V.; Morozov, V.F.

    2005-07-01

    In this paper we summarize some results of our theoretical investigations of helix-coil transition both in single-strand (polypeptides) and two-strand (polynucleotides) macromolecules. The Hamiltonian of the Generalized Model of Polypeptide Chain (GMPC) is introduced to describe the system in which the conformations are correlated over some dimensional range Δ (it equals 3 for polypeptide, because one H-bond fixes three pairs of rotation, for double strand DNA it equals to one chain rigidity because of impossibility of loop formation on the scale less than Δ). The Hamiltonian does not contain any parameter designed especially for helix-coil transition and uses pure molecular microscopic parameters (the energy of hydrogen bond formation, reduced partition function of repeated unit, the number of repeated units fixed by one hydrogen bond, the energies of interaction between the repeated units and the solvent molecules). To calculate averages we evaluate the partition function using the transfer-matrix approach. The GMPC allowed to describe the influence of a number of factors, affecting the transition, basing on a unified microscopic approach. Thus we obtained, that solvents change transition temperature and interval in different ways, depending on type of solvent and on energy of solvent- macromolecule interaction; stacking on the background of H-bonding increases stability and decreases cooperativity of melting. For heterogeneous DNA we could analytically derive well known formulae for transition temperature and interval. In the framework of GMPC we calculate and show the difference of two order parameters of helix-coil transition - the helicity degree, and the average fraction of repeated units in helical conformation. Given article has the aim to review the results obtained during twenty years in the context of GMPC. (author)

  15. PACAP has anti-apoptotic effect in the salivary gland of an invertebrate species, Helix pomatia.

    Science.gov (United States)

    Pirger, Zsolt; Nemeth, Jozsef; Hiripi, Laszlo; Toth, Gabor; Kiss, Peter; Lubics, Andrea; Tamas, Andrea; Hernadi, Laszlo; Kiss, Tibor; Reglodi, Dora

    2008-11-01

    Pituitary adenylate cyclase activating polypeptide (PACAP) shows a remarkable sequence similarity among species and several studies provide evidence that the functions of PACAP have also been conserved among vertebrate species. Relatively little is known about its presence and functions in invertebrates. The aim of the present study was to investigate whether the well-known anti-apoptotic effect of PACAP can also be demonstrated in invertebrates. This effect was studied in the salivary gland of a molluscan species, Helix pomatia. In this work, we first showed the presence of PACAP-like immunoreactivity in the Helix salivary gland by means of immunohistochemistry. Radioimmunoassay measurements showed that PACAP38-like immunoreactivity dominated in the salivary gland of both active and inactive snails and its concentration was higher in active than in inactive animals in contrast to PACAP27-like immunoreactivity, which did not show activity-dependent changes. PACAP induced a significant elevation of cAMP level in salivary gland extracts. Application of apoptosis-inducing agents, dopamine and colchicine, led to a marked increase in the number of terminal uridine deoxynucleotidyl transferase dUTP nick end labeling (TUNEL)-positive apoptotic cells in the salivary gland, which was significantly attenuated by PACAP treatment. In a similar manner, the number of caspase-positive cells was reduced after co-application of dopamine and PACAP. Taken together, the data indicate that PACAP activates cAMP in a molluscan species and we show, for the first time, that PACAP is anti-apoptotic in the invertebrate Helix pomatia.

  16. Geometry of the toroidal N-helix: optimal-packing and zero-twist

    DEFF Research Database (Denmark)

    Olsen, Kasper; Bohr, Jakob

    2012-01-01

    in the other direction under pull. The packing problem for tubular N-helices is extended to bent helices where the strands are coiled on toruses. We analyze the geometry of open circular helices and develop criteria for the strands to be in contact. The analysis is applied to a single, a double and a triple...... helix. General N-helices are discussed, as well as zero-twist helices for N > 1. The derived geometrical restrictions are gradually modified by changing the aspect ratio of the torus....

  17. Modified quad helix appliance for thumb sucking and cross bite correction

    Directory of Open Access Journals (Sweden)

    C Vinay

    2013-01-01

    Full Text Available Digit sucking habit is a learned pattern of behavior commonly seen in children of preschool age. Prolonged digit sucking beyond the preschool age, lead to the development of malocclusion such as anterior open bite, maxillary constriction and posterior crossbite. Treatment strategies include interception of habit and correction of the malocclusion. The present case report describes a modified quad helix appliance used successfully in a 9-year-old child to intercept thumb sucking habit and simultaneous correction of posterior crossbite. The appliance has the advantage of easy fabrication, being versatile and more patients compliant.

  18. CAXIAS DO SUL TECHNOLOGY INCUBATOR: TECHNOLOGICAL INNOVATION THROUGH THE TRIPLE HELIX

    Directory of Open Access Journals (Sweden)

    Priscila Bresolin Tisott

    2014-09-01

    on investment in the form of registered patents, attracting new students and recognition as a technological, entrepreneurial and innovative university. The perceived benefits for the industrial sector are training and development for businesses and entrepreneurs, while the municipal government perceives the reduced number of businesses failing and the financial returns, in the form of taxes, as benefits. The findings show that the Triple Helix concept developed by Etzkowitz (2009 plays a cooperative and participatory role in the Technology Incubator, even though it could be applied to more segments of society.

  19. Effect of microclimatic conditions on the reproductive behavior of the Helix aspersa

    Directory of Open Access Journals (Sweden)

    Hugo Ángel Fernández Cabrera

    2013-12-01

    Full Text Available The aim of this study was to evaluate the effect of microclimatic conditions (temperature and relative humidity on the reproduction of land snails (Helix aspersa. For this, incubation’s chambers were conditioned with different four levels of relative humidity and temperature. Posture, length of incubation and hatching rate were evaluated. It was concluded that 81.0 ± 0.63% of relative humidity and 23.9 ± 0.39 °C of temperature the best results are achieved.

  20. An evolutionary triple helix to strengthen energy regulation: Implications for management

    Energy Technology Data Exchange (ETDEWEB)

    Rizzi, Francesco; Borzoni, Matteo

    2010-09-15

    Regulation is the basic tool to implement energy policy. The evolution of the regulation is influenced by its impacts on the industrial activities. Consequently, entrepreneurs acts in a continuously adapting-by-interacting environment. Both from a systemic and an atomistic perspective, this paper provides a theoretical framework for energy regulation development in order to support management implications. This work builds on the triple helix model and extends it to energy regulation development processes. It concludes that the analysis of intangible resources and their related services at inter-organizational level is fundamental to guide companies in designing win-win corporate strategies and in their operazionalization.

  1. Novel triplet of flexor muscles in the posterior tentacles of the snail, Helix pomatia.

    Science.gov (United States)

    Hernádi, L; Teyke, T

    2012-01-01

    The anatomy of three novel flexor muscles in the posterior tentacles of Helix pomatia is described. The muscles originate from the ventral side of the sensory pad and are anchored at different sites in the base of the tentacle stem. The muscles span the tentacle and always take the length of the stem which depends on the rate of tentacle protrusion indicating that the muscles are both contractile and extremely stretchable. The three anchoring points at the base of the stem determine three space axes along which the contraction of a muscle or the synchronous contraction of the muscles can move the tentacle in space.

  2. Mathematical modeling and design parameters of crushing machines with variable-pitch helix of the screw

    Directory of Open Access Journals (Sweden)

    Pelenko V. V.

    2017-11-01

    Full Text Available From the point of view of the effectiveness of the top cutting unit, the helix angle in the end portion of the screw is the most important and characteristic parameter, as it determines the pressure of the meat material in the zone of interaction of a knife and grate. The importance of solving the problem of mathematical modeling of geometry is due to the need to address the problem of minimizing the reverse flow of the food material when injecting into the cutting zone, as the specified effect of "locking" significantly reduces the performance of the transfer process, increases energy consumption of the equipment and entails the deterioration of the quality of the raw materials output. The problem of determining the length of the helix variable pitch for screw chopper food materials has been formulated and solved by methods of differential geometry. The task of correct description of the law of changing the angle of helix inclination along its length has been defined in this case as a key to provide the required dependence of this angle tangent on the angle of the radius-vector of the circle. It has been taken into account that the reduction in the pitch of the screw in the direction of the product delivery should occur at a decreasing rate. The parametric equation of the helix has been written in the form of three functional dependencies of the corresponding cylindrical coordinates. Based on the wide range analysis and significant number of models of tops from different manufacturers the boundaries of possible changes in the angles of inclination of the helical line of the first and last turns of the screw have been identified. The auger screw length is determined mathematically in the form of an analytical relationship and both as a function of the variable angle of its rise, and as a function of the rotation angle of the radius-vector of the circle generatrix, which makes it possible to expand the design possibilities of this node. Along

  3. Modified Quad Helix for Correction of Severely Rotated Incisors in Cleft Cases

    Directory of Open Access Journals (Sweden)

    Aneesha Ashok Shetty

    2014-01-01

    Full Text Available Clefts of the lip, alveolus and/or palate are often associated with several dental anomalies, the most common of which are severely rotated maxillary incisors. Patients with such congenital deformities seek orthodontic treatment as early as possible, for an esthetic as well as psychosocial benefit. Here, a new clinical technique is demonstrated for correction of a severely rotated maxillary central incisor which can be carried out in the mixed dentition phase itself, by modifying a commonly used palatal expansion appliance: the quad helix.

  4. Speckle noise suppression using a helix-free ferroelectric liquid crystal cell

    Energy Technology Data Exchange (ETDEWEB)

    Andreev, A L; Andreeva, T B; Kompanets, I N [P N Lebedev Physics Institute, Russian Academy of Sciences, Moscow (Russian Federation); Zalyapin, N V [National Research Nuclear University ' ' MEPhI' ' (Russian Federation)

    2014-12-31

    We have studied the method for suppressing speckle noise in patterns produced by a laser based on a fast-response electro-optical cell with a ferroelectric liquid crystal (FLC) in which helicoid is absent, i.e., compensated for. The character of smectic layer deformation in an electric field is considered along with the mechanism of spatially inhomogeneous phase modulation of a laser beam passing through the cell which is accompanied by the destruction of phase relations in the beam. Advantages of a helix-free FLC cell are pointed out as compared to helical crystal cells studied previously. (liquid crystal devices)

  5. Peran Quadruple Helix dalam Meningkatkan Kreativitas dan Kapabilitas Inovasi (Studi pada Industri Kreatif Sektor Fashion)

    OpenAIRE

    -, Mulyana; -, Sutapa

    2015-01-01

    Creative industries are significantly contribution to economic development, however the development of   creative industries are still many problems, especially human resources, so it has an impact on the innovation capability. This study aims to examine the effect of quadruple helix (intellectual, government, business, civil soceity) to enhance the crativity and innovation capability, on sectors of fashion in Central Java. Sampling methods using purposive sampling and dat...

  6. Kerr effect and Kerr constant enhancement in vertically aligned deformed helix ferroelectric liquid crystals

    Science.gov (United States)

    Shi, Liangyu; Srivastava, Abhishek Kumar; Chigrinov, Vladimir G.; Kwok, Hoi-Sing

    2016-09-01

    In this article, we review recently achieved Kerr effect progress in novel liquid crystal (LC) material: vertically aligned deformed helix ferroelectric liquid crystal (VADHFLC). With an increasing applied electric field, the induced inplane birefringence of LCs shows quadratic nonlinearity. The theoretical calculations and experimental details are illustrated. With an enhanced Kerr constant to 130 nm/V2, this VADHFLC cell can achieve a 2π modulation by a small efficient electric field with a fast response around 100 μs and thus can be employed in both display and photonics devices.

  7. Efecto de las condiciones micro-climáticas en el comportamiento reproductivo del Helix aspersa

    Directory of Open Access Journals (Sweden)

    Hugo Ángel Fernández Cabrera

    2013-01-01

    Full Text Available El objetivo del presente estudio fue evaluar el efecto factores microclimáticos (temperatura y humedad relativa en la reproducción de caracoles de tierra (Helix aspersa. Para esto se acondicionaron cámaras de incubación con cuatro niveles de humedad relativa y de temperatura. Se evaluaron como respuestas la postura, duración de la incubación y porcentaje de eclosión. Se concluye que a 81,0 ± 0,63% de humedad relativa y 23,9 ± 0,39 °C de temperatura se logran los mejores resultados.

  8. A benchmark server using high resolution protein structure data, and benchmark results for membrane helix predictions.

    Science.gov (United States)

    Rath, Emma M; Tessier, Dominique; Campbell, Alexander A; Lee, Hong Ching; Werner, Tim; Salam, Noeris K; Lee, Lawrence K; Church, W Bret

    2013-03-27

    Helical membrane proteins are vital for the interaction of cells with their environment. Predicting the location of membrane helices in protein amino acid sequences provides substantial understanding of their structure and function and identifies membrane proteins in sequenced genomes. Currently there is no comprehensive benchmark tool for evaluating prediction methods, and there is no publication comparing all available prediction tools. Current benchmark literature is outdated, as recently determined membrane protein structures are not included. Current literature is also limited to global assessments, as specialised benchmarks for predicting specific classes of membrane proteins were not previously carried out. We present a benchmark server at http://sydney.edu.au/pharmacy/sbio/software/TMH_benchmark.shtml that uses recent high resolution protein structural data to provide a comprehensive assessment of the accuracy of existing membrane helix prediction methods. The server further allows a user to compare uploaded predictions generated by novel methods, permitting the comparison of these novel methods against all existing methods compared by the server. Benchmark metrics include sensitivity and specificity of predictions for membrane helix location and orientation, and many others. The server allows for customised evaluations such as assessing prediction method performances for specific helical membrane protein subtypes.We report results for custom benchmarks which illustrate how the server may be used for specialised benchmarks. Which prediction method is the best performing method depends on which measure is being benchmarked. The OCTOPUS membrane helix prediction method is consistently one of the highest performing methods across all measures in the benchmarks that we performed. The benchmark server allows general and specialised assessment of existing and novel membrane helix prediction methods. Users can employ this benchmark server to determine the most

  9. Trace metallic elements in Helix aspersa terrestrial snails of a semiarid ecosystem

    International Nuclear Information System (INIS)

    Gaso P, M.I.; Segovia, N.; Zarazua, G.; Montes, F.; Morton, O.; Armienta, M.A.; Hernandez, E.

    2001-01-01

    The concentration of some major elements and traces in soil samples and of Helix aspersa eatable terrestrial snails were analysed at the Radioactive Wastes Storage Center (CADER) and in other reference sites. The methodology includes the use of an atomic absorption spectrophotometer, an X-ray fluorescence equipment and an Icp-mass spectroscope. The concentrations of some toxic elements (Ba, Cd, Cr, Ni, Pb and V) in the soft tissue of the snails were greater than the toxic levels reported in the literature for such trace elements. The snails compared with another wild eatable foods present transfer coefficients soil-snail high relatively. (Author)

  10. The effect of C-terminal helix on the stability of FF domain studied by molecular dynamics simulation.

    Science.gov (United States)

    Zhao, Liling; Cao, Zanxia; Wang, Jihua

    2012-01-01

    To investigate the effect of C-terminal helix on the stability of the FF domain, we studied the native domain FF3-71 from human HYPA/FBP11 and the truncated version FF3-60 with C-terminal helix being deleted by molecular dynamics simulations with GROMACS package and GROMOS 43A1 force field. The results indicated that the structures of truncated version FF3-60 were evident different from those of native partner FF3-71. Compared with FF3-71, the FF3-60 lost some native contacts and exhibited some similar structural characters to those of intermediate state. The C-terminal helix played a major role in stabilizing the FF3-71 domain. To a certain degree, the FF domain had a tendency to form an intermediate state without the C-terminal helix. In our knowledge, this was the first study to examine the role of C-terminal helix of FF domain in detail by molecular dynamics simulations, which was useful to understand the three-state folding mechanism of the small FF domain.

  11. Functional diversity of human basic helix-loop-helix transcription factor TCF4 isoforms generated by alternative 5' exon usage and splicing.

    Directory of Open Access Journals (Sweden)

    Mari Sepp

    Full Text Available BACKGROUND: Transcription factor 4 (TCF4 alias ITF2, E2-2, ME2 or SEF2 is a ubiquitous class A basic helix-loop-helix protein that binds to E-box DNA sequences (CANNTG. While involved in the development and functioning of many different cell types, recent studies point to important roles for TCF4 in the nervous system. Specifically, human TCF4 gene is implicated in susceptibility to schizophrenia and TCF4 haploinsufficiency is the cause of the Pitt-Hopkins mental retardation syndrome. However, the structure, expression and coding potential of the human TCF4 gene have not been described in detail. PRINCIPAL FINDINGS: In the present study we used human tissue samples to characterize human TCF4 gene structure and TCF4 expression at mRNA and protein level. We report that although widely expressed, human TCF4 mRNA expression is particularly high in the brain. We demonstrate that usage of numerous 5' exons of the human TCF4 gene potentially yields in TCF4 protein isoforms with 18 different N-termini. In addition, the diversity of isoforms is increased by alternative splicing of several internal exons. For functional characterization of TCF4 isoforms, we overexpressed individual isoforms in cultured human cells. Our analysis revealed that subcellular distribution of TCF4 isoforms is differentially regulated: Some isoforms contain a bipartite nuclear localization signal and are exclusively nuclear, whereas distribution of other isoforms relies on heterodimerization partners. Furthermore, the ability of different TCF4 isoforms to regulate E-box controlled reporter gene transcription is varied depending on whether one or both of the two TCF4 transcription activation domains are present in the protein. Both TCF4 activation domains are able to activate transcription independently, but act synergistically in combination. CONCLUSIONS: Altogether, in this study we have described the inter-tissue variability of TCF4 expression in human and provided evidence

  12. Temperature, salinity, conductivity, pressure, transmissivity measurements collected using CTD from the Alpha Helix in the Chukchi Sea during 1996 (NODC Accession 0061042)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Temperature, salinity, conductivity, pressure, and transmissivity data gathered by CTD from the Alpha Helix (cruise HX194), September 1996

  13. Therapy-Emergent Drug Resistance to Integrase Strand Transfer Inhibitors in HIV-1 Patients: A Subgroup Meta-Analysis of Clinical Trials.

    Directory of Open Access Journals (Sweden)

    Jiangzhou You

    Full Text Available Integrase strand transfer inhibitors (INSTIs are a novel class of anti-HIV agents that show high activity in inhibiting HIV-1 replication. Currently, licensed INSTIs include raltegravir (RAL, elvitegravir (EVG and dolutegravir (DTG; these drugs have played a critical role in AIDS therapy, serving as additional weapons in the arsenal for treating patients infected with HIV-1. To date, long-term data regarding clinical experience with INSTI use and the emergence of resistance remain scarce. However, the literature is likely now sufficiently comprehensive to warrant a meta-analysis of resistance to INSTIs.Our team implemented a manuscript retrieval protocol using Medical Subject Headings (MeSH via the Web of Science, MEDLINE, EMBASE, and Cochrane Central Register of Controlled Trials databases. We screened the literature based on inclusion and exclusion criteria and then performed a quality analysis and evaluation using RevMan software, Stata software, and the Strengthening the Reporting of Observational Studies in Epidemiology (STROBE. We also performed a subgroup analysis. Finally, we calculated resistance rates and risk ratios (RRs for the three types of drugs.We identified 26 references via the database search. A meta-analysis of the RAL data revealed that the resistance rate was 3.9% (95% CI = 2.9%-4.9% for the selected randomized controlled trials (RCTs. However, the RAL resistance rate reached 40.9% (95% CI = 8.8%-72.9% for the selected observational studies (OBSs. The rates of resistance to RAL that were associated with HIV subtypes A, B, and C as well as with more complex subtypes were 0.1% (95% CI = -0.7%-0.9%, 2.5% (95% CI = 0.5%-4.5%, 4.6% (95% CI = 2.7%-6.6% and 2.2% (95% CI = 0.7%-3.7%, respectively. The rates of resistance to EVG and DTG were 1.2% (95% CI = 0.2%-2.2% and 0.1% (95% CI = -0.2%-0.5%, respectively. Furthermore, we found that the RRs for antiviral resistance were 0.414 (95% CI = 0.210-0.816 between DTG and RAL and 0

  14. Identification of helix capping and β-turn motifs from NMR chemical shifts

    International Nuclear Information System (INIS)

    Shen Yang; Bax, Ad

    2012-01-01

    We present an empirical method for identification of distinct structural motifs in proteins on the basis of experimentally determined backbone and 13 C β chemical shifts. Elements identified include the N-terminal and C-terminal helix capping motifs and five types of β-turns: I, II, I′, II′ and VIII. Using a database of proteins of known structure, the NMR chemical shifts, together with the PDB-extracted amino acid preference of the helix capping and β-turn motifs are used as input data for training an artificial neural network algorithm, which outputs the statistical probability of finding each motif at any given position in the protein. The trained neural networks, contained in the MICS (motif identification from chemical shifts) program, also provide a confidence level for each of their predictions, and values ranging from ca 0.7–0.9 for the Matthews correlation coefficient of its predictions far exceed those attainable by sequence analysis. MICS is anticipated to be useful both in the conventional NMR structure determination process and for enhancing on-going efforts to determine protein structures solely on the basis of chemical shift information, where it can aid in identifying protein database fragments suitable for use in building such structures.

  15. Protein roadblocks and helix discontinuities are barriers to the initiation of mismatch repair

    Science.gov (United States)

    Pluciennik, Anna; Modrich, Paul

    2007-01-01

    The hemimethylated d(GATC) sequence that directs Escherichia coli mismatch repair can reside on either side of a mismatch at a separation distance of 1,000 bp or more. Initiation of repair involves the mismatch-, MutS-, and MutL-dependent activation of MutH endonuclease, which incises the unmethylated strand at the d(GATC) sequence, with the ensuing strand break serving as the loading site for the appropriate 3′-to-5′ or 5′-to-3′ excision system. However, the mechanism responsible for the coordinated recognition of the mismatch and a hemimodified d(GATC) site is uncertain. We show that a protein roadblock (EcoRIE111Q, a hydrolytically defective form of EcoRI endonuclease) placed on the helix between the two DNA sites inhibits MutH activation by 70–80% and that events that escape inhibition are attributable, at least in part, to diffusion of EcoRIE111Q away from its recognition site. We also demonstrate that a double-strand break located within the shorter path linking the mismatch and a d(GATC) site in a circular heteroduplex abolishes MutH activation, whereas a double-strand break within the longer path is without effect. These findings support the idea that initiation of mismatch repair involves signaling along the helix contour. PMID:17620611

  16. Advanced bionics thin lateral and Helix II electrodes: a temporal bone study.

    Science.gov (United States)

    Wright, Charles G; Roland, Peter S; Kuzma, Janusz

    2005-11-01

    This study was performed to evaluate the insertional properties of two cochlear implant electrodes recently developed by Advanced Bionics Corporation. Anatomic study using human cadaveric temporal bones. The electrode prototypes we tested are the Thin Lateral and Helix II arrays, which incorporate features designed to minimize insertional trauma. A total of eight electrodes (4 of each prototype) were evaluated after insertion into freshly fixed temporal bones. The electrodes were inserted by way of standard cochleostomies, and the specimens were subsequently dissected to assess electrode position, insertion depth, and intracochlear trauma. Quantitative data regarding insertion depths and contact distances from the modiolus are presented for all electrodes tested. The mean insertion depths were 368 degrees for the Thin Lateral electrodes, which are designed to approximate the lateral cochlear wall, and 436 degrees for the Helix II electrodes, which occupy a more medial position in the scala tympani. No evidence of insertional trauma was observed with either electrode. The ease of insertion and absence of trauma were confirmed during additional trials in which electrode behavior was directly observed during insertion into previously opened cochleas. Both electrodes performed favorably in our human temporal bone trials, and both arrays appear promising for clinical use, especially in patients with residual hearing in whom atraumatic insertion is an important objective.

  17. A triple-helix forming oligonucleotide targeting genomic DNA fails to induce mutation.

    Science.gov (United States)

    Reshat, Reshat; Priestley, Catherine C; Gooderham, Nigel J

    2012-11-01

    Purine tracts in duplex DNA can bind oligonucleotide strands in a sequence specific manner to form triple-helix structures. Triple-helix forming oligonucleotides (TFOs) targeting supFG1 constructs have previously been shown to be mutagenic raising safety concerns for oligonucleotide-based pharmaceuticals. We have engineered a TFO, TFO27, to target the genomic Hypoxanthine-guanine phosphoribosyltransferase (HPRT) locus to define the mutagenic potential of such structures at genomic DNA. We report that TFO27 was resistant to nuclease degradation and readily binds to its target motif in a cell free system. Contrary to previous studies using the supFG1 reporter construct, TFO27 failed to induce mutation within the genomic HPRT locus. We suggest that it is possible that previous reports of triplex-mediated mutation using the supFG1 reporter construct could be confounded by DNA quadruplex formation. Although the present study indicates that a TFO targeting a genomic locus lacks mutagenic activity, it is unclear if this finding can be generalised to all TFOs and their targets. For the present, we suggest that it is prudent to avoid large purine stretches in oligonucleotide pharmaceutical design to minimise concern regarding off-target genotoxicity.

  18. Antibacterial Activity of Silver Nanoparticles Synthesized by Using Extracts of Hedera helix

    Directory of Open Access Journals (Sweden)

    Ahmadreza Abbasifar

    2017-01-01

    Full Text Available Background Silver nanoparticles (AgNPs are one of the most widely applicable particles whose application is increasing in Nano world daily. Silver nanoparticles have expressed significant advances owing to wide range of applications in the field of bio-medical, sensors, antimicrobials, catalysts, electronics, optical fibers, agricultural, bio-labeling and the other areas. Green synthesis is the safe and easiest method of producing silver nanoparticles. Because of the production of the silver ions, silver nanoparticles are found to have the antibacterial activity. Objectives The aim of this study was to investigate antibacterial activity of silver nanoparticles synthesized by using extracts of Hedera helix against Bacillus subtilis and Klebsiella pneumoniae. Methods In this experimental study AgNPs were prepared by the reaction of 1mM silver nitrate and extracts of Hedera helix. Antibacterial activity of AgNPs was assessed by using disc diffusion method against Bacillus subtilis and Klebsiella pneumoniae. The AgNPs were characterized by UV-visible (vis spectrophotometer, particle size analyzer by dynamic light scattering (DLS method, transmission electron microscopy (TEM. Results AgNPs obtained showed significantly higher antimicrobial activities against B. subtilis and K. pneumonia in comparison to both AgNO3 and raw plant extracts. Conclusions Biological methods are a good competent for the chemical procedures, which are environment friendly and convenient.

  19. One-electron second-order optical activity of a helix

    Science.gov (United States)

    Maki, Jeffery J.; Persoons, André

    1996-06-01

    The second-order nonlinear-optical response of a chiral molecule is calculated. We model the optical response classically using a single electron bound to a helical path. The helical motion of the electron causes optical activity in the second-order response. The hyperpolarizability tensor of a single helix and the susceptibility tensor for a thin film of helices are given. We examine the process of second-harmonic generation from a chiral surface using the calculated susceptibility tensor. The efficiency of the harmonic generation is different for left- and right-hand circularly polarized fundamental light, which is ascribed to be a form of nonlinear optical activity. The roles of pitch and radius of the helix are readily seen in the microscopic and macroscopic second-order optical responses and in the surface second-harmonic generation, which may provide some insight for synthesizing new chiral compounds. Our results also allow us to draw conclusions about the relative strength and importance to second-order optical activity of electric- and magnetic-dipole transitions. For instance, we confirm that optical activity can occur in surface second-harmonic generation from electric-only response, but we find that magnetic response can make a similar contribution and thus should not be ignored.

  20. Stakeholder engagement in quattro helix model for mobile phone reverse logistics in Indonesia: a conceptual framework

    Science.gov (United States)

    Maheswari, H.; Yudoko, G.; Adhiutama, A.

    2017-12-01

    The number of e-waste from mobile phone industry is still dominating until now. This is happened because there is no mutual commitment from all of parties i.e. businesses, government, and societies to reduce the use of mobile phone that has the shortest product life cycle. There are many researches study about firms’ motivation and government’s role, other discuss about actions of communities in supporting reverse logistics implementation. Unfortunately, research about engagement mechanism that involving all parties is still rare. Therefore, it is important to find the engagement model through this conceptual paper and it is expected useful to build the novel model. Through literature review, the results of this research are establishing the Quattro helix model as the appropriate structure to build the robust team by exploring stakeholder theories; mapping the engagement model either in form of collaboration or participation that consider stakeholders’ role and motivation and finding six types of engagement that consider their interest; and determining the novel model of engagement through Quattro helix model for implementing reverse logistics in handling e-waste by describing the linkage and the gaps among existing model.

  1. Adjustment of metabolite composition in the haemolymph to seasonal variations in the land snail Helix pomatia.

    Science.gov (United States)

    Nicolai, Annegret; Filser, Juliane; Lenz, Roman; Bertrand, Carole; Charrier, Maryvonne

    2011-05-01

    In temperate regions, land snails are subjected to subzero temperatures in winter and hot temperatures often associated to drought in summer. The response to these environmental factors is usually a state of inactivity, hibernation and aestivation, respectively, in a temperature and humidity buffered refuge, accompanied by physiological adjustments to resist cold or heat stress. We investigated how environmental factors in the microhabitat and body condition influence the metabolite composition of haemolymph of the endangered species Helix pomatia. We used UPLC and GC-MS techniques and analyzed annual biochemical variations in a multivariate model. Hibernation and activity months differed in metabolite composition. Snails used photoperiod as cue for seasonal climatic variations to initiate a physiological state and were also highly sensitive to temperature variations, therefore constantly adjusting their physiological processes. Galactose levels gave evidence for the persistence of metabolic activity with energy expenditure during hibernation and for high reproductive activity in June. Triglycerides accumulated prior to hibernation might act as cryoprotectants or energy reserves. During the last month of hibernation snails activated physiological processes related to arousal. During activity, protein metabolism was reflected by high amino acid level. An exceptional aestivation period was observed in April giving evidence for heat stress responses, like the protection of cells from dehydration by polyols and saccharides, the membrane stabilization by cholesterol and enhanced metabolism using the anaerobic succinic acid pathway to sustain costly stress responses. In conclusion, physiological adjustments to environmental variations in Helix pomatia involve water loss regulation, cryoprotectant or heatprotectant accumulation.

  2. Physical model of voltage sensing in sodium channels based on the sliding helix complex

    Science.gov (United States)

    Chancey, C. C.; George, S. A.

    1996-05-01

    We have modeled voltage sensing in the sodium channel by evaluating forces on the S4 α-helix portion of the channel molecule, which we assume moves outward during activation. The interaction between the S4 α-helix segment and its environment was modeled by (i) nearest-neighbor Coulombic forces, (ii) the electric force due to an external electric field, and (iii) static mechanical and electrostatic forces. These terms collectively describe a depolarization-dependent effective potential within which the S4 segment moves. Thermal transitions between center-of-mass energy states of the segment were modeled starting from the Boltzmann distribution, and the time evolution of the segment's position relative to the membrane was simulated. Combining the histories of four such processes models the activation history of the channel molecule. The model simulation is in good qualitative agreement with batrachotoxin-modified single channel open and closed dwell time distributions and with such a channel's open probability as a function of depolarization. The model also qualitatively agrees with site-specific mutagenesis experiments, which show the different effects of eliminating positive charges on the cytoplasmic and extracellular ends of the S4 segment.

  3. Identification of helix capping and {beta}-turn motifs from NMR chemical shifts

    Energy Technology Data Exchange (ETDEWEB)

    Shen Yang; Bax, Ad, E-mail: bax@nih.gov [National Institutes of Health, Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases (United States)

    2012-03-15

    We present an empirical method for identification of distinct structural motifs in proteins on the basis of experimentally determined backbone and {sup 13}C{sup {beta}} chemical shifts. Elements identified include the N-terminal and C-terminal helix capping motifs and five types of {beta}-turns: I, II, I Prime , II Prime and VIII. Using a database of proteins of known structure, the NMR chemical shifts, together with the PDB-extracted amino acid preference of the helix capping and {beta}-turn motifs are used as input data for training an artificial neural network algorithm, which outputs the statistical probability of finding each motif at any given position in the protein. The trained neural networks, contained in the MICS (motif identification from chemical shifts) program, also provide a confidence level for each of their predictions, and values ranging from ca 0.7-0.9 for the Matthews correlation coefficient of its predictions far exceed those attainable by sequence analysis. MICS is anticipated to be useful both in the conventional NMR structure determination process and for enhancing on-going efforts to determine protein structures solely on the basis of chemical shift information, where it can aid in identifying protein database fragments suitable for use in building such structures.

  4. Detergent properties influence the stability of the glycophorin A transmembrane helix dimer in lysophosphatidylcholine micelles.

    Science.gov (United States)

    Stangl, Michael; Veerappan, Anbazhagan; Kroeger, Anja; Vogel, Peter; Schneider, Dirk

    2012-12-19

    Detergents might affect membrane protein structures by promoting intramolecular interactions that are different from those found in native membrane bilayers, and fine-tuning detergent properties can be crucial for obtaining structural information of intact and functional transmembrane proteins. To systematically investigate the influence of the detergent concentration and acyl-chain length on the stability of a transmembrane protein structure, the stability of the human glycophorin A transmembrane helix dimer has been analyzed in lyso-phosphatidylcholine micelles of different acyl-chain length. While our results indicate that the transmembrane protein is destabilized in detergents with increasing chain-length, the diameter of the hydrophobic micelle core was found to be less crucial. Thus, hydrophobic mismatch appears to be less important in detergent micelles than in lipid bilayers and individual detergent molecules appear to be able to stretch within a micelle to match the hydrophobic thickness of the peptide. However, the stability of the GpA TM helix dimer linearly depends on the aggregation number of the lyso-PC detergents, indicating that not only is the chemistry of the detergent headgroup and acyl-chain region central for classifying a detergent as harsh or mild, but the detergent aggregation number might also be important. Copyright © 2012 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  5. Helix unfolding/refolding characterizes the functional dynamics of Staphylococcus aureus Clp protease.

    Science.gov (United States)

    Ye, Fei; Zhang, Jie; Liu, Hongchuan; Hilgenfeld, Rolf; Zhang, Ruihan; Kong, Xiangqian; Li, Lianchun; Lu, Junyan; Zhang, Xinlei; Li, Donghai; Jiang, Hualiang; Yang, Cai-Guang; Luo, Cheng

    2013-06-14

    The ATP-dependent Clp protease (ClpP) plays an essential role not only in the control of protein quality but also in the regulation of bacterial pathogen virulence, making it an attractive target for antibacterial treatment. We have previously determined the crystal structures of Staphylococcus aureus ClpP (SaClpP) in two different states, extended and compressed. To investigate the dynamic switching of ClpP between these states, we performed a series of molecular dynamics simulations. During the structural transition, the long and straight helix E in the extended SaClpP monomer underwent an unfolding/refolding process, resulting in a kinked helix very similar to that in the compressed monomer. As a stable intermediate in the molecular dynamics simulation, the compact state was suggested and subsequently identified in x-ray crystallographic experiment. Our combined studies also determined that Ala(140) acted as a "hinge" during the transition between the extended and compressed states, and Glu(137) was essential for stabilizing the compressed state. Overall, this study provides molecular insights into the dynamics and mechanism of the functional conformation changes of SaClpP. Given the highly conserved sequences of ClpP proteins among different species, these findings potentially reflect a switching mechanism for the dynamic process shared in the whole ClpP family in general and thus aid in better understand the principles of Clp protease assembly and function.

  6. Opposite Displacement of Helix F in Attractant and Repellent Signaling by Sensory Rhodopsin-Htr Complexes*

    Science.gov (United States)

    Sasaki, Jun; Tsai, Ah-lim; Spudich, John L.

    2011-01-01

    Two forms of the phototaxis receptor sensory rhodopsin I distinguished by differences in its photoactive site have been shown to be directly correlated with attractant and repellent signaling by the dual-signaling protein. In prior studies, differences in the photoactive site defined the two forms, namely the direction of light-induced proton transfer from the chromophore and the pKa of an Asp counterion to the protonated chromophore. Here, we show by both in vivo and in vitro measurements that the two forms are distinct protein conformers with structural similarities to two conformers seen in the light-driven proton transport cycle of the related protein bacteriorhodopsin. Measurements of spontaneous cell motility reversal frequencies, an in vivo measure of histidine kinase activity in the phototaxis system, indicate that the two forms are a photointerconvertible pair, with one conformer activating and the other inhibiting the kinase. Protein conformational changes in these photoconversions monitored by site-directed spin labeling show that opposite structural changes in helix F, distant from the photoactive site, correspond to the opposite phototaxis signals. The results provide the first direct evidence that displacements of helix F are directly correlated with signaling and impact our understanding of the sensory rhodopsin I signaling mechanism and the evolution of diverse functionality in this protein family. PMID:21454480

  7. Opposite displacement of helix F in attractant and repellent signaling by sensory rhodopsin-Htr complexes.

    Science.gov (United States)

    Sasaki, Jun; Tsai, Ah-lim; Spudich, John L

    2011-05-27

    Two forms of the phototaxis receptor sensory rhodopsin I distinguished by differences in its photoactive site have been shown to be directly correlated with attractant and repellent signaling by the dual-signaling protein. In prior studies, differences in the photoactive site defined the two forms, namely the direction of light-induced proton transfer from the chromophore and the pK(a) of an Asp counterion to the protonated chromophore. Here, we show by both in vivo and in vitro measurements that the two forms are distinct protein conformers with structural similarities to two conformers seen in the light-driven proton transport cycle of the related protein bacteriorhodopsin. Measurements of spontaneous cell motility reversal frequencies, an in vivo measure of histidine kinase activity in the phototaxis system, indicate that the two forms are a photointerconvertible pair, with one conformer activating and the other inhibiting the kinase. Protein conformational changes in these photoconversions monitored by site-directed spin labeling show that opposite structural changes in helix F, distant from the photoactive site, correspond to the opposite phototaxis signals. The results provide the first direct evidence that displacements of helix F are directly correlated with signaling and impact our understanding of the sensory rhodopsin I signaling mechanism and the evolution of diverse functionality in this protein family.

  8. Mechanical evaluation of quad-helix appliance made of low-nickel stainless steel wire

    Directory of Open Access Journals (Sweden)

    Rogério Lacerda dos Santos

    2013-06-01

    Full Text Available OBJECTIVE: The objective of this study was to test the hypothesis that there is no difference between stainless steel and low-nickel stainless steel wires as regards mechanical behavior. Force, resilience, and elastic modulus produced by Quad-helix appliances made of 0.032-inch and 0.036-inch wires were evaluated. METHODS: Sixty Quad-helix appliances were made, thirty for each type of alloy, being fifteen for each wire thickness, 0.032-in and 0.036-in. All the archwires were submitted to mechanical compression test using an EMIC DL-10000 machine simulating activations of 4, 6, 9, and 12 mm. Analysis of variance (ANOVA with multiple comparisons and Tukey's test were used (p < 0.05 to assess force, resilience, and elastic modulus. RESULTS: Statistically significant difference in the forces generated, resilience and elastic modulus were found between the 0.032-in 0.036-in thicknesses (p < 0.05. CONCLUSIONS: Appliances made of low-nickel stainless steel alloy had force, resilience, and elastic modulus similar to those made of stainless steel alloy.

  9. Hierarchical cascades of instability govern the mechanics of coiled coils: helix unfolding precedes coil unzipping.

    Science.gov (United States)

    Hamed, Elham; Keten, Sinan

    2014-07-15

    Coiled coils are a fundamental emergent motif in proteins found in structural biomaterials, consisting of α-helical secondary structures wrapped in a supercoil. A fundamental question regarding the thermal and mechanical stability of coiled coils in extreme environments is the sequence of events leading to the disassembly of individual oligomers from the universal coiled-coil motifs. To shed light on this phenomenon, here we report atomistic simulations of a trimeric coiled coil in an explicit water solvent and investigate the mechanisms underlying helix unfolding and coil unzipping in the assembly. We employ advanced sampling techniques involving steered molecular dynamics and metadynamics simulations to obtain the free-energy landscapes of single-strand unfolding and unzipping in a three-stranded assembly. Our comparative analysis of the free-energy landscapes of instability pathways shows that coil unzipping is a sequential process involving multiple intermediates. At each intermediate state, one heptad repeat of the coiled coil first unfolds and then unzips due to the loss of contacts with the hydrophobic core. This observation suggests that helix unfolding facilitates the initiation of coiled-coil disassembly, which is confirmed by our 2D metadynamics simulations showing that unzipping of one strand requires less energy in the unfolded state compared with the folded state. Our results explain recent experimental findings and lay the groundwork for studying the hierarchical molecular mechanisms that underpin the thermomechanical stability/instability of coiled coils and similar protein assemblies. Copyright © 2014 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  10. Poly(A) Tail Recognition by a Viral RNA Element Through Assembly of a Triple Helix

    Energy Technology Data Exchange (ETDEWEB)

    M Mitton-Fry; S DeGregorio; J Wang; T Steitz; J Steitz

    2011-12-31

    Kaposi's sarcoma-associated herpesvirus produces a highly abundant, nuclear noncoding RNA, polyadenylated nuclear (PAN) RNA, which contains an element that prevents its decay. The 79-nucleotide expression and nuclear retention element (ENE) was proposed to adopt a secondary structure like that of a box H/ACA small nucleolar RNA (snoRNA), with a U-rich internal loop that hybridizes to and protects the PAN RNA poly(A) tail. The crystal structure of a complex between the 40-nucleotide ENE core and oligo(A){sub 9} RNA at 2.5 angstrom resolution reveals that unlike snoRNAs, the U-rich loop of the ENE engages its target through formation of a major-groove triple helix. A-minor interactions extend the binding interface. Deadenylation assays confirm the functional importance of the triple helix. Thus, the ENE acts as an intramolecular RNA clamp, sequestering the PAN poly(A) tail and preventing the initiation of RNA decay.

  11. A "bulged" double helix in a RNA-protein contact site

    DEFF Research Database (Denmark)

    Peattie, D A; Douthwaite, S; Garrett, R A

    1981-01-01

    The binding of ribosomal protein L18 affects specific nucleotides in Escherichia coli 5S RNA as detected by dimethyl sulfate alkylation and RNase A digestion of the 5S-L18 complex. Most of the affected nucleotides are clustered and localize a site of RNA-protein interaction in and around...... the defined central helix [Fox, G. E. & Woese, C. (1975) Nature (London) 256, 505-507] of 5S RNA. Chemical carbethoxylation of the native 5S RNA with diethyl pyrocarbonate shows that a striking feature of this region is an unstacked adenosine residue at position 66. We propose that this residue exists...... as a singly bulged nucleotide extending the Fox and Woese central helix by two base pairs in the E. coli sequence (to positions 16-23/60-68) as well as in each of 61 (prokaryotic and eukaryotic) aligned 5S RNA sequences. In each case, the single bulged nucleotide is at the relative position of adenosine-66...

  12. Nuclear receptor ligand-binding domains: reduction of helix H12 dynamics to favour crystallization

    Energy Technology Data Exchange (ETDEWEB)

    Nahoum, Virginie; Lipski, Alexandra; Quillard, Fabien; Guichou, Jean-François [INSERM, U554, 34090 Montpellier (France); Université de Montpellier, CNRS, UMR5048, Centre de Biochimie Structurale (CBS), 34090 Montpellier (France); Boublik, Yvan [CNRS, UMR5237, Centre de Recherche de Biochimie Macromoléculaire (CRBM), 34293 Montpellier (France); Pérez, Efrèn [Universidade de Vigo, Departamento de Quimica Organica, Facultad de Química, 36310 Vigo (Spain); Germain, Pierre [Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), BP 10142, 67404 Illkirch CEDEX (France); Lera, Angel R. de [Universidade de Vigo, Departamento de Quimica Organica, Facultad de Química, 36310 Vigo (Spain); Bourguet, William, E-mail: bourguet@cbs.cnrs.fr [INSERM, U554, 34090 Montpellier (France); Université de Montpellier, CNRS, UMR5048, Centre de Biochimie Structurale (CBS), 34090 Montpellier (France)

    2008-07-01

    Attempts have been made to crystallize the ligand-binding domain of the human retinoid X receptor in complex with a variety of newly synthesized ligands. An inverse correlation was observed between the ‘crystallizability’ and the structural dynamics of the various receptor–ligand complexes. Crystallization trials of the human retinoid X receptor α ligand-binding domain (RXRα LBD) in complex with various ligands have been carried out. Using fluorescence anisotropy, it has been found that when compared with agonists these small-molecule effectors enhance the dynamics of the RXRα LBD C-terminal helix H12. In some cases, the mobility of this helix could be dramatically reduced by the addition of a 13-residue co-activator fragment (CoA). In keeping with these observations, crystals have been obtained of the corresponding ternary RXRα LBD–ligand–CoA complexes. In contrast, attempts to crystallize complexes with a highly mobile H12 remained unsuccessful. These experimental observations substantiate the previously recognized role of co-regulator fragments in facilitating the crystallization of nuclear receptor LBDs.

  13. Role of the future creative universities in the triple helix of science and technology corridors

    Directory of Open Access Journals (Sweden)

    Iraj nabipour

    2015-01-01

    Full Text Available The science and technology corridor is a complex cluster containing universities, science parks, research centers, high-tech companies, venture capital, institutional and physical infrastructures, and human capital in a defined geography with its unique management and legal structure in association with the business space and knowledge-based products. In fact, the science and technology corridor reflects the concept of development based on the knowledge region (the especial region for science and technology. The knowledge region is clearly a triple helix phenomenon par excellence: universities, governments and businesses combine their efforts to construct a common advantage which they would not be able to offer on their own. The future creative universities in connection with the knowledge city-regions not only will deal with innovation and entrepreneurial training but also produce a competitive, vibrant environment with high indices for quality of life and full of green technologies. In this article, we will present functional interactions of the creative universities in the triple helix, particularly the missions for the Iranian universities of medical sciences. As a theoretical model, the complex interactions of Bushehr University of Medical Sciences and Health Services with Bushehr Science and Technology Corridor will be discussed.

  14. Deer mouse hemoglobin exhibits a lowered oxygen affinity owing to mobility of the E helix

    Science.gov (United States)

    Inoguchi, Noriko; Oshlo, Jake R.; Natarajan, Chandrasekhar; Weber, Roy E.; Fago, Angela; Storz, Jay F.; Moriyama, Hideaki

    2013-01-01

    The deer mouse, Peromyscus maniculatus, exhibits altitude-associated variation in hemoglobin oxygen affinity. To examine the structural basis of this functional variation, the structure of the hemoglobin was solved. Recombinant hemoglobin was expressed in Escherichia coli and was purified by ion-exchange chromatography. Recombinant hemoglobin was crystallized by the hanging-drop vapor-diffusion method using polyethylene glycol as a precipitant. The obtained orthorhombic crystal contained two subunits in the asymmetric unit. The refined structure was interpreted as the aquo-met form. Structural comparisons were performed among hemoglobins from deer mouse, house mouse and human. In contrast to human hemoglobin, deer mouse hemoglobin lacks the hydrogen bond between α1Trp14 in the A helix and α1Thr67 in the E helix owing to the Thr67Ala substitution. In addition, deer mouse hemoglobin has a unique hydrogen bond at the α1β1 interface between residues α1Cys34 and β1Ser128. PMID:23545644

  15. ELISA for Determination of Human Growth Hormone: Recognition of Helix 4 Epitopes

    Directory of Open Access Journals (Sweden)

    Moura Juliana F.

    2004-01-01

    Full Text Available Human growth hormone (hGH signal transduction initiates with a receptor dimerization in which one molecule binds to the receptor through sites 1 and 2. A sandwich enzyme-linked immunosorbent assay was developed for quantifying hGH molecules that present helix 4 from binding site 1. For this, horse anti-rhGH antibodies were eluted by an immunoaffinity column constituted by sepharose-rhGH. These antibodies were purified through a second column with synthetic peptide correspondent to hGH helix 4, immobilized to sepharose, and used as capture antibodies. Those that did not recognize synthetic peptide were used as a marker antibody. The working range was of 1.95 to 31.25 ng/mL of hGH. The intra-assay coefficient of variation (CV was between 4.53% and 6.33%, while the interassay CV was between 6.00% and 8.27%. The recovery range was between 96.0% to 103.8%. There was no cross-reactivity with human prolactin. These features show that our assay is an efficient method for the determination of hGH.

  16. The new Audi A4; Der neue Audi A4

    Energy Technology Data Exchange (ETDEWEB)

    Basshuysen, R. van [ed.; Siebenpfeiffer, W. [comp.

    2000-11-01

    Details of the new Audi A4 are presented. The new car has a c{sub w} value of 0.28 in spite of a serial air conditioning system and a 'multitronic' control system for the 2.5 l V6 TDI engine with a rotary momentum of up to 310 Nm. A number of engines are available including a new spark ignition engine with a cubic capacity of 2 l and a five-valve technology. The high comfort of the car is illustrated by the examples of the voice-operated car phone and telematic system. Electrionic communication systems and infotainment are given high priority. The front window pane is heated by a 42 V heating system in order to improve the driving safety, and the air conditioning system was improved by two-zone contorl. This special issue of ATZ presents further details. [German] Mit der Weiterentwicklung des A4 hat Audi erneut seine Bereitschaft zum harten Wettbewerb im Segment der Mittelklasse signalisiert. Um die Kundenbeduerfnisse hinsichtlich sportlicher Dynamik und Komfort zu erfuellen, sind den Audi-Ingenieuren in vielen Details Loesungen gelungen, die den neuen A4 unverwechselbar machen. Hinter dem weiterentwickelten Audi-typischen Styling stecken innovative technische Entwicklungen. Die neu konstruierte Trapezlenker-Hinterachse beispielsweise verbessert die Fahreigenschaften. Ein c{sub w}-Wert von 0,28 bei den Basismotorisierungen - trotz serienmaessiger Klimaanlage - ist das Ergebnis umfangreicher Massnahmen bei der Aerodynamik. Die als vorbildlich geltende 'multitronic' ist nun auch fuer den 2,5-l-V6-TDI-Motor erhaeltlich und fuer ein Drehmoment von bis zu 310 Nm ausgelegt. Die Auswahl der Motoren - darunter ein neuer Ottomotor mit 2 l Hubraum und Fuenfventiltechnik - laesst kaum Wuensche offen. Was die Komfortausstattung betrifft, zeigt sich Audi mit dem neuen A4 wieder einmal als Trendsetter. Der Einsatz eines Sprachbediensystems fuer das Autotelefon und die angebotene Telematic sind Beispiele dafuer. Die gesamte Kommunikationselektronik und das

  17. The Penta Helix Model of Innovation in Oman: An HEI Perspective

    Directory of Open Access Journals (Sweden)

    Alrence S Halibas

    2017-05-01

    Full Text Available Aim/Purpose: Countries today strategically pursue regional development and economic diversification to compete in the world market. Higher Education Institutions (HEIs are at the crux of this political strategy. The paper reviews how HEIs can propel regional socio-economic growth and development by way of research innovation and entrepreneurship. Background: Offering an academic perspective about the role of HEIs using the Penta Helix innovation network for business and social innovation, the paper discusses opportunities and challenges in gestating an innovation culture. It likewise seeks, identifies and details strategies and workable programs. Methodology: Best-practice innovation campaigns initiated by Omani HEIs in collaboration with capstone programs organized by the government were parsed from selected local and international literature. The study includes a causal analysis of innovation information contained in 40 out of 44 published OAAA Quality Audit reports about HEIs from 2009 to 2016. The best-practice programs serve as success indicators and will be used as a field metric effect a Penta Helix blueprint for innovation. Contribution: The paper discusses how HEIs can engender, nurture, drive, and sustain innovation and entrepreneurial activity by using an innovation strategic blueprint like the Penta Helix model. It gathers together the recent historical attempts at promoting innovation by HEIs. It likewise suggests the creation of a network channel to allow key players in the innovation network to share innovation information and to collaborate with each other. Furthermore, it contributes to the development of innovation culture in HEIs. Findings: Expectations run high in academia. For one, universities believe that all innovations embryonically begin within their halls. Universities–too–believe it is naturally incumbent on them to stimulate and advance innovation despite that most innovation programs are initiated by the

  18. Human immunodeficiency virus integrase inhibitors efficiently suppress feline immunodeficiency virus replication in vitro and provide a rationale to redesign antiretroviral treatment for feline AIDS

    Science.gov (United States)

    Savarino, Andrea; Pistello, Mauro; D'Ostilio, Daniela; Zabogli, Elisa; Taglia, Fabiana; Mancini, Fabiola; Ferro, Stefania; Matteucci, Donatella; De Luca, Laura; Barreca, Maria Letizia; Ciervo, Alessandra; Chimirri, Alba; Ciccozzi, Massimo; Bendinelli, Mauro

    2007-01-01

    Background Treatment of feline immunodeficiency virus (FIV) infection has been hampered by the absence of a specific combination antiretroviral treatment (ART). Integrase strand transfer inhibitors (INSTIs) are emerging as a promising new drug class for HIV-1 treatment, and we evaluated the possibility of inhibiting FIV replication using INSTIs. Methods Phylogenetic analysis of lentiviral integrase (IN) sequences was carried out using the PAUP* software. A theoretical three-dimensional structure of the FIV IN catalytic core domain (CCD) was obtained by homology modeling based on a crystal structure of HIV-1 IN CCD. The interaction of the transferred strand of viral DNA with the catalytic cavity of FIV IN was deduced from a crystal structure of a structurally similar transposase complexed with transposable DNA. Molecular docking simulations were conducted using a genetic algorithm (GOLD). Antiviral activity was tested in feline lymphoblastoid MBM cells acutely infected with the FIV Petaluma strain. Circular and total proviral DNA was quantified by real-time PCR. Results The calculated INSTI-binding sites were found to be nearly identical in FIV and HIV-1 IN CCDs. The close similarity of primate and feline lentivirus IN CCDs was also supported by phylogenetic analysis. In line with these bioinformatic analyses, FIV replication was efficiently inhibited in acutely infected cell cultures by three investigational INSTIs, designed for HIV-1 and belonging to different classes. Of note, the naphthyridine carboxamide INSTI, L-870,810 displayed an EC50 in the low nanomolar range. Inhibition of FIV integration in situ was shown by real-time PCR experiments that revealed accumulation of circular forms of FIV DNA within cells treated with L-870,810. Conclusion We report a drug class (other than nucleosidic reverse transcriptase inhibitors) that is capable of inhibiting FIV replication in vitro. The present study helped establish L-870,810, a compound successfully tested in

  19. Human immunodeficiency virus integrase inhibitors efficiently suppress feline immunodeficiency virus replication in vitro and provide a rationale to redesign antiretroviral treatment for feline AIDS

    Directory of Open Access Journals (Sweden)

    Ciervo Alessandra

    2007-10-01

    Full Text Available Abstract Background Treatment of feline immunodeficiency virus (FIV infection has been hampered by the absence of a specific combination antiretroviral treatment (ART. Integrase strand transfer inhibitors (INSTIs are emerging as a promising new drug class for HIV-1 treatment, and we evaluated the possibility of inhibiting FIV replication using INSTIs. Methods Phylogenetic analysis of lentiviral integrase (IN sequences was carried out using the PAUP* software. A theoretical three-dimensional structure of the FIV IN catalytic core domain (CCD was obtained by homology modeling based on a crystal structure of HIV-1 IN CCD. The interaction of the transferred strand of viral DNA with the catalytic cavity of FIV IN was deduced from a crystal structure of a structurally similar transposase complexed with transposable DNA. Molecular docking simulations were conducted using a genetic algorithm (GOLD. Antiviral activity was tested in feline lymphoblastoid MBM cells acutely infected with the FIV Petaluma strain. Circular and total proviral DNA was quantified by real-time PCR. Results The calculated INSTI-binding sites were found to be nearly identical in FIV and HIV-1 IN CCDs. The close similarity of primate and feline lentivirus IN CCDs was also supported by phylogenetic analysis. In line with these bioinformatic analyses, FIV replication was efficiently inhibited in acutely infected cell cultures by three investigational INSTIs, designed for HIV-1 and belonging to different classes. Of note, the naphthyridine carboxamide INSTI, L-870,810 displayed an EC50 in the low nanomolar range. Inhibition of FIV integration in situ was shown by real-time PCR experiments that revealed accumulation of circular forms of FIV DNA within cells treated with L-870,810. Conclusion We report a drug class (other than nucleosidic reverse transcriptase inhibitors that is capable of inhibiting FIV replication in vitro. The present study helped establish L-870,810, a compound

  20. Contribution of the C-terminal region within the catalytic core domain of HIV-1 integrase to yeast lethality, chromatin binding and viral replication.

    Science.gov (United States)

    Xu, Zaikun; Zheng, Yingfeng; Ao, Zhujun; Clement, Martin; Mouland, Andrew J; Kalpana, Ganjam V; Belhumeur, Pierre; Cohen, Eric A; Yao, Xiaojian

    2008-11-14

    HIV-1 integrase (IN) is a key viral enzymatic molecule required for the integration of the viral cDNA into the genome. Additionally, HIV-1 IN has been shown to play important roles in several other steps during the viral life cycle, including reverse transcription, nuclear import and chromatin targeting. Interestingly, previous studies have demonstrated that the expression of HIV-1 IN induces the lethal phenotype in some strains of Saccharomyces cerevisiae. In this study, we performed mutagenic analyses of the C-terminal region of the catalytic core domain of HIV-1 IN in order to delineate the critical amino acid(s) and/or motif(s) required for the induction of the lethal phenotype in the yeast strain HP16, and to further elucidate the molecular mechanism which causes this phenotype. Our study identified three HIV-1 IN mutants, V165A, A179P and KR186,7AA, located in the C-terminal region of the catalytic core domain of IN that do not induce the lethal phenotype in yeast. Chromatin binding assays in yeast and mammalian cells demonstrated that these IN mutants were impaired for the ability to bind chromatin. Additionally, we determined that while these IN mutants failed to interact with LEDGF/p75, they retained the ability to bind Integrase interactor 1. Furthermore, we observed that VSV-G-pseudotyped HIV-1 containing these IN mutants was unable to replicate in the C8166 T cell line and this defect was partially rescued by complementation with the catalytically inactive D64E IN mutant. Overall, this study demonstrates that three mutations located in the C-terminal region of the catalytic core domain of HIV-1 IN inhibit the IN-induced lethal phenotype in yeast by inhibiting the binding of IN to the host chromatin. These results demonstrate that the C-terminal region of the catalytic core domain of HIV-1 IN is important for binding to host chromatin and is crucial for both viral replication and the promotion of the IN-induced lethal phenotype in yeast.

  1. Contribution of the C-terminal region within the catalytic core domain of HIV-1 integrase to yeast lethality, chromatin binding and viral replication

    Directory of Open Access Journals (Sweden)

    Belhumeur Pierre

    2008-11-01

    Full Text Available Abstract Background HIV-1 integrase (IN is a key viral enzymatic molecule required for the integration of the viral cDNA into the genome. Additionally, HIV-1 IN has been shown to play important roles in several other steps during the viral life cycle, including reverse transcription, nuclear import and chromatin targeting. Interestingly, previous studies have demonstrated that the expression of HIV-1 IN induces the lethal phenotype in some strains of Saccharomyces cerevisiae. In this study, we performed mutagenic analyses of the C-terminal region of the catalytic core domain of HIV-1 IN in order to delineate the critical amino acid(s and/or motif(s required for the induction of the lethal phenotype in the yeast strain HP16, and to further elucidate the molecular mechanism which causes this phenotype. Results Our study identified three HIV-1 IN mutants, V165A, A179P and KR186,7AA, located in the C-terminal region of the catalytic core domain of IN that do not induce the lethal phenotype in yeast. Chromatin binding assays in yeast and mammalian cells demonstrated that these IN mutants were impaired for the ability to bind chromatin. Additionally, we determined that while these IN mutants failed to interact with LEDGF/p75, they retained the ability to bind Integrase interactor 1. Furthermore, we observed that VSV-G-pseudotyped HIV-1 containing these IN mutants was unable to replicate in the C8166 T cell line and this defect was partially rescued by complementation with the catalytically inactive D64E IN mutant. Conclusion Overall, this study demonstrates that three mutations located in the C-terminal region of the catalytic core domain of HIV-1 IN inhibit the IN-induced lethal phenotype in yeast by inhibiting the binding of IN to the host chromatin. These results demonstrate that the C-terminal region of the catalytic core domain of HIV-1 IN is important for binding to host chromatin and is crucial for both viral replication and the promotion of

  2. The Neurogenic Basic Helix-Loop-Helix Transcription Factor NeuroD6 Concomitantly Increases Mitochondrial mass and Regulates Cytoskeletal Organization in the Early Stages of Neuronal Differentiation

    Directory of Open Access Journals (Sweden)

    Kristin Kathleen Baxter

    2009-08-01

    Full Text Available Mitochondria play a central role during neurogenesis by providing energy in the form of ATP for cytoskeletal remodelling, outgrowth of neuronal processes, growth cone activity and synaptic activity. However, the fundamental question of how differentiating neurons control mitochondrial biogenesis remains vastly unexplored. Since our previous studies have shown that the neurogenic bHLH (basic helix–loop–helix transcription factor NeuroD6 is sufficient to induce differentiation of the neuronal progenitor-like PC12 cells and that it triggers expression of mitochondrial-related genes, we investigated whether NeuroD6 could modulate the mitochondrial biomass using our PC12-ND6 cellular paradigm. Using a combination of flow cytometry, confocal microscopy and mitochondrial fractionation, we demonstrate that NeuroD6 stimulates maximal mitochondrial mass at the lamellipodia stage, thus preceding axonal growth. NeuroD6 triggers remodelling of the actin and microtubule networks in conjunction with increased expression of the motor protein KIF5B, thus promoting mitochondrial movement in developing neurites with accumulation in growth cones. Maintenance of the NeuroD6-induced mitochondrial mass requires an intact cytoskeletal network, as its disruption severely reduces mitochondrial mass. The present study provides the first evidence that NeuroD6 plays an integrative role in co-ordinating increase in mitochondrial mass with cytoskeletal remodelling, suggestive of a role of this transcription factor as a co-regulator of neuronal differentiation and energy metabolism.

  3. Structural studies of polypeptides: Mechanism of immunoglobin catalysis and helix propagation in hybrid sequence, disulfide containing peptides

    Energy Technology Data Exchange (ETDEWEB)

    Storrs, R.W.

    1992-08-01

    Catalytic immunoglobin fragments were studied Nuclear Magnetic Resonance spectroscopy to identify amino acid residues responsible for the catalytic activity. Small, hybrid sequence peptides were analyzed for helix propagation following covalent initiation and for activity related to the protein from which the helical sequence was derived. Hydrolysis of p-nitrophenyl carbonates and esters by specific immunoglobins is thought to involve charge complementarity. The pK of the transition state analog P-nitrophenyl phosphate bound to the immunoglobin fragment was determined by [sup 31]P-NMR to verify the juxtaposition of a positively charged amino acid to the binding/catalytic site. Optical studies of immunoglobin mediated photoreversal of cis, syn cyclobutane thymine dimers implicated tryptophan as the photosensitizing chromophore. Research shows the chemical environment of a single tryptophan residue is altered upon binding of the thymine dimer. This tryptophan residue was localized to within 20 [Angstrom] of the binding site through the use of a nitroxide paramagnetic species covalently attached to the thymine dimer. A hybrid sequence peptide was synthesized based on the bee venom peptide apamin in which the helical residues of apamin were replaced with those from the recognition helix of the bacteriophage 434 repressor protein. Oxidation of the disufide bonds occured uniformly in the proper 1-11, 3-15 orientation, stabilizing the 434 sequence in an [alpha]-helix. The glycine residue stopped helix propagation. Helix propagation in 2,2,2-trifluoroethanol mixtures was investigated in a second hybrid sequence peptide using the apamin-derived disulfide scaffold and the S-peptide sequence. The helix-stop signal previously observed was not observed in the NMR NOESY spectrum. Helical connectivities were seen throughout the S-peptide sequence. The apamin/S-peptide hybrid binded to the S-protein (residues 21-166 of ribonuclease A) and reconstituted enzymatic activity.

  4. Structural studies of polypeptides: Mechanism of immunoglobin catalysis and helix propagation in hybrid sequence, disulfide containing peptides

    Energy Technology Data Exchange (ETDEWEB)

    Storrs, Richard Wood [Univ. of California, Berkeley, CA (United States)

    1992-08-01

    Catalytic immunoglobin fragments were studied Nuclear Magnetic Resonance spectroscopy to identify amino acid residues responsible for the catalytic activity. Small, hybrid sequence peptides were analyzed for helix propagation following covalent initiation and for activity related to the protein from which the helical sequence was derived. Hydrolysis of p-nitrophenyl carbonates and esters by specific immunoglobins is thought to involve charge complementarity. The pK of the transition state analog P-nitrophenyl phosphate bound to the immunoglobin fragment was determined by 31P-NMR to verify the juxtaposition of a positively charged amino acid to the binding/catalytic site. Optical studies of immunoglobin mediated photoreversal of cis, syn cyclobutane thymine dimers implicated tryptophan as the photosensitizing chromophore. Research shows the chemical environment of a single tryptophan residue is altered upon binding of the thymine dimer. This tryptophan residue was localized to within 20 Å of the binding site through the use of a nitroxide paramagnetic species covalently attached to the thymine dimer. A hybrid sequence peptide was synthesized based on the bee venom peptide apamin in which the helical residues of apamin were replaced with those from the recognition helix of the bacteriophage 434 repressor protein. Oxidation of the disufide bonds occured uniformly in the proper 1-11, 3-15 orientation, stabilizing the 434 sequence in an α-helix. The glycine residue stopped helix propagation. Helix propagation in 2,2,2-trifluoroethanol mixtures was investigated in a second hybrid sequence peptide using the apamin-derived disulfide scaffold and the S-peptide sequence. The helix-stop signal previously observed was not observed in the NMR NOESY spectrum. Helical connectivities were seen throughout the S-peptide sequence. The apamin/S-peptide hybrid binded to the S-protein (residues 21-166 of ribonuclease A) and reconstituted enzymatic activity.

  5. Right- and left-handed three-helix proteins. I. Experimental and simulation analysis of differences in folding and structure.

    Science.gov (United States)

    Glyakina, Anna V; Pereyaslavets, Leonid B; Galzitskaya, Oxana V

    2013-09-01

    Despite the large number of publications on three-helix protein folding, there is no study devoted to the influence of handedness on the rate of three-helix protein folding. From the experimental studies, we make a conclusion that the left-handed three-helix proteins fold faster than the right-handed ones. What may explain this difference? An important question arising in this paper is whether the modeling of protein folding can catch the difference between the protein folding rates of proteins with similar structures but with different folding mechanisms. To answer this question, the folding of eight three-helix proteins (four right-handed and four left-handed), which are similar in size, was modeled using the Monte Carlo and dynamic programming methods. The studies allowed us to determine the orders of folding of the secondary-structure elements in these domains and amino acid residues which are important for the folding. The obtained data are in good correlation with each other and with the experimental data. Structural analysis of these proteins demonstrated that the left-handed domains have a lesser number of contacts per residue and a smaller radius of cross section than the right-handed domains. This may be one of the explanations of the observed fact. The same tendency is observed for the large dataset consisting of 332 three-helix proteins (238 right- and 94 left-handed). From our analysis, we found that the left-handed three-helix proteins have some less-dense packing that should result in faster folding for some proteins as compared to the case of right-handed proteins. Copyright © 2013 Wiley Periodicals, Inc.

  6. Is the orientation of the fibrillar helix in the main layer of cell walls constant or variable within a tree?

    Directory of Open Access Journals (Sweden)

    W. Pyszyński

    2015-01-01

    Full Text Available The study material consisted of samples from 270 Picea excelsa and 54 Abies alba trees with variable type of wood grain. No correlation was found between the orientation of the fibrils and the type of grain. In all the samples in which the fibrillar helix formed an angle with the cell axis greater than 10° (80% of the total the helix was of Z type. In the other samples undulations commonly occured in the orientation of fibrils within a cell. These were in the form of local deviations to the left or right of the general trend to develop the Z type of spiral.

  7. HELIX128S-2 - A readout chip for the silicon vertex detector and inner tracker detector of HERA-B

    International Nuclear Information System (INIS)

    Trunk, U.; Fallot-Burghardt, W.; Sexauer, E.; Knoepfle, K-T.; Hofmann, W.; Cuje, M.; Glass, B.; Feuerstack-Raible, M.; Eisele, F.; Straumann, U.

    1998-01-01

    HERA-B is a fixed target experiment at the HERA proton storage ring dedicated to examine CP-violation in the B-Meson system. Based on the RD20-FElix concept a readout chip has been designed in AMS's 0.8 μm CMOS process for the HERA-B silicon vertex and inner tracker (MSGC) detectors. Various test chips have been submitted and successfully tested since '95, thus enabling the submission of a fully integrated 128 channel version in April '97. Design features of this chip (HELIX128S-2) and test results of its predecessor HELIX128 are presented

  8. The Microbiota and Abundance of the Class 1 Integron-Integrase Gene in Tropical Sewage Treatment Plant Influent and Activated Sludge.

    Directory of Open Access Journals (Sweden)

    Magna C Paiva

    Full Text Available Bacteria are assumed to efficiently remove organic pollutants from sewage in sewage treatment plants, where antibiotic-resistance genes can move between species via mobile genetic elements known as integrons. Nevertheless, few studies have addressed bacterial diversity and class 1 integron abundance in tropical sewage. Here, we describe the extant microbiota, using V6 tag sequencing, and quantify the class 1 integron-integrase gene (intI1 in raw sewage (RS and activated sludge (AS. The analysis of 1,174,486 quality-filtered reads obtained from RS and AS samples revealed complex and distinct bacterial diversity in these samples. The RS sample, with 3,074 operational taxonomic units, exhibited the highest alpha-diversity indices. Among the 25 phyla, Proteobacteria, Bacteroidetes and Firmicutes represented 85% (AS and 92% (RS of all reads. Increased relative abundance of Micrococcales, Myxococcales, and Sphingobacteriales and reduced pathogen abundance were noted in AS. At the genus level, differences were observed for the dominant genera Simplicispira and Diaphorobacter (AS as well as for Enhydrobacter (RS. The activated sludge process decreased (55% the amount of bacteria harboring the intI1 gene in the RS sample. Altogether, our results emphasize the importance of biological treatment for diminishing pathogenic bacteria and those bearing the intI1 gene that arrive at a sewage treatment plant.

  9. Effect of temperature on the fate of genes encoding tetracycline resistance and the integrase of class 1 integrons within anaerobic and aerobic digesters treating municipal wastewater solids.

    Science.gov (United States)

    Diehl, David L; LaPara, Timothy M

    2010-12-01

    The objective of this research was to investigate the ability of anaerobic and aerobic digesters to reduce the quantity of antibiotic resistant bacteria in wastewater solids. Lab-scale digesters were operated at different temperatures (22 °C, 37 °C, 46 °C, and 55 °C) under both anaerobic and aerobic conditions and fed wastewater solids collected from a full-scale treatment facility. Quantitative PCR was used to track five genes encoding tetracycline resistance (tet(A), tet(L), tet(O), tet(W), and tet(X)) and the gene encoding the integrase (intI1) of class 1 integrons. Statistically significant reductions in the quantities of these genes occurred in the anaerobic reactors at 37 °C, 46 °C, and 55 °C, with the removal rates and removal efficiencies increasing as a function of temperature. The aerobic digesters, in contrast, were generally incapable of significantly decreasing gene quantities, although these digesters were operated at much shorter mean hydraulic residence times. This research suggests that high temperature anaerobic digestion of wastewater solids would be a suitable technology for eliminating various antibiotic resistance genes, an emerging pollutant of concern.

  10. 1-Hydroxypyrido[2,3-d]pyrimidin-2(1H)-ones as novel selective HIV integrase inhibitors obtained via privileged substructure-based compound libraries.

    Science.gov (United States)

    Gao, Ping; Zhang, Lingzi; Sun, Lin; Huang, Tianguang; Tan, Jing; Zhang, Jian; Zhou, Zhongxia; Zhao, Tong; Menéndez-Arias, Luis; Pannecouque, Christophe; Clercq, Erik De; Zhan, Peng; Liu, Xinyong

    2017-10-15

    A small library containing 3-hydroxyquinazoline-2,4(1H,3H)-dione and 1-hydroxypyrido[2,3-d]pyrimidin-2(1H)-one scaffolds was obtained via the copper(I)-catalyzed azidealkyne cycloaddition (CuAAC) reaction and evaluated for their anti-HIV activity in MT-4 cells. Among the synthesized compounds, several 1-hydroxypyrido[2,3-d]pyrimidin-2(1H)-one derivatives showed remarkable anti-HIV potency with EC 50 values ranging from 0.92 to 26.85µM. The most active one, IIA-2, also showed remarkable and selective potency against HIV type 1 integrase (IN). To the best of our knowledge, this is the first report showing that 1-hydroxypyrido[2,3-d]pyrimidin-2(1H)-ones are selective HIV IN inhibitors. Preliminary structure-activity relationship (SAR) studies suggested that the divalent metal ion chelators and the nature and position of substituents around the core are important for antiviral potency. Molecular modeling has been used to predict the binding site of the pyrido[2,3-d]pyrimidin-2(1H)-one core in HIV type 1 IN and suggestions are made for improvement of its inhibitory activity. Copyright © 2017 Elsevier Ltd. All rights reserved.

  11. L-Chicoric acid inhibits human immunodeficiency virus type 1 integration in vivo and is a noncompetitive but reversible inhibitor of HIV-1 integrase in vitro

    International Nuclear Information System (INIS)

    Reinke, Ryan A.; Lee, Deborah J.; McDougall, Brenda R.; King, Peter J.; Victoria, Joseph; Mao Yingqun; Lei Xiangyang; Reinecke, Manfred G.; Robinson, W. Edward

    2004-01-01

    The human immunodeficiency virus (HIV) integrase (IN) must covalently join the viral cDNA into a host chromosome for productive HIV infection. L-Chicoric acid (L-CA) enters cells poorly but is a potent inhibitor of IN in vitro. Using quantitative real-time polymerase chain reaction (PCR), L-CA inhibits integration at concentrations from 500 nM to 10 μM but also inhibits entry at concentrations above 1 μM. Using recombinant HIV IN, steady-state kinetic analyses with L-CA were consistent with a noncompetitive or irreversible mechanism of inhibition. IN, in the presence or absence of L-CA, was successively washed. Inhibition of IN diminished, demonstrating that L-CA was reversibly bound to the protein. These data demonstrate that L-CA is a noncompetitive but reversible inhibitor of IN in vitro and of HIV integration in vivo. Thus, L-CA likely interacts with amino acids other than those which bind substrate

  12. The Microbiota and Abundance of the Class 1 Integron-Integrase Gene in Tropical Sewage Treatment Plant Influent and Activated Sludge.

    Science.gov (United States)

    Paiva, Magna C; Ávila, Marcelo P; Reis, Mariana P; Costa, Patrícia S; Nardi, Regina M D; Nascimento, Andréa M A

    2015-01-01

    Bacteria are assumed to efficiently remove organic pollutants from sewage in sewage treatment plants, where antibiotic-resistance genes can move between species via mobile genetic elements known as integrons. Nevertheless, few studies have addressed bacterial diversity and class 1 integron abundance in tropical sewage. Here, we describe the extant microbiota, using V6 tag sequencing, and quantify the class 1 integron-integrase gene (intI1) in raw sewage (RS) and activated sludge (AS). The analysis of 1,174,486 quality-filtered reads obtained from RS and AS samples revealed complex and distinct bacterial diversity in these samples. The RS sample, with 3,074 operational taxonomic units, exhibited the highest alpha-diversity indices. Among the 25 phyla, Proteobacteria, Bacteroidetes and Firmicutes represented 85% (AS) and 92% (RS) of all reads. Increased relative abundance of Micrococcales, Myxococcales, and Sphingobacteriales and reduced pathogen abundance were noted in AS. At the genus level, differences were observed for the dominant genera Simplicispira and Diaphorobacter (AS) as well as for Enhydrobacter (RS). The activated sludge process decreased (55%) the amount of bacteria harboring the intI1 gene in the RS sample. Altogether, our results emphasize the importance of biological treatment for diminishing pathogenic bacteria and those bearing the intI1 gene that arrive at a sewage treatment plant.

  13. The Microbiota and Abundance of the Class 1 Integron-Integrase Gene in Tropical Sewage Treatment Plant Influent and Activated Sludge

    Science.gov (United States)

    Paiva, Magna C.; Ávila, Marcelo P.; Reis, Mariana P.; Costa, Patrícia S.; Nardi, Regina M. D.; Nascimento, Andréa M. A.

    2015-01-01

    Bacteria are assumed to efficiently remove organic pollutants from sewage in sewage treatment plants, where antibiotic-resistance genes can move between species via mobile genetic elements known as integrons. Nevertheless, few studies have addressed bacterial diversity and class 1 integron abundance in tropical sewage. Here, we describe the extant microbiota, using V6 tag sequencing, and quantify the class 1 integron-integrase gene (intI1) in raw sewage (RS) and activated sludge (AS). The analysis of 1,174,486 quality-filtered reads obtained from RS and AS samples revealed complex and distinct bacterial diversity in these samples. The RS sample, with 3,074 operational taxonomic units, exhibited the highest alpha-diversity indices. Among the 25 phyla, Proteobacteria, Bacteroidetes and Firmicutes represented 85% (AS) and 92% (RS) of all reads. Increased relative abundance of Micrococcales, Myxococcales, and Sphingobacteriales and reduced pathogen abundance were noted in AS. At the genus level, differences were observed for the dominant genera Simplicispira and Diaphorobacter (AS) as well as for Enhydrobacter (RS). The activated sludge process decreased (55%) the amount of bacteria harboring the intI1 gene in the RS sample. Altogether, our results emphasize the importance of biological treatment for diminishing pathogenic bacteria and those bearing the intI1 gene that arrive at a sewage treatment plant. PMID:26115093

  14. HIV type 1 coreceptor tropism, CCR5 genotype, and integrase inhibitor resistance profiles in Vietnam: implications for the introduction of new antiretroviral regimens.

    Science.gov (United States)

    Luu, Quynh Phuong; Dean, Jonathan; Do, Trinh Thi Diem; Carr, Michael J; Dunford, Linda; Coughlan, Suzie; Connell, Jeff; Nguyen, Hien Tran; Hall, William W; Nguyen Thi, Lan Anh

    2012-10-01

    In Vietnam, where an estimated 280,000 people will be HIV-positive by 2012, recommended antiretroviral regimens do not include more recently developed therapeutics, such as Integrase inhibitors (INI) and coreceptor antagonists. This study examined HIV-1 coreceptor tropism and INI drug resistance profiles, in parallel with CCR5 genotypes, in a cohort of 60 HIV-positive individuals from different regions of Vietnam. No evidence of INI resistance was detected. Some 40% of individuals had X4-tropic HIV-1, making them unsuitable for treatment with CCR5 antagonists. We identified a novel CCR5 variant-S272P-along with other, previously reported variants: G106R, C178R, W153C, R223Q, and S336I. Interestingly, CCR5 variants known to affect HIV-1 infectivity were observed only in individuals harboring X4-tropic virus. Together, this study presents valuable baseline information on HIV-1 INI resistance, coreceptor tropism, and CCR5 variants in HIV-positive individuals in Vietnam. This should help inform policy on the future use of novel antiretrovirals in Vietnam.

  15. The structure of the XPF-ssDNA complex underscores the distinct roles of the XPF and ERCC1 helix- hairpin-helix domains in ss/ds DNA recognition

    NARCIS (Netherlands)

    Das, Devashish; Folkers, Gert; van Dijk, Marc; Jaspers, Nicolaas G.J.; Hoeijmakers, Jan H.J.; Kaptein, Robert; Boelens, Rolf

    2012-01-01

    Human XPF/ERCC1 is a structure-specific DNA endonuclease that nicks the damaged DNA strand at the 5' end during nucleotide excision repair. We determined the structure of the complex of the C-terminal domain of XPF with 10 nt ssDNA. A positively charged region within the second helix of the first

  16. Postembryonic neurogenesis in the procerebrum of the terrestrial snail, Helix lucorum L

    Science.gov (United States)

    Zakharov, I. S.; Hayes, N. L.; Ierusalimsky, V. N.; Nowakowski, R. S.; Balaban, P. M.

    1998-01-01

    Neuronogenesis during posthatching development of the procerebrum of the terrestrial snail Helix lucorum was analyzed using bromodeoxyuridine immunohistochemistry to label proliferating cells. Comparison of the distribution of labeled cells in a series of animals which differed in age at the time of incubation with bromodeoxyuridine, in survival time after incubation, and in age at sacrifice reveals a clear pattern and developmental sequence in neuron origin. First, the proliferating cells are located only at the apical portion of the procerebrum. Second, cells which are produced at any particular age remain, for the most part, confined to a single layer in the procerebrum. Third, as development proceeds, each layer of previously produced neurons is displaced toward the basal part of the procerebrum by the production of additional neurons. Our results suggest that the vast majority of the neurons (probably about 70-80%) of the snail procerebrum are produced during the first 1-2 months of posthatching development.

  17. Class-B GPCR activation: is ligand helix-capping the key?

    Science.gov (United States)

    Neumann, Jean-Michel; Couvineau, Alain; Murail, Samuel; Lacapère, Jean-Jacques; Jamin, Nadège; Laburthe, Marc

    2008-07-01

    The class B family of G-protein-coupled receptors (GPCRs) regulates essential physiological functions such as exocrine and endocrine secretions, feeding behaviour, metabolism, growth, and neuro- and immuno-modulations. These receptors are activated by endogenous peptide hormones including secretin, glucagon, vasoactive intestinal peptide, corticotropin-releasing factor and parathyroid hormone. We have identified a common structural motif that is encoded in all class B GPCR-ligand N-terminal sequences. We propose that this local structure, a helix N-capping motif, is formed upon receptor binding and constitutes a key element underlying class B GPCR activation. The folded backbone conformation imposed by the capping structure could serve as a template for a rational design of drugs targeting class B GPCRs in several diseases.

  18. Stomatal Responses to Light and Drought Stress in Variegated Leaves of Hedera helix1

    Science.gov (United States)

    Aphalo, Pedro J.; Sánchez, Rodolfo A.

    1986-01-01

    Direct and indirect mechanisms underlying the light response of stomata were studied in variegated leaves of the juvenile phase of Hedera helix L. Dose response curves of leaf conductance were measured with blue and red light in leaves kept in normal or in an inverted position. In the green portions of the leaves, the sensitivity to blue light was nearly 100 times higher than that to red light. No response to red light was observed in the white portions of the leaves up to 90 micromoles per square meter per second. Red light indirectly affected leaf conductance while blue light had a direct effect. Leaf conductance was found to be more sensitive to drought stress and showed a more persistent aftereffect in the white portions of the leaves. A differential effect of drought stress on the responses to blue and red light was also observed. PMID:16664900

  19. Solvent-Exposed Salt Bridges Influence the Kinetics of α-Helix Folding and Unfolding.

    Science.gov (United States)

    Meuzelaar, Heleen; Tros, Martijn; Huerta-Viga, Adriana; van Dijk, Chris N; Vreede, Jocelyne; Woutersen, Sander

    2014-03-06

    Salt bridges are known to play an essential role in the thermodynamic stability of the folded conformation of many proteins, but their influence on the kinetics of folding remains largely unknown. Here, we investigate the effect of Glu-Arg salt bridges on the kinetics of α-helix folding using temperature-jump transient-infrared spectroscopy and steady-state UV circular dichroism. We find that geometrically optimized salt bridges (Glu - and Arg + are spaced four peptide units apart, and the Glu/Arg order is such that the side-chain rotameric preferences favor salt-bridge formation) significantly speed up folding and slow down unfolding, whereas salt bridges with unfavorable geometry slow down folding and slightly speed up unfolding. Our observations suggest a possible explanation for the surprising fact that many biologically active proteins contain salt bridges that do not stabilize the native conformation: these salt bridges might have a kinetic rather than a thermodynamic function.

  20. Bridge helix bending promotes RNA polymerase II backtracking through a critical and conserved threonine residue

    KAUST Repository

    Da, Lin-Tai

    2016-04-19

    The dynamics of the RNA polymerase II (Pol II) backtracking process is poorly understood. We built a Markov State Model from extensive molecular dynamics simulations to identify metastable intermediate states and the dynamics of backtracking at atomistic detail. Our results reveal that Pol II backtracking occurs in a stepwise mode where two intermediate states are involved. We find that the continuous bending motion of the Bridge helix (BH) serves as a critical checkpoint, using the highly conserved BH residue T831 as a sensing probe for the 3′-terminal base paring of RNA:DNA hybrid. If the base pair is mismatched, BH bending can promote the RNA 3′-end nucleotide into a frayed state that further leads to the backtracked state. These computational observations are validated by site-directed mutagenesis and transcript cleavage assays, and provide insights into the key factors that regulate the preferences of the backward translocation.

  1. [Neurochemical mechanisms of food aversion conditioning consolidation in snail Helix lucorum].

    Science.gov (United States)

    Solntseva, S V; Nikitin, v P

    2008-11-01

    Effects of cycloheximide, protein synthesis inhibitors, as well as serotonin receptor antagonist and NMDA receptor antagonist on food aversion conditioning consolidation were studied in snail Helix lucorum. Food aversion conditioning was absent in snails after application of cycloheximide. Repeated produced no food aversion conditioning for the same type of food in these snails without cycloheximide application. Food aversion conditioning was absent in snails after applications of metiotepin, nonselective serotonin receptors antagonist, or after MK-801, NMDA glutamate receptors antagonist. At the same time, repeated training produced facilitated food aversion conditioning for the same type of food in these snails. Our experiments were the first which showed that effect on different molecular mechanisms evoked reversible or irreversible disruption of long-term memory consolidation during the same learning. It was suggested that suppression of retrieval produced reversible effect, whereas disruption of memory storage initiated irreversible effect on long-term memory consolidation.

  2. In vitro metabolism of androstenedione and identification of endogenous steroids in Helix aspersa

    International Nuclear Information System (INIS)

    Le Guellec, D.; Thiard, M.C.; Remy-Martin, J.P.; Deray, A.; Gomot, L.; Adessi, G.L.

    1987-01-01

    In vitro metabolism of androstenedione in gonads of juvenile and adult Helix aspersa has been investigated. The conversion of [ 3 H]androstenedione into testosterone, 5 alpha-dihydrotestosterone, androsterone, and estriol was demonstrated. In juvenile animals testosterone (59.8%) is the major metabolite whereas in adult animals androsterone (18.8%) is. The following endogenous steroids have been identified by gas chromatography-mass spectrometry in adult gonads: androsterone, dehydroepiandrosterone, androstenedione, 3 alpha-androstanediol, estrone, estradiol-17 beta, and estriol. The levels of testosterone, 5 alpha-dihydrotestosterone, androstenedione, and progesterone have been measured by RIAs in gonads and hemolymph. Their levels vary with the physiological stage: the gonadal and circulating levels of testosterone decrease with the sexual maturation whereas the 5 alpha-dihydrotestosterone increases. These differences observed in metabolism and in level of steroids between the juvenile and the adult snails allow us to suppose that these steroids have a biological role

  3. Broadband and high-efficiency vortex beam generator based on a hybrid helix array.

    Science.gov (United States)

    Fang, Chaoqun; Wu, Chao; Gong, Zhijie; Zhao, Song; Sun, Anqi; Wei, Zeyong; Li, Hongqiang

    2018-04-01

    The vortex beam which carries the orbital angular momentum has versatile applications, such as high-resolution imaging, optical communications, and particle manipulation. Generating vortex beams with the Pancharatnam-Berry (PB) phase has drawn considerable attention for its unique spin-to-orbital conversion features. Despite the PB phase being frequency independent, an optical element with broadband high-efficiency circular polarization conversion feature is still needed for the broadband high-efficiency vortex beam generation. In this work, a broadband and high-efficiency vortex beam generator based on the PB phase is built with a hybrid helix array. Such devices can generate vortex beams with arbitrary topological charge. Moreover, vortex beams with opposite topological charge can be generated with an opposite handedness incident beam that propagates backward. The measured efficiency of our device is above 65% for a wide frequency range, with the relative bandwidth of 46.5%.

  4. Spontaneously amplified homochiral organic-inorganic nano-helix complexes via self-proliferation

    Science.gov (United States)

    Zhai, Halei; Quan, Yan; Li, Li; Liu, Xiang-Yang; Xu, Xurong; Tang, Ruikang

    2013-03-01

    Most spiral coiled biomaterials in nature, such as gastropod shells, are homochiral, and the favoured chiral feature can be precisely inherited. This inspired us that selected material structures, including chirality, could be specifically replicated into the self-similar populations; however, a physicochemical understanding of the material-based heritage is unknown. We study the homochirality by using calcium phosphate mineralization in the presence of racemic amphiphilic molecules and biological protein. The organic-inorganic hybrid materials with spiral coiling characteristics are produced at the nanoscale. The resulted helixes are chiral with the left- and right-handed characteristics, which are agglomerated hierarchically to from clusters and networks. It is interesting that each cluster or network is homochiral so that the enantiomorphs can be separated readily. Actually, each homochiral architecture is evolved from an original chiral helix, demonstrating the heritage of the matrix chirality during the material proliferation under a racemic condition. By using the Ginzburg-Landaue expression we find that the chiral recognition in the organic-inorganic hybrid formation may be determined by a spontaneous chiral separation and immobilization of asymmetric amphiphilic molecules on the mineral surface, which transferred the structural information from the mother matrix to the descendants by an energetic control. This study shows how biomolecules guide the selective amplification of chiral materials via spontaneous self-replication. Such a strategy can be applied generally in the design and production of artificial materials with self-similar structure characteristics.Most spiral coiled biomaterials in nature, such as gastropod shells, are homochiral, and the favoured chiral feature can be precisely inherited. This inspired us that selected material structures, including chirality, could be specifically replicated into the self-similar populations; however, a

  5. Structural Dynamics Of The S4 Voltage-Sensor Helix In Lipid Bilayers Lacking Lipid Phosphates

    Science.gov (United States)

    Andersson, Magnus; Freites, J. Alfredo; Tobias, Douglas J.; White, Stephen H.

    2011-01-01

    Voltage-dependent K+ (Kv) channels require lipid phosphates for functioning. The S4 helix, which carries the gating charges in the voltage-sensing domain (VSD), inserts into membranes while being stabilized by a protein-lipid interface in which lipid phosphates play an essential role. To examine the physical basis of the protein-lipid interface in the absence of lipid phosphates, we performed molecular dynamics (MD) simulations of a KvAP S4 variant (S4mut) in bilayers with and without lipid phosphates. We find that in dioleoyltrimethylammoniumpropane (DOTAP) bilayers lacking lipid phosphates, the gating charges are solvated by anionic counterions and, hence, lack the bilayer support provided by phosphate-containing palmitoyloleoylglycerophosphocholine (POPC) bilayers. The result is a water-permeable bilayer with a significantly smaller deformations around the peptide. Together, these results provide an explanation for the non-functionality of VSDs in terms of a destabilizing protein-lipid interface. PMID:21692541

  6. Transient Non-Native Helix Formation during the Folding of b-Lactoglobulin

    Directory of Open Access Journals (Sweden)

    Masamichi Ikeguchi

    2014-02-01

    Full Text Available In ideal proteins, only native interactions are stabilized step-by-step in a smooth funnel-like energy landscape. In real proteins, however, the transient formation of non-native structures is frequently observed. In this review, the transient formation of non-native structures is described using the non-native helix formation during the folding of b-lactoglobulin as a prominent example. Although b-lactoglobulin is a predominantly b-sheet protein, it has been shown to form non-native helices during the early stage of folding. The location of non-native helices, their stabilization mechanism, and their role in the folding reaction are discussed.

  7. Communication: Role of explicit water models in the helix folding/unfolding processes

    Science.gov (United States)

    Palazzesi, Ferruccio; Salvalaglio, Matteo; Barducci, Alessandro; Parrinello, Michele

    2016-09-01

    In the last years, it has become evident that computer simulations can assume a relevant role in modelling protein dynamical motions for their ability to provide a full atomistic image of the processes under investigation. The ability of the current protein force-fields in reproducing the correct thermodynamics and kinetics systems behaviour is thus an essential ingredient to improve our understanding of many relevant biological functionalities. In this work, employing the last developments of the metadynamics framework, we compare the ability of state-of-the-art all-atom empirical functions and water models to consistently reproduce the folding and unfolding of a helix turn motif in a model peptide. This theoretical study puts in evidence that the choice of the water models can influence the thermodynamic and the kinetics of the system under investigation, and for this reason cannot be considered trivial.

  8. The double helix revisited: a paradox of science and a paradigm of human behaviour

    Directory of Open Access Journals (Sweden)

    Argüelles, Juan Carlos

    2007-06-01

    Full Text Available In the modern history of Science, few breakthroughs have caused an impact comparative to the Double Helix, the three-dimensional structure of DNA proposed by Watson & Crick in 1953, an event whose 50th anniversary was widely celebrated in the non-specialist media, three years ago. Although the discovery had little transcendence at the time, it has unquestionably been of great importance ever since. The Double Helix has underlined the true biological value of nucleic acids compared with proteins, demonstrating that genes are not amorphous entities but have a specific chemical composition and adopt an ordered spatial folding pattern. Elucidation of this key configuration made it possible to establish a direct relationship between the structure and the function of macromolecules, a relationship which is not so clear in the case of proteins. During these last fifty years much has been written and argued about the circumstances surrounding the discovery and about the behaviour and attitudes of many of the protagonists. Besides Watson & Crick, other scientists, whose contribution has not been adequately recognised, played an important part in solving the Double Helix mystery. This article contains some ethical and scientific reflections which revise some of these essential contributions and throws light on the role played in history by these comparatively «unknown soldiers» of science. The Double Helix story is undoubtedly a manifestation of the human side of science and many scientists believe that the available evidence taken as a whole permits an alternative story to be written.

    En la desarrollo histórico de la Ciencia moderna, pocos descubrimientos han causado un impacto comparativo a las repercusiones de la Doble Hélice, la estructura tridimensional del ADN, propuesta por Watson y Crick en 1953. El 50º aniversario de aquel evento fue ampliamente celebrado hace tres años, incluso por los medios no especializados en informaci

  9. Positioning and Imaging Detection of Corona Discharge in Air with Double Helix Acoustic Sensors Array

    Directory of Open Access Journals (Sweden)

    Ming Dong

    2017-12-01

    Full Text Available Corona discharge could be a serious hazard to the outdoor insulation of a high-voltage (HV system and also is an important detection issue for insulation status diagnosis. In this study, a double-helix-ultrasonic-array (DHUA sensor was developed for corona positioning, which showed a good sensitivity to the ultrasonic signal originating from corona discharge and a satisfied directional response in static beam patterns. Based on the matting-based image fusion method, the acoustic intensity matrix was fused to a visual scene in real-time. This ultrasonic visualization technology was then evaluated with a simulation test and performed in a real air insulated substation (AIS, and showed a satisfied performance in terms of sensitivity and spatial location resolution.

  10. Blocking of conditioning of tentacle lowering in the snail (Helix aspersa).

    Science.gov (United States)

    Acebes, Félix; Solar, Patricia; Carnero, Susana; Loy, Ignacio

    2009-07-01

    Experiment 1 established the effectiveness of an appetitive conditioning of odours procedure with snails (Helix aspersa) that was subsequently used for the study of blocking. In this important phenomenon, the conditioning of a CS1 (where CS is the conditioned stimulus) prior to conditioning of a compound, CS1CS2, blocked the conditioning to the CS2. Experiments 2a, 2b, and 2c demonstrated this associative effect using three different experimental controls. Experiments 3a and 3b replicated the blocking effect and allowed us to reject an explanation of blocking based on generalized effects of several treatments of diverse stimuli in blocking and control groups (the pseudoblocking effect). The implications of these results for the study of invertebrate cognition by means of conditioning techniques are discussed.

  11. Single-molecule observation of helix staggering, sliding, and coiled coil misfolding

    Science.gov (United States)

    Xi, Zhiqun; Gao, Ying; Sirinakis, George; Guo, Honglian; Zhang, Yongli

    2012-01-01

    The biological functions of coiled coils generally depend on efficient folding and perfect pairing of their α-helices. Dynamic changes in the helical registry that lead to staggered helices have only been proposed for a few special systems and not found in generic coiled coils. Here, we report our observations of multiple staggered helical structures of two canonical coiled coils. The partially folded structures are formed predominantly by coiled coil misfolding and occasionally by helix sliding. Using high-resolution optical tweezers, we characterized their energies and transition kinetics at a single-molecule level. The staggered states occur less than 2% of the time and about 0.1% of the time at zero force. We conclude that dynamic changes in helical registry may be a general property of coiled coils. Our findings should have broad and unique implications in functions and dysfunctions of proteins containing coiled coils. PMID:22451899

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

    Science.gov (United States)

    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.

  13. Innovative Development of Kazakhstan on The Basis of Triple Helix and Cluster Approach

    Directory of Open Access Journals (Sweden)

    Farkhat Musayevich Dnishev

    2015-06-01

    Full Text Available The aim of the research is to study the Triple Helix model feasibility in developing innovations and using cluster approach in Kazakhstan. There are possible points of the emergence of clusters in Kazakhstan. However, there are a lot of constraining factors. First of all, institutional and social factors: the culture of business, unfair competition, low trust of economic agents to each other and to power institutes, low psychological readiness for cooperation of the enterprises of various branches and regions, poor development of chambers of commerce, and industrial associations. For the time being, the majority of regions of Kazakhstan are characterized by a limited set of high technology industrial branches, and a sharp shortage of universities generating innovation and research institutes. The research results show that the open innovation model is realized in a limited scale that does not allow to export innovations into external markets, to participate in global technology chains and international research networks. At the same time, some interaction schemes and preconditions for the development of the Triple Helix model are emerging. However, in general, the innovation policy is not systemic; it does not unite actions in the sphere of science and technology, education, industry, and regional initiatives. As the result of the research, some policy implications are given. For the development of clusters in Kazakhstan, it is desirable to use such a way, as integration into global cluster networks. It is necessary to make use of foreign experience at which various specialized state agencies become participants of clusters. It is necessary to focus not only on science but also industry, which should play the central role in the innovation process.

  14. Oxidation of Helix-3 methionines precedes the formation of PK resistant PrP.

    Directory of Open Access Journals (Sweden)

    Tamar Canello

    2010-07-01

    Full Text Available While elucidating the peculiar epitope of the alpha-PrP mAb IPC2, we found that PrPSc exhibits the sulfoxidation of residue M213 as a covalent signature. Subsequent computational analysis predicted that the presence of sulfoxide groups at both Met residues 206 and 213 destabilize the alpha-fold, suggesting oxidation may facilitate the conversion of PrPC into PrPSc. To further study the effect of oxidation on prion formation, we generated pAbs to linear PrP peptides encompassing the Helix-3 region, as opposed to the non-linear complexed epitope of IPC2. We now show that pAbs, whose epitopes comprise Met residues, readily detected PrPC, but could not recognize most PrPSc bands unless they were vigorously reduced. Next, we showed that the alpha-Met pAbs did not recognize newly formed PrPSc, as is the case for the PK resistant PrP present in lines of prion infected cells. In addition, these reagents did not detect intermediate forms such as PK sensitive and partially aggregated PrPs present in infected brains. Finally, we show that PrP molecules harboring the pathogenic mutation E200K, which is linked to the most common form of familial CJD, may be spontaneously oxidized. We conclude that the oxidation of methionine residues in Helix-3 represents an early and important event in the conversion of PrPC to PrPSc. We believe that further investigation into the mechanism and role of PrP oxidation will be central in finally elucidating the mechanism by which a normal cell protein converts into a pathogenic entity that causes fatal brain degeneration.

  15. NMR studies of abasic sites in DNA duplexes: Deoxyadenosine stacks into the helix opposite acyclic lesions

    International Nuclear Information System (INIS)

    Kalnik, M.W.; Chang, Chienneng; Johnson, F.; Grollman, A.P.; Patel, D.J.

    1989-01-01

    Proton and phosphorus NMR studies are reported for two complementary nonanucleotide duplexes containing acyclic abasic sites. The first duplex, d(C-A-T-G-A-G-T-A-C)·d(G-T-A-C-P-C-A-T-G), contains an acyclic propanyl moiety, P, located opposite a deoxyadenosine at the center of the helix (designated AP P 9-mer duplex). The second duplex, d(C-A-T-G-A-G-T-A-C-)·d(G-T-A-C-E-C-A-T-G), contains a similarly located acyclic ethanyl moiety, E (designated AP E 9-mer duplex). The ethanyl moiety is one carbon shorter than the natural carbon-phosphodiester backbone of a single nucleotide unit of DNA. The majority of the exchangeable and nonexchangeable base and sugar protons in both the AP P 9-mer and AP E 9-mer duplexes, including those at the abasic site, have been assigned by recording and analyzing two-dimensional phase-sensitive NOESY data sets in H 2 O and D 2 O solution between -5 and 5 degree C. These spectroscopic observations establish that A5 inserts into the helix opposite the abasic site (P14 and El14) and stacks between the flanking G4·C15 and G6·C13 Watson-Crick base pairs in both the AP P 9-mer and AP E 9-mer duplexes. Proton NMR parameters for the Ap P 9-mer and AP E 9-mer duplexes are similar to those reported previously. These proton NMR experiments demonstrate that the structures at abasic sites are very similar whether the five-membered ring is open or closed or whether the phosphodiester backbone is shortened by one carbon atom. Phosphorus spectra of the AP P 9-mer and AP E 9-mer duplexes (5 degree C) indicate that the backbone conformation is similarly perturbed at three phosphodiester backbone torsion angles

  16. National and international dimensions of the Triple Helix in Japan: University-industry-government versus international coauthorship relations

    NARCIS (Netherlands)

    Leydesdorff, L.; Sun, Y.

    2009-01-01

    International co-authorship relations and university-industry-government (Triple Helix) relations have hitherto been studied separately. Using Japanese publication data for the 1981-2004 period, we were able to study both kinds of relations in a single design. In the Japanese file, 1,277,030

  17. Estudio del hepatopáncreas de Helix Aspersa en situación normal y tras ayuno

    OpenAIRE

    Almendros Gallego, Antonio Manuel

    2013-01-01

    Se hace un estudio a microscopia electrónica de los diversos tipos celulares que componen el hepatopancreas de Helix Aspersa atendiendo a los componentes celulares característicos de cada uno de ellos tanto en situación normal como a diversos periodos de ayuno

  18. Cooperative formation of native-like tertiary contacts in the ensemble of unfolded states of a four-helix protein

    DEFF Research Database (Denmark)

    Bruun, Susanne W; Iesmantavicius, Vytautas; Danielsson, Jens

    2010-01-01

    In studies of the ensembles of unfolded structures of a four-helix bundle protein, we have detected the presence of potential precursors of native tertiary structures. These observations were based on the perturbation of NMR chemical shifts of the protein backbone atoms by single site mutations. ...

  19. Synthesis and studies of modified oligonucleotides-directed triple helix formation at the purine-pyrimidine interrupted site.

    Science.gov (United States)

    Jazouli, Mohamed; Guianvarc'h, Dominique; Bougrin, Khalid; Soufiaoui, Mohamed; Vierling, Pierre; Benhida, Rachid

    2003-01-01

    Triple helix formation is still restricted to oligopurine-oligopyrimidine double stranded DNA target. Herein we focus on our progress achieved in nucleobase and oligonucleotide modifications area to address the chemical challenge to circumvent the recognition of a purine-pyrimidine base pair interruption in an oligopyrimidine-oligopurine DNA sequence.

  20. Measuring the knowledge base of an economy in terms of triple-helix relations among 'technology, organization, and territory'

    NARCIS (Netherlands)

    Leydesdorff, L.; Dolfsma, W.; van der Panne, G.; Viale, R.; Etzkowitz, H.

    2010-01-01

    Can the knowledge base of an economy be measured? In this study, we combine the perspective of regional economics on the interrelationships among technology, organization, and territory with the triple-helix model, and offer the mutual information in three dimensions as an indicator of the

  1. Chain length dependence of the helix orientation in Langmuir-Blodgett monolayers of alpha-helical diblock copolypeptides

    NARCIS (Netherlands)

    Nguyen, Le-Thu T.; Ardana, Aditya; Vorenkamp, Eltjo J.; ten Brinke, Gerrit; Schouten, Arend J.

    2010-01-01

    The effect of chain length on the helix orientation of alpha-helical diblock copolypeptides in Langmuir and Langmuir-Blodgett monolayers is reported for the first time. Amphiphilic diblock copolypeptides (PLGA-b-PMLGSLGs) of poly(alpha-L-glutamic acid) (PLGA) and

  2. A Triple Helix Strategy for Promoting SME Development: The Case of a Dried Banana Community Enterprise in Thailand

    Science.gov (United States)

    Yuwawutto, Sauwapa; Smitinont, Thitapha; Charoenanong, Numtip; Yokakul, Nattaka; Chatratana, Sonchai; Zawdie, Girma

    2010-01-01

    This paper examines the university-industry-government relationship as a mechanism for enhancing the efficiency and competitiveness of small and medium-sized enterprises (SMEs). The case of a community enterprise producing dried banana products in the north of Thailand is used to demonstrate the significance of the Triple Helix model for business…

  3. Changes in the reproductive system of the snail Helix aspersa caused by mucus from the love dart

    NARCIS (Netherlands)

    Koene, J M; Chase, R.

    The function of the love dart in certain species of terrestrial snails is unknown. In Helix aspersa, the dart is a sharp calcareous structure that is used to pierce the partner's skin during courtship. When expelled, the dart is covered with a thick mucus. The hypothesis tested here is that the

  4. What can triple helix frameworks offer to the analysis of eco-innovation dynamics? Theoretical and methodological considerations

    DEFF Research Database (Denmark)

    Yang, Yan; Holgaard, Jette Egelund; Remmen, Arne

    2012-01-01

    Bringing environmental concerns into focus of innovation processes will in several cases also expand the numbers of actors involved. Eco-innovation and triple helix are often frameworks applied to analyse how environmental concerns are integrated in the innovation processes and how different stak...

  5. Modulating immunogenic properties of HIV-1 gp41 membrane-proximal external region by destabilizing six-helix bundle structure.

    Science.gov (United States)

    Banerjee, Saikat; Shi, Heliang; Habte, Habtom H; Qin, Yali; Cho, Michael W

    2016-03-01

    The C-terminal alpha-helix of gp41 membrane-proximal external region (MPER; (671)NWFDITNWLWYIK(683)) encompassing 4E10/10E8 epitopes is an attractive target for HIV-1 vaccine development. We previously reported that gp41-HR1-54Q, a trimeric protein comprised of the MPER in the context of a stable six-helix bundle (6HB), induced strong immune responses against the helix, but antibodies were directed primarily against the non-neutralizing face of the helix. To better target 4E10/10E8 epitopes, we generated four putative fusion intermediates by introducing double point mutations or deletions in the heptad repeat region 1 (HR1) that destabilize 6HB in varying degrees. One variant, HR1-∆10-54K, elicited antibodies in rabbits that targeted W672, I675 and L679, which are critical for 4E10/10E8 recognition. Overall, the results demonstrated that altering structural parameters of 6HB can influence immunogenic properties of the MPER and antibody targeting. Further exploration of this strategy could allow development of immunogens that could lead to induction of 4E10/10E8-like antibodies. Copyright © 2016 Elsevier Inc. All rights reserved.

  6. When Triple Helix Unravels: A Multi-Case Analysis of Failures in Industry-University Cooperative Research Centres

    Science.gov (United States)

    Gray, Denis; Sundstrom, Eric; Tornatzky, Louis G.; McGowen, Lindsey

    2011-01-01

    Cooperative research centres (CRCs) increasingly foster Triple Helix (industry-university-government) collaboration and represent significant vehicles for cooperation across sectors, the promotion of knowledge and technology transfer and ultimately the acceleration of innovation. A growing social science literature on CRCs focuses on their…

  7. Behavioural modification of the optic tentacle of Helix pom atia; effect of puromycin, activity of S-100

    DEFF Research Database (Denmark)

    Christoffersen, Gert Rene Juul; Frederiksen, K; Johansen, Jørgen

    1981-01-01

    1. 1. A long-term learning phenomenon in the tentacle of Helix pomatia has been observed.   2. 2. Repeated mechanical stimulation of the optic tentacle led to habituation of the associated withdrawal-extension action pattern whereas repeated combined mechanical and electrical stimulation...

  8. Peptide–oligonucleotide conjugates as nanoscale building blocks for assembly of an artificial three-helix protein mimic

    DEFF Research Database (Denmark)

    Lou, Chenguang; Martos Maldonado, Manuel Cristo; Madsen, Charlotte Stahl

    2016-01-01

    (CD) spectroscopy, small-angle X-ray scattering and transmission electron microscopy. These studies revealed the formation of the desired triple helix and coiled coil domains at low concentrations, while a dimer of trimers was dominating at high concentration. CD spectroscopy showed an extraordinarily...

  9. Temperature, salinity and transmissivity data from the Alpha Helix in the Arctic Ocean, 2000 - 2004 (NODC Accession 0059005)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — This is calibrated CTD downcast data from five Alpha Helix cruises: HX235, 8/1/2000 - 9/30/2000 HX250, 9/1/2001 - 9/30/2001 HX260, 6/1/2002 - 6/30/2002 HX274,...

  10. Salt bridge interactions within the β2integrin α7helix mediate force-induced binding and shear resistance ability.

    Science.gov (United States)

    Zhang, Xiao; Li, Linda; Li, Ning; Shu, Xinyu; Zhou, Lüwen; Lü, Shouqin; Chen, Shenbao; Mao, Debin; Long, Mian

    2018-01-01

    The functional performance of the αI domain α 7 helix in β 2 integrin activation depends on the allostery of the α 7 helix, which axially slides down; therefore, it is critical to elucidate what factors regulate the allostery. In this study, we determined that there were two conservative salt bridge interaction pairs that constrain both the upper and bottom ends of the α 7 helix. Molecular dynamics (MD) simulations for three β 2 integrin members, lymphocyte function-associated antigen-1 (LFA-1; α L β 2 ), macrophage-1 antigen (Mac-1; α M β 2 ) and α x β 2 , indicated that the magnitude of the salt bridge interaction is related to the stability of the αI domain and the strength of the corresponding force-induced allostery. The disruption of the salt bridge interaction, especially with double mutations in both salt bridges, significantly reduced the force-induced allostery time for all three members. The effects of salt bridge interactions of the αI domain α 7 helix on β 2 integrin conformational stability and allostery were experimentally validated using Mac-1 constructs. The results demonstrated that salt bridge mutations did not alter the conformational state of Mac-1, but they did increase the force-induced ligand binding and shear resistance ability, which was consistent with MD simulations. This study offers new insight into the importance of salt bridge interaction constraints of the αI domain α 7 helix and external force for β 2 integrin function. © 2017 Federation of European Biochemical Societies.

  11. A Single-Center Retrospective Cohort Analysis of Maternal and Infant Outcomes in HIV-Infected Mothers Treated with Integrase Inhibitors During Pregnancy.

    Science.gov (United States)

    Mounce, Monique L; Pontiggia, Laura; Adams, Jessica L

    2017-12-01

    Integrase strand transfer inhibitors (INSTI) are currently being investigated for the treatment of HIV in pregnancy. The purpose of this study is to evaluate the differences in maternal and infant outcomes in HIV-positive mothers treated with INSTI-containing antiretroviral therapy (ART) during pregnancy compared to protease inhibitor (PI)-containing ART. A retrospective, cohort study of INSTI- and PI-based ART used in pregnancy between 2007 and 2015 was performed. The primary objective was to evaluate the differences in viral load (VL) suppression prior to delivery. Secondary endpoints included time to and duration of VL suppression and safety parameters in both mothers and infants. For the primary analysis, the two arms were matched 1:2 INSTI to PI based on the presence or absence of viremia at the time of pregnancy determination. Additional analysis was performed on the entire matched and unmatched dataset. Twenty-one patients were matched (7 INSTI and 14 PI). There were no significant differences between groups with respect to the proportion of patients with VL suppression prior to delivery (71.4% INSTI vs. 92.9% PI, p = 0.247), and there were no significant differences in any of the secondary endpoints. Patients with documented adherence issues were statistically more likely to not be virologically suppressed prior to delivery (p = 0.002). No differences in efficacy or safety were found between patients treated with INSTIs compared to PIs. This study supports the further investigation of the use of INSTIs during pregnancy to reduce HIV transmission.

  12. Marine integrons containing novel integrase genes, attachment sites, attI, and associated gene cassettes in polluted sediments from Suez and Tokyo Bays.

    Science.gov (United States)

    Elsaied, Hosam; Stokes, Hatch W; Kitamura, Keiko; Kurusu, Yasurou; Kamagata, Yoichi; Maruyama, Akihiko

    2011-07-01

    In order to understand the structure and biological significance of integrons and associated gene cassettes in marine polluted sediments, metagenomic DNAs were extracted from sites at Suez and Tokyo Bays. PCR amplicons containing new integrase genes, intI, linked with novel gene cassettes, were recovered and had sizes from 1.8 to 2.5 kb. This approach uncovered, for the first time, the structure and diversity of both marine integron attachment site, attI, and the first gene cassette, the most efficiently expressed integron-associated gene cassette. The recovered 13 and 20 intI phylotypes, from Suez and Tokyo Bay samples, respectively, showed a highly divergence, suggesting a difference in integron composition between the sampling sites. Some intI phylotypes showed similarity with that from Geobacter metallireducens, belonging to Deltaproteobacteria, the dominant class in both sampling sites, as determined by 16S rRNA gene analysis. Thirty distinct families of putative attI site, as determined by the presence of an attI-like simple site, were recovered. A total of 146 and 68 gene cassettes represented Suez and Tokyo Bay unsaturated cassette pools, respectively. Gene cassettes, including a first cassette, from both sampling sites encoded two novel families of glyoxalase/bleomycin antibiotic-resistance protein. Gene cassettes from Suez Bay encoded proteins similar to haloacid dehalogenases, protein disulfide isomerases and death-on-curing and plasmid maintenance system killer proteins. First gene cassettes from Tokyo Bay encoded a xenobiotic-degrading protein, cardiolipin synthetase, esterase and WD40-like β propeller protein. Many of the first gene cassettes encoded proteins with no ascribable function but some of them were duplicated and possessed signal functional sites, suggesting efficient adaptive functions to their bacterial sources. Thus, each sampling site had a specific profile of integrons and cassette types consistent with the hypothesis that the

  13. Development of pharmacophore similarity-based quantitative activity hypothesis and its applicability domain: applied on a diverse data-set of HIV-1 integrase inhibitors.

    Science.gov (United States)

    Kumar, Sivakumar Prasanth; Jasrai, Yogesh T; Mehta, Vijay P; Pandya, Himanshu A

    2015-01-01

    Quantitative pharmacophore hypothesis combines the 3D spatial arrangement of pharmacophore features with biological activities of the ligand data-set and predicts the activities of geometrically and/or pharmacophoric similar ligands. Most pharmacophore discovery programs face difficulties in conformational flexibility, molecular alignment, pharmacophore features sampling, and feature selection to score models if the data-set constitutes diverse ligands. Towards this focus, we describe a ligand-based computational procedure to introduce flexibility in aligning the small molecules and generating a pharmacophore hypothesis without geometrical constraints to define pharmacophore space, enriched with chemical features necessary to elucidate common pharmacophore hypotheses (CPHs). Maximal common substructure (MCS)-based alignment method was adopted to guide the alignment of carbon molecules, deciphered the MCS atom connectivity to cluster molecules in bins and subsequently, calculated the pharmacophore similarity matrix with the bin-specific reference molecules. After alignment, the carbon molecules were enriched with original atoms in their respective positions and conventional pharmacophore features were perceived. Distance-based pharmacophoric descriptors were enumerated by computing the interdistance between perceived features and MCS-aligned 'centroid' position. The descriptor set and biological activities were used to develop support vector machine models to predict the activities of the external test set. Finally, fitness score was estimated based on pharmacophore similarity with its bin-specific reference molecules to recognize the best and poor alignments and, also with each reference molecule to predict outliers of the quantitative hypothesis model. We applied this procedure to a diverse data-set of 40 HIV-1 integrase inhibitors and discussed its effectiveness with the reported CPH model.

  14. Helix Nebula - the Science Cloud: a public-private partnership to build a multidisciplinary cloud platform for data intensive science

    Science.gov (United States)

    Jones, Bob; Casu, Francesco

    2013-04-01

    The feasibility of using commercial cloud services for scientific research is of great interest to research organisations such as CERN, ESA and EMBL, to the suppliers of cloud-based services and to the national and European funding agencies. Through the Helix Nebula - the Science Cloud [1] initiative and with the support of the European Commission, these stakeholders are driving a two year pilot-phase during which procurement processes and governance issues for a framework of public/private partnership will be appraised. Three initial flagship use cases from high energy physics, molecular biology and earth-observation are being used to validate the approach, enable a cost-benefit analysis to be undertaken and prepare the next stage of the Science Cloud Strategic Plan [2] to be developed and approved. The power of Helix Nebula lies in a shared set of services for initially 3 very different sciences each supporting a global community and thus building a common e-Science platform. Of particular relevance is the ESA sponsored flagship application SuperSites Exploitation Platform (SSEP [3]) that offers the global geo-hazard community a common platform for the correlation and processing of observation data for supersites monitoring. The US-NSF Earth Cube [4] and Ocean Observatory Initiative [5] (OOI) are taking a similar approach for data intensive science. The work of Helix Nebula and its recent architecture model [6] has shown that is it technically feasible to allow publicly funded infrastructures, such as EGI [7] and GEANT [8], to interoperate with commercial cloud services. Such hybrid systems are in the interest of the existing users of publicly funded infrastructures and funding agencies because they will provide "freedom of choice" over the type of computing resources to be consumed and the manner in which they can be obtained. But to offer such freedom-of choice across a spectrum of suppliers, various issues such as intellectual property, legal responsibility

  15. Infrared study of synthetic peptide analogues of the calcium-binding site III of troponin C: The role of helix F of an EF-hand motif.

    Science.gov (United States)

    Nara, Masayuki; Morii, Hisayuki; Tanokura, Masaru

    2013-05-01

    The EF-hand motif (helix-loop-helix) is a Ca(2+)-binding domain that is common among many intracellular Ca(2+)-binding proteins. We applied Fourier-transform infrared spectroscopy to study the synthetic peptide analogues of site III of rabbit skeletal muscle troponin C (helix E-loop-helix F). The 17-residue peptides corresponding to loop-helix F (DRDADGYIDAEELAEIF), where one residue is substituted by the D-type amino acid, were investigated to disturb the α-helical conformation of helix F systematically. These D-type-substituted peptides showed no band at about 1555 cm(-1) even in the Ca(2+)-loaded state although the native peptide (L-type only) showed a band at about 1555 cm(-1) in the Ca(2+)-loaded state, which is assigned to the side-chain COO(-) group of Glu at the 12th position, serving as the ligand for Ca(2+) in the bidentate coordination mode. Therefore, helix F is vital to the interaction between the Ca(2+) and the side-chain COO(-) group of Glu at the 12th position. Implications of the COO(-) antisymmetric stretch and the amide-I' of the synthetic peptide analogues of the Ca(2+)-binding sites are discussed. Copyright © 2012 Wiley Periodicals, Inc.

  16. Technology Entreprenurship in the Changing Business Environment – A Triple Helix Performance Model

    Directory of Open Access Journals (Sweden)

    Maja Levi Jaksić

    2015-02-01

    Full Text Available In this paper the contribution of technology management and entrepreneurship to sustainable development is emphasized and the Triple Helix (TH model is used to analyse the performance of different actors in accomplishing the activities of Technology Innovation Management and Entrepreneurship (TIME. By analysing TH model (Government – University – Industry in relation to accomplishing TIME main functions: Planning, Organizing and Control (POC, we created a general model which measures TIME effectiveness related to the key elements of the TH model. The general model – TMD-TH (Technology Management and Development – Triple Helix represents the framework for further more specific research into the relations of the observed dimensions. From the general model, three sub-models are excluded: TMD-G (Government, TMD-U (University and TMD-I (Industry, and each of TH dimensions is observed by a set of indicators classified from the perspective of the specific function (POC of TIME. This provides better categorization of TIME indicators and linkage with the actors in the TH model. The applicability of the suggested general model was tested by a set of indicators at the example of Serbia, Austria and Finland and comparison of these countries was made from a perspective of innovativeness and sustainable development. Since technology, innovation and entrepreneurship are considered as the main forces leading to sustainable development at different levels of the economy and society, it is of paramount importance to develop our capacities to better monitor, analyse and develop these forces. A model is developed with a set of indicators that enables the systematic analysis in concrete situations in practice. In this paper, the model is applied at the level of the national economy, the results obtained point to the most critical activities of the actors in the TH model in accomplishing TIME. The performance model represents a base for developing policies

  17. Overlapping effector interfaces define the multiple functions of the HIV-1 Nef polyproline helix

    Directory of Open Access Journals (Sweden)

    Kuo Lillian S

    2012-05-01

    proteins. To the N- and C- terminal sides of the polyproline helix are multifunctional protein interaction sites. The polyproline segment is also adapted to downregulate MHCI with a non-canonical binding surface. Our results demonstrate that Nef polyproline helix is highly adapted to directly interact with multiple host cell proteins.

  18. The Nup62 Coiled-Coil Motif Provides Plasticity for Triple-Helix Bundle Formation.

    Science.gov (United States)

    Dewangan, Pravin S; Sonawane, Parshuram J; Chouksey, Ankita R; Chauhan, Radha

    2017-06-06

    The central transport channel of the vertebrate nuclear pore complex (NPC) consists of nucleoporins: Nup62, Nup54, and Nup58. The coiled-coil domains in α-helical regions of these nucleoporins are thought to be crucial for several protein-protein interactions in the NPC subcomplexes. In this study, we determined the crystal structure of the coiled-coil domain of rat Nup62 fragment (residues 362-425) to 2.4 Å resolution. The crystal structure shows the conserved coiled-coil domain as a parallel three-helix bundle for the Nup62(362-425) fragment. On the basis of our size exclusion chromatography coupled to multiangle light scattering analysis and glutaraldehyde cross-linking experiments, we conclude that the Nup62(362-425) fragment displays dynamic behavior in solution and can also exist in either homodimeric or homotrimeric states. Our comparative analysis of the rat Nup62(362-425) homotrimeric structure with previously reported heterotrimeric structures [rat Nup62(362-425)·Nup54(346-407) and Xenopus Nup62(358-485)·Nup54(315-450)·Nup58(283-406) complexes] demonstrates the structural basis for parallel triple-helix bundle formation for Nup62 with different partners. Moreover, we show that the coiled-coil domain of Nup62 is sufficient for interaction with the coiled-coil domain of rat Exo70, a protein in an exocyst complex. On the basis of these observations, we suggest the plausible chain replacement mechanism that yields to diverse protein assemblies with Nup62. In summary, the coiled-coil motif present in Nup62 imparts the ability to form a homotrimer and heterotrimers either with Nup54 or with Nup54-Nup58 within the NPCs as well as with Exo70 beyond the NPCs. These complexes of Nup62 suggest the crucial role of the coiled-coil motifs in providing plasticity to various modular assemblies.

  19. An amphipathic alpha-helix controls multiple roles of brome mosaic virus protein 1a in RNA replication complex assembly and function.

    Directory of Open Access Journals (Sweden)

    Ling Liu

    2009-03-01

    Full Text Available Brome mosaic virus (BMV protein 1a has multiple key roles in viral RNA replication. 1a localizes to perinuclear endoplasmic reticulum (ER membranes as a peripheral membrane protein, induces ER membrane invaginations in which RNA replication complexes form, and recruits and stabilizes BMV 2a polymerase (2a(Pol and RNA replication templates at these sites to establish active replication complexes. During replication, 1a provides RNA capping, NTPase and possibly RNA helicase functions. Here we identify in BMV 1a an amphipathic alpha-helix, helix A, and use NMR analysis to define its structure and propensity to insert in hydrophobic membrane-mimicking micelles. We show that helix A is essential for efficient 1a-ER membrane association and normal perinuclear ER localization, and that deletion or mutation of helix A abolishes RNA replication. Strikingly, mutations in helix A give rise to two dramatically opposite 1a function phenotypes, implying that helix A acts as a molecular switch regulating the intricate balance between separable 1a functions. One class of helix A deletions and amino acid substitutions markedly inhibits 1a-membrane association and abolishes ER membrane invagination, viral RNA template recruitment, and replication, but doubles the 1a-mediated increase in 2a(Pol accumulation. The second class of helix A mutations not only maintains efficient 1a-membrane association but also amplifies the number of 1a-induced membrane invaginations 5- to 8-fold and enhances viral RNA template recruitment, while failing to stimulate 2a(Pol accumulation. The results provide new insights into the pathways of RNA replication complex assembly and show that helix A is critical for assembly and function of the viral RNA replication complex, including its central role in targeting replication components and controlling modes of 1a action.

  20. Molecular analysis of the human SLC13A4 sulfate transporter gene promoter

    Energy Technology Data Exchange (ETDEWEB)

    Jefferis, J. [Mater Medical Research Institute, South Brisbane, Queensland (Australia); Rakoczy, J. [Mater Medical Research Institute, South Brisbane, Queensland (Australia); School of Biomedical Sciences, University of Queensland, St. Lucia, Queensland (Australia); Translational Research Institute, Woolloongabba, Queensland (Australia); Simmons, D.G. [School of Biomedical Sciences, University of Queensland, St. Lucia, Queensland (Australia); Dawson, P.A., E-mail: paul.dawson@mmri.mater.org.au [Mater Medical Research Institute, South Brisbane, Queensland (Australia); Translational Research Institute, Woolloongabba, Queensland (Australia)

    2013-03-29

    Highlights: ► Basal promoter activity of SLC13A4 −57 to −192 nt upstream of transcription initiation site. ► Human SLC13A4 5′-flanking region has conserved motifs with other placental species. ► Putative NFY, SP1 and KLF7 motifs in SLC13A4 5′-flanking region enhance transcription. -- Abstract: The human solute linked carrier (SLC) 13A4 gene is primarily expressed in the placenta where it is proposed to mediate the transport of nutrient sulfate from mother to fetus. The molecular mechanisms involved in the regulation of SLC13A4 expression remain unknown. To investigate the regulation of SLC13A4 gene expression, we analysed the transcriptional activity of the human SLC13A4 5′-flanking region in the JEG-3 placental cell line using luciferase reporter assays. Basal transcriptional activity was identified in the region −57 to −192 nucleotides upstream of the SLC13A4 transcription initiation site. Mutational analysis of the minimal promoter region identified Nuclear factor Y (NFY), Specificity protein 1 (SP1) and Krüppel like factor 7 (KLF7) motifs which conferred positive transcriptional activity, as well as Zinc finger protein of the cerebellum 2 (ZIC2) and helix–loop–helix protein 1 (HEN1) motifs that repressed transcription. The conserved NFY, SP1, KLF7, ZIC2 and HEN1 motifs in the SLC13A4 promoter of placental species but not in non-placental species, suggests a potential role for these putative transcriptional factor binding motifs in the physiological control of SLC13A4 mRNA expression.

  1. The Impact of the ‘Austrian’ Mutation of the Amyloid Precursor Protein Transmembrane Helix is Communicated to the Hinge Region

    DEFF Research Database (Denmark)

    Stelzer, Walter; Scharnagl, Christina; Leurs, Ulrike

    2016-01-01

    The transmembrane helix of the amyloid precursor protein is subject to proteolytic cleavages by γ-secretase at different sites resulting in Aβ peptides of different length and toxicity. A number of point mutations within this transmembrane helix alter the cleavage pattern thus enhancing production...... destabilizes amide hydrogen bonds in the hinge which connects dimerization and cleavage regions. Weaker intrahelical hydrogen bonds at the hinge may enhance helix bending and thereby affect recognition of the transmembrane substrate by the enzyme and/or presentation of its cleavage sites to the catalytic cleft....

  2. Biochemical characterization and helix stabilizing properties of HSNP-C' from the thermoacidophilic archaeon Sulfolobus acidocaldarius.

    Science.gov (United States)

    Celestina, F; Suryanarayana, T

    2000-01-19

    Helix stabilizing nucleoid protein HSNP-C' from the thermophilic archaeon Sulfolobus acidocaldarius has been characterized with respect to its interactions with nucleic acids by gel retardation assay, affinities to immobilized matrices, electron microscopy, and fluorescence titration. The amino acids implicated in the DNA binding site of the protein have been shown by selectively modifying specific amino acyl functional groups and looking at their effects on the DNA binding properties of the protein. Lysine, arginine, tryptophan, and tyrosine residues of the protein HSNP-C' were modified with pyridoxal-5-phosphate; 2,3-butanedione; BNPS-skatole; and tetranitromethane, respectively. The modification of residues was assessed according to standard procedures. The effect of the chemical modification on the function of the protein HSNP-C' with respect to DNA protein interactions was studied and the results indicate the definite involvement of tyrosines and also the significant involvement of the flanking tryptophan residues in the DNA binding domain on the protein. Copyright 2000 Academic Press.

  3. Isolation and chemical analysis of nanoparticles from English ivy (Hedera helix L.).

    Science.gov (United States)

    Lenaghan, Scott C; Burris, Jason N; Chourey, Karuna; Huang, Yujian; Xia, Lijin; Lady, Belinda; Sharma, Ritin; Pan, Chongle; LeJeune, Zorabel; Foister, Shane; Hettich, Robert L; Stewart, C Neal; Zhang, Mingjun

    2013-10-06

    Bio-inspiration for novel adhesive development has drawn increasing interest in recent years with the discovery of the nanoscale morphology of the gecko footpad and mussel adhesive proteins. Similar to these animal systems, it was discovered that English ivy (Hedera helix L.) secretes a high strength adhesive containing uniform nanoparticles. Recent studies have demonstrated that the ivy nanoparticles not only contribute to the high strength of this adhesive, but also have ultraviolet (UV) protective abilities, making them ideal for sunscreen and cosmetic fillers, and may be used as nanocarriers for drug delivery. To make these applications a reality, the chemical nature of the ivy nanoparticles must be elucidated. In the current work, a method was developed to harvest bulk ivy nanoparticles from an adventitious root culture system, and the chemical composition of the nanoparticles was analysed. UV/visible spectroscopy, inductively coupled plasma mass spectrometry, Fourier transform infrared spectroscopy and electrophoresis were used in this study to identify the chemical nature of the ivy nanoparticles. Based on this analysis, we conclude that the ivy nanoparticles are proteinaceous.

  4. 4-twist helix snake to maintain polarization in multi-GeV proton rings

    Directory of Open Access Journals (Sweden)

    F. Antoulinakis

    2017-09-01

    Full Text Available Solenoid Siberian snakes have successfully maintained polarization in particle rings below 1 GeV, but never in multi-GeV rings, because the spin rotation by a solenoid is inversely proportional to the beam momentum. High energy rings, such as Brookhaven’s 255 GeV Relativistic Heavy Ion Collider (RHIC, use only odd multiples of pairs of transverse B-field Siberian snakes directly opposite each other. When it became impractical to use a pair of Siberian Snakes in Fermilab’s 120  GeV/c Main Injector, we searched for a new type of single Siberian snake that could overcome all depolarizing resonances in the 8.9–120  GeV/c range. We found that a snake made of one 4-twist helix and 2 dipoles could maintain the polarization. This snake design could solve the long-standing problem of significant polarization loss during acceleration of polarized protons from a few GeV to tens of GeV, such as in the AGS, before injecting them into multi-hundred GeV rings, such as RHIC.

  5. Measurements of Drag Coefficients and Rotation Rates of Free-Falling Helixes

    KAUST Repository

    Al-Omari, Abdulrhaman A.

    2016-05-01

    The motion of bacteria in the environment is relevant to several fields. At very small scales and with simple helical shapes, we are able to describe experimentally and mathematically the motion of solid spirals falling freely within a liquid pool. Using these shapes we intend to mimic the motion of bacteria called Spirochetes. We seek to experimentally investigate the linear and the rotational motion of such shapes. A better understanding of the dynamics of this process will be practical not only on engineering and physics, but the bioscience and environmental as well. In the following pages, we explore the role of the shape on the motion of passive solid helixes in different liquids. We fabricate three solid helical shapes and drop them under gravity in water, glycerol and a mixture of 30% glycerol in water. That generated rotation due to helical angle in water. However, we observe the rotation disappear in glycerol. The movement of the solid helical shapes is imaged using a high-speed video camera. Then, the images are analyzed using the supplied software and a computer. Using these simultaneous measurements, we examine the terminal velocity of solid helical shapes. Using this information we computed the drag coefficient and the drag force. We obtain the helical angular velocity and the torque applied to the solid. The results of this study will allow us to more accurately predict the motion of solid helical shape. This analysis will also shed light onto biological questions of bacteria movement.

  6. Physics and application of persistent spin helix state in semiconductor heterostructures

    Science.gov (United States)

    Kohda, Makoto; Salis, Gian

    2017-07-01

    In order to utilize the spin degree of freedom in semiconductors, control of spin states and transfer of the spin information are fundamental requirements for future spintronic devices and quantum computing. Spin orbit (SO) interaction generates an effective magnetic field for moving electrons and enables spin generation, spin manipulation and spin detection without using external magnetic field and magnetic materials. However, spin relaxation also takes place due to a momentum dependent SO-induced effective magnetic field. As a result, SO interaction is considered to be a double-edged sword facilitating spin control but preventing spin transport over long distances. The persistent spin helix (PSH) state solves this problem since uniaxial alignment of the SO field with SU(2) symmetry enables the suppression of spin relaxation while spin precession can still be controlled. Consequently, understanding the PSH becomes an important step towards future spintronic technologies for classical and quantum applications. Here, we review recent progress of PSH in semiconductor heterostructures and its device application. Fundamental physics of SO interaction and the conditions of a PSH state in semiconductor heterostructures are discussed. We introduce experimental techniques to observe a PSH and explain both optical and electrical measurements for detecting a long spin relaxation time and the formation of a helical spin texture. After emphasizing the bulk Dresselhaus SO coefficient γ, the application of PSH states for spin transistors and logic circuits are discussed.

  7. Role of Side-Chain Conformational Entropy in Transmembrane Helix Dimerization of Glycophorin A

    Science.gov (United States)

    Liu, Wei; Crocker, Evan; Siminovitch, David J.; Smith, Steven O.

    2003-01-01

    Dimerization of the transmembrane domain of glycophorin A is mediated by a seven residue motif LIxxGVxxGVxxT through a combination of van der Waals and hydrogen bonding interactions. One of the unusual features of the motif is the large number of β-branched amino acids that may limit the entropic cost of dimerization by restricting side-chain motion in the monomeric transmembrane helix. Deuterium NMR spectroscopy is used to characterize the dynamics of fully deuterated Val80 and Val84, two essential amino acids of the dimerization motif. Deuterium spectra of the glycophorin A transmembrane dimer were obtained using synthetic peptides corresponding to the transmembrane sequence containing either perdeuterated Val80 or Val84. These data were compared with spectra of monomeric glycophorin A peptides deuterated at Val84. In all cases, the deuterium line shapes are characterized by fast methyl group rotation with virtually no motion about the Cα-Cβ bond. This is consistent with restriction of the side chain in both the monomer and dimer due to intrahelical packing interactions involving the β-methyl groups, and indicates that there is no energy cost associated with dimerization due to loss of conformational entropy. In contrast, deuterium NMR spectra of Met81 and Val82, in the lipid interface, reflected greater motional averaging and fast exchange between different side-chain conformers. PMID:12547806

  8. Effects of dietary exposure to forest pesticides on the brown garden snail Helix aspersa mueller

    Energy Technology Data Exchange (ETDEWEB)

    Schuytema, G.S.; Nebeker, A.V.; Griffis, W.L.

    1994-01-01

    Brown garden snails, Helix aspersa, were fed prepared diets with 12 pesticides used in forest spraying practices where endangered arboreal and terrestrial snails may be at risk. Acephate, atrazine, glyphosate, hexazinone, and picloram were not lethal at concentrations of 5,000 mg/kg in 14-day screening tests. The remaining seven pesticides, lethal to 13-100% of the tested snails at 5,000 mg/kg, were evaluated in 10-day definitive feeding tests. Azinphosmethyl (Guthion) and aminocarb were the most toxic, with 10-day LC50s of 188 and 313 mg/kg, respectively. Paraquat, trichlorfon and fenitrothion had 10-day LC50s of 659, 664, and 7,058 mg/kg respectively. Avoidance of pesticide-containing foods occurred, e.g., 10-day LC50s of >10,000 mg/kg for carbaryl and ethyl parathion. Significant descreases (p<0.05) in snail weight (total, shell-only, body-only) or shell diameter were accompanied by a significant decrease in the amount of food consumed/snail/day. Concentrations of pesticide in tissues were measured in snails exposed to atrazine and azinphosmethyl; there was no bioaccumulation. (Copyright (c) 1994 Springer-Verlag New York Inc.)

  9. Dust particulate absorption by ivy (Hedera helix L) on historic walls in urban environments

    International Nuclear Information System (INIS)

    Sternberg, Troy; Viles, Heather; Cathersides, Alan; Edwards, Mona

    2010-01-01

    The potential bio-protective role of urban greenery and how it interacts with airborne dust and pollutants has been the subject of much recent research. As particulate pollution has been implicated in both the deterioration of building materials and in damaging human health, understanding how it interacts with urban greenery is of great applied interest. Common or English Ivy (Hedera helix L) grows widely on urban walls in many parts of the world, and thus any bio-protective role it might play is of broad relevance. Using Scanning Electron Microscopy ivy leaves collected on roadways were examined to determine if ivy can absorb dust and pollutants that can instigate decay processes on stone walls and impact human health in urban environments. Results showed that ivy acts as a 'particle sink', absorbing particulate matter, particularly in high-traffic areas. It was effective in adhering fine ( 10 per m 2 . Our findings suggest that through absorbing pollutant particles ivy can retard bio-deteriorative processes on historic walls and reduce human exposure to respiratory problems caused by vehicle pollutants.

  10. Chaotic mixing in a helix-like pipe with periodic variations in curvature and torsion

    International Nuclear Information System (INIS)

    Jang, Bongkyun; Funakoshi, Mitsuaki

    2010-01-01

    Chaotic motion of fluid particles due to a steady viscous flow in a helix-like circular pipe caused by an axial pressure gradient, and the mixing efficiency of this flow are numerically examined here. The pipe is wound around a circular or elliptic cylinder with a constant pitch so that the curvature κ and torsion τ of the centerline of this pipe vary continuously and periodically. If both κ and τ are small and slowly varying, the cross-sectional motion of fluid particles is expected to be approximately governed by the sum of Dean's flow and the flow of rigid rotation. From Poincare sections and the values of an index of the extent of mixing, it is found that there is an intermediate range of Reynolds number Re of flow within which chaotic regions in Poincare sections are large and mixing efficiency over a short time is high. Moreover, larger chaotic regions and higher mixing efficiency are observed for pipes wound around a circular cylinder of smaller radius and for pipes wound around a thinner elliptic cylinder. These results can be explained by the variation in characteristic ratio λ=12τ/(κRe) in one period.

  11. Field-induced Gap and Quantized Charge Pumping in Nano-helix

    Energy Technology Data Exchange (ETDEWEB)

    Qi, Xiao-Liang; /Stanford U., Phys. Dept. /Tsinghua U., Beijing; Zhang, Shou-Cheng; /Stanford U., Phys. Dept.

    2010-02-15

    We propose several novel physical phenomena based on nano-scale helical wires. Applying a static electric field transverse to the helical wire induces a metal to insulator transition, with the band gap determined by the applied voltage. Similar idea can be applied to 'geometrically' constructing one-dimensional systems with arbitrary external potential. With a quadrupolar electrode configuration, the electric field could rotate in the transverse plane, leading to a quantized dc charge current proportional to the frequency of the rotation. Such a device could be used as a new standard for the high precession measurement of the electric current. The inverse effect implies that passing an electric current through a helical wire in the presence of a transverse static electric field can lead to a mechanical rotation of the helix. This effect can be used to construct nano-scale electro-mechanical motors. Finally, our methodology also enables new ways of controlling and measuring the electronic properties of helical biological molecules such as the DNA.

  12. After the double helix: Rosalind Franklin's research on Tobacco mosaic virus.

    Science.gov (United States)

    Creager, Angela N H; Morgan, Gregory J

    2008-06-01

    Rosalind Franklin is best known for her informative X-ray diffraction patterns of DNA that provided vital clues for James Watson and Francis Crick's double-stranded helical model. Her scientific career did not end when she left the DNA work at King's College, however. In 1953 Franklin moved to J. D. Bernal's crystallography laboratory at Birkbeck College, where she shifted her focus to the three-dimensional structure of viruses, obtaining diffraction patterns of Tobacco mosaic virus (TMV) of unprecedented detail and clarity. During the next five years, while making significant headway on the structural determination of TMV, Franklin maintained an active correspondence with both Watson and Crick, who were also studying aspects of virus structure. Developments in TMV research during the 1950s illustrate the connections in the emerging field of molecular biology between structural studies of nucleic acids and of proteins and viruses. They also reveal how the protagonists of the "race for the double helix" continued to interact personally and professionally during the years when Watson and Crick's model for the double-helical structure of DNA was debated and confirmed.

  13. The double helix of cultural assimilationism and neo-liberalism: citizenship in contemporary governmentality.

    Science.gov (United States)

    Schinkel, Willem; Van Houdt, Friso

    2010-12-01

    In this article the recent transformations of citizenship in the Netherlands are analysed in relation to a developing form of governmentality. We regard citizenship as a state regulated technique of in- and exclusion and a crucial instrument in the management of populations. Taking the Dutch contexts of immigration and integration as our case, we argue that cultural assimilationism and neo-liberalism appear in a double helix: they combine to form a new governmental strategy we call neo-liberal communitarianism. Neo-liberal communitarianism is the underlying rationale of a population management that operates both in an individualizing (citizenship as individual participation and responsibility) and a de-individualizing way ('community' at various aggregate and localized levels as frame of 'integration'). It thus combines a communitarian care of a Dutch culturally grounded national community - conceived as traditionally'enlightened' and 'liberal'- with a neo-liberal emphasis on the individual's responsibility to achieve membership of that community. 'Community' is thereby selectively seen as mobilized and present (when immigrant integration is concerned) or as latently present and still in need of mobilization (when indigenous Dutch are concerned). Concomitantly, a repressive responsibilization and a facilitative responsibilization are aimed at these two governmentally differentiated populations.

  14. Larval development of Angiostrongylus vasorum in the land snail Helix aspersa.

    Science.gov (United States)

    Di Cesare, Angela; Crisi, Paolo Emidio; Bartolini, Roberto; Iorio, Raffaella; Talone, Tonino; Filippi, Laura; Traversa, Donato

    2015-10-01

    The metastrongyloid nematode Angiostrongylus vasorum affects the heart and pulmonary arteries of dogs and wild animals. Over the recent years, dog angiostrongylosis has gained great attention in the veterinary community for the expansion of its geographic range and for a rise in the number of clinical cases. Global warming, changes in phenology of mollusc intermediate hosts and movements of wild reservoirs have been evocated in the spreading of mollusc-borne parasites, including A. vasorum. The land snail Helix aspersa, a vector of other respiratory metastrongyloids, is endemic in most regions of the World, where it is a pest outside its native Mediterranean range. In the present study, the susceptibility and suitability of H. aspersa as an intermediate host of A. vasorum were investigated along with the characteristics of larval recovery and development following two different ways of inoculation, i.e. experimental (group A) vs natural infection (group B). After infections, the snails were kept at environmental conditions for 2 months. Five snails from groups A and B were randomly selected, digested and examined at 15-day intervals for 2 months. L1s, L2s and L3s were microscopically identified based on key morphological and morphometric characteristics and their identity was genetically confirmed. The results showed that A. vasorum may reach the infective stage in H. aspersa and that uptake of larvae and parasitic burden within the snails depend on the grazing capability of the molluscs. Biological and epidemiological implications are discussed.

  15. Use of the land snail Helix aspersa for monitoring heavy metal soil contamination in Northeast Algeria.

    Science.gov (United States)

    Larba, R; Soltani, N

    2014-08-01

    The objective of this study was to investigate the impact of anthropogenic activities on soil quality using the land snail Helix aspersa as a bioindicator. Soil samples and snails were collected from several sites in Northeast Algeria during the summer and winter of 2010. All of the sites were chosen due to their proximity to industrial factories-a potential source of soil pollution via heavy metal contamination. The concentration of heavy metals (Pb, Cd, Mn, and Fe) in soil samples was analyzed using atomic absorption spectrophotometry. Activity levels of glutathione S-transferase (GST) and acetylcholinesterase (AChE), indicators of oxidative stress and neurotoxicity, respectively, were measured in snails collected from each site. GST and AChE activity were found to vary between sites and by season. The highest levels of GST activity were registered during the summer at sites closest to potential sources of pollution. AChE activity levels also peaked during the summer with the highest values recorded at the site in El Hadjar. These increased levels of bioindicative stress response correlated with increasing metal concentration in soil samples collected at each site.

  16. Imidacloprid induced alterations in enzyme activities and energy reserves of the land snail, Helix aspersa.

    Science.gov (United States)

    Radwan, M A; Mohamed, M S

    2013-09-01

    The in vivo sublethal toxic effects (0.2 and 0.6 LD50) of topically applied imidacloprid on biochemical biomarkers in the land snail, Helix aspersa was examined. Biochemical perturbations were assessed by measuring the three enzymatic (Acetylcholinesterase, AChE; catalase, CAT and glutathione-S-transferase, GST) activities and three energy reserves (protein, glycogen and lipids) in the snails. Snail samples were taken from each sublethal dose and control groups at 1, 3 and 7 days after treatment. The results revealed that there were overall decrease in AChE activity as well as depletion of lipids and glycogen contents in the imidacloprid-treated snails compared to control groups. The CAT and GST activities of treated snails with the sublethal doses of imidacloprid were significantly higher than those of untreated controls along the three times of exposure. Moreover, an increase in the level of total proteins was observed in animals treated with 0.6 LD50 imidacloprid compared to control groups. The alterations in all tested biochemical perturbations were most pronounced with the 0.6 LD50 than 0.2 LD50. This study suggests that alterations of the enzyme activities and energy reserves in this species that could be useful as biomarkers of imidacloprid exposure in the evaluation of terrestrial impacts of this insecticide. Copyright © 2013 Elsevier Inc. All rights reserved.

  17. Evaluation of the genotoxic potential of soil contaminated with mineral coal tailings on snail Helix aspersa.

    Science.gov (United States)

    de Souza, Melissa Rosa; da Silva, Fernanda Rabaioli; de Souza, Claudia Telles; Niekraszewicz, Liana; Dias, Johnny Ferraz; Premoli, Suziane; Corrêa, Dione Silva; Soares, Mariana do Couto; Marroni, Norma Possa; Morgam-Martins, Maria Isabel; da Silva, Juliana

    2015-11-01

    Coal remains an important source of energy, although the fuel is a greater environmental pollutant. Coal is a mixture of several chemicals, especially inorganic elements and polycyclic aromatic hydrocarbons (PAH). Many of these compounds have mutagenic and carcinogenic effects on organisms exposed to this mineral. In the town of Charqueadas (Brazil), the tailings from mining were used for landfill in the lower areas of the town, and the consequence is the formation of large deposits of this material. The purpose of this study was to evaluate the genotoxic potential of soil samples contaminated by coal waste in different sites at Charqueadas, using the land snail Helix aspersa as a biomonitor organism. Thirty terrestrial snails were exposed to different treatments: 20 were exposed to the soil from two different sites in Charqueadas (site 1 and 2; 10 in each group) and 10 non-exposed (control group). Hemolymph cells were collected after 24h, 5days and 7days of exposure and comet assay, micronucleus test, oxidative stress tests were performed. Furthermore, this study quantified the inorganic elements present in soil samples by the PIXE technique and polycyclic aromatic hydrocarbons (PAH) by HPLC. This evaluation shows that, in general, soils from sites in Charqueadas, demonstrated a genotoxic effect associated with increased oxidative stress, inorganic and PAH content. These results demonstrate that the coal pyrite tailings from Charqueadas are potentially genotoxic and that H. aspersa is confirmed to be a sensitive instrument for risk assessment of environmental pollution. Copyright © 2015 Elsevier Ltd. All rights reserved.

  18. Natural selection reduces energy metabolism in the garden snail, helix aspersa (cornu aspersum).

    Science.gov (United States)

    Artacho, Paulina; Nespolo, Roberto F

    2009-04-01

    Phenotypic selection is widely recognized as the primary cause of adaptive evolution in natural populations, a fact that has been documented frequently over the last few decades, mainly in morphological and life-history traits. The energetic definition of fitness predicts that natural selection will maximize the residual energy available for growth and reproduction, suggesting that energy metabolism could be a target of selection. To address this problem, we chose the garden snail, Helix aspersa (Cornu aspersum). We performed a seminatural experiment for measuring phenotypic selection on standard metabolic rate (SMR), the minimum cost of maintenance in ectotherm organisms. To discount selection on correlated traits, we included two additional whole-organism performance traits (mean speed and maximum force of dislodgement). We found a combination of linear (negative directional selection, beta=-0.106 +/- 0.06; P= 0.001) and quadratic (stabilizing selection, gamma=-0.012 +/- 0.033; P= 0.061) selection on SMR. Correlational selection was not significant for any possible pair of traits. This suggests that individuals with average-to-reduced SMRs were promoted by selection. To the best of our knowledge, this is the first study showing significant directional selection on the obligatory cost of maintenance in an animal, providing support for the energetic definition of fitness.

  19. Mechanisms of triggering H1 helix in prion proteins unfolding revealed by molecular dynamic simulation

    Science.gov (United States)

    Tseng, Chih-Yuan; Lee, H. C.

    2006-03-01

    In template-assistance model, normal Prion protein (PrP^C), the pathogen to cause several prion diseases such as Creutzfeldt-Jakob (CJD) in human, Bovine Spongiform Encephalopathy (BSE) in cow, and scrapie in sheep, converts to infectious prion (PrP^Sc) through a transient interaction with PrP^Sc. Furthermore, conventional studies showed S1-H1-S2 region in PrP^C to be the template of S1-S2 β-sheet in PrP^Sc, and Prion protein's conformational conversion may involve an unfolding of H1 and refolding into β-sheet. Here we prepare several mouse prion peptides that contain S1-H1-S2 region with specific different structures, which are corresponding to specific interactions, to investigate possible mechanisms to trigger H1 α-helix unfolding process via molecular dynamic simulation. Three properties, conformational transition, salt-bridge in H1, and hydrophobic solvent accessible surface (SAS) are analyzed. From these studies, we found the interaction that triggers H1 unfolding to be the one that causes dihedral angle at residue Asn^143 changes. Whereas interactions that cause S1 segment's conformational changes play a minor in this process. These studies offers an additional evidence for template-assistance model.

  20. 4-twist helix snake to maintain polarization in multi-GeV proton rings

    International Nuclear Information System (INIS)

    Antoulinakis, F.; Chen, Y.; Dutton, A.; Rossi De La Fuente, E.; Haupert, S.

    2017-01-01

    Solenoid Siberian snakes have successfully maintained polarization in particle rings below 1 GeV, but never in multi-GeV rings, because the spin rotation by a solenoid is inversely proportional to the beam momentum. High energy rings, such as Brookhaven’s 255 GeV Relativistic Heavy Ion Collider (RHIC), use only odd multiples of pairs of transverse B-field Siberian snakes directly opposite each other. When it became impractical to use a pair of Siberian Snakes in Fermilab’s 120 GeV/c Main Injector, we searched for a new type of single Siberian snake that could overcome all depolarizing resonances in the 8.9–120 GeV/c range. We found that a snake made of one 4-twist helix and 2 dipoles could maintain the polarization. Here, this snake design could solve the long-standing problem of significant polarization loss during acceleration of polarized protons from a few GeV to tens of GeV, such as in the AGS, before injecting them into multi-hundred GeV rings, such as RHIC.

  1. 4-twist helix snake to maintain polarization in multi-GeV proton rings

    Science.gov (United States)

    Antoulinakis, F.; Chen, Y.; Dutton, A.; Rossi De La Fuente, E.; Haupert, S.; Ljungman, E. A.; Myers, P. D.; Thompson, J. K.; Tai, A.; Aidala, C. A.; Courant, E. D.; Krisch, A. D.; Leonova, M. A.; Lorenzon, W.; Raymond, R. S.; Sivers, D. W.; Wong, V. K.; Yang, T.; Derbenev, Y. S.; Morozov, V. S.; Kondratenko, A. M.

    2017-09-01

    Solenoid Siberian snakes have successfully maintained polarization in particle rings below 1 GeV, but never in multi-GeV rings, because the spin rotation by a solenoid is inversely proportional to the beam momentum. High energy rings, such as Brookhaven's 255 GeV Relativistic Heavy Ion Collider (RHIC), use only odd multiples of pairs of transverse B-field Siberian snakes directly opposite each other. When it became impractical to use a pair of Siberian Snakes in Fermilab's 120 GeV /c Main Injector, we searched for a new type of single Siberian snake that could overcome all depolarizing resonances in the 8.9 - 120 GeV /c range. We found that a snake made of one 4-twist helix and 2 dipoles could maintain the polarization. This snake design could solve the long-standing problem of significant polarization loss during acceleration of polarized protons from a few GeV to tens of GeV, such as in the AGS, before injecting them into multi-hundred GeV rings, such as RHIC.

  2. Mirror symmetry breaking of silicon polymers--from weak bosons to artificial helix.

    Science.gov (United States)

    Fujiki, Michiya

    2009-01-01

    From elemental particles to human beings, matter and living worlds in our universe are dissymmetric with respect to mirror symmetry. Since the early 19th century, the origin of biomolecular handedness has been puzzling scientists. Nature's elegant bottom-up preference, however, sheds light on new concepts of generating, amplifying, and switching artificial polymers, supramolecules, liquid crystals, and organic crystals that can exhibit ambidextrous circular dichroism in the UV/Visible region with efficiency in production under milder ambient conditions. In the 1920s, Kipping, who first synthesized polysilanes with phenyl groups, had much interest in the handedness of inorganic and organic substances from 1898 to 1909 in his early research life. Polysilanes--which are soluble Si-Si bonded chain-like near-UV chromophores that carry a rich variety of organic groups--may become a bridge between animate and inanimate polymer systems. The present account focuses on several mirror symmetry breaking phenomena exemplified in polysilanes carrying chiral and/or achiral side groups, which are in isotropic dilute solution, as polymer particles dispersed in solution, and in a double layer film immobilized at the solid surface, and subtle differences in the helix, by dictating ultimately ultraweak chiral forces at subatomic, atomic, and molecular levels. Copyright 2009 The Japan Chemical Journal Forum and Wiley Periodicals, Inc.

  3. CFD analysis and flow model reduction for surfactant production in helix reactor

    Directory of Open Access Journals (Sweden)

    Nikačević N.M.

    2015-01-01

    Full Text Available Flow pattern analysis in a spiral Helix reactor is conducted, for the application in the commercial surfactant production. Step change response curves (SCR were obtained from numerical tracer experiments by three-dimensional computational fluid dynamics (CFD simulations. Non-reactive flow is simulated, though viscosity is treated as variable in the direction of flow, as it increases during the reaction. The design and operating parameters (reactor diameter, number of coils and inlet velocity are varied in CFD simulations, in order to examine the effects on the flow pattern. Given that 3D simulations are not practical for fast computations needed for optimization, scale-up and control, CFD flow model is reduced to one-dimensional axial dispersion (AD model with spatially variable dispersion coefficient. Dimensionless dispersion coefficient (Pe is estimated under different conditions and results are analyzed. Finally, correlation which relates Pe number with Reynolds number and number of coils from the reactor entrance is proposed for the particular reactor application and conditions.

  4. Helix and Drugs: Snails for Western Health Care From Antiquity to the Present.

    Science.gov (United States)

    Bonnemain, Bruno

    2005-03-01

    The land helix, or snail, has been used in medicine since antiquity and prepared according to several formulations. This historical report traces the understanding of their properties from the time of Hippocrates, who proposed the use of snail mucus against protoccle and Pliny who thought that the snail increased the speed of delivery and was "a sovereign remedy to treat pain related to burns, abscesses and other wounds", Galien recommended snails against hydrops foetails. In the 18th century, various snail "preparations" were also recommended for external use with dermatological disorders and internally for symptoms associated with tuberculosis and nephritis. Surprisingly, the 19th century saw a renewed interest in the pharmaceutical and medical use of snails with numerous indications for snail preparations. This interest in snails did not stop at the end of the 19th century. The 1945 edition of Dorvault devotes an entire paragraph to snails, indicating that the therapeutic usage of snails was still alive at that time. Recently the FDA has also shown an interest in snails. Ziconotide (SNXIII), a synthetic peptide coming from snail venom, has been under FDA review since 1999. Pre-clinical and clinical studies of this new drug are promising.

  5. Helix and Drugs: Snails for Western Health Care From Antiquity to the Present

    Directory of Open Access Journals (Sweden)

    Bruno Bonnemain

    2005-01-01

    Full Text Available The land helix, or snail, has been used in medicine since antiquity and prepared according to several formulations. This historical report traces the understanding of their properties from the time of Hippocrates, who proposed the use of snail mucus against protoccle and Pliny who thought that the snail increased the speed of delivery and was “a sovereign remedy to treat pain related to burns, abscesses and other wounds”, Galien recommended snails against hydrops foetails. In the 18th century, various snail “preparations” were also recommended for external use with dermatological disorders and internally for symptoms associated with tuberculosis and nephritis. Surprisingly, the 19th century saw a renewed interest in the pharmaceutical and medical use of snails with numerous indications for snail preparations. This interest in snails did not stop at the end of the 19th century. The 1945 edition of Dorvault devotes an entire paragraph to snails, indicating that the therapeutic usage of snails was still alive at that time. Recently the FDA has also shown an interest in snails. Ziconotide (SNXIII, a synthetic peptide coming from snail venom, has been under FDA review since 1999. Pre-clinical and clinical studies of this new drug are promising.

  6. Flexoelectro-optic properties of chiral nematic liquid crystals in the uniform standing helix configuration

    Science.gov (United States)

    Castles, F.; Morris, S. M.; Coles, H. J.

    2009-09-01

    The flexoelectro-optic effect describes the rotation of the optic axis of a short-pitch chiral nematic liquid crystal under the application of an electric field. We investigate the effect in the uniform standing helix, or “Grandjean” configuration. An in-plane electric field is applied. The director profile is determined numerically using a static one-dimensional continuum model with strong surface anchoring. The Berreman method is used to solve for plane-wave solutions to Maxwell’s equations, and predict the optical properties of the resulting structure in general cases. By using a chiral nematic with short pitch between crossed polarizers an optical switch may be generated. With no applied field the configuration is nontransmissive at normal incidence, but becomes transmissive with an applied field. For this case, numerical results using the Berreman method are supplemented with an analytic theory and found to be in good agreement. The transmitted intensity as a function of tilt, the contrast ratio, and the tilt required for full intensity modulation are presented. The angular dependence of the transmission is calculated and the isocontrast curves are plotted. For typical material and cell parameters a switching speed of 0.017 ms and contrast ratio of 1500:1 at normal incidence are predicted, at a switch-on tilt of 41.5 degrees. Experimental verification of the analytic and numerical models is provided.

  7. Knowledge-enhancing Helix: An Approach for Developing Key Academic Skills at Universities. A Case Study

    Directory of Open Access Journals (Sweden)

    Nadja Boeller

    2008-08-01

    Full Text Available In an increasingly e-literate society, new and media technologies are proliferating and traditional teaching approaches are challenged to meet new requirements. Other aspects such as teamwork and knowledge exchange are also becoming more important. Collaborative working methods are more dominant in the networked business environment. The vocational training at universities is therefore continuously facing with new challenges. The objective of this paper is to show how the implementation of a holistic teaching approach including the idea of blended learning can be an effective way to train students in collaborative and academic working skills, methodological expertise, information literacy, as well as making students aware of new developments in media literacy. This approach follows our proposed concept DIAMOND (Didactical Approach for Media Competence Development with a main focus on the implementation of the "knowledgeenhancing helix." This pedagogical concept was developed corresponding the requirements for "good online learning." The paper accordingly discusses the theoretical framework and presents a case study where the concept was implemented in practice.

  8. Role of collagen triple helix repeat containing-1 in tumor and inflammatory diseases

    Directory of Open Access Journals (Sweden)

    Qian Wu

    2017-01-01

    Full Text Available Initially, collagen triple helix repeat containing-1 (CTHRC1 is expressed mainly in adventitial fibroblasts and neointimal smooth muscle cells of balloon-injured vessels, and increases cell migration, promotes tissue repair in response to injury. A variety of studies demonstrated that over-expression of CTHRC1 in solid tumors results in enhancement of migration and invasion of tumor cells, and is associated with decreased overall survival and disease-free survival. CTHRC1 expression is elevated in hepatitis B virus-infected patients and highly correlated with hepatocellular carcinoma progression as well. Furthermore, CTHRC1 plays a pivotal role in a great many fields, including increases bone mass, prevents myelination, reverses collagen synthesis in keloid fibroblasts, and increases fibroblast-like synoviocytes migration speed and abundant production of arthritic pannus in rheumatoid arthritis. Therefore, it will provide new insight into the pathogenesis of tumor and autoimmune diseases, and will shed new light on the therapy of related clinical diseases.

  9. Humoral serotonin and dopamine modulate the feeding in the snail, Helix pomatia.

    Science.gov (United States)

    Hernádi, L; Kárpáti, L; Gyori, J; Vehovszky, Agnes; Hiripi, L

    2008-01-01

    We investigated the effect of elevated levels of humoral 5HT and DA on the feeding latency of Helix pomatia, 1 day, 3 days and 10 days following satiation, by injecting monoamines into the haemocoel. HPLC assay of monoamines showed that both 5HT and DA are present in pmol/ml concentrations in the haemolymph of both starved and non-starved animals. Elevated levels of 5HT and DA were most effective at decreasing the feeding latency 10 days following satiation when DA decreased the feeding latency in a concentration dependent manner between 10(-7) and 10(-5) M whereas 5HT levels decreased the feeding latency only at 10(-6) M but increased it at 10(-5) M. Immunocytochemistry revealed that both 5HT3 and D1 receptor-like immuno-reactivity are present in cell bodies located in the same areas of the buccal ganglia. Our observations suggest that both humoral DA and 5HT mutually modulate the activity of the feeding CPG through neurons which have these receptors.

  10. Modification-dependent restriction endonuclease, MspJI, flips 5-methylcytosine out of the DNA helix.

    Science.gov (United States)

    Horton, John R; Wang, Hua; Mabuchi, Megumu Yamada; Zhang, Xing; Roberts, Richard J; Zheng, Yu; Wilson, Geoffrey G; Cheng, Xiaodong

    2014-10-29

    MspJI belongs to a family of restriction enzymes that cleave DNA containing 5-methylcytosine (5mC) or 5-hydroxymethylcytosine (5hmC). MspJI is specific for the sequence 5(h)mC-N-N-G or A and cleaves with some variability 9/13 nucleotides downstream. Earlier, we reported the crystal structure of MspJI without DNA and proposed how it might recognize this sequence and catalyze cleavage. Here we report its co-crystal structure with a 27-base pair oligonucleotide containing 5mC. This structure confirms that MspJI acts as a homotetramer and that the modified cytosine is flipped from the DNA helix into an SRA-like-binding pocket. We expected the structure to reveal two DNA molecules bound specifically to the tetramer and engaged with the enzyme's two DNA-cleavage sites. A coincidence of crystal packing precluded this organization, however. We found that each DNA molecule interacted with two adjacent tetramers, binding one specifically and the other non-specifically. The latter interaction, which prevented cleavage-site engagement, also involved base flipping and might represent the sequence-interrogation phase that precedes specific recognition. MspJI is unusual in that DNA molecules are recognized and cleaved by different subunits. Such interchange of function might explain how other complex multimeric restriction enzymes act. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

  11. Lipid Binding of the Amphipathic Helix Serving as Membrane Anchor of Pestivirus Glycoprotein Erns.

    Science.gov (United States)

    Aberle, Daniel; Oetter, Kay-Marcus; Meyers, Gregor

    2015-01-01

    Pestiviruses express a peculiar protein named Erns representing envelope glycoprotein and RNase, which is important for control of the innate immune response and persistent infection. The latter functions are connected with secretion of a certain amount of Erns from the infected cell. Retention/secretion of Erns is most likely controlled by its unusual membrane anchor, a long amphipathic helix attached in plane to the membrane. Here we present results of experiments conducted with a lipid vesicle sedimentation assay able to separate lipid-bound from unbound protein dissolved in the water phase. Using this technique we show that a protein composed of tag sequences and the carboxyterminal 65 residues of Erns binds specifically to membrane vesicles with a clear preference for compositions containing negatively charged lipids. Mutations disturbing the helical folding and/or amphipathic character of the anchor as well as diverse truncations and exchange of amino acids important for intracellular retention of Erns had no or only small effects on the proteins membrane binding. This result contrasts the dramatically increased secretion rates observed for Erns proteins with equivalent mutations within cells. Accordingly, the ratio of secreted versus cell retained Erns is not determined by the lipid affinity of the membrane anchor.

  12. Lipid Binding of the Amphipathic Helix Serving as Membrane Anchor of Pestivirus Glycoprotein Erns.

    Directory of Open Access Journals (Sweden)

    Daniel Aberle

    Full Text Available Pestiviruses express a peculiar protein named Erns representing envelope glycoprotein and RNase, which is important for control of the innate immune response and persistent infection. The latter functions are connected with secretion of a certain amount of Erns from the infected cell. Retention/secretion of Erns is most likely controlled by its unusual membrane anchor, a long amphipathic helix attached in plane to the membrane. Here we present results of experiments conducted with a lipid vesicle sedimentation assay able to separate lipid-bound from unbound protein dissolved in the water phase. Using this technique we show that a protein composed of tag sequences and the carboxyterminal 65 residues of Erns binds specifically to membrane vesicles with a clear preference for compositions containing negatively charged lipids. Mutations disturbing the helical folding and/or amphipathic character of the anchor as well as diverse truncations and exchange of amino acids important for intracellular retention of Erns had no or only small effects on the proteins membrane binding. This result contrasts the dramatically increased secretion rates observed for Erns proteins with equivalent mutations within cells. Accordingly, the ratio of secreted versus cell retained Erns is not determined by the lipid affinity of the membrane anchor.

  13. Triple Helix and European Union (EU Funding: The case of Latin America, especially Mexico and the Seventh European Framework Program

    Directory of Open Access Journals (Sweden)

    Jürgen Haberleithner

    2010-05-01

    Full Text Available The following analysis not only seeks to develop new potential intervention models; it also aims to create a detailed analysis of the existing problems with regards to communication among the active participants of Triple Helix (especially in Mexico. The special situation in Latin America with regards to existing corruption, the unequal distribution of power between the government and the private sector, dependence on other economies and other social issues will be analysed in accordance with the main focus of the investigation. The subsequent linking of potential partners to the development of an initiative for submitting a future Triple Helix/FP7 project represent an important contribution to a longer-term perspective on the preceding investigation. The potential partnerships between Europe and Latin America (in addition to other possible world regions, such as, for example, Pacific Asia will create an initial project draft within the scope of the conference.

  14. Conservation of a helix-stabilizing dipole moment in the PP-fold family of regulatory peptides

    DEFF Research Database (Denmark)

    Bjørnholm, B; Jørgensen, Flemming Steen; Schwartz, T W

    1993-01-01

    of the electrostatic properties of five representative members of the PP-fold family of peptides (human neuropeptide Y, human peptide YY, human pancreatic polypeptide, avian PP, and lamprey peptide methionine tyrosine) shows that this characteristic charge clustering gives rise to a common dipole moment of 325-450 D...... directed from the beta-turn region toward the receptor-binding region. This overall dipole moment is antiparallel to the dipole moment of the alpha-helix caused by alignment of the peptide dipoles parallel to the helix. Calculations of the stabilization energy for this antiparallel dipole moment...... forces alone when the screening effect is considered. This energy is of the same order of magnitude as the enthalpy change for the unfolding of avian PP (approximately 30 kcal/mol), strongly indicating that the charge-dipole interactions are of significant importance for the stability of the three...

  15. Development of a simplified one-dimensional nonlinear helix traveling wave tube code based on the Lagrangian disk model

    International Nuclear Information System (INIS)

    Joo, Y. D.; Park, G. S.; Sinha, A. K.

    2004-01-01

    A stationary one-dimensional nonlinear code, as a design tool, for a helix traveling wave tube (TWT) is developed using Lagrangian large-signal approach. The analytic solutions of the electronic and the circuit equation are derived and combined to establish a set of simple expressions for developing a simple code without using any numerical method. The present simple code quickly finds the TWT performance parameters, such as the output power, gain, and efficiency, in an analytic and straightforward manner. The results from the present simple code are compared with those from numerical work and the particle-in-cell (PIC) code and are found to be in good agreement. Also, design of a helix TWT with over 200 watts of output power in frequency range of 2-18 GHz is carried out using present code.

  16. Improving membrane binding as a design strategy for amphipathic peptide hormones: 2-helix variants of PYY3-36.

    Science.gov (United States)

    Pedersen, Søren L; Bhatia, Vikram K; Jurt, Simon; Paulsson, Johan F; Pedersen, Maria H; Jorgensen, Rasmus; Holst, Birgitte; Stamou, Dimitrios; Vrang, Niels; Zerbe, Oliver; Jensen, Knud J

    2012-09-01

    It has been hypothesized that amphipathic peptides might bind to membranes prior to activating their cognate receptors, but this has proven difficult to test. The peptide hormone PYY3-36 is believed to perform its appetite-suppressing actions through binding to hypothalamic Y2 receptors. It has been proposed that PYY3-36 via its amphipathic α-helix binds to the plasma membrane prior to receptor docking. Here, our aim was to study the implication of this hypothesis using new analogs of PYY3-36. We first studied membrane binding of PYY3-36. Next, we designed a series of PYY3-36 analogs to increase membrane-binding affinity by substituting the N-terminal segment with a de novo designed α-helical, amphipathic sequence. These 2-helix variants of PYY3-36 were assembled by solid-phase peptide synthesis. Pharmacological studies demonstrated that even though the native peptide sequence was radically changed, highly active Y2 receptor agonists were generated. A potent analog, with a Kd of 4 nM for membranes, was structurally characterized by NMR in the membrane-bound state, which clearly showed that it formed the expected 2-helix. The topology of the peptide-micelle association was studied by paramagnetic relaxation enhancement using a spin label, which confirmed that the hydrophobic residues bound to the membrane. Our studies further support the hypothesis that PYY3-36 associates with the membrane and indicate that this can be used in the design of novel molecules with high receptor binding potency. These observations are likely to be generally important for peptide hormones and biopharmaceutical drugs derived from them. This new 2-helix variant of PYY3-36 will be useful as a tool compound for studying peptide-membrane interactions. Copyright © 2012 European Peptide Society and John Wiley & Sons, Ltd.

  17. Specific features of the planarian Dugesia tigrina regeneration and mollusk Helix albescens nociception under weak electromagnetic shielding

    Science.gov (United States)

    Temur'yants, N. A.; Demtsun, N. A.; Kostyuk, A. S.; Yarmolyuk, N. S.

    2012-12-01

    It has been demonstrated that weak electromagnetic shielding stimulates regeneration in the planarian Dugesia tigrina, the stimulating intensity being dependent on both the initial state of the animals, which is determined by season, and their functional asymmetry. As has been shown, the effect of a weak electromagnetic field induces phasic changes in the nociceptive sensitivity of the mollusk Helix albescens: an increase in the sensitivity to a thermal stimulus is replaced by the development of the hypalgesic effect.

  18. Modulating immunogenic properties of HIV-1 gp41 membrane-proximal external region by destabilizing six-helix bundle structure

    Energy Technology Data Exchange (ETDEWEB)

    Banerjee, Saikat; Shi, Heliang; Habte, Habtom H.; Qin, Yali; Cho, Michael W., E-mail: mcho@iastate.edu

    2016-03-15

    The C-terminal alpha-helix of gp41 membrane-proximal external region (MPER; {sup 671}NWFDITNWLWYIK{sup 683}) encompassing 4E10/10E8 epitopes is an attractive target for HIV-1 vaccine development. We previously reported that gp41-HR1-54Q, a trimeric protein comprised of the MPER in the context of a stable six-helix bundle (6HB), induced strong immune responses against the helix, but antibodies were directed primarily against the non-neutralizing face of the helix. To better target 4E10/10E8 epitopes, we generated four putative fusion intermediates by introducing double point mutations or deletions in the heptad repeat region 1 (HR1) that destabilize 6HB in varying degrees. One variant, HR1-∆10-54K, elicited antibodies in rabbits that targeted W672, I675 and L679, which are critical for 4E10/10E8 recognition. Overall, the results demonstrated that altering structural parameters of 6HB can influence immunogenic properties of the MPER and antibody targeting. Further exploration of this strategy could allow development of immunogens that could lead to induction of 4E10/10E8-like antibodies. - Highlights: • Four gp41 MPER-based immunogens that resemble fusion intermediates were generated. • C-terminal region of MPER that contains 4E10/10E8 epitopes was highly immunogenic. • Altering 6HB structure can influence immunogenic properties of the MPER. • Induced antibodies targeted multiple residues critical for 4E10/10E8 binding. • Development of immunogens based on fusion intermediates is a promising strategy.

  19. Nanomechanics of single keratin fibres: A Raman study of the alpha helix -> beta sheet transition and water effect

    OpenAIRE

    Colomban, Philippe; Paquin, Raphaël

    2007-01-01

    International audience; The use of micro-Raman spectroscopy, through chemical bond nano-scale probes, allows the changes in conformations (α helix -> β sheet), chain orientation, disconnection of disulfide bonds (-20%) and the increase of intra and inter-chain distances during the strain to be distinguished. The combination of micro-Raman spectroscopy and a allows a quantitative measure of the extension of chemical bonds in the peptidic chain during loading. The nano-structural transformation...

  20. On the mechanism of irradiation effect on the function of Helix pomatia neuron Na+, K+-pump

    International Nuclear Information System (INIS)

    Ajrapetyan, S.N.; Egorova, E.G.; Sagiyan, A.A.; Dadalyan, S.S.; Dvoretskij, A.I.; Sulejmonyan, M.A.

    1987-01-01

    Mechanism of irradiation effect on passive permeability, Na + /Ca 2+ exchange, Na + , K + -pump function intensity, the number of membrane functionally active pump units (Na + , K + -ATP-ase molecules) was determined using Helix pomatia and nervous ganglions isolated from them and irradiated by 5.16 Kl/kg dose. The data obtained show that ionizing radiation leads to obvious destructions in the mechanisms of neuron Na + , K + -pump functioning

  1. Glass-like dynamics of the strain-induced coil/helix transition on a permanent polymer network.

    Science.gov (United States)

    Ronsin, O; Caroli, C; Baumberger, T

    2016-02-14

    We study the stress response to a step strain of covalently bonded gelatin gels in the temperature range where triple helix reversible crosslink formation is prohibited. We observe slow stress relaxation towards a T-dependent finite asymptotic level. We show that this is assignable to the strain-induced coil → helix transition, previously evidenced by Courty et al. [Proc. Natl. Acad. Sci. U. S. A. 102, 13457 (2005)], of a fraction of the polymer strands. Relaxation proceeds, in a first stage, according to a stretched exponential dynamics, then crosses over to a terminal simple exponential decay. The respective characteristic times τK and τf exhibit an Arrhenius-like T-dependence with an associated energy E incompatibly larger than the activation barrier height for the isomerisation process which sets the clock for an elementary coil → helix transformation event. We tentatively assign this glass-like slowing down of the dynamics to the long-range couplings due to the mechanical noise generated by the local elementary events in this random elastic medium.

  2. Microbiological and chemical evaluation of Helix spp. snails from local and non-EU markets, utilised as food in Sardinia

    Directory of Open Access Journals (Sweden)

    Arianna Corda

    2014-04-01

    Full Text Available With this study, 28 pools of snails of the genus Helix, respectively Helix aspersa (n=24 and Helix vermiculata (n=4 were analysed. They were taken from snail farming and stores. The snails were from Sardinia, other regions of Italy, and from abroad. All the samples were examined as pool looking for these microbiological target: Salmonella spp., Listeria monocytogenes, Escherichia coli O157, Clostridium perfringens, Norovirus and Hepatitis A Virus (HAV. In the same pools, the concentration of cadmium and lead by inductively coupled plasma mass spectrometry was also determined. The levels of these heavy metals were quite high, especially for cadmium. Two samples were positive for Salmonella spp., while no sample was positive for Escherichia coli O157, HAV and Norovirus. Two samples were positive for Clostridium perfringens and 8 for Listeria monocytogenes. The microrganisms related to Listeria monocytogenes were identified using biochemical techniques, then serotyped and gene sequenced by multiple loci sequence typing technique. Furthermore, antimicrobial restistence was tested on the same samples.

  3. Efficiency of diatomaceous earth against Helix aspersa in “in vitro” conditions in Pamplona, Norte de Santander

    Directory of Open Access Journals (Sweden)

    Andrés Camilo Méndez Otero

    2017-11-01

    Full Text Available Aims: To determine the effectiveness of the diatomaceous earth against Helix aspersa by contact and ingestion with sprinkling and powdering treatments “in vitro”. Settings and Design: Assays with completely randomized designs with five treatments and four replicates were carried out. The treatments were three doses of diatomaceous earth (at 1 kg / ha, 2 kg / ha, 4 kg / ha, agricultural lime at 2 kg / ha and one control. Methods and Material: In the assays adults of Helix aspersa Müller fed with lettuce (Lactuca sativa L. were used. The percentage of mortality, effect on mobility and shell affectation were determined with the treatments by sprinkling and powdering both contact and ingestion. Statistical analysis used: The data were transformed into 2 arcsin √% / 100, and were processed by means of an analysis of variance, using the statistical package SPSS version 21. The means were compared by the Tukey test for P <0.05 Conclusions: The effects of diatomaceous earth both by contact and by ingestion with spraying and powdering treatments against the mollusc Helix aspersa are low at doses between 1 and 4 kg / ha under "in vitro" conditions.

  4. Chondrocutaneous helix rim advancement flap: two classical and two new modifications for very large defects of the ear.

    Science.gov (United States)

    Valesky, E M; Kaufmann, R; Meissner, M

    2015-11-01

    The plastic reconstruction of the ear after microscopically controlled tumor surgery is a particular challenge. The chondrocutaneous helix rim advancement flap (CHRAF) is perfectly suited for the repair of different defects of the helix and even defects beyond. Here, we describe two known and two new modifications of the CHRAF that enlarge the scope of application in tumor surgery of the ear. We demonstrate the different techniques and practical application of the repair and evaluate the benefits and limitations. The CHRAF and its modifications is an excellent method for repair of various defects of the helical region of the ear. The CHRAF and its modifications proves to be an good alternative to other methods of closure by preserving the anatomical contour and mechanical and acoustic functions in a single-staged procedure with excellent aesthetic results. The two new modifications we introduced here, enlarge the known armentarium for very large defects of the upper pole and the mid-helix of the ear. © 2014 European Academy of Dermatology and Venereology.

  5. Glass-like stress relaxation of a permanent gelatin network as a signature of pretransitional helix-coil fluctuations

    Science.gov (United States)

    Caroli, Christiane; Ronsin, Olivier; Lemaître, Anaël

    2018-02-01

    The stress response of permanently crosslinked gelatin gels was recently observed to display glass-like features, namely, a stretched-exponential behavior terminated by an exponential decay, the characteristic time scales of which increase dramatically with decreasing temperature. This phenomenon is studied here using a model of flexible polymer gel network where relaxation proceeds via elementary monomer exchanges between helix and coil segments. The relaxation dynamics of a full network simulation is found to be nearly identical to that of a model of independent strands, which shows that for flexible polymer gels in the range of elastic moduli of interest, both strand contour length disorder and elastic couplings are irrelevant. We thus focus on the independent strand model and find it not only to explain the observed functional form of the stress relaxation curves but also to yield predictions that match very satisfactorily the experimental measurements of final relaxation time and total stress drop. The system under study thus constitutes a rare case where the origin of glass-like behavior can be unambiguously identified, namely, as the signature of the enhancement of helix content fluctuations when approaching from above the mean-field helix-coil transition of strands.

  6. BAROMETRIC PRESSURE and Other Data from ALPHA HELIX From North Pacific Ocean and Others from 19931016 to 19931103 (NODC Accession 9400051)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — The Conductivity, Temperature and Depth (CTD) and other data were collected in Bering Sea and North Pacific Ocean. Data was collected from Ship ALPHA HELIX cruise...

  7. The type II collagen fragments Helix-II and CTX-II reveal different enzymatic pathways of human cartilage collagen degradation

    DEFF Research Database (Denmark)

    Charni-Ben Tabassi, N; Desmarais, S; Jensen, Anne-Christine Bay

    2008-01-01

    OBJECTIVE: Cartilage degradation in osteoarthritis (OA) generates the type II collagen fragments, Helix-II and CTX-II that can be used as clinical biological markers. Helix-II and C-telopeptide of type II collagen (CTX-II) levels are associated independently with progression of OA suggesting...... that they may be generated through different collagenolytic pathways. In this study we analyzed the release of Helix-II and CTX-II from human cartilage collagen by the proteinases reported to play a role in cartilage degradation. METHODS: In vitro, human articular cartilage extract was incubated with activated...... pathways. Helix-II and CTX-II alone reflect only partially overall cartilage collagen degradation. These findings may explain why these two biological markers could provide complementary information on disease progression in OA....

  8. Nutrients data collected from the ALPHA HELIX from the coastal waters of Alaska from 03 March 1998 to 07 December 1999 (NODC Accession 0000637)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Nutrients data were collected from bottle casts from the ALPHA HELIX in the coastal waters of Alaska from 08 March 1998 to 07 December 1999. Data include...

  9. Chemical and depth data from the ALPHA HELIX using bottle casts in the Bering Sea from 1987-09-07 to 1988-06-11 (NODC Accession 0000263)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Chemical and depth data were collected from the ALPHA HELIX from September 7, 1987 to June 11, 1988. Data were submitted by the University of Alaska - Fairbanks;...

  10. A novel Bxb1 integrase RMCE system for high fidelity site-specific integration of mAb expression cassette in CHO Cells.

    Science.gov (United States)

    Inniss, Mara C; Bandara, Kalpanie; Jusiak, Barbara; Lu, Timothy K; Weiss, Ron; Wroblewska, Liliana; Zhang, Lin

    2017-08-01

    As CHO cell line development for biotherapeutic production becomes more sophisticated through the availability of the CHO genome sequence, the ability to accurately and reproducibly engineer the host cell genome has become increasingly important. Multiple well characterized systems for site-specific integration will enable more complex cell line engineering to generate cell lines with desirable attributes. We built and characterized a novel recombinase mediated cassette exchange (RMCE) system using Bxb1 integrase and compared it to the commonly used Flp/FRT RMCE system. We first integrated a DNA construct flanked by either Bxb1 attachment sites or FRT sequences (referred to as a landing pad) into the Fer1L4 genomic locus of CHO-S cells using CRISPR/Cas9 mediated homologous recombination. We characterized the resulting clones harboring either the Bxb1 or Flp/FRT landing pad using whole genome resequencing to compare their genomes with the parental host cell line. We determined that each landing pad was specifically integrated into the Fer1L4 locus in the selected clones and observed no major structural changes in the genome or variations in copy number as a result of CRISPR/Cas9 modification. We subsequently tested the ability of the Bxb1 and Flp/FRT landing pad clones to perform proper RMCE with donor vectors containing identical mAb expression cassettes flanked by either Bxb1 attachment sites or FRT sites. We demonstrated that both RMCE systems were able to generate stable pools in a similar time frame with comparable mAb expression. Through genetic characterization of up to 24 clones derived from either system, we determined that the BxB1 RMCE system yielded higher fidelity RMCE events than the Flp/FRT system as evidenced by a higher percentage of clones with expected integration of the mAb cassette into the landing pad in the respective cell lines. We conclude that Bxb1 RMCE is an excellent alternative to Flp/FRT RMCE and valuable addition to our toolbox

  11. Emerging contaminants and nutrients synergistically affect the spread of class 1 integron-integrase (intI1) and sul1 genes within stable streambed bacterial communities.

    Science.gov (United States)

    Subirats, Jèssica; Timoner, Xisca; Sànchez-Melsió, Alexandre; Balcázar, José Luis; Acuña, Vicenç; Sabater, Sergi; Borrego, Carles M

    2018-03-10

    Wastewater effluents increase the nutrient load of receiving streams while introducing a myriad of anthropogenic chemical pollutants that challenge the resident aquatic (micro)biota. Disentangling the effects of both kind of stressors and their potential interaction on the dissemination of antibiotic resistance genes in bacterial communities requires highly controlled manipulative experiments. In this work, we investigated the effects of a combined regime of nutrients (at low, medium and high concentrations) and a mixture of emerging contaminants (ciprofloxacin, erythromycin, sulfamethoxazole, diclofenac, and methylparaben) on the bacterial composition, abundance and antibiotic resistance profile of biofilms grown in artificial streams. In particular, we investigated the effect of this combined stress on genes encoding resistance to ciprofloxacin (qnrS), erythromycin (ermB), sulfamethoxazole (sul1 and sul2) as well as the class 1 integron-integrase gene (intI1). Only genes conferring resistance to sulfonamides (sul1 and sul2) and intI1 gene were detected in all treatments during the study period. Besides, bacterial communities exposed to emerging contaminants showed higher copy numbers of sul1 and intI1 genes than those not exposed, whereas nutrient amendments did not affect their abundance. However, bacterial communities exposed to both emerging contaminants and a high nutrient concentration (1, 25 and 1 mg L -1 of phosphate, nitrate and ammonium, respectively) showed the highest increase on the abundance of sul1 and intI1 genes thus suggesting a factors synergistic effect of both stressors. Since none of the treatments caused a significant change on the composition of bacterial communities, the enrichment of sul1 and intI1 genes within the community was caused by their dissemination under the combined pressure exerted by nutrients and emerging contaminants. To the best of our knowledge, this is the first study demonstrating the contribution of nutrients on

  12. Human Immunodeficiency Virus Type 1 Employs the Cellular Dynein Light Chain 1 Protein for Reverse Transcription through Interaction with Its Integrase Protein

    Science.gov (United States)

    Jayappa, Kallesh Danappa; Ao, Zhujun; Wang, Xiaoxia; Mouland, Andrew J.; Shekhar, Sudhanshu; Yang, Xi

    2015-01-01

    ABSTRACT In this study, we examined the requirement for host dynein adapter proteins such as dynein light chain 1 (DYNLL1), dynein light chain Tctex-type 1 (DYNLT1), and p150Glued in early steps of human immunodeficiency virus type 1 (HIV-1) replication. We found that the knockdown (KD) of DYNLL1, but not DYNLT1 or p150Glued, resulted in significantly lower levels of HIV-1 reverse transcription in cells. Following an attempt to determine how DYNLL1 could impact HIV-1 reverse transcription, we detected the DYNLL1 interaction with HIV-1 integrase (IN) but not with capsid (CA), matrix (MA), or reverse transcriptase (RT) protein. Furthermore, by mutational analysis of putative DYNLL1 interaction motifs in IN, we identified the motifs 52GQVD and 250VIQD in IN as essential for DYNLL1 interaction. The DYNLL1 interaction-defective IN mutant HIV-1 (HIV-1INQ53A/Q252A) exhibited impaired reverse transcription. Through further investigations, we have also detected relatively smaller amounts of particulate CA in DYNLL1-KD cells or in infections with HIV-1INQ53A/Q252A mutant virus. Overall, our study demonstrates the novel interaction between HIV-1 IN and cellular DYNLL1 proteins and suggests the requirement of this virus-cell interaction for proper uncoating and efficient reverse transcription of HIV-1. IMPORTANCE Host cellular DYNLL1, DYNLT1, and p150Glued proteins have been implicated in the replication of several viruses. However, their roles in HIV-1 replication have not been investigated. For the first time, we demonstrated that during viral infection, HIV-1 IN interacts with DYNLL1, and their interaction was found to have a role in proper uncoating and efficient reverse transcription of HIV-1. Thus, interaction of IN and DYNLL1 may be a potential target for future anti-HIV therapy. Moreover, while our study has evaluated the involvement of IN in HIV-1 uncoating and reverse transcription, it also predicts a possible mechanism by which IN contributes to these early viral

  13. Sensitive Deep-Sequencing-Based HIV-1 Genotyping Assay To Simultaneously Determine Susceptibility to Protease, Reverse Transcriptase, Integrase, and Maturation Inhibitors, as Well as HIV-1 Coreceptor Tropism

    Science.gov (United States)

    Gibson, Richard M.; Meyer, Ashley M.; Winner, Dane; Archer, John; Feyertag, Felix; Ruiz-Mateos, Ezequiel; Leal, Manuel; Robertson, David L.; Schmotzer, Christine L.

    2014-01-01

    With 29 individual antiretroviral drugs available from six classes that are approved for the treatment of HIV-1 infection, a combination of different phenotypic and genotypic tests is currently needed to monitor HIV-infected individuals. In this study, we developed a novel HIV-1 genotypic assay based on deep sequencing (DeepGen HIV) to simultaneously assess HIV-1 susceptibilities to all drugs targeting the three viral enzymes and to predict HIV-1 coreceptor tropism. Patient-derived gag-p2/NCp7/p1/p6/pol-PR/RT/IN- and env-C2V3 PCR products were sequenced using the Ion Torrent Personal Genome Machine. Reads spanning the 3′ end of the Gag, protease (PR), reverse transcriptase (RT), integrase (IN), and V3 regions were extracted, truncated, translated, and assembled for genotype and HIV-1 coreceptor tropism determination. DeepGen HIV consistently detected both minority drug-resistant viruses and non-R5 HIV-1 variants from clinical specimens with viral loads of ≥1,000 copies/ml and from B and non-B subtypes. Additional mutations associated with resistance to PR, RT, and IN inhibitors, previously undetected by standard (Sanger) population sequencing, were reliably identified at frequencies as low as 1%. DeepGen HIV results correlated with phenotypic (original Trofile, 92%; enhanced-sensitivity Trofile assay [ESTA], 80%; TROCAI, 81%; and VeriTrop, 80%) and genotypic (population sequencing/Geno2Pheno with a 10% false-positive rate [FPR], 84%) HIV-1 tropism test results. DeepGen HIV (83%) and Trofile (85%) showed similar concordances with the clinical response following an 8-day course of maraviroc monotherapy (MCT). In summary, this novel all-inclusive HIV-1 genotypic and coreceptor tropism assay, based on deep sequencing of the PR, RT, IN, and V3 regions, permits simultaneous multiplex detection of low-level drug-resistant and/or non-R5 viruses in up to 96 clinical samples. This comprehensive test, the first of its class, will be instrumental in the development of new

  14. Exploring the molecular mechanism of cross-resistance to HIV-1 integrase strand transfer inhibitors by molecular dynamics simulation and residue interaction network analysis.

    Science.gov (United States)

    Xue, Weiwei; Jin, Xiaojie; Ning, Lulu; Wang, Meixia; Liu, Huanxiang; Yao, Xiaojun

    2013-01-28

    The rapid emergence of cross-resistance to the integrase strand transfer inhibitors (INSTIs) has become a serious problem in the therapy of human immunodeficiency virus type 1 (HIV-1) infection. Understanding the detailed molecular mechanism of INSTIs cross-resistance is therefore critical for the development of new effective therapy against cross-resistance. On the basis of the homology modeling constructed structure of tetrameric HIV-1 intasome, the detailed molecular mechanism of the cross-resistance mutation E138K/Q148K to three important INSTIs (Raltegravir (RAL, FDA approved in 2007), Elvitegravir (EVG, FDA approved in 2012), and Dolutegravir (DTG, phase III clinical trials)) was investigated by using molecular dynamics (MD) simulation and residue interaction network (RIN) analysis. The results from conformation analysis and binding free energy calculation can provide some useful information about the detailed binding mode and cross-resistance mechanism for the three INSTIs to HIV-1 intasome. Binding free energy decomposition analysis revealed that Pro145 residue in the 140s 1oop (Gly140 to Gly149) of the HIV-1 intasome had strong hydrophobic interactions with INSTIs and played an important role in the binding of INSTIs to HIV-1 intasome active site. A systematic comparison and analysis of the RIN proves that the communications between the residues in the resistance mutant is increased when compared with that of the wild-type HIV-1 intasome. Further analysis indicates that residue Pro145 may play an important role and is relevant to the structure rearrangement in HIV-1 intasome active site. In addition, the chelating ability of the oxygen atoms in INSTIs (e.g., RAL and EVG) to Mg(2+) in the active site of the mutated intasome was reduced due to this conformational change and is also responsible for the cross-resistance mechanism. Notably, the cross-resistance mechanism we proposed could give some important information for the future rational design of novel

  15. A 'FIREWORK' OF H2 KNOTS IN THE PLANETARY NEBULA NGC 7293 (THE HELIX NEBULA)

    International Nuclear Information System (INIS)

    Matsuura, M.; Speck, A. K.; McHunu, B. M.; Tanaka, I.; Wright, N. J.; Viti, S.; Wesson, R.; Smith, M. D.; Zijlstra, A. A.

    2009-01-01

    We present a deep and wide field-of-view (4' x 7') image of the planetary nebula (PN) NGC 7293 (the Helix Nebula) in the 2.12 μm H 2 v = 1 → 0 S(1) line. The excellent seeing (0.''4) at the Subaru Telescope, allows the details of cometary knots to be examined. The knots are found at distances of 2.'2-6.'4 from the central star (CS). At the inner edge and in the inner ring (up to 4.'5 from the CS), the knot often show a 'tadpole' shape, an elliptical head with a bright crescent inside and a long tail opposite to the CS. In detail, there are variations in the tadpole shapes, such as narrowing tails, widening tails, meandering tails, or multipeaks within a tail. In the outer ring (4.'5-6.'4 from the CS), the shapes are more fractured, and the tails do not collimate into a single direction. The transition in knot morphology from the inner edge to the outer ring is clearly seen. The number density of knots governs the H 2 surface brightness in the inner ring: H 2 exists only within the knots. Possible mechanisms which contribute to the shaping of the knots are discussed, including photoionization and streaming motions. A plausible interpretation of our images is that inner knots are being overrun by a faster wind, but that this has not (yet) reached the outer knots. Based on H 2 formation and destruction rates, H 2 gas can survive in knots from formation during the late asymptotic giant branch phase throughout the PN phase. These observations provide new constraints on the formation and evolution of knots, and on the physics of molecular gas embedded within ionized gas.

  16. BuD, a helix–loop–helix DNA-binding domain for genome modification

    Energy Technology Data Exchange (ETDEWEB)

    Stella, Stefano [Spanish National Cancer Research Centre (CNIO), Calle de Melchor Fernández Almagro 3, 28029 Madrid (Spain); University of Copenhagen, Blegdamsvej 3B, 2200 Copenhagen (Denmark); Molina, Rafael; López-Méndez, Blanca [Spanish National Cancer Research Centre (CNIO), Calle de Melchor Fernández Almagro 3, 28029 Madrid (Spain); Juillerat, Alexandre; Bertonati, Claudia; Daboussi, Fayza [Cellectis, 8 Rue de la Croix Jarry, 75013 Paris (France); Campos-Olivas, Ramon [Spanish National Cancer Research Centre (CNIO), Calle de Melchor Fernández Almagro 3, 28029 Madrid (Spain); Duchateau, Phillippe [Cellectis, 8 Rue de la Croix Jarry, 75013 Paris (France); Montoya, Guillermo, E-mail: guillermo.montoya@cpr.ku.dk [Spanish National Cancer Research Centre (CNIO), Calle de Melchor Fernández Almagro 3, 28029 Madrid (Spain); University of Copenhagen, Blegdamsvej 3B, 2200 Copenhagen (Denmark)

    2014-07-01

    Crystal structures of BurrH and the BurrH–DNA complex are reported. DNA editing offers new possibilities in synthetic biology and biomedicine for modulation or modification of cellular functions to organisms. However, inaccuracy in this process may lead to genome damage. To address this important problem, a strategy allowing specific gene modification has been achieved through the addition, removal or exchange of DNA sequences using customized proteins and the endogenous DNA-repair machinery. Therefore, the engineering of specific protein–DNA interactions in protein scaffolds is key to providing ‘toolkits’ for precise genome modification or regulation of gene expression. In a search for putative DNA-binding domains, BurrH, a protein that recognizes a 19 bp DNA target, was identified. Here, its apo and DNA-bound crystal structures are reported, revealing a central region containing 19 repeats of a helix–loop–helix modular domain (BurrH domain; BuD), which identifies the DNA target by a single residue-to-nucleotide code, thus facilitating its redesign for gene targeting. New DNA-binding specificities have been engineered in this template, showing that BuD-derived nucleases (BuDNs) induce high levels of gene targeting in a locus of the human haemoglobin β (HBB) gene close to mutations responsible for sickle-cell anaemia. Hence, the unique combination of high efficiency and specificity of the BuD arrays can push forward diverse genome-modification approaches for cell or organism redesign, opening new avenues for gene editing.

  17. Collagen triple helix repeat containing 1 is a new promigratory marker of arthritic pannus.

    Science.gov (United States)

    Shekhani, Mohammed Talha; Forde, Toni S; Adilbayeva, Altynai; Ramez, Mohamed; Myngbay, Askhat; Bexeitov, Yergali; Lindner, Volkhard; Adarichev, Vyacheslav A

    2016-07-19

    The formation of destructive hypercellular pannus is critical to joint damage in rheumatoid arthritis (RA). The collagen triple helix repeat containing 1 (CTHRC1) protein expressed by activated stromal cells of diverse origin has previously been implicated in tissue remodeling and carcinogenesis. We recently discovered that the synovial Cthrc1 mRNA directly correlates with arthritis severity in mice. This study characterizes the role of CTHRC1 in arthritic pannus formation. Synovial joints of mice with collagen antibody-induced arthritis (CAIA) and human RA-fibroblast-like synoviocytes (FLS) were immunostained for CTHRC1, FLS and macrophage-specific markers. CTHRC1 levels in plasma from patients with RA were measured using sandwich ELISA. The migratory response of fibroblasts was studied with a transwell migration assay and time-lapse microscopy. Velocity and directness of cell migration was analyzed by recording the trajectories of cells treated with rhCTHRC1. Immunohistochemical analysis of normal and inflamed synovium revealed highly inducible expression of CTHRC1 in arthritis (10.9-fold). At the tissue level, CTHRC1-expressing cells occupied the same niche as large fibroblast-like cells positive for α-smooth muscle actin (α-SMA) and cadherin 11 (CDH11). CTHRC1 was produced by activated FLS predominantly located at the synovial intimal lining and at the bone-pannus interface. Cultured RA-FLS expressed CDH11, α-SMA, and CTHRC1. Upon treatment with exogenous rhCTHRC1, embryonic fibroblasts and RA-FLS significantly increased migration velocity, directness, and cell length along the front-tail axis (1.4-fold, p marker of activated synoviocytes in murine experimental arthritis and RA. The pro-migratory effect of CTHRC1 on synoviocytes is considered one of the mechanisms promoting hypercellularity of the arthritic pannus.

  18. Identification of SNARE complex modulators that inhibit exocytosis from an alpha-helix-constrained combinatorial library.

    Science.gov (United States)

    Blanes-Mira, Clara; Pastor, Maria T; Valera, Elvira; Fernández-Ballester, Gregorio; Merino, Jaime M; Gutierrez, Luis M; Perez-Payá, Enrique; Ferrer-Montiel, Antonio

    2003-01-01

    Synthetic peptides patterned after the proteins involved in vesicle fusion [the so-called SNARE (soluble N -ethylmaleimide-sensitive fusion protein attachment protein receptor) proteins] are potent inhibitors of SNARE complex assembly and neuronal exocytosis. It is noteworthy that the identification of peptide sequences not related to the SNARE proteins has not been accomplished yet; this is due, in part, to the structural constraints and the specificity of the protein interactions that govern the formation of the SNARE complex. Here we have addressed this question and used a combinatorial approach to identify peptides that modulate the assembly of the SNARE core complex and inhibit neuronal exocytosis. An alpha-helix-constrained, mixture-based, 17-mer combinatorial peptide library composed of 137180 sequences was synthesized in a positional scanning format. Peptide mixtures were assayed for their ability to prevent the formation of the in vitro -reconstituted SDS-resistant SNARE core complex. Library deconvolution identified eight peptides that inhibited the assembly of the SNARE core complex. Notably, the most potent 17-mer peptide (acetyl-SAAEAFAKLYAEAFAKG-NH2) abolished both Ca2+-evoked catecholamine secretion from detergent-permeabilized chromaffin cells and L-glutamate release from intact hippocampal primary cultures. Collectively, these findings indicate that amino acid sequences that prevent SNARE complex formation are not restricted to those that mimic domains of SNARE proteins, thus expanding the diversity of molecules that target neuronal exocytosis. Because of the implication of neurosecretion in the aetiology of several human neurological disorders, these newly identified peptides may be considered hits for the development of novel anti-spasmodic drugs. PMID:12852787

  19. Non-linearity of the collagen triple helix in solution and implications for collagen function.

    Science.gov (United States)

    Walker, Kenneth T; Nan, Ruodan; Wright, David W; Gor, Jayesh; Bishop, Anthony C; Makhatadze, George I; Brodsky, Barbara; Perkins, Stephen J

    2017-06-16

    Collagen adopts a characteristic supercoiled triple helical conformation which requires a repeating (Xaa-Yaa-Gly) n sequence. Despite the abundance of collagen, a combined experimental and atomistic modelling approach has not so far quantitated the degree of flexibility seen experimentally in the solution structures of collagen triple helices. To address this question, we report an experimental study on the flexibility of varying lengths of collagen triple helical peptides, composed of six, eight, ten and twelve repeats of the most stable Pro-Hyp-Gly (POG) units. In addition, one unblocked peptide, (POG) 10unblocked , was compared with the blocked (POG) 10 as a control for the significance of end effects. Complementary analytical ultracentrifugation and synchrotron small angle X-ray scattering data showed that the conformations of the longer triple helical peptides were not well explained by a linear structure derived from crystallography. To interpret these data, molecular dynamics simulations were used to generate 50 000 physically realistic collagen structures for each of the helices. These structures were fitted against their respective scattering data to reveal the best fitting structures from this large ensemble of possible helix structures. This curve fitting confirmed a small degree of non-linearity to exist in these best fit triple helices, with the degree of bending approximated as 4-17° from linearity. Our results open the way for further studies of other collagen triple helices with different sequences and stabilities in order to clarify the role of molecular rigidity and flexibility in collagen extracellular and immune function and disease. © 2017 The Author(s).

  20. FMRF-amide is a glucose-lowering hormone in the snail Helix aspersa.

    Science.gov (United States)

    Rőszer, Tamás; Kiss-Tóth, Éva D

    2014-11-01

    Although glucose is metabolically the most important carbohydrate in almost all living organisms, still little is known about the evolution of the hormonal control of cellular glucose uptake. In this study, we identify Phe-Met-Arg-Phe-amide (FMRFa), also known as molluscan cardioexcitatory tetrapeptide, as a glucose-lowering hormone in the snail Helix aspersa. FMRFa belongs to an evolutionarily conserved neuropeptide family and is involved in the neuron-to-muscle signal transmission in the snail digestive system. This study shows that, beyond this function, FMRFa also has glucose-lowering activity. We found neuronal transcription of genes encoding FMRFa and its receptor and moreover the hemolymph FMRFa levels were peaking at metabolically active periods of the snails. In turn, hypometabolism of the dormant periods was associated with abolished FMRFa production. In the absence of FMRFa, the midintestinal gland ("hepatopancreas") cells were deficient in their glucose uptake, contributing to the development of glucose intolerance. Exogenous FMRFa restored the absorption of hemolymph glucose by the midintestinal gland cells and improved glucose tolerance in dormant snails. We show that FMRFa was released to the hemolymph in response to glucose challenge. FMRFa-containing nerve terminals reach the interstitial sinusoids between the chondroid cells in the artery walls. We propose that, in addition to the known sites of possible FMRFa secretion, these perivascular sinusoids serve as neurohemal organs and allow FMRFa release. This study suggests that in evolution, not only the insulin-like peptides have adopted the ability to increase cellular glucose uptake and can act as hypoglycemic hormones.

  1. The cDNA sequence of three hemocyanin subunits from the garden snail Helix lucorum.

    Science.gov (United States)

    De Smet, Lina; Dimitrov, Ivan; Debyser, Griet; Dolashka-Angelova, Pavlina; Dolashki, Aleksandar; Van Beeumen, Jozef; Devreese, Bart

    2011-11-10

    Hemocyanins are blue copper containing respiratory proteins residing in the hemolymph of many molluscs and arthropods. They can have different molecular masses and quaternary structures. Moreover, several molluscan hemocyanins are isolated with one, two or three isoforms occurring as decameric, didecameric, multidecameric or tubule aggregates. We could recently isolate three different hemocyanin isopolypeptides from the hemolymph of the garden snail Helix lucorum (HlH). These three structural subunits were named α(D)-HlH, α(N)-HlH and β-HlH. We have cloned and sequenced their cDNA which is the first result ever reported for three isoforms of a molluscan hemocyanin. Whereas the complete gene sequence of α(D)-HlH and β-HlH was obtained, including the 5' and 3' UTR, 180bp of the 5' end and around 900bp at the 3' end are missing for the third subunit. The subunits α(D)-HlH and β-HlH comprise a signal sequence of 19 amino acids plus a polypeptide of 3409 and 3414 amino acids, respectively. We could determine 3031 residues of the α(N)-HLH subunit. Sequence comparison with other molluscan hemocyanins shows that α(D)-HlH is more related to Aplysia californicum hemocyanin than to each of its own isopolypeptides. The structural subunits comprise 8 different functional units (FUs: a, b, c, d, e, f, g, h) and each functional unit possesses a highly conserved copper-A and copper-B site for reversible oxygen binding. Potential N-glycosylation sites are present in all three structural subunits. We confirmed that all three different isoforms are effectively produced and secreted in the hemolymph of H. lucorum by analyzing a tryptic digest of the purified native hemocyanin by MALDI-TOF and LC-FTICR mass spectrometry. Copyright © 2011 Elsevier B.V. All rights reserved.

  2. Mass spectrometric analysis of activity-dependent changes of neuropeptide profile in the snail, Helix pomatia.

    Science.gov (United States)

    Pirger, Z; Lubics, A; Reglodi, D; Laszlo, Z; Mark, L; Kiss, T

    2010-12-01

    Terrestrial snails are able to transform themselves into inactivity ceasing their behavioral activity under unfavorable environmental conditions. In the present study, we report on the activity-dependent changes of the peptide and/or polypeptide profile in the brain and hemolymph of the snail, Helix pomatia, using MALDI TOF and quadrupole mass spectrometry. The present data indicate that the snails respond to low temperature by increasing or decreasing the output of selected peptides. Average mass spectra of the brain and hemolymph revealed numerous peaks predominantly present during the active state (19 and 10 peptides/polypeptides, respectively), while others were observed only during hibernation (11 and 13). However, there were peptides and/or polypeptides or their fragments present irrespective of the activity states (49 and 18). The intensity of fourteen peaks that correspond to previously identified neuropeptides varied in the brain of active snails compared to those of hibernating animals. Among those the intensity of eight peptides increased significantly in active animals while in hibernated animals the intensity of another six peptides increased significantly. A new peptide or peptide fragment at m/z 1110.7 was identified in a brain of the snail with the following suggested amino acid sequence: GSGASGSMPATTS. This peptide was found to be more abundant in active animals because the intensity of the peptide was significantly higher compared to hibernating animals. In summary, our results revealed substantial differences in the peptide/polypeptide profile of the brain and hemolymph of active and hibernating snails suggesting a possible contribution of peptides in the process of hibernation. Copyright © 2010 Elsevier Ltd. All rights reserved.

  3. [Neuronal mechanisms of associative food aversion conditioning reconsolidation in snail Helix lucorum].

    Science.gov (United States)

    Kozyrev, S A; Nikitin, V P

    2009-06-01

    We have previously showed that reactivation of long-term memory during protin synthesis inhibitor application initiated disruption of memory recalling in snails Helix lucorum with food aversion conditioning reflex. In present work cellular mechanisms of memory reactivation were studied in snail LP11 and RP11 command neurons of defense behavior. In first trial experiments mechanisms of amnesia induction were investigated in semiintact preparations 24 hours after aversion conditioning with single type of food. It was found that application of conditioned food stimulus on snail lip during CNS perfusion with cycloheximide (protein synthesis inhibitor) initiated depression of synaptic response evoked by conditioned stimulus 2.5 hours after reminding. In second tria experiments neuronal mechanisms of amnesia development were studied. Snails were conditioned with two types of food. Cycloheximide was injected into mantle cavity and conditioned stimulus of one type of food was presented 24 hours after snail learning. Semiintact preparations were prepared 1,3, 7 and 15 days after cycloheximide injection + reminding procedure. It was found that neural responses evoked by conditioned food stimulus which was used as reminding stimulus gradually decreased during 1, 3 and 7 days. Neural responses evoked by the conditioned stimulus at 7 and 15 days were not significantly differed from control differentiated food stimulus and were significantly weaker then neural responses evoked by second conditioned food stimulus which was not used as a reminding stimulus. It was suggested that specific and protein synthesis-dependent changes in synaptic connections effectiveness in LP11 and RP11 neurons is one of the cellular mechanisms of amnesia obtained after disruption of long-term memory reconsolidation in snail.

  4. Role of helix 44 of 16S rRNA in the fidelity of translation initiation.

    Science.gov (United States)

    Qin, Daoming; Liu, Qi; Devaraj, Aishwarya; Fredrick, Kurt

    2012-03-01

    The molecular mechanisms that govern translation initiation to ensure accuracy remain unclear. Here, we provide evidence that the subunit-joining step of initiation is controlled in part by a conformational change in the 1408 region of helix h44. First, chemical probing of 30S initiation complexes formed with either a cognate (AUG) or near-cognate (AUC) start codon shows that an IF1-dependent enhancement at A1408 is reduced in the presence of AUG. This change in reactivity is due to a conformational change rather than loss of IF1, because other portions of the IF1 footprint are unchanged and high concentrations of IF1 fail to diminish the reactivity difference seen at A1408. Second, mutations in h44 such as A1413C stimulate 50S docking and cause reduced reactivity at A1408. Third, streptomycin, which has been shown by Rodnina and coworkers to stimulate 50S docking by reversing the inhibitory effects of IF1, also causes reduced reactivity at A1408. Collectively, these data support a model in which IF1 alters the A1408 region of h44 in a way that makes 50S docking unfavorable, and canonical codon-anticodon pairing in the P site restores h44 to a docking-favorable conformation. We also find that, in the absence of factors, the cognate 30S•AUG•fMet-tRNA ternary complex is >1000-fold more stable than the near-cognate 30S•AUC•fMet-tRNA complex. Hence, the selectivity of ternary complex formation is inherently high, exceeding that of initiation in vivo by more than 10-fold.

  5. Predicting RNA 3D structure using a coarse-grain helix-centered model.

    Science.gov (United States)

    Kerpedjiev, Peter; Höner Zu Siederdissen, Christian; Hofacker, Ivo L

    2015-06-01

    A 3D model of RNA structure can provide information about its function and regulation that is not possible with just the sequence or secondary structure. Current models suffer from low accuracy and long running times and either neglect or presume knowledge of the long-range interactions which stabilize the tertiary structure. Our coarse-grained, helix-based, tertiary structure model operates with only a few degrees of freedom compared with all-atom models while preserving the ability to sample tertiary structures given a secondary structure. It strikes a balance between the precision of an all-atom tertiary structure model and the simplicity and effectiveness of a secondary structure representation. It provides a simplified tool for exploring global arrangements of helices and loops within RNA structures. We provide an example of a novel energy function relying only on the positions of stems and loops. We show that coupling our model to this energy function produces predictions as good as or better than the current state of the art tools. We propose that given the wide range of conformational space that needs to be explored, a coarse-grain approach can explore more conformations in less iterations than an all-atom model coupled to a fine-grain energy function. Finally, we emphasize the overarching theme of providing an ensemble of predicted structures, something which our tool excels at, rather than providing a handful of the lowest energy structures. © 2015 Kerpedjiev et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

  6. Computational study of elements of stability of a four-helix bundle protein biosurfactant

    Science.gov (United States)

    Schaller, Andrea; Connors, Natalie K.; Dwyer, Mirjana Dimitrijev; Oelmeier, Stefan A.; Hubbuch, Jürgen; Middelberg, Anton P. J.

    2015-01-01

    Biosurfactants are surface-active molecules produced principally by microorganisms. They are a sustainable alternative to chemically-synthesized surfactants, having the advantages of being non-toxic, highly functional, eco-friendly and biodegradable. However they are currently only used in a few industrial products due to costs associated with production and purification, which exceed those for commodity chemical surfactants. DAMP4, a member of a four-helix bundle biosurfactant protein family, can be produced in soluble form and at high yield in Escherichia coli, and can be recovered using a facile thermal phase-separation approach. As such, it encompasses an interesting synergy of biomolecular and chemical engineering with prospects for low-cost production even for industrial sectors. DAMP4 is highly functional, and due to its extraordinary thermal stability it can be purified in a simple two-step process, in which the combination of high temperature and salt leads to denaturation of all contaminants, whereas DAMP4 stays stable in solution and can be recovered by filtration. This study aimed to characterize and understand the fundamental drivers of DAMP4 stability to guide further process and surfactant design studies. The complementary use of experiments and molecular dynamics simulation revealed a broad pH and temperature tolerance for DAMP4, with a melting point of 122.4 °C, suggesting the hydrophobic core as the major contributor to thermal stability. Simulation of systematically created in silico variants of DAMP4 showed an influence of number and location of hydrophilic mutations in the hydrophobic core on stability, demonstrating a tolerance of up to three mutations before a strong loss in stability occurred. The results suggest a consideration of a balance of stability, functionality and kinetics for new designs according to their application, aiming for maximal functionality but at adequate stability to allow for cost-efficient production using thermal

  7. Trypsin-kallikrein isoinhibitor K (type Kunitz) from snails (Helix pomatia). Purification and characterization.

    Science.gov (United States)

    Dietl, T; Tschesche, H

    1975-10-15

    A basic proteinase inhibitor, isoinhibitor K, was purified by SE-Sephadex C-25 column chromatography from the mixture of acid-stable and heat-stable isoinhibitors of the snail (Helix pomatia). Isoinhibitor K is homogeneous in polyacrylamide gel, cellulose acetate and polyacrylamide-dodecylsulfate electrophoresis. From the electrophoretic mobility in dodecylsulfate-polyacrylamide gel and apparent molecular weight of 6500 +/- 200 was estimated. From the amino acid composition the inhibitor consists of 58 amino acid residues. It contains three disulfide bridges, a C-terminal valine and a lysine residue at the reactive site. Isoinhibitor K inhibits the enzymes: bovine trypsin and chymotrypsin, porcine plasmin and pancreatic kallikrein, the trypsin-like component of Streptomyces griseus proteinase-pronase E, and fungi proteinase K from Tritirachium album Limber, which is only inhibited very slightly in contrast to the effect of the mixture of isoinhibitors. The inhibitory effect of isoinhibitor K against these enzymes is compared to that of the mixture or of other isoinhibitors. The following enzymes are not inhibited by isoinhibitor K: Aspergillus proteinase P and alkaline bacillus proteinase 2231 (Röhm), which both are inhibited by the mixture of isoinhibitors. Porcine elastase, bacterial proteinase N (M) (Röhm), and a trypsin-like proteinase from wheat are not inhibited, porcine acrosin and porcine serum kallikrein only to a very minor extent by the mixture of isoinhibitors. Reactive-site peptide-bond cleavage during inhibition could not be detected. Thus, the inhibitory behaviour is just as broad in specificity and as unusual as that of the trypsin-kallikrein inhibitor (Kunitz) from bovine organs. The N-terminus is blocked by pyroglutamic acid. Isoinhibitor K is the main component of the isoinhibitors secreted into the mucus and amounts to 35-40% of the mixture.

  8. Antimicrobial properties of mucus from the brown garden snail Helix aspersa.

    Science.gov (United States)

    Pitt, S J; Graham, M A; Dedi, C G; Taylor-Harris, P M; Gunn, A

    2015-01-01

    Research into naturally occurring antimicrobial substances has yielded effective treatments. One area of interest is peptides and proteins produced by invertebrates as part of their defence system, including the contents of mollusc mucus. Mucus produced by the African giant land snail, Achatina fulica has been reported to contain two proteins with broad-spectrum antibacterial activity. Mucus from the brown garden snail, Helix aspersa, appears to have skin regeneration properties. This study sought to investigate the antimicrobial properties of H. aspersa mucus. Mucus was collected from H. aspersa snails, diluted in phosphate-buffered saline (PBS), with the supernatant tested against a wide range of organisms in a disc-diffusion antimicrobial assay. This was followed with comparative experiments involving A. fulica, including bacteriophage assays. Mucus from both species of snail was passed through a series of protein size separation columns in order to determine the approximate size of the antimicrobial substance. Electrophoresis was also carried out on the H. aspersa mucus. Results indicated that H. aspersa mucus had a strong antibacterial effect against several strains of Pseudomonas aeruginosa and a weak effect against Staphylococcus aureus. Mucus from A. fulica also inhibited the growth of S. aureus, but the broad spectrum of activity reported by other workers was not observed. Antimicrobial activity was not caused by bacteriophage. Size separation experiments indicated that the antimicrobial substance(s) in H. aspersa were between 30 and 100 kDa. Electrophoresis revealed two proteins in this region (30-40 kDa and 50-60 kDa). These do not correspond with antimicrobial proteins previously reported in A. fulica. This study found one or more novel antimicrobial agents in H. aspersa mucus, with a strong effect against P. aeruginosa.

  9. Triple Helix Model in Developing Technological Innovation: The Case of Computer Based Interlocking

    Directory of Open Access Journals (Sweden)

    Erry Ricardo Nurzal

    2013-01-01

    Full Text Available Computer Based Interlocking (CBI is the most important part of the electric railway signalingsystem which serves as the “brain” that controls the operation of electrical signaling system thatreplaces the role of the electromagnetic relay which has gradually been abandoned. Developmentof CBI is based on the fact that until recently the signaling system in Indonesia is still reliedheavily on the CBI products from foreign vendors. Therefore, the development of domestic CBIproducts is really necessary in order to decrease the dependence on technology from foreignvendors that at the same time is also be able to support the accelerated development of railwayinfrastructure in Indonesia.The CBI development is conducted in the form of a collaboration involving the government,universities, and industry. This paper explores and analyzes the roles and interactions betweendifferent actors in the triple helix perspective, and identifies how the innovation ecosystemfunctioning with support from the government.Based on the data collected through in-depth interview with the actors involved in the CBI casestudy, this paper gives some key findings. First, government role is very important in establishingan innovation ecosystem for CBI development. Second, the leader in the development of CBItechnological innovation should be the industry supported by governmental R & D institutionsand universities. Third, R & D consortium is an effective vehicle for creating an interactionamong industry, academia, and government.Such findings may provide conceptual direction which is important for the development oftechnological innovation in Indonesia. In addition, in providing support for R&D activities, thegovernment needs to direct its R & D incentives to the industry as a business practitioner with itsown technology roadmap. The possible further research may include some research issues. First,is the pattern occurred in the CBI case also applied in the other cases of

  10. Helix Geometry, Hydration, and G\\cdot A Mismatch in a B-DNA Decamer

    Science.gov (United States)

    Prive, Gilbert G.; Heinemann, Udo; Chandrasegaran, Srinivasan; Kan, Lou-Sing; Kopka, Mary L.; Dickerson, Richard E.

    1987-10-01

    The DNA double helix is not a regular, featureless barberpole molecule. Different base sequences have their own special signature, in the way that they influence groove width, helical twist, bending, and mechanical rigidity or resistance to bending. These special features probably help other molecules such as repressors to read and recognize one base sequence in preference to another. Single crystal x-ray structure analysis is beginning to show us the various structures possible in the B-DNA family. The DNA decamer C-C-A-A-G-A-T-T-G-G appears to be a better model for mixed-sequence B-DNA than was the earlier C-G-C-G-A-A-T-T-C-G-C-G, which is more akin to regions of poly (dA)\\cdot poly(dT). The G\\cdot A mismatch base pairs at the center of the decamer are in the anti-anti conformation about their bonds from base to sugar, in agreement with nuclear magnetic resonance evidence on this and other sequences, and in contrast to the anti-syn geometry reported for G\\cdot A pairs in C-G-C-G-A-A-T-T-A-G-C-G. The ordered spine of hydration seen earlier in the narrow-grooved dodecamer has its counterpart, in this wide-grooved decamer, in two strings of water molecules lining the walls of the minor groove, bridging from purine N3 or pyrimidine O2, to the following sugar O4'. The same strings of hydration are present in the phosphorothioate analog of G-C-G-C-G-C. Unlike the spine, which is broken up by the intrusion of amine groups at guanines, these water strings are found in general, mixed-sequence DNA because they can pass by unimpeded to either side of a guanine N2 amine. The spine and strings are perceived as two extremes of a general pattern of hydration of the minor groove, which probably is the dominant factor in making B-DNA the preferred form at high hydration.

  11. SU-E-T-564: Multi-Helix Rotating Shield Brachytherapy for Cervical Cancer

    Energy Technology Data Exchange (ETDEWEB)

    Dadkhah, H; Wu, X [University of Iowa, Iowa City, IA (United States); Flynn, R; Kim, Y [University of Iowa Hospitals and Clinics, Iowa City, IA (United States)

    2015-06-15

    Purpose: To present a novel and practical brachytherapy technique, called multi-helix rotating shield brachytherapy (H-RSBT), for the precise positioning of a partial shield in a curved applicator. H-RSBT enables RSBT delivery using only translational motion of the radiation source/shield combination. H-RSBT overcomes the challenges associated with previously proposed RSBT approaches based on a serial (S-RSBT) step-and-shoot delivery technique, which required independent translational and rotational motion. Methods: A Fletcher-type applicator, compatible with the combination of a Xoft Axxent™ electronic brachytherapy source and a 0.5 mm thick tungsten shield, is proposed. The wall of the applicator contains six evenly-spaced helical keyways that rigidly define the emission direction of the shield as a function of depth. The shield contains three protruding keys and is attached to the source such that it rotates freely. S-RSBT and H-RSBT treatment plans with 180° and 45° azimuthal emission angles were generated for five cervical cancer patients representative of a wide range of high-risk clinical target volume (HR-CTV) shapes and applicator positions. The number of beamlets used in the treatment planning process was nearly constant for S-RSBT and H-RSBT by using dwell positions separated by 5 and 1.7 mm, respectively, and emission directions separated by 22.5° and 60°, respectively. For all the treatment plans the EQD2 of the HR-CTV was escalated until the EQD{sub 2cc} tolerance of either the bladder, rectum, or sigmoid colon was reached. Results: Treatment times for H-RSBT tended to be shorter than for S-RSBT, with changes of −38.47% to 1.12% with an average of −8.34%. The HR-CTV D{sub 90} changed by −8.81% to 2.08% with an average of −2.46%. Conclusion: H-RSBT is a mechanically feasible technique in the curved applicators needed for cervical cancer brachytherapy. S-RSBT and H-RSBT dose distributions were clinically equivalent for all patients

  12. 45 CFR 12a.4 - Suitability determination.

    Science.gov (United States)

    2010-10-01

    ... 45 Public Welfare 1 2010-10-01 2010-10-01 false Suitability determination. 12a.4 Section 12a.4... PROPERTY TO ASSIST THE HOMELESS § 12a.4 Suitability determination. (a) Suitability determination. Within 30... determination to the landholding agency. Properties that are under lease, contract, license, or agreement by...

  13. 22 CFR 9a.4 - Classification.

    Science.gov (United States)

    2010-04-01

    ... 22 Foreign Relations 1 2010-04-01 2010-04-01 false Classification. 9a.4 Section 9a.4 Foreign... ENERGY PROGRAMS; RELATED MATERIAL § 9a.4 Classification. (a) Section 1 of E.O. 11932, August 4, 1976.... If the officer determines that the information or material warrants classification, he shall assign...

  14. SuperBiHelix method for predicting the pleiotropic ensemble of G-protein-coupled receptor conformations.

    Science.gov (United States)

    Bray, Jenelle K; Abrol, Ravinder; Goddard, William A; Trzaskowski, Bartosz; Scott, Caitlin E

    2014-01-07

    There is overwhelming evidence that G-protein-coupled receptors (GPCRs) exhibit several distinct low-energy conformations, each of which might favor binding to different ligands and/or lead to different downstream functions. Understanding the function of such proteins requires knowledge of the ensemble of low-energy configurations that might play a role in this pleiotropic functionality. We earlier reported the BiHelix method for efficiently sampling the (12)(7) = 35 million conformations resulting from 30° rotations about the axis (η) of all seven transmembrane helices (TMHs), showing that the experimental structure is reliably selected as the best conformation from this ensemble. However, various GPCRs differ sufficiently in the tilts of the TMHs that this method need not predict the optimum conformation starting from any other template. In this paper, we introduce the SuperBiHelix method in which the tilt angles (θ, ϕ) are optimized simultaneously with rotations (η) efficiently enough that it is practical and sufficient to sample (5 × 3 × 5)(7) = 13 trillion configurations. This method can correctly identify the optimum structure of a GPCR starting with the template from a different GPCR. We have validated this method by predicting known crystal structure conformations starting from the template of a different protein structure. We find that the SuperBiHelix conformational ensemble includes the higher energy conformations associated with the active protein in addition to those associated with the more stable inactive protein. This methodology was then applied to design and experimentally confirm structures of three mutants of the CB1 cannabinoid receptor associated with different functions.

  15. A double-helix and cross-patterned solenoid used as a wirelessly powered receiver for medical implants

    Science.gov (United States)

    Mao, Shitong; Wang, Hao; Mao, Zhi-Hong; Sun, Mingui

    2018-05-01

    Many medical implants need to be designed in the shape of a cylinder (rod), a cuboid or a capsule in order to adapt to a specific site within the human body or facilitate the implantation procedure. In order to wirelessly power these types of implants, a pair of coils, one is located inside the human body and one is outside, is often used. Since most organs such as major muscles, blood vessels, and nerve bundles are anatomically parallel to the body surface, the most desired wireless power transfer (WPT) direction is from the external power transmission pad (a planar coil) to the lateral surface of the implant. However, to obtain optimal coupling, the currently used solenoid coil requires being positioned perpendicular to the body surface, which is often medically or anatomically unacceptable. In this research, a concentric double-helix (DH) coil with an air core is presented for use in implantable devices. Two helical coils are tilted at opposite angles (±45 degrees) to form a cross pattern. The WPT system is designed using the magnetic resonance concept for wireless power transfer (MR-WPT). The power transfer efficiency (PTE) relies on the near-field magnetic coupling which is closely related to the location and orientation of the DH coil. We explain how the novel structure of the DH solenoid magnifies the mutual inductance with the widely adopted circular planner coil and how the PTE is improved in comparison to the case of the conventional solenoid coil. We also study an important case where the double-helix power reception coil is laterally and angularly misaligned with the transmitter. Finally, our computational study using the finite element method and experimental study with actually constructed prototypes are presented which have proven our new double-helix coil design.

  16. Helix Nebula: Enabling federation of existing data infrastructures and data services to an overarching cross-domain e-infrastructure

    Science.gov (United States)

    Lengert, Wolfgang; Farres, Jordi; Lanari, Riccardo; Casu, Francesco; Manunta, Michele; Lassalle-Balier, Gerard

    2014-05-01

    Helix Nebula has established a growing public private partnership of more than 30 commercial cloud providers, SMEs, and publicly funded research organisations and e-infrastructures. The Helix Nebula strategy is to establish a federated cloud service across Europe. Three high-profile flagships, sponsored by CERN (high energy physics), EMBL (life sciences) and ESA/DLR/CNES/CNR (earth science), have been deployed and extensively tested within this federated environment. The commitments behind these initial flagships have created a critical mass that attracts suppliers and users to the initiative, to work together towards an "Information as a Service" market place. Significant progress in implementing the following 4 programmatic goals (as outlined in the strategic Plan Ref.1) has been achieved: × Goal #1 Establish a Cloud Computing Infrastructure for the European Research Area (ERA) serving as a platform for innovation and evolution of the overall infrastructure. × Goal #2 Identify and adopt suitable policies for trust, security and privacy on a European-level can be provided by the European Cloud Computing framework and infrastructure. × Goal #3 Create a light-weight governance structure for the future European Cloud Computing Infrastructure that involves all the stakeholders and can evolve over time as the infrastructure, services and user-base grows. × Goal #4 Define a funding scheme involving the three stake-holder groups (service suppliers, users, EC and national funding agencies) into a Public-Private-Partnership model to implement a Cloud Computing Infrastructure that delivers a sustainable business environment adhering to European level policies. Now in 2014 a first version of this generic cross-domain e-infrastructure is ready to go into operations building on federation of European industry and contributors (data, tools, knowledge, ...). This presentation describes how Helix Nebula is being used in the domain of earth science focusing on geohazards. The

  17. [Immunocytochemical detection of substances related to methionine enkephalin in the tentacles and food of the snail (Helix aspersa)].

    Science.gov (United States)

    Marchand, C R; Dubois, M P

    1986-01-01

    Immunocytochemistry reveals the presence of methionine-enkephalin-like substance(s) in the collar cells of the two kinds of tentacles and in the foot of the snail Helix aspersa. The density of the immunoreactive material is higher in young animals than in adults. The greater part of the substance(s) is released at the surface of the epidermis and probably mixed with the mucus. A possible neuroendocrine and/or neuromodulatory function can be considered especially for the collar cells connected with the tentacular ganglia.

  18. Structure, dynamics, and elasticity of free 16S rRNA helix 44 studied by molecular dynamics simulations

    Czech Academy of Sciences Publication Activity Database

    Réblová, Kamila; Lankaš, F.; Rázga, Filip; Krasovská, Maryna V.; Koča, J.; Šponer, Jiří

    2006-01-01

    Roč. 82, č. 5 (2006), s. 504-520 ISSN 0006-3525 R&D Projects: GA ČR(CZ) GA203/05/0388; GA ČR(CZ) GA203/05/0009; GA ČR(CZ) GD204/03/H016; GA MŠk(CZ) LC512 Institutional research plan: CEZ:AV0Z50040507 Keywords : molecular dynamics * helix 44 * 16S rRNA Subject RIV: BO - Biophysics Impact factor: 2.480, year: 2006

  19. Recent advances in fast-switching liquid-crystal mode based on uniform lying helix (ULH) (Conference Presentation)

    Science.gov (United States)

    Wittek, Michael; Siemianowski, Simon; Fiebranz, Bernd; Klasen-Memmer, Melanie

    2017-02-01

    Uniform lying Helix (ULH) is one of the most promising new LC modes for fast switching applications with alternative driving schemes such as FSC (field sequential colour). ULH operates via flexoelectric switching, unlike current commercially available liquid crystal modes that operate via dielectric switching. The realisation of mixtures with suitable flexoelectric properties requires unique materials composed of specially designed ´bi-mesogenś and also the addition of a chiral dopant. Recent advances, updated mixture performance and other challenges such as alignment and driving for this new mode are discussed.

  20. Molecular structure of r/GCG/d/TATACGC/ - A DNA-RNA hybrid helix joined to double helical DNA

    Science.gov (United States)

    Wang, A. H.-J.; Fujii, S.; Rich, A.; Van Boom, J. H.; Van Der Marel, G. A.; Van Boeckel, S. A. A.

    1982-01-01

    The molecule r(GCG)d(TATACGC) is self-complementary and forms two DNA-RNA hybrid segments surrounding a central region of double helical DNA; its molecular structure has been solved by X-ray analysis. All three parts of the molecule adopt a conformation which is close to that seen in the 11-fold RNA double helix. The conformation of the ribonucleotides is partly determined by water molecules bridging between the ribose O2' hydroxyl group and cytosine O2. The hybrid-DNA duplex junction contains no structural discontinuities. However, the central DNA TATA sequence has some structural irregularities.