WorldWideScience

Sample records for model sumo substrates

  1. Sumo-dependent substrate targeting of the SUMO protease Ulp1

    Directory of Open Access Journals (Sweden)

    Westerbeck Jason W

    2011-10-01

    Full Text Available Abstract Background In the yeast Saccharomyces cerevisiae, the essential small ubiquitin-like modifier (SUMO protease Ulp1 is responsible for both removing SUMO/Smt3 from specific target proteins and for processing precursor SUMO into its conjugation-competent form. Ulp1 localizes predominantly to nuclear pore complexes but has also been shown to deconjugate sumoylated septins at the bud-neck of dividing cells. How Ulp1 is directed to bud-neck localized septins and other cytoplasmic deconjugation targets is not well understood. Results Using a structure/function approach, we set out to elucidate features of Ulp1 that are required for substrate targeting. To aid our studies, we took advantage of a catalytically inactive mutant of Ulp1 that is greatly enriched at the septin ring of dividing yeast cells. We found that the localization of Ulp1 to the septins requires both SUMO and specific structural features of Ulp1's catalytic domain. Our analysis identified a 218-amino acid, substrate-trapping mutant of the catalytic domain of Ulp1, Ulp1(3(C580S, that is necessary and sufficient for septin localization. We also used the targeting and SUMO-binding properties of Ulp1(3(C580S to purify Smt3-modified proteins from cell extracts. Conclusions Our study provides novel insights into how the Ulp1 SUMO protease is actively targeted to its substrates in vivo and in vitro. Furthermore, we found that a substrate-trapping Ulp1(3(C580S interacts robustly with human SUMO1, SUMO2 and SUMO2 chains, making it a potentially useful tool for the analysis and purification of SUMO-modified proteins.

  2. Proteome-wide screens for small ubiquitin-like modifier (SUMO) substrates identify Arabidopsis proteins implicated in diverse biological processes

    National Research Council Canada - National Science Library

    Nabil Elrouby; George Coupland

    2010-01-01

    .... Despite this, little is known about SUMO targets in plants. Here we identified 238 Arabidopsis proteins as potential SUMO substrates because they interacted with SUMO-conjugating enzyme and/or SUMO protease (ESD4...

  3. Advances on SUMO substrates in Arabidopsis%拟南芥SUMO 底物的研究进展

    Institute of Scientific and Technical Information of China (English)

    郭敏霞; 傅永福

    2013-01-01

    SUMO(Small ubiquitin-related modifier)化修饰是普遍存在于真核生物中的一种翻译后修饰,在很多细胞过程中发挥重要作用.文章阐述了拟南芥中SUMO 底物的最新研究进展.首先介绍SUMO 化修饰过程,SUMO底物的鉴定,包括鉴定方法的进展和鉴定成果.然后依据底物的亚细胞定位、生理功能和参与的生理生化过程对其进行分类和分析,以期更好地理解蛋白质SUMO 化修饰方式在植物生长发育中的功能.%SUMO (Small ubiquitin-related modifier) modification, one of the essential posttranslational modifications in eukaryotes, plays an important role in various cellular processes. This review summarized the recent progresses on SUMO substrates in Arabidopsis. We firstly described the pathway of SUMO conjugation, and then focused on screening for SUMO substrates, including the methods of identification and SUMO substrates identified. Finally, we classified these substrates on the basis of their subcellular localization, functions, and biological processes. It is expected to provide the basis for better understanding the roles of sumoylation of proteins in plants.

  4. SUMO-1 gene transfer improves cardiac function in a large-animal model of heart failure.

    Science.gov (United States)

    Tilemann, Lisa; Lee, Ahyoung; Ishikawa, Kiyotake; Aguero, Jaume; Rapti, Kleopatra; Santos-Gallego, Carlos; Kohlbrenner, Erik; Fish, Kenneth M; Kho, Changwon; Hajjar, Roger J

    2013-11-13

    Recently, the impact of small ubiquitin-related modifier 1 (SUMO-1) on the regulation and preservation of sarcoplasmic reticulum calcium adenosine triphosphatase (SERCA2a) function was discovered. The amount of myocardial SUMO-1 is decreased in failing hearts, and its knockdown results in severe heart failure (HF) in mice. In a previous study, we showed that SUMO-1 gene transfer substantially improved cardiac function in a murine model of pressure overload-induced HF. Toward clinical translation, we evaluated in this study the effects of SUMO-1 gene transfer in a swine model of ischemic HF. One month after balloon occlusion of the proximal left anterior descending artery followed by reperfusion, the animals were randomized to receive either SUMO-1 at two doses, SERCA2a, or both by adeno-associated vector type 1 (AAV1) gene transfer via antegrade coronary infusion. Control animals received saline infusions. After gene delivery, there was a significant increase in the maximum rate of pressure rise [dP/dt(max)] that was most pronounced in the group that received both SUMO-1 and SERCA2a. The left ventricular ejection fraction (LVEF) improved after high-dose SUMO-1 with or without SERCA2a gene delivery, whereas there was a decline in LVEF in the animals receiving saline. Furthermore, the dilatation of LV volumes was prevented in the treatment groups. SUMO-1 gene transfer therefore improved cardiac function and stabilized LV volumes in a large-animal model of HF. These results support the critical role of SUMO-1 in SERCA2a function and underline the therapeutic potential of SUMO-1 for HF patients.

  5. SUMO: regulating the regulator

    Directory of Open Access Journals (Sweden)

    Bossis Guillaume

    2006-06-01

    Full Text Available Abstract Post-translational modifiers of the SUMO (Small Ubiquitin-related Modifier family have emerged as key regulators of protein function and fate. While the past few years have seen an enormous increase in knowledge on SUMO enzymes, substrates, and consequences of modification, regulation of SUMO conjugation is far from being understood. This brief review will provide an overview on recent advances concerning (i the interplay between sumoylation and other post-translational modifications at the level of individual targets and (ii global regulation of SUMO conjugation and deconjugation.

  6. Modellering van begrazing in SUMO : verbetering van de vegetatiemodellering in de Natuurplanner

    NARCIS (Netherlands)

    Wamelink, G.W.W.; Wegman, R.M.A.; Slim, P.A.; Dirksen, J.; Mol-Dijkstra, J.P.; Dobben, van H.F.

    2001-01-01

    In samenwerking met het RIVM en WU wordt het vegetatievoorspellingsmodel SUMO ontwikkeld. Het model vormt een integraal onderdeel met SMART en wordt onder andere gebruikt in de natuurplanner van het RIVM. Om de modellering van de effecten van begrazing op de vegetatieontwikkeling en successie mogeli

  7. Identification of SUMO target proteins by quantitative proteomics

    DEFF Research Database (Denmark)

    Andersen, Jens S; Matic, Ivan; Vertegaal, Alfred C O

    2009-01-01

    The identification of target proteins for small ubiquitin-like modifiers (SUMOs) is a critical step towards a detailed understanding of the cellular functions of SUMOs. Substrate protein identification for SUMOs is hampered by the low abundance of SUMO targets, the finding that only a small fract...

  8. Crystal Structure of UBA2[superscript ufd]-Ubc9: Insights into E1-E2 Interactions in Sumo Pathways

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Jing; Taherbhoy, Asad M.; Hunt, Harold W.; Seyedin, Steven N.; Miller, David W.; Miller, Darcie J.; Huang, Danny T.; Schulman, Brenda A. (SJCH)

    2012-04-30

    Canonical ubiquitin-like proteins (UBLs) such as ubiquitin, Sumo, NEDD8, and ISG15 are ligated to targets by E1-E2-E3 multienzyme cascades. The Sumo cascade, conserved among all eukaryotes, regulates numerous biological processes including protein localization, transcription, DNA replication, and mitosis. Sumo conjugation is initiated by the heterodimeric Aos1-Uba2 E1 enzyme (in humans called Sae1-Uba2), which activates Sumo's C-terminus, binds the dedicated E2 enzyme Ubc9, and promotes Sumo C-terminal transfer between the Uba2 and Ubc9 catalytic cysteines. To gain insights into details of E1-E2 interactions in the Sumo pathway, we determined crystal structures of the C-terminal ubiquitin fold domain (ufd) from yeast Uba2 (Uba2{sup ufd}), alone and in complex with Ubc9. The overall structures of both yeast Uba2{sup ufd} and Ubc9 superimpose well on their individual human counterparts, suggesting conservation of fundamental features of Sumo conjugation. Docking the Uba2{sup ufd}-Ubc9 and prior full-length human Uba2 structures allows generation of models for steps in Sumo transfer from Uba2 to Ubc9, and supports the notion that Uba2 undergoes remarkable conformational changes during the reaction. Comparisons to previous structures from the NEDD8 cascade demonstrate that UBL cascades generally utilize some parallel E1-E2 interaction surfaces. In addition, the structure of the Uba2{sup ufd}-Ubc9 complex reveals interactions unique to Sumo E1 and E2. Comparison with a previous Ubc9-E3 complex structure demonstrates overlap between Uba2 and E3 binding sites on Ubc9, indicating that loading with Sumo and E3-catalyzed transfer to substrates are strictly separate steps. The results suggest mechanisms establishing specificity and order in Sumo conjugation cascades.

  9. GPS-SUMO: a tool for the prediction of sumoylation sites and SUMO-interaction motifs.

    Science.gov (United States)

    Zhao, Qi; Xie, Yubin; Zheng, Yueyuan; Jiang, Shuai; Liu, Wenzhong; Mu, Weiping; Liu, Zexian; Zhao, Yong; Xue, Yu; Ren, Jian

    2014-07-01

    Small ubiquitin-like modifiers (SUMOs) regulate a variety of cellular processes through two distinct mechanisms, including covalent sumoylation and non-covalent SUMO interaction. The complexity of SUMO regulations has greatly hampered the large-scale identification of SUMO substrates or interaction partners on a proteome-wide level. In this work, we developed a new tool called GPS-SUMO for the prediction of both sumoylation sites and SUMO-interaction motifs (SIMs) in proteins. To obtain an accurate performance, a new generation group-based prediction system (GPS) algorithm integrated with Particle Swarm Optimization approach was applied. By critical evaluation and comparison, GPS-SUMO was demonstrated to be substantially superior against other existing tools and methods. With the help of GPS-SUMO, it is now possible to further investigate the relationship between sumoylation and SUMO interaction processes. A web service of GPS-SUMO was implemented in PHP+JavaScript and freely available at http://sumosp.biocuckoo.org.

  10. Lamin A/C mutants disturb sumo1 localization and sumoylation in vitro and in vivo.

    Directory of Open Access Journals (Sweden)

    Émilie Boudreau

    Full Text Available A-type lamins A and C are nuclear intermediate filament proteins in which mutations have been implicated in multiple disease phenotypes commonly known as laminopathies. A few studies have implicated sumoylation in the regulation of A-type lamins. Sumoylation is a post-translational protein modification that regulates a wide range of cellular processes through the attachment of small ubiquitin-related modifier (sumo to various substrates. Here we showed that laminopathy mutants result in the mislocalization of sumo1 both in vitro (C2C12 cells overexpressing mutant lamins A and C and in vivo (primary myoblasts and myopathic muscle tissue from the Lmna(H222P/H222P mouse model. In C2C12 cells, we showed that the trapping of sumo1 in p.Asp192Gly, p.Gln353Lys, and p.Arg386Lys aggregates of lamin A/C correlated with an increased steady-state level of sumoylation. However, lamin A and C did not appear to be modified by sumo1. Our results suggest that mutant lamin A/C alters the dynamics of sumo1 and thus misregulation of sumoylation may be contributing to disease progression in laminopathies.

  11. Regulation of SUMO Modification

    NARCIS (Netherlands)

    P.M. Knipscheer (Puck Maria)

    2007-01-01

    textabstractThe small ubiquitin related modifier SUMO is a posttranslational modifier that functions in a wide range of cellular processes like intracellular transport, cell cycle regulation, DNA repair and regulation of transcription. SUMO is an 11 kDa protein and is ligated to its target proteins

  12. SUMO and KSHV Replication

    Energy Technology Data Exchange (ETDEWEB)

    Chang, Pei-Ching [Institute of Microbiology and Immunology, National Yang-Ming University, Taipei 112, Taiwan (China); Kung, Hsing-Jien, E-mail: hkung@nhri.org.tw [Institute for Translational Medicine, College of Medical Science and Technology, Taipei Medical University, Taipei 110, Taiwan (China); Department of Biochemistry and Molecular Medicine, University of California, Davis, CA 95616 (United States); UC Davis Cancer Center, University of California, Davis, CA 95616 (United States); Division of Molecular and Genomic Medicine, National Health Research Institutes, 35 Keyan Road, Zhunan, Miaoli County 35053, Taiwan (China)

    2014-09-29

    Small Ubiquitin-related MOdifier (SUMO) modification was initially identified as a reversible post-translational modification that affects the regulation of diverse cellular processes, including signal transduction, protein trafficking, chromosome segregation, and DNA repair. Increasing evidence suggests that the SUMO system also plays an important role in regulating chromatin organization and transcription. It is thus not surprising that double-stranded DNA viruses, such as Kaposi’s sarcoma-associated herpesvirus (KSHV), have exploited SUMO modification as a means of modulating viral chromatin remodeling during the latent-lytic switch. In addition, SUMO regulation allows the disassembly and assembly of promyelocytic leukemia protein-nuclear bodies (PML-NBs), an intrinsic antiviral host defense, during the viral replication cycle. Overcoming PML-NB-mediated cellular intrinsic immunity is essential to allow the initial transcription and replication of the herpesvirus genome after de novo infection. As a consequence, KSHV has evolved a way as to produce multiple SUMO regulatory viral proteins to modulate the cellular SUMO environment in a dynamic way during its life cycle. Remarkably, KSHV encodes one gene product (K-bZIP) with SUMO-ligase activities and one gene product (K-Rta) that exhibits SUMO-targeting ubiquitin ligase (STUbL) activity. In addition, at least two viral products are sumoylated that have functional importance. Furthermore, sumoylation can be modulated by other viral gene products, such as the viral protein kinase Orf36. Interference with the sumoylation of specific viral targets represents a potential therapeutic strategy when treating KSHV, as well as other oncogenic herpesviruses. Here, we summarize the different ways KSHV exploits and manipulates the cellular SUMO system and explore the multi-faceted functions of SUMO during KSHV’s life cycle and pathogenesis.

  13. Characterizing the N- and C-terminal Small Ubiquitin-like Modifier (SUMO)-interacting Motifs of the Scaffold Protein DAXX

    NARCIS (Netherlands)

    Escobar-Cabrera, E.; Okon, M.; Lau, D.K.W.; Dart, C.F.; Bonvin, A.M.J.J.; McIntosh, L.P.

    2011-01-01

    DAXX is a scaffold protein with diverse roles that often depend upon binding SUMO via its N- and/or C-terminal SUMO-interacting motifs (SIM-N and SIM-C). Using NMR spectroscopy, we characterized the in vitro binding properties of peptide models of SIM-N and SIM-C to SUMO-1 and SUMO-2. In each case,

  14. Regulation of REGγ cellular distribution and function by SUMO modification

    Institute of Scientific and Technical Information of China (English)

    Yan Wu; Honglin Luo; Xiaotao Li; Lu Wang; Ping Zhou; Guangqiang Wang; Yu Zeng; Ying Wang; Jian Liu; Bianhong Zhang; Shuang Liu

    2011-01-01

    Discovery of emerging REGy-regulated proteins has accentuated the RECry-proteasome as an important pathway in multiple biological processes, including cell growth, cell cycle regulation, and apoptosis. However, little is known about the regulation of the REGy-proteasome pathway. Here we demonstrate that REGγ can be SUMOylated in vitro and in vivo by SUMO-1, SUMO-2, and SUMO-3. The SUMO-E3 protein inhibitor of activated STAT(PIAS)1physically associates with REGy and promotes SUMOylation of REGy. SUMOylation of RECry was found to occur at multiple sites, including K6, K14, and K12. Mutation analysis indicated that these SUMO sites simultaneously contributed to the SUMOylation status of REGy in cells. Posttranslational modification of REGγ by SUMO conjugation was revealed to mediate cytosolic translocation of REGγ and to cause increased stability of this proteasome activator.SUMOylation-deficient REGγ displayed attenuated ability to degrade p21waf//Cipl due to reduced affinity of the REGγ SUMOylation-defective mutant for p21. Taken together, we report a previously unrecognized mechanism regulating the activity of the proteasome activator REGy. This regulatory mechanism may enable REGy to function as a more potent factor in protein degradation with a broader substrate spectrum.

  15. USP7 is a SUMO deubiquitinase essential for DNA replication

    DEFF Research Database (Denmark)

    Lecona, Emilio; Rodriguez-Acebes, Sara; Specks, Julia

    2016-01-01

    Post-translational modification of proteins by ubiquitin (Ub) and Ub-like modifiers regulates DNA replication. We have previously shown that chromatin around replisomes is rich in SUMO and poor in Ub, whereas mature chromatin exhibits an opposite pattern. How this SUMO-rich, Ub-poor environment...... is maintained at sites of DNA replication in mammalian cells remains unexplored. Here we identify USP7 as a replisome-enriched SUMO deubiquitinase that is essential for DNA replication. By acting on SUMO and SUMOylated proteins, USP7 counteracts their ubiquitination. Inhibition or genetic deletion of USP7 leads...... to the accumulation of Ub on SUMOylated proteins, which are displaced away from replisomes. Our findings provide a model explaining the differential accumulation of SUMO and Ub at replication forks and identify an essential role of USP7 in DNA replication that should be considered in the development of USP7...

  16. USP7 is a SUMO deubiquitinase essential for DNA replication.

    Science.gov (United States)

    Lecona, Emilio; Rodriguez-Acebes, Sara; Specks, Julia; Lopez-Contreras, Andres J; Ruppen, Isabel; Murga, Matilde; Muñoz, Javier; Mendez, Juan; Fernandez-Capetillo, Oscar

    2016-04-01

    Post-translational modification of proteins by ubiquitin (Ub) and Ub-like modifiers regulates DNA replication. We have previously shown that chromatin around replisomes is rich in SUMO and poor in Ub, whereas mature chromatin exhibits an opposite pattern. How this SUMO-rich, Ub-poor environment is maintained at sites of DNA replication in mammalian cells remains unexplored. Here we identify USP7 as a replisome-enriched SUMO deubiquitinase that is essential for DNA replication. By acting on SUMO and SUMOylated proteins, USP7 counteracts their ubiquitination. Inhibition or genetic deletion of USP7 leads to the accumulation of Ub on SUMOylated proteins, which are displaced away from replisomes. Our findings provide a model explaining the differential accumulation of SUMO and Ub at replication forks and identify an essential role of USP7 in DNA replication that should be considered in the development of USP7 inhibitors as anticancer agents.

  17. Identification and molecular properties of SUMO-binding proteins in arabidopsis

    KAUST Repository

    Park, Hyeongcheol

    2011-05-20

    Reversible conjugation of the small ubiquitin modifier (SUMO) peptide to proteins (SUMOylation) plays important roles in cellular processes in animals and yeasts. However, little is known about plant SUMO targets. To identify SUMO substrates in Arabidopsis and to probe for biological functions of SUMO proteins, we constructed 6xHis-3xFLAG fused AtSUMO1 (HFAtSUMO1) controlled by the CaMV35S promoter for transformation into Arabidopsis Col-0. After heat treatment, an increased sumoylation pattern was detected in the transgenic plants. SUMO1-modified proteins were selected after two-dimensional gel electrophoresis (2-DE) image analysis and identified using matrix-assisted laser-desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). We identified 27 proteins involved in a variety of processes such as nucleic acid metabolism, signaling, metabolism, and including proteins of unknown functions. Binding and sumoylation patterns were confirmed independently. Surprisingly, MCM3 (At5G46280), a DNA replication licensing factor, only interacted with and became sumoylated by AtSUMO1, but not by SUMO1ΔGG or AtSUMO3. The results suggest specific interactions between sumoylation targets and particular sumoylation enzymes. ©2011 KSMCB.

  18. Meiosis and SUMO

    DEFF Research Database (Denmark)

    Holm, Lærke Rebekka

    to target proteins can be catalyzed by the SUMO E3 ligase Pli1. In this study we investigate the role of Pli1 and Pmt3 during meiotic differentiation and at repetitive DNA during mitotic growth. Target proteins for Pmt3 are many; however, Pli1 has a meiosis-specic function regulating meiotic recombination...

  19. Meiosis and SUMO

    DEFF Research Database (Denmark)

    Holm, Lærke Rebekka

    to target proteins can be catalyzed by the SUMO E3 ligase Pli1. In this study we investigate the role of Pli1 and Pmt3 during meiotic differentiation and at repetitive DNA during mitotic growth. Target proteins for Pmt3 are many; however, Pli1 has a meiosis-specic function regulating meiotic recombination...

  20. Divergent signaling via SUMO modification: potential for CFTR modulation.

    Science.gov (United States)

    Ahner, Annette; Gong, Xiaoyan; Frizzell, Raymond A

    2016-02-01

    The cystic fibrosis transmembrane conductance regulator (CFTR) is generally responsible for the cAMP/PKA regulated anion conductance at the apical membranes of secretory epithelial cells. Mutations in CFTR underlie cystic fibrosis (CF), in which the most common variant, F508del, causes protein misfolding and its proteasome-mediated degradation. A new pathway that contributes to mutant CFTR degradation is mediated by the small heat shock protein, Hsp27, which cooperates with Ubc9, the E2 enzyme for SUMOylation, to selectively conjugate mutant CFTR with SUMO-2/3. This SUMO paralog can form polychains, which are recognized by the ubiquitin E3 enzyme, RNF4, leading to CFTR ubiquitylation and recognition by the proteasome. We found also that F508del CFTR could be modified by SUMO-1, a paralog that does not support SUMO polychain formation. The use of different SUMO paralogs to modify and target a single substrate for divergent purposes is not uncommon. In this short review we discuss the possibility that conjugation with SUMO-1 could protect mutant CFTR from disposal by RNF4 and similar ubiquitin ligases. We hypothesize that such a pathway could contribute to therapeutic efforts to stabilize immature mutant CFTR and thereby enhance the action of therapeutics that correct CFTR trafficking to the apical membranes.

  1. SUMO Wrestles with Recombination

    Directory of Open Access Journals (Sweden)

    Lumír Krejčí

    2012-07-01

    Full Text Available DNA double-strand breaks (DSBs comprise one of the most toxic DNA lesions, as the failure to repair a single DSB has detrimental consequences on the cell. Homologous recombination (HR constitutes an error-free repair pathway for the repair of DSBs. On the other hand, when uncontrolled, HR can lead to genome rearrangements and needs to be tightly regulated. In recent years, several proteins involved in different steps of HR have been shown to undergo modification by small ubiquitin-like modifier (SUMO peptide and it has been suggested that deficient sumoylation impairs the progression of HR. This review addresses specific effects of sumoylation on the properties of various HR proteins and describes its importance for the homeostasis of DNA repetitive sequences. The article further illustrates the role of sumoylation in meiotic recombination and the interplay between SUMO and other post-translational modifications.

  2. 2nd SUMO Conference

    CERN Document Server

    Weber, Melanie

    2015-01-01

    This contributed volume contains the conference proceedings of the Simulation of Urban Mobility (SUMO) conference 2014, Berlin. The included research papers cover a wide range of topics in traffic planning and simulation, including open data, vehicular communication, e-mobility, urban mobility, multimodal traffic as well as usage approaches. The target audience primarily comprises researchers and experts in the field, but the book may also be beneficial for graduate students.  

  3. Quantitative Förster resonance energy transfer analysis for kinetic determinations of SUMO-specific protease.

    Science.gov (United States)

    Liu, Yan; Song, Yang; Madahar, Vipul; Liao, Jiayu

    2012-03-01

    Förster resonance energy transfer (FRET) technology has been widely used in biological and biomedical research, and it is a very powerful tool for elucidating protein interactions in either dynamic or steady state. SUMOylation (the process of SUMO [small ubiquitin-like modifier] conjugation to substrates) is an important posttranslational protein modification with critical roles in multiple biological processes. Conjugating SUMO to substrates requires an enzymatic cascade. Sentrin/SUMO-specific proteases (SENPs) act as an endopeptidase to process the pre-SUMO or as an isopeptidase to deconjugate SUMO from its substrate. To fully understand the roles of SENPs in the SUMOylation cycle, it is critical to understand their kinetics. Here, we report a novel development of a quantitative FRET-based protease assay for SENP1 kinetic parameter determination. The assay is based on the quantitative analysis of the FRET signal from the total fluorescent signal at acceptor emission wavelength, which consists of three components: donor (CyPet-SUMO1) emission, acceptor (YPet) emission, and FRET signal during the digestion process. Subsequently, we developed novel theoretical and experimental procedures to determine the kinetic parameters, k(cat), K(M), and catalytic efficiency (k(cat)/K(M)) of catalytic domain SENP1 toward pre-SUMO1. Importantly, the general principles of this quantitative FRET-based protease kinetic determination can be applied to other proteases. Copyright © 2011 Elsevier Inc. All rights reserved.

  4. Joosep ja sumo / Mart Velsker

    Index Scriptorium Estoniae

    Velsker, Mart, 1966-

    2008-01-01

    Arvustus: Kivisildnik. Sumo. Pärnu : Jumalikud Ilmutused, 2007 ; Rilke, Rainer Maria. Joosepi kahtlused / tõlk. Kivisildnik. Pärnu : Jumalikud Ilmutused, 2008. - Artikkel sisaldab bibliograafiat joonealustes viidetes

  5. SUMO-1 regulates the conformational dynamics of Thymine-DNA Glycosylase regulatory domain and competes with its DNA binding activity

    Directory of Open Access Journals (Sweden)

    Eilebrecht Sebastian

    2011-02-01

    Full Text Available Abstract Background The human thymine-DNA glycosylase (TDG plays a dual role in base excision repair of G:U/T mismatches and in transcription. Regulation of TDG activity by SUMO-1 conjugation was shown to act on both functions. Furthermore, TDG can interact with SUMO-1 in a non-covalent manner. Results Using NMR spectroscopy we have determined distinct conformational changes in TDG upon either covalent sumoylation on lysine 330 or intermolecular SUMO-1 binding through a unique SUMO-binding motif (SBM localized in the C-terminal region of TDG. The non-covalent SUMO-1 binding induces a conformational change of the TDG amino-terminal regulatory domain (RD. Such conformational dynamics do not exist with covalent SUMO-1 attachment and could potentially play a broader role in the regulation of TDG functions for instance during transcription. Both covalent and non-covalent processes activate TDG G:U repair similarly. Surprisingly, despite a dissociation of the SBM/SUMO-1 complex in presence of a DNA substrate, SUMO-1 preserves its ability to stimulate TDG activity indicating that the non-covalent interactions are not directly involved in the regulation of TDG activity. SUMO-1 instead acts, as demonstrated here, indirectly by competing with the regulatory domain of TDG for DNA binding. Conclusions SUMO-1 increases the enzymatic turnover of TDG by overcoming the product-inhibition of TDG on apurinic sites. The mechanism involves a competitive DNA binding activity of SUMO-1 towards the regulatory domain of TDG. This mechanism might be a general feature of SUMO-1 regulation of other DNA-bound factors such as transcription regulatory proteins.

  6. SUMO-2 and PIAS1 modulate insoluble mutant huntingtin protein accumulation.

    Science.gov (United States)

    O'Rourke, Jacqueline Gire; Gareau, Jaclyn R; Ochaba, Joseph; Song, Wan; Raskó, Tamás; Reverter, David; Lee, John; Monteys, Alex Mas; Pallos, Judit; Mee, Lisa; Vashishtha, Malini; Apostol, Barbara L; Nicholson, Thomas Peter; Illes, Katalin; Zhu, Ya-Zhen; Dasso, Mary; Bates, Gillian P; Difiglia, Marian; Davidson, Beverly; Wanker, Erich E; Marsh, J Lawrence; Lima, Christopher D; Steffan, Joan S; Thompson, Leslie M

    2013-07-25

    A key feature in Huntington disease (HD) is the accumulation of mutant Huntingtin (HTT) protein, which may be regulated by posttranslational modifications. Here, we define the primary sites of SUMO modification in the amino-terminal domain of HTT, show modification downstream of this domain, and demonstrate that HTT is modified by the stress-inducible SUMO-2. A systematic study of E3 SUMO ligases demonstrates that PIAS1 is an E3 SUMO ligase for both HTT SUMO-1 and SUMO-2 modification and that reduction of dPIAS in a mutant HTT Drosophila model is protective. SUMO-2 modification regulates accumulation of insoluble HTT in HeLa cells in a manner that mimics proteasome inhibition and can be modulated by overexpression and acute knockdown of PIAS1. Finally, the accumulation of SUMO-2-modified proteins in the insoluble fraction of HD postmortem striata implicates SUMO-2 modification in the age-related pathogenic accumulation of mutant HTT and other cellular proteins that occurs during HD progression.

  7. SUMO-2 and PIAS1 Modulate Insoluble Mutant Huntingtin Protein Accumulation

    Directory of Open Access Journals (Sweden)

    Jacqueline Gire O’Rourke

    2013-07-01

    Full Text Available A key feature in Huntington disease (HD is the accumulation of mutant Huntingtin (HTT protein, which may be regulated by posttranslational modifications. Here, we define the primary sites of SUMO modification in the amino-terminal domain of HTT, show modification downstream of this domain, and demonstrate that HTT is modified by the stress-inducible SUMO-2. A systematic study of E3 SUMO ligases demonstrates that PIAS1 is an E3 SUMO ligase for both HTT SUMO-1 and SUMO-2 modification and that reduction of dPIAS in a mutant HTT Drosophila model is protective. SUMO-2 modification regulates accumulation of insoluble HTT in HeLa cells in a manner that mimics proteasome inhibition and can be modulated by overexpression and acute knockdown of PIAS1. Finally, the accumulation of SUMO-2-modified proteins in the insoluble fraction of HD postmortem striata implicates SUMO-2 modification in the age-related pathogenic accumulation of mutant HTT and other cellular proteins that occurs during HD progression.

  8. SUMO-fusion, purification, and characterization of a (+)-zizaene synthase from Chrysopogon zizanioides

    Energy Technology Data Exchange (ETDEWEB)

    Hartwig, S.; Frister, T.; Alemdar, S.; Li, Z.; Scheper, T.; Beutel, S., E-mail: beutel@iftc.uni-hannover.de

    2015-03-20

    An uncharacterized plant cDNA coding for a polypeptide presumably having sesquiterpene synthase activity, was expressed in soluble and active form. Two expression strategies were evaluated in Escherichia coli. The enzyme was fused to a highly soluble SUMO domain, in addition to being produced in an unfused form by a cold-shock expression system. Yields up to ∼325 mg/L{sup −1} were achieved in batch cultivations. The 6x-His-tagged enzyme was purified employing an Ni{sup 2+}-IMAC-based procedure. Identity of the protein was established by Western Blot analysis as well as peptide mass fingerprinting. A molecular mass of 64 kDa and an isoelectric point of pI 4.95 were determined by 2D gel electrophoresis. Cleavage of the fusion domain was possible by digestion with specific SUMO protease. The synthase was active in Mg{sup 2+} containing buffer and catalyzed the production of (+)-zizaene (syn. khusimene), a precursor of khusimol, from farnesyl diphosphate. Product identity was confirmed by GC–MS and comparison of retention indices. Enzyme kinetics were determined by measuring initial reaction rates for the product, using varying substrate concentrations. By assuming a Michaelis–Menten model, kinetic parameters of K{sub M} = 1.111 μM (±0.113), v{sub max} = 0.3245 μM min{sup −1} (±0.0035), k{sub cat} = 2.95 min{sup −1}, as well as a catalytic efficiency k{sub cat}/K{sub M} = 4.43 × 10{sup 4} M{sup −1} s{sup −1} were calculated. Fusion to a SUMO moiety can substantially increase soluble expression levels of certain hard to express terpene synthases in E. coli. The kinetic data determined for the recombinant synthase are comparable to other described plant sesquiterpene synthases and in the typical range of enzymes belonging to the secondary metabolism. This leaves potential for optimizing catalytic parameters through methods like directed evolution. - Highlights: • Uncharacterized (+)-zizaene synthase from C. zizanoides was cloned

  9. SUMO-fusion, purification, and characterization of a (+)-zizaene synthase from Chrysopogon zizanioides.

    Science.gov (United States)

    Hartwig, S; Frister, T; Alemdar, S; Li, Z; Scheper, T; Beutel, S

    2015-03-20

    An uncharacterized plant cDNA coding for a polypeptide presumably having sesquiterpene synthase activity, was expressed in soluble and active form. Two expression strategies were evaluated in Escherichia coli. The enzyme was fused to a highly soluble SUMO domain, in addition to being produced in an unfused form by a cold-shock expression system. Yields up to ∼325 mg/L(-1) were achieved in batch cultivations. The 6x-His-tagged enzyme was purified employing an Ni(2+)-IMAC-based procedure. Identity of the protein was established by Western Blot analysis as well as peptide mass fingerprinting. A molecular mass of 64 kDa and an isoelectric point of pI 4.95 were determined by 2D gel electrophoresis. Cleavage of the fusion domain was possible by digestion with specific SUMO protease. The synthase was active in Mg(2+) containing buffer and catalyzed the production of (+)-zizaene (syn. khusimene), a precursor of khusimol, from farnesyl diphosphate. Product identity was confirmed by GC-MS and comparison of retention indices. Enzyme kinetics were determined by measuring initial reaction rates for the product, using varying substrate concentrations. By assuming a Michaelis-Menten model, kinetic parameters of KM = 1.111 μM (±0.113), vmax = 0.3245 μM min(-1) (±0.0035), kcat = 2.95 min(-1), as well as a catalytic efficiency kcat/KM = 4.43 × 10(4) M(-1)s(-1) were calculated. Fusion to a SUMO moiety can substantially increase soluble expression levels of certain hard to express terpene synthases in E. coli. The kinetic data determined for the recombinant synthase are comparable to other described plant sesquiterpene synthases and in the typical range of enzymes belonging to the secondary metabolism. This leaves potential for optimizing catalytic parameters through methods like directed evolution. Copyright © 2015 Elsevier Inc. All rights reserved.

  10. SUMO-1 possesses DNA binding activity

    Directory of Open Access Journals (Sweden)

    Wieruszeski Jean-Michel

    2010-05-01

    Full Text Available Abstract Background Conjugation of small ubiquitin-related modifiers (SUMOs is a frequent post-translational modification of proteins. SUMOs can also temporally associate with protein-targets via SUMO binding motifs (SBMs. Protein sumoylation has been identified as an important regulatory mechanism especially in the regulation of transcription and the maintenance of genome stability. The precise molecular mechanisms by which SUMO conjugation and association act are, however, not understood. Findings Using NMR spectroscopy and protein-DNA cross-linking experiments, we demonstrate here that SUMO-1 can specifically interact with dsDNA in a sequence-independent fashion. We also show that SUMO-1 binding to DNA can compete with other protein-DNA interactions at the example of the regulatory domain of Thymine-DNA Glycosylase and, based on these competition studies, estimate the DNA binding constant of SUMO1 in the range 1 mM. Conclusion This finding provides an important insight into how SUMO-1 might exert its activity. SUMO-1 might play a general role in destabilizing DNA bound protein complexes thereby operating in a bottle-opener way of fashion, explaining its pivotal role in regulating the activity of many central transcription and DNA repair complexes.

  11. A high-yield double-purification proteomics strategy for the identification of SUMO sites.

    Science.gov (United States)

    Hendriks, Ivo A; Vertegaal, Alfred C O

    2016-09-01

    The small ubiquitin-like modifier (SUMO) is a protein modifier that is post-translationally coupled to thousands of lysines in more than a thousand proteins. An understanding of which lysines are modified by SUMO is critical in unraveling its function as a master regulator of all nuclear processes, as well as its involvement in diseases such as cancer. Here we describe a protocol for the lysine-deficient (K0) method for efficient identification of SUMOylated lysines by mass spectrometry (MS). To our knowledge, the K0 method is the only currently available method that can routinely identify >1,000 SUMO sites in mammalian cells under standard growth conditions. The K0 strategy relies on introducing a His10-tagged SUMO wherein all lysines have been substituted to arginines. Lysine deficiency renders the SUMO immune to digestion by the endoproteinase Lys-C, which in turn allows for stringent and high-yield tandem purification through the His10 tag. In addition, the His10-tagged SUMO also contains a C-terminal Q87R mutation, which accommodates generation of SUMO-site peptides with a QQTGG mass remnant after digestion with trypsin. This remnant possesses a unique mass signature and readily generates diagnostic ions in the fragment ion scans, which increases SUMO-site identification confidence. The K0 method can be applied in any mammalian cell line or in any model system that allows for integration of the K0-SUMO construct. From the moment of cell lysis, the K0 method takes ∼7 d to perform.

  12. The SUMO Pathway in Mitosis.

    Science.gov (United States)

    Mukhopadhyay, Debaditya; Dasso, Mary

    2017-01-01

    Mitosis is the stage of the cell cycle during which replicated chromosomes must be precisely divided to allow the formation of two daughter cells possessing equal genetic material. Much of the careful spatial and temporal organization of mitosis is maintained through post-translational modifications, such as phosphorylation and ubiquitination, of key cellular proteins. Here, we will review evidence that sumoylation, conjugation to the SUMO family of small ubiquitin-like modifiers, also serves essential regulatory roles during mitosis. We will discuss the basic biology of sumoylation, how the SUMO pathway has been implicated in particular mitotic functions, including chromosome condensation, centromere/kinetochore organization and cytokinesis, and what cellular proteins may be the targets underlying these phenomena.

  13. SUMO wrestling in cell movement

    Institute of Scientific and Technical Information of China (English)

    Miia M Rytinki; Jorma J Palvimo

    2011-01-01

    @@ Small ubiquitin-like modifier(SUMO)proteins 1,2 and 3 can be covalently conjugated to specific lysine residues on target proteins in a process dubbed SUMOylation.This conserved posttranslational modification that was reported for the first time in 1996 has emerged as an important regulatory mechanism in cell physiology,especially in nuclear signaling,transport,transcription and DNA replication/repair[1,2].

  14. Defining the SUMO System in Maize: SUMOylation Is Up-Regulated during Endosperm Development and Rapidly Induced by Stress.

    Science.gov (United States)

    Augustine, Robert C; York, Samuel L; Rytz, Thérèse C; Vierstra, Richard D

    2016-07-01

    In response to abiotic and biotic challenges, plants rapidly attach small ubiquitin-related modifier (SUMO) to a large collection of nuclear proteins, with studies in Arabidopsis (Arabidopsis thaliana) linking SUMOylation to stress tolerance via its modification of factors involved in chromatin and RNA dynamics. Despite this importance, little is known about SUMOylation in crop species. Here, we describe the plant SUMO system at the phylogenetic, biochemical, and transcriptional levels with a focus on maize (Zea mays). In addition to canonical SUMOs, land plants encode a loosely constrained noncanonical isoform and a variant containing a long extension upstream of the signature β-grasp fold, with cereals also expressing a novel diSUMO polypeptide bearing two SUMO β-grasp domains in tandem. Maize and other cereals also synthesize a unique SUMO-conjugating enzyme variant with more restricted expression patterns that is enzymatically active despite a distinct electrostatic surface. Maize SUMOylation primarily impacts nuclear substrates, is strongly induced by high temperatures, and displays a memory that suppresses subsequent conjugation. Both in-depth transcript and conjugate profiles in various maize organs point to tissue/cell-specific functions for SUMOylation, with potentially significant roles during embryo and endosperm maturation. Collectively, these studies define the organization of the maize SUMO system and imply important functions during seed development and stress defense.

  15. Senp1 Is Essential for Desumoylating Sumo1-Modified Proteins but Dispensable for Sumo2 and Sumo3 Deconjugation in the Mouse Embryo

    Directory of Open Access Journals (Sweden)

    Prashant Sharma

    2013-05-01

    Full Text Available Posttranslational modification with small ubiquitin-like modifier (Sumo regulates numerous cellular and developmental processes. Sumoylation is dynamic with deconjugation by Sumo-specific proteases (Senps regulating steady-state levels. Different Senps are found in distinct subcellular domains, which may limit their deconjugation activity to colocalizing Sumo-modified proteins. In vitro, Senps can discriminate between the different Sumo paralogs: Sumo1 versus the highly related Sumo2 and Sumo3 (Sumo2/3, which can form poly-Sumo chains. However, a full understanding of Senp specificity in vivo is still lacking. Here, using biochemical and genetic approaches, we establish that Senp1 has an essential, nonredundant function to desumoylate Sumo1-modified proteins during mouse embryonic development. Senp1 specificity for Sumo1 conjugates represents an intrinsic function and not simply a product of colocalization. In contrast, Senp1 has only a limited role in Sumo2/3 desumoylation, although it may regulate Sumo1-mediated termination of poly-Sumo2/3 chains.

  16. Modeling graphene-substrate interactions

    NARCIS (Netherlands)

    Amlaki, T.

    2016-01-01

    In this thesis I focussed on the interactions between graphene-like materials (grapheme and germanene) and various substrates. The attractive properties of graphene like a high carrier mobility, its single-atomic thickness and its theoretical magic have made graphene a very popular and promising can

  17. Modeling graphene-substrate interactions

    NARCIS (Netherlands)

    Amlaki, Taher

    2016-01-01

    In this thesis I focussed on the interactions between graphene-like materials (grapheme and germanene) and various substrates. The attractive properties of graphene like a high carrier mobility, its single-atomic thickness and its theoretical magic have made graphene a very popular and promising can

  18. SUMO: A small unmanned meteorological observer for atmospheric boundary layer research

    Energy Technology Data Exchange (ETDEWEB)

    Reuder, J; Jonassen, M; Mayer, S [Geophysical Institute, University of Bergen, Allegaten 70, 5009 Bergen (Norway); Brisset, P [Ecole Nationale de l' Aviation Civile (ENAC), 7 avenue Edouard Belin, 31055 Toulouse (France); Mueller, M [Orleansstrasse 26a, 31135 Hildesheim (Germany)], E-mail: joachim.reuder@gfi.uib.no, E-mail: pascal.brisset@enac.fr, E-mail: marius.jonassen@gfi.uib.no, E-mail: martin@pfump.org, E-mail: stephanie.mayer@gfi.uib.no

    2008-05-01

    A new system for atmospheric measurements in the lower troposphere has been developed and successfully tested. The presented Small Unmanned Meteorological Observer (SUMO) is based on a light-weighted commercially available model airplane, equipped with an autopilot and meteorological sensors for temperature, humidity and pressure. During the 5 week field campaign FLOHOF (Flow over and around HofsjoUkull) in Central Iceland the system has been successfully tested in July/August 2007. Atmospheric profiles of temperature, humidity, wind speed and wind direction have been determined up to 3500 m above ground. In addition the applicability of SUMO for horizontal surveys up to 4 km away from the launch site has been approved. During a 3 week campaign on and around Spitsbergen in February/March 2008 the SUMO system also proved its functionality under harsh polar conditions, reaching altitudes above 1500 m at ground temperatures of -20 deg. C and wind speeds up to 15 m s{sup -1}. With its wingspan of 80 cm, its length of 75 cm and its weight of below 600 g, SUMO is easy to transport and operate even in remote areas. The direct material costs for one SUMO unit, including airplane, autopilot and sensors are below 1200 Euro. Assuming at least several tenths of flights for each airframe, SUMO provides a cost-efficient measurement system with a large potential to close the existing observational gap of reasonable atmospheric measurement systems in between meteorological masts/towers and radiosondes.

  19. Functional characterization of a SUMO deconjugating protease of Plasmodium falciparum using newly identified small molecule inhibitors.

    Science.gov (United States)

    Ponder, Elizabeth L; Albrow, Victoria E; Leader, Brittany A; Békés, Miklós; Mikolajczyk, Jowita; Fonović, Urša Pečar; Shen, Aimee; Drag, Marcin; Xiao, Junpeng; Deu, Edgar; Campbell, Amy J; Powers, James C; Salvesen, Guy S; Bogyo, Matthew

    2011-06-24

    Small ubiquitin-related modifier (SUMO) is implicated in the regulation of numerous biological processes including transcription, protein localization, and cell cycle control. Protein modification by SUMO is found in Plasmodium falciparum; however, its role in the regulation of the parasite life cycle is poorly understood. Here we describe functional studies of a SUMO-specific protease (SENP) of P. falciparum, PfSENP1 (PFL1635w). Expression of the catalytic domain of PfSENP1 and biochemical profiling using a positional scanning substrate library demonstrated that this protease has unique cleavage sequence preference relative to the human SENPs. In addition, we describe a class of small molecule inhibitors of this protease. The most potent lead compound inhibited both recombinant PfSENP1 activity and P. falciparum replication in infected human blood. These studies provide valuable new tools for the study of SUMOylation in P. falciparum. Copyright © 2011 Elsevier Ltd. All rights reserved.

  20. Analysis of the SUMO2 Proteome during HSV-1 Infection.

    Directory of Open Access Journals (Sweden)

    Elizabeth Sloan

    2015-07-01

    Full Text Available Covalent linkage to members of the small ubiquitin-like (SUMO family of proteins is an important mechanism by which the functions of many cellular proteins are regulated. Sumoylation has roles in the control of protein stability, activity and localization, and is involved in the regulation of transcription, gene expression, chromatin structure, nuclear transport and RNA metabolism. Sumoylation is also linked, both positively and negatively, with the replication of many different viruses both in terms of modification of viral proteins and modulation of sumoylated cellular proteins that influence the efficiency of infection. One prominent example of the latter is the widespread reduction in the levels of cellular sumoylated species induced by herpes simplex virus type 1 (HSV-1 ubiquitin ligase ICP0. This activity correlates with relief from intrinsic immunity antiviral defence mechanisms. Previous work has shown that ICP0 is selective in substrate choice, with some sumoylated proteins such the promyelocytic leukemia protein PML being extremely sensitive, while RanGAP is completely resistant. Here we present a comprehensive proteomic analysis of changes in the cellular SUMO2 proteome during HSV-1 infection. Amongst the 877 potentially sumoylated species detected, we identified 124 whose abundance was decreased by a factor of 3 or more by the virus, several of which were validated by western blot and expression analysis. We found many previously undescribed substrates of ICP0 whose degradation occurs by a range of mechanisms, influenced or not by sumoylation and/or the SUMO2 interaction motif within ICP0. Many of these proteins are known or are predicted to be involved in the regulation of transcription, chromatin assembly or modification. These results present novel insights into mechanisms and host cell proteins that might influence the efficiency of HSV-1 infection.

  1. Characterization of the loss of SUMO pathway function on cancer cells and tumor proliferation.

    Science.gov (United States)

    He, Xingyue; Riceberg, Jessica; Pulukuri, Sai M; Grossman, Steve; Shinde, Vaishali; Shah, Pooja; Brownell, James E; Dick, Larry; Newcomb, John; Bence, Neil

    2015-01-01

    SUMOylation is a post-translational ubiquitin-like protein modification pathway that regulates important cellular processes including chromosome structure, kinetochore function, chromosome segregation, nuclear and sub-nuclear organization, transcription and DNA damage repair. There is increasing evidence that the SUMO pathway is dysregulated in cancer, raising the possibility that modulation of this pathway may have therapeutic potential. To investigate the importance of the SUMO pathway in the context of cancer cell proliferation and tumor growth, we applied lentivirus-based short hairpin RNAs (shRNA) to knockdown SUMO pathway genes in human cancer cells. shRNAs for SAE2 and UBC9 reduced SUMO conjugation activity and inhibited proliferation of human cancer cells. To expand upon these observations, we generated doxycycline inducible conditional shRNA cell lines for SAE2 to achieve acute and reversible SAE2 knockdown. Conditional SAE2 knockdown in U2OS and HCT116 cells slowed cell growth in vitro, and SAE2 knockdown induced multiple terminal outcomes including apoptosis, endoreduplication and senescence. Multinucleated cells became senescent and stained positive for the senescence marker, SA-β Gal, and displayed elevated levels of p53 and p21. In an attempt to explain these phenotypes, we confirmed that loss of SUMO pathway activity leads to a loss of SUMOylated Topoisomerase IIα and the appearance of chromatin bridges which can impair proper cytokinesis and lead to multinucleation. Furthermore, knockdown of SAE2 induces disruption of PML nuclear bodies which may further promote apoptosis or senescence. In an in vivo HCT116 xenograft tumor model, conditional SAE2 knockdown strongly impaired tumor growth. These data demonstrate that the SUMO pathway is required for cancer cell proliferation in vitro and tumor growth in vivo, implicating the SUMO pathway as a potential cancer therapeutic target.

  2. Characterization of the loss of SUMO pathway function on cancer cells and tumor proliferation.

    Directory of Open Access Journals (Sweden)

    Xingyue He

    Full Text Available SUMOylation is a post-translational ubiquitin-like protein modification pathway that regulates important cellular processes including chromosome structure, kinetochore function, chromosome segregation, nuclear and sub-nuclear organization, transcription and DNA damage repair. There is increasing evidence that the SUMO pathway is dysregulated in cancer, raising the possibility that modulation of this pathway may have therapeutic potential. To investigate the importance of the SUMO pathway in the context of cancer cell proliferation and tumor growth, we applied lentivirus-based short hairpin RNAs (shRNA to knockdown SUMO pathway genes in human cancer cells. shRNAs for SAE2 and UBC9 reduced SUMO conjugation activity and inhibited proliferation of human cancer cells. To expand upon these observations, we generated doxycycline inducible conditional shRNA cell lines for SAE2 to achieve acute and reversible SAE2 knockdown. Conditional SAE2 knockdown in U2OS and HCT116 cells slowed cell growth in vitro, and SAE2 knockdown induced multiple terminal outcomes including apoptosis, endoreduplication and senescence. Multinucleated cells became senescent and stained positive for the senescence marker, SA-β Gal, and displayed elevated levels of p53 and p21. In an attempt to explain these phenotypes, we confirmed that loss of SUMO pathway activity leads to a loss of SUMOylated Topoisomerase IIα and the appearance of chromatin bridges which can impair proper cytokinesis and lead to multinucleation. Furthermore, knockdown of SAE2 induces disruption of PML nuclear bodies which may further promote apoptosis or senescence. In an in vivo HCT116 xenograft tumor model, conditional SAE2 knockdown strongly impaired tumor growth. These data demonstrate that the SUMO pathway is required for cancer cell proliferation in vitro and tumor growth in vivo, implicating the SUMO pathway as a potential cancer therapeutic target.

  3. Direct and/or Indirect Roles for SUMO in Modulating Alpha-Synuclein Toxicity

    Directory of Open Access Journals (Sweden)

    Shamini Vijayakumaran

    2015-07-01

    Full Text Available α-Synuclein inclusion bodies are a pathological hallmark of several neurodegenerative diseases, including Parkinson’s disease, and contain aggregated α-synuclein and a variety of recruited factors, including protein chaperones, proteasome components, ubiquitin and the small ubiquitin-like modifier, SUMO-1. Cell culture and animal model studies suggest that misfolded, aggregated α-synuclein is actively translocated via the cytoskeletal system to a region of the cell where other factors that help to lessen the toxic effects can also be recruited. SUMO-1 covalently conjugates to various intracellular target proteins in a way analogous to ubiquitination to alter cellular distribution, function and metabolism and also plays an important role in a growing list of cellular pathways, including exosome secretion and apoptosis. Furthermore, SUMO-1 modified proteins have recently been linked to cell stress responses, such as oxidative stress response and heat shock response, with increased SUMOylation being neuroprotective in some cases. Several recent studies have linked SUMOylation to the ubiquitin-proteasome system, while other evidence implicates the lysosomal pathway. Other reports depict a direct mechanism whereby sumoylation reduced the aggregation tendency of α-synuclein, and reduced the toxicity. However, the precise role of SUMO-1 in neurodegeneration remains unclear. In this review, we explore the potential direct or indirect role(s of SUMO-1 in the cellular response to misfolded α-synuclein in neurodegenerative disorders.

  4. Elevated salicylic acid levels conferred by increased expression of ISOCHORISMATE SYNTHASE 1 contribute to hyperaccumulation of SUMO1 conjugates in the Arabidopsis mutant early in short days 4.

    Science.gov (United States)

    Villajuana-Bonequi, Mitzi; Elrouby, Nabil; Nordström, Karl; Griebel, Thomas; Bachmair, Andreas; Coupland, George

    2014-07-01

    Post-translational modification of proteins by attachment of small ubiquitin-like modifier (SUMO) is essential for plant growth and development. Mutations in the SUMO protease early in short days 4 (ESD4) cause hyperaccumulation of conjugates formed between SUMO and its substrates, and phenotypically are associated with extreme early flowering and impaired growth. We performed a suppressor mutagenesis screen of esd4 and identified a series of mutants called suppressor of esd4 (sed), which delay flowering, enhance growth and reduce hyperaccumulation of SUMO conjugates. Genetic mapping and genome sequencing indicated that one of these mutations (sed111) is in the gene salicylic acid induction-deficient 2 (SID2), which encodes ISOCHORISMATE SYNTHASE I, an enzyme required for biosynthesis of salicylic acid (SA). Analyses showed that compared with wild-type plants, esd4 contains higher levels of SID2 mRNA and about threefold more SA, whereas sed111 contains lower SA levels. Other sed mutants also contain lower SA levels but are not mutant for SID2, although most reduce SID2 mRNA levels. Therefore, higher SA levels contribute to the small size, early flowering and elevated SUMO conjugate levels of esd4. Our results support previous data indicating that SUMO homeostasis influences SA biosynthesis in wild-type plants, and also demonstrate that elevated levels of SA strongly increase the abundance of SUMO conjugates.

  5. Kaposi's sarcoma-associated herpesvirus K-Rta exhibits SUMO-targeting ubiquitin ligase (STUbL like activity and is essential for viral reactivation.

    Directory of Open Access Journals (Sweden)

    Yoshihiro Izumiya

    Full Text Available The small ubiquitin-like modifier (SUMO is a protein that regulates a wide variety of cellular processes by covalent attachment of SUMO moieties to a diverse array of target proteins. Sumoylation also plays an important role in the replication of many viruses. Previously, we showed that Kaposi's sarcoma-associated herpesvirus (KSHV encodes a SUMO-ligase, K-bZIP, which catalyzes sumoylation of host and viral proteins. We report here that this virus also encodes a gene that functions as a SUMO-targeting ubiquitin-ligase (STUbL which preferentially targets sumoylated proteins for degradation. K-Rta, the major transcriptional factor which turns on the entire lytic cycle, was recently found to have ubiquitin ligase activity toward a selected set of substrates. We show in this study that K-Rta contains multiple SIMs (SUMO interacting motif and binds SUMOs with higher affinity toward SUMO-multimers. Like RNF4, the prototypic cellular STUbL, K-Rta degrades SUMO-2/3 and SUMO-2/3 modified proteins, including promyelocytic leukemia (PML and K-bZIP. PML-NBs (nuclear bodies or ND-10 are storage warehouses for sumoylated proteins, which negatively regulate herpesvirus infection, as part of the intrinsic immune response. Herpesviruses have evolved different ways to degrade or disperse PML bodies, and KSHV utilizes K-Rta to inhibit PML-NBs formation. This process depends on K-Rta's ability to bind SUMO, as a K-Rta SIM mutant does not effectively degrade PML. Mutations in the K-Rta Ring finger-like domain or SIM significantly inhibited K-Rta transactivation activity in reporter assays and in the course of viral reactivation. Finally, KSHV with a mutation in the Ring finger-like domain or SIM of K-Rta replicates poorly in culture, indicating that reducing SUMO-conjugates in host cells is important for viral replication. To our knowledge, this is the first virus which encodes both a SUMO ligase and a SUMO-targeting ubiquitin ligase that together may generate

  6. A comprehensive compilation of SUMO proteomics

    DEFF Research Database (Denmark)

    Hendriks, Ivo A; Vertegaal, Alfred C O

    2016-01-01

    Small ubiquitin-like modifiers (SUMOs) are essential for the regulation of several cellular processes and are potential therapeutic targets owing to their involvement in diseases such as cancer and Alzheimer disease. In the past decade, we have witnessed a rapid expansion of proteomic approaches...... for identifying sumoylated proteins, with recent advances in detecting site-specific sumoylation. In this Analysis, we combined all human SUMO proteomics data currently available into one cohesive database. We provide proteomic evidence for sumoylation of 3,617 proteins at 7,327 sumoylation sites, and insight...

  7. MaxSUMO: A New Expert Approach for Evaluating Mobility Management Projects

    Directory of Open Access Journals (Sweden)

    Veronique Van Acker

    2013-06-01

    Full Text Available There is need for better understanding of how mobility management interventions work and how they affect the individuals’ modal choice decisions, as well as need for robust evaluation techniques allowing any behavioural changes to be observed. Changing individual’s behaviour is not a one-step process and any evaluation methodology should account for this. A new standardized expert evaluation resource MaxSUMO takes this step-wise process into account. MaxSUMO is based on a new theoretical behavioural change model MaxSEM which measures individuals’ stage positions (their susceptibility to change behaviour and stage movement (progression towards actual behavioural change. This paper illustrates the use of MaxSUMO by the evaluation of the mobility campaign “I keep moving, even without my car” undertaken by the City of Ghent.

  8. Cooperativity of the SUMO and Ubiquitin Pathways in Genome Stability

    Directory of Open Access Journals (Sweden)

    Minghua Nie

    2016-02-01

    Full Text Available Covalent attachment of ubiquitin (Ub or SUMO to DNA repair proteins plays critical roles in maintaining genome stability. These structurally related polypeptides can be viewed as distinct road signs, with each being read by specific protein interaction motifs. Therefore, via their interactions with selective readers in the proteome, ubiquitin and SUMO can elicit distinct cellular responses, such as directing DNA lesions into different repair pathways. On the other hand, through the action of the SUMO-targeted ubiquitin ligase (STUbL family proteins, ubiquitin and SUMO can cooperate in the form of a hybrid signal. These mixed SUMO-ubiquitin chains recruit “effector” proteins such as the AAA+ ATPase Cdc48/p97-Ufd1-Npl4 complex that contain both ubiquitin and SUMO interaction motifs. This review will summarize recent key findings on collaborative and distinct roles that ubiquitin and SUMO play in orchestrating DNA damage responses.

  9. Small ubiquitin-like modifier (SUMO) modification of zinc finger protein 131 potentiates its negative effect on estrogen signaling.

    Science.gov (United States)

    Oh, Yohan; Chung, Kwang Chul

    2012-05-18

    Like ubiquitin, small ubiquitin-like modifier (SUMO) covalently attaches to specific target proteins and modulates their functional properties, including subcellular localization, protein dimerization, DNA binding, and transactivation of transcription factors. Diverse transcriptional co-regulator complexes regulate the ability of estrogen receptors to respond to positive and negative acting hormones. Zinc finger protein 131 (ZNF131) is poorly characterized but may act as a repressor of estrogen receptor α (ERα)-mediated trans-activation. Here, we identify ZNF131 as a target for SUMO modification and as a substrate for the SUMO E3 ligase human polycomb protein 2 (hPc2). We report that the SUMO-interacting motif 1 (SIM1) and the C-box of hPc2 are critical regions required for ZNF131 SUMOylation and define the ZNF131 SUMOylation site as lysine 567. We further show that SUMO modification potentiates the negative effect of ZNF131 on estrogen signaling and consequently attenuates estrogen-induced cell growth in a breast cancer cell line. Our findings suggest that SUMOylation is a novel regulator of ZNF131 action in estrogen signaling and breast cancer cell proliferation.

  10. A sparse scattering model for nanoparticles on rough substrates

    DEFF Research Database (Denmark)

    Karamehmedovic, Mirza; Hansen, Poul-Erik; Wriedt, Thomas

    2013-01-01

    We present and validate an efficient forward scattering model for nanoparticles on rough contaminated substrates.......We present and validate an efficient forward scattering model for nanoparticles on rough contaminated substrates....

  11. Analysis list: SUMO2 [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available SUMO2 Blood,Prostate + hg19 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/target/...SUMO2.1.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/target/SUMO2.5.tsv http://dbarchive.biosciencedb...c.jp/kyushu-u/hg19/target/SUMO2.10.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/colo/SUMO2.Blood.tsv,http://dbarchive.bioscien...cedbc.jp/kyushu-u/hg19/colo/SUMO2.Prostate.tsv http://dbarchive.bioscience...dbc.jp/kyushu-u/hg19/colo/Blood.gml,http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/colo/Prostate.gml ...

  12. Modelling substrate specificity and enantioselectivity for lipases and esterases by substrate-imprinted docking

    Directory of Open Access Journals (Sweden)

    Tyagi Sadhna

    2009-06-01

    Full Text Available Abstract Background Previously, ways to adapt docking programs that were developed for modelling inhibitor-receptor interaction have been explored. Two main issues were discussed. First, when trying to model catalysis a reaction intermediate of the substrate is expected to provide more valid information than the ground state of the substrate. Second, the incorporation of protein flexibility is essential for reliable predictions. Results Here we present a predictive and robust method to model substrate specificity and enantioselectivity of lipases and esterases that uses reaction intermediates and incorporates protein flexibility. Substrate-imprinted docking starts with covalent docking of reaction intermediates, followed by geometry optimisation of the resulting enzyme-substrate complex. After a second round of docking the same substrate into the geometry-optimised structures, productive poses are identified by geometric filter criteria and ranked by their docking scores. Substrate-imprinted docking was applied in order to model (i enantioselectivity of Candida antarctica lipase B and a W104A mutant, (ii enantioselectivity and substrate specificity of Candida rugosa lipase and Burkholderia cepacia lipase, and (iii substrate specificity of an acetyl- and a butyrylcholine esterase toward the substrates acetyl- and butyrylcholine. Conclusion The experimentally observed differences in selectivity and specificity of the enzymes were reproduced with an accuracy of 81%. The method was robust toward small differences in initial structures (different crystallisation conditions or a co-crystallised ligand, although large displacements of catalytic residues often resulted in substrate poses that did not pass the geometric filter criteria.

  13. Construction, Expression and Purification of SUMO1-GST Fusion Protein

    Institute of Scientific and Technical Information of China (English)

    QIAO Xiao-fang; FANG Xue-dong; LIU Jun

    2011-01-01

    Sumoylation is an important protein modification discovered recently. SUMO(small ubiquitin-related modifier) pathway regulates the protein stability and transcriptional activity with a 12-kDa small molecular protein,SUMO, ligated to the target protein. The purification of SUMO proteins is a key step to reveal their function. The purpose of this study was to construct the recombinant SUMO1 gene cloned to a pGEX-4T-1 vector to express and purify the SUMO1-GST fusion protein in Escherichia coli. First, the full length DNA sequence of SUMO1 gene was amplified by PCR and was ligated to pMD18-T vector. Then the SUMO1 gene was subcloned to pGEX-4T-1 prokaryotic expression vector between BamHI and XhoI sites, and transformed in Escherichia coli DH5a cells. The right colonies were identified by restrictive enzyme digestion and sequencing. The correct rebombinant plasmid of pGEX-4T-1-SUMO1 was transformed in Escherichia coli BL21 cells and then induced by IPTG(isopropyl-β-D-lthiogalacto-pyranoside) to express the SUMO1-GST fusion protein. The highly purified SUMOl-GST(glutathione S-transferase) fusion protein was obtained by affinity chromatography. Finally, the properties of SUMO1-GST fusion protein were confirmed by Coomassie brilliant blue strain and Western blot analysis. The recombinant plasmid of pGEX-4T-1-SUMO 1 was successfully constructed, and SUMO1-GST fusion proteins were successfully expressed.

  14. SUMO modulation of protein aggregation and degradation

    Directory of Open Access Journals (Sweden)

    Marco Feligioni

    2015-09-01

    Full Text Available Small ubiquitin-like modifier (SUMO conjugation and binding to target proteins regulate a wide variety of cellular pathways. The functional aspects of SUMOylation include changes in protein-protein interactions, intracellular trafficking as well as protein aggregation and degradation. SUMO has also been linked to specialized cellular pathways such as neuronal development and synaptic transmission. In addition, SUMOylation is associated with neurological diseases associated with abnormal protein accumulations. SUMOylation of the amyloid and tau proteins involved in Alzheimer's disease and other tauopathies may contribute to changes in protein solubility and proteolytic processing. Similar events have been reported for α-synuclein aggregates found in Parkinson's disease, polyglutamine disorders such as Huntington's disease as well as protein aggregates found in amyotrophic lateral sclerosis (ALS. This review provides a detailed overview of the impact SUMOylation has on the etiology and pathology of these related neurological diseases.

  15. The Small Unmanned Meteorological Observer SUMO: A new tool for atmospheric boundary layer research

    Directory of Open Access Journals (Sweden)

    Joachim Reuder

    2009-05-01

    Full Text Available The Small Unmanned Meteorological Observer SUMO has been developed as a cost-efficient measurement system with the aim to close the existing observational gap of atmospheric measurement systems in between meteorological masts/towers and radiosondes. The system is highly flexible and has the capability for in-situ ABL measurements with unique spatial and temporal resolution. SUMO is based on a light-weighted styrofoam model airplane, equipped with an autopilot system for autonomous flight missions and in its recent version with meteorological sensors for temperature, humidity and pressure. With its wingspan of 80 cm, its length of 75 cm and a total lift-off weight of 580 g, SUMO is easy to transport and operate even in remote areas with limited infrastructure. During several field campaigns in 2007 and 2008 the system has been successfully tested and operated. Atmospheric profiles of temperature, humidity, wind speed and wind direction have been determined up to 3500 m above ground during the FLOHOF (FLOw over and around HOFsjökull field campaign in Central Iceland in July/August 2007. During a 3 week campaign on and around Spitsbergen in February/March 2008 the SUMO system also proved its functionality under polar conditions, reaching altitudes above 1500 m even at ground temperatures of -20° C and wind speeds up to 15 m s-1.

  16. The Small Unmanned Meteorological Observer SUMO. A new tool for atmospheric boundary layer research

    Energy Technology Data Exchange (ETDEWEB)

    Reuder, Joachim; Jonassen, Marius; Mayer, Stephanie [Bergen Univ. (Norway). Geophysical Inst.; Brisset, Pascal [Ecole Nationale de l' Aviation Civile (ENAC), Toulouse (France); Mueller, Martin [Martin Mueller Engineering, Hildesheim (Germany)

    2009-04-15

    The Small Unmanned Meteorological Observer SUMO has been developed as a cost-efficient measurement system with the aim to close the existing observational gap of atmospheric measurement systems in between meteorological masts/towers and radiosondes. The system is highly flexible and has the capability for in-situ ABL measurements with unique spatial and temporal resolution. SUMO is based on a light-weighted styrofoam model airplane, equipped with an autopilot system for autonomous flight missions and in its recent version with meteorological sensors for temperature, humidity and pressure. With its wingspan of 80 cm, its length of 75 cm and a total lift-off weight of 580 g, SUMO is easy to transport and operate even in remote areas with limited infrastructure. During several field campaigns in 2007 and 2008 the system has been successfully tested and operated. Atmospheric profiles of temperature, humidity, wind speed and wind direction have been determined up to 3500 m above ground during the FLOHOF (FLOw over and around HOFsjoekull) field campaign in Central Iceland in July/August 2007. During a 3 week campaign on and around Spitsbergen in February/March 2008 the SUMO system also proved its functionality under polar conditions, reaching altitudes above 1500 m even at ground temperatures of -20 C and wind speeds up to 15 m s{sup -1}. (orig.)

  17. The small ubiquitin-like modifier (SUMO) is required for gonadal and uterine-vulval morphogenesis in Caenorhabditis elegans.

    Science.gov (United States)

    Broday, Limor; Kolotuev, Irina; Didier, Christine; Bhoumik, Anindita; Gupta, Bhagwati P; Sternberg, Paul W; Podbilewicz, Benjamin; Ronai, Ze'ev

    2004-10-01

    The small ubiquitin-like modifier (SUMO) modification alters the subcellular distribution and function of its substrates. Here we show the major role of SUMO during the development of the Caenorhabditis elegans reproductive system. smo-1 deletion mutants develop into sterile adults with abnormal somatic gonad, germ line, and vulva. SMO-1::GFP reporter is highly expressed in the somatic reproductive system. smo-1 animals lack a vulval-uterine connection as a result of impaired ventral uterine pi-cell differentiation and anchor cell fusion. Mutations in the LIN-11 LIM domain transcription factor lead to a uterine phenotype that resembles the smo-1 phenotype. LIN-11 is sumoylated, and its sumoylation is required for its activity during uterine morphogenesis. Expression of a SUMO-modified LIN-11 in the smo-1 background partially rescued pi-cell differentiation and retained LIN-11 in nuclear bodies. Thus, our results identify the reproductive system as the major SUMO target during postembryonic development and highlight LIN-11 as a physiological substrate whose sumoylation is associated with the formation of a functional vulval-uterine connection.

  18. SUMO: Solar Ultraviolet Monitor and Ozone Nanosatellite

    Science.gov (United States)

    Damé, L.; Meftah, M.; Irbah, A.; Hauchecorne, A.; Keckhut, P.; Sarkissian, A.; Godin-Beekman, S.; Rogers, D. J.; Bove, P.; Lagage, P. O.; DeWitte, S.

    2014-12-01

    SUMO is an innovative proof-of-concept nanosatellite aiming to measure on the same platform the different components of the Earth radiation budget (ERB), the solar energy input and the energy reemitted at the top of the Earth atmosphere, with a particular focus on the far UV (FUV) part of the spectrum and on the ozone layer. The FUV is the only wavelength band with energy absorbed in the high atmosphere (stratosphere), in the ozone (Herzberg continuum, 200-220 nm) and oxygen bands, and its high variability is most probably at the origin of a climate influence (UV affects stratospheric dynamics and temperatures, altering interplanetary waves and weather patterns both poleward and downward to the lower stratosphere and tropopause). A simultaneous observation of incoming FUV and ozone production would bring an invaluable information on this process of solar-climate forcing. Space instruments have already measured the different components of the ERB but this is the first time that all instruments will operate on the same platform. This characteristic by itself guarantees original scientific results. SUMO is a 3.6 kg, 3W, 10x10x30 cm3 nanosatellite ("3U"), with a "1U" payload of definition has been completed (platform and payload AIT are possible in 24 months). SUMO is proposed for the nanosatellite program of Polytechnic School and CNES (following QB50) for a flight in 2018. Follow-up is 2 fold: on one part more complete measurements using SUMO miniaturized instruments on a larger satellite; on the other part, increase of the coverage in local time and latitude using a constellation of SUMO nanosatellites around the Earth to further geolocalize the Sun influence on our planet. Nanosatellites, with cost and risk limited, are also excellent platforms to evaluate technologies for future missions, e.g. nanotechnology ZnO protection barriers to limit contamination from solar panels in the UV and reduce reflection losses in the visible, or MgZnO solar blind detectors (R

  19. Puromycin induces SUMO and ubiquitin redistribution upon proteasome inhibition

    Energy Technology Data Exchange (ETDEWEB)

    Matsumoto, Hotaru [Course for Biological Sciences, Faculty of Science, Kumamoto University, Kumamoto (Japan); Saitoh, Hisato, E-mail: hisa@kumamoto-u.ac.jp [Course for Biological Sciences, Faculty of Science, Kumamoto University, Kumamoto (Japan); Department of Biological Sciences, Graduate School of Science and Technology, Kumamoto University, Kumamoto (Japan)

    2016-07-29

    We have previously reported the co-localization of O-propargyl-puromycin (OP-Puro) with SUMO-2/3 and ubiquitin at promyelocytic leukemia-nuclear bodies (PML-NBs) in the presence of the proteasome inhibitor MG132, implying a role for the ubiquitin family in sequestering OP-puromycylated immature polypeptides to the nucleus during impaired proteasome activity. Here, we found that as expected puromycin induced SUMO-1/2/3 accumulation with ubiquitin at multiple nuclear foci in HeLa cells when co-exposed to MG132. Co-administration of puromycin and MG132 also facilitated redistribution of PML and the SUMO-targeted ubiquitin ligase RNF4 concurrently with SUMO-2/3. As removal of the drugs from the medium led to disappearance of the SUMO-2/3-ubiquitin nuclear foci, our findings indicated that nuclear assembly/disassembly of SUMO-2/3 and ubiquitin was pharmacologically manipulable, supporting our previous observation on OP-Puro, which predicted the ubiquitin family function in sequestrating aberrant proteins to the nucleus. -- Highlights: •Puromycin exhibits the O-propargyl-puromycin effect. •Puromycin induces SUMO redistribution upon proteasome inhibition. •Ubiquitin and RNF4 accumulate at PML-nuclear bodies with SUMO-2/3. •The ubiquitin family may function in nuclear sequestration of immature proteins.

  20. Mutation in SUMO E3 ligase, SIZ1, disrupts the mature female gametophyte in Arabidopsis

    KAUST Repository

    Ling, Yu

    2012-01-09

    Female gametophyte is the multicellular haploid structure that can produce embryo and endosperm after fertilization, which has become an attractive model system for investigating molecular mechanisms in nuclei migration, cell specification, cell-to-cell communication and many other processes. Previous reports found that the small ubiquitin-like modifier (SUMO) E3 ligase, SIZ1, participated in many processes depending on particular target substrates and suppression of salicylic acid (SA) accumulation. Here, we report that SIZ1 mediates the reproductive process. SIZ1 showed enhanced expression in female organs, but was not detected in the anther or pollen. A defect in the siz1-2 maternal source resulted in reduced seed-set regardless of high SA concentration within the plant. Moreover, aniline blue staining and scanning electron microscopy revealed that funicular and micropylar pollen tube guidance was arrested in siz1-2 plants. Some of the embryo sacs of ovules in siz1-2 were also disrupted quickly after stage FG7. There was no significant affects of the siz1-2 mutation on expression of genes involved in female gametophyte development- or pollen tube guidance in ovaries. Together, our results suggest that SIZ1 sustains the stability and normal function of the mature female gametophyte which is necessary for pollen tube guidance. © 2012 Ling et al.

  1. Global SUMO proteome responses guide gene regulation, mRNA biogenesis, and plant stress responses.

    NARCIS (Netherlands)

    M.J. Mazur; H.A. van den Burg

    2012-01-01

    Small Ubiquitin-like MOdifier (SUMO) is a key regulator of abiotic stress, disease resistance, and development in plants. The identification of >350 plant SUMO targets has revealed many processes modulated by SUMO and potential consequences of SUMO on its targets. Importantly, highly related protein

  2. Effect of substrate competition in kinetic models of metabolic networks.

    Science.gov (United States)

    Schäuble, Sascha; Stavrum, Anne Kristin; Puntervoll, Pål; Schuster, Stefan; Heiland, Ines

    2013-09-02

    Substrate competition can be found in many types of biological processes, ranging from gene expression to signal transduction and metabolic pathways. Although several experimental and in silico studies have shown the impact of substrate competition on these processes, it is still often neglected, especially in modelling approaches. Using toy models that exemplify different metabolic pathway scenarios, we show that substrate competition can influence the dynamics and the steady state concentrations of a metabolic pathway. We have additionally derived rate laws for substrate competition in reversible reactions and summarise existing rate laws for substrate competition in irreversible reactions.

  3. Identification and developmental expression of Xenopus laevis SUMO proteases.

    Directory of Open Access Journals (Sweden)

    Yonggang Wang

    Full Text Available SUMO proteins are small ubiquitin-related modifiers. All SUMOs are synthesized as propeptides that are post-translationally cleaved prior to conjugation. After processing, SUMOs become covalently conjugated to cellular targets through a pathway that is similar to ubiquitination. Ubiquitin like protein proteases/Sentrin specific proteases (Ulp/SENPs mediate both processing and deconjugation of SUMOs. The action of Ulp/SENPs makes SUMOylation a highly dynamic post-translational modification. To investigate how Ulp/SENPs are regulated in a developmental context, we isolated and characterized all Ulp/SENPs in Xenopus laevis. Xenopus possess homologues of mammalian SENP3, 5, 6 and 7. All of these enzymes reacted with HA-tagged vinyl sulfone derivatives of SUMO-2 (HA-SU2-VS but not SUMO-1 (HA-SU1-VS, suggesting that they act primarily on SUMO-2 and -3. In contrast, Xenopus possess a single member of the SENP1/SENP2 subfamily of Ulp/SENPs, most closely related to mammalian SENP1. Xenopus SENP1 reacted with HA-SU1-VS and HA-SU2-VS, suggesting that it acts on all SUMO paralogues. We analyzed the mRNA and protein levels for each of the Ulp/SENPs through development; we found that they show distinct patterns of expression that may involve both transcriptional and post-transcriptional regulation. Finally, we have characterized the developmental function of the most abundant Ulp/SENP found within Xenopus eggs, SENP3. Depletion of SENP3 using morpholino antisense oligonucleotides (morpholinos caused accumulation of high molecular weight SUMO-2/3 conjugated species, defects in developing embryos and changes in the expression of some genes regulated by the transforming growth factor beta (TGF-beta pathway. These findings collectively indicate that SUMO proteases are both highly regulated and essential for normal development.

  4. Structure of the Siz/PIAS SUMO E3 Ligase Siz1 and Determinants Required for SUMO Modification of PCNA

    Energy Technology Data Exchange (ETDEWEB)

    Yunus, Ali A.; Lima, Christopher D.; (SKI)

    2010-01-12

    Siz1 is a founding member of the Siz/PIAS RING family of SUMO E3 ligases. The X-ray structure of an active Siz1 ligase revealed an elongated tripartite architecture comprised of an N-terminal PINIT domain, a central zinc-containing RING-like SP-RING domain, and a C-terminal domain we term the SP-CTD. Structure-based mutational analysis and biochemical studies show that the SP-RING and SP-CTD are required for activation of the E2SUMO thioester, while the PINIT domain is essential for redirecting SUMO conjugation to the proliferating cell nuclear antigen (PCNA) at lysine 164, a nonconsensus lysine residue that is not modified by the SUMO E2 in the absence of Siz1. Mutational analysis of Siz1 and PCNA revealed surfaces on both proteins that are required for efficient SUMO modification of PCNA in vitro and in vivo.

  5. SUMO-Enriched Proteome for Drosophila Innate Immune Response

    Science.gov (United States)

    Handu, Mithila; Kaduskar, Bhagyashree; Ravindranathan, Ramya; Soory, Amarendranath; Giri, Ritika; Elango, Vijay Barathi; Gowda, Harsha; Ratnaparkhi, Girish S.

    2015-01-01

    Small ubiquitin-like modifier (SUMO) modification modulates the expression of defense genes in Drosophila, activated by the Toll/nuclear factor-κB and immune-deficient/nuclear factor-κB signaling networks. We have, however, limited understanding of the SUMO-modulated regulation of the immune response and lack information on SUMO targets in the immune system. In this study, we measured the changes to the SUMO proteome in S2 cells in response to a lipopolysaccharide challenge and identified 1619 unique proteins in SUMO-enriched lysates. A confident set of 710 proteins represents the immune-induced SUMO proteome and analysis suggests that specific protein domains, cellular pathways, and protein complexes respond to immune stress. A small subset of the confident set was validated by in-bacto SUMOylation and shown to be bona-fide SUMO targets. These include components of immune signaling pathways such as Caspar, Jra, Kay, cdc42, p38b, 14-3-3ε, as well as cellular proteins with diverse functions, many being components of protein complexes, such as prosß4, Rps10b, SmD3, Tango7, and Aats-arg. Caspar, a human FAF1 ortholog that negatively regulates immune-deficient signaling, is SUMOylated at K551 and responds to treatment with lipopolysaccharide in cultured cells. Our study is one of the first to describe SUMO proteome for the Drosophila immune response. Our data and analysis provide a global framework for the understanding of SUMO modification in the host response to pathogens. PMID:26290570

  6. Sentrin/SUMO specific proteases as novel tissue-selective modulators of vitamin D receptor-mediated signaling.

    Directory of Open Access Journals (Sweden)

    Wai-Ping Lee

    Full Text Available Vitamin D receptor (VDR is a substrate for modification with small ubiquitin-like modifier (SUMO. To further assess the role of reversible SUMOylation within the vitamin D hormonal response, we evaluated the effects of sentrin/SUMO-specific proteases (SENPs that can function to remove small ubiquitin-like modifier (SUMO from target proteins upon the activities of VDR and related receptors. We report that SENP1 and SENP2 strikingly potentiate ligand-mediated transactivation of VDR and also its heterodimeric partner, retinoid X receptor (RXRα with depletion of cellular SENP1 significantly diminishing the hormonal responsiveness of the endogenous vitamin D target gene CYP24A1. We find that SENP-directed modulation of VDR activity is cell line-dependent, achieving potent modulatory effects in Caco-2 and HEK-293 cells, while in MCF-7 cells the vitamin D signal is unaffected by any tested SENP. In support of their function as novel modulators of the vitamin D hormonal pathway we demonstrate that both SENP1 and SENP2 can interact with VDR and reverse its modification with SUMO2. In a preliminary analysis we identify lysine 91, a residue known to be critical for formation and DNA binding of the VDR-RXR heterodimer, as a minor SUMO acceptor site within VDR. In combination, our results support a repressor function for SUMOylation of VDR and reveal SENPs as a novel class of VDR/RXR co-regulatory protein that significantly modulate the vitamin D response and which could also have important impact upon the functionality of both RXR-containing homo and heterodimers.

  7. An Analytical Model for Spectral Peak Frequency Prediction of Substrate Noise in CMOS Substrates

    DEFF Research Database (Denmark)

    Shen, Ming; Mikkelsen, Jan H.

    2013-01-01

    This paper proposes an analytical model describing the generation of switching current noise in CMOS substrates. The model eliminates the need for SPICE simulations in existing methods by conducting a transient analysis on a generic CMOS inverter and approximating the switching current waveform us...

  8. Modeling substrate-bacteria-grazer interactions coupled to substrate transport in groundwater

    Science.gov (United States)

    Bajracharya, Bijendra M.; Lu, Chuanhe; Cirpka, Olaf A.

    2014-05-01

    Models of microbial dynamics coupled to solute transport in aquifers typically require the introduction of a bacterial capacity term to prevent excessive microbial growth close to substrate-injection boundaries. The factors controlling this carrying capacity, however, are not fully understood. In this study, we propose that grazers or bacteriophages may control the density of bacterial biomass in continuously fed porous media. We conceptualize the flow-through porous medium as a series of retentostats, in which the dissolved substrate is advected with water flow whereas the biomasses of bacteria and grazers are considered essentially immobile. We first model a single retentostat with Monod kinetics of bacterial growth and a second-order grazing law, which shows that the system oscillates but approaches a stable steady state with nonzero concentrations of substrate, bacteria, and grazers. The steady state concentration of the bacteria biomass is independent of the substrate concentration in the inflow. When coupling several retentostats in a series to mimic a groundwater column, the steady state bacteria concentrations thus remain at a constant level over a significant travel distance. The one-dimensional reactive transport model also accounts for substrate dispersion and a random walk of grazers influenced by the bacteria concentration. These dispersive-diffusive terms affect the oscillations until steady state is reached, but hardly the steady state value itself. We conclude that grazing, or infection by bacteriophages, is a possible explanation of the maximum biomass concentration frequently needed in bioreactive transport models. Its value depends on parameters related to the grazers or bacteriophages and is independent of bacterial growth parameters or substrate concentration, provided that there is enough substrate to sustain bacteria and grazers.

  9. SUMO-2 Orchestrates Chromatin Modifiers in Response to DNA Damage

    DEFF Research Database (Denmark)

    Hendriks, Ivo A; Treffers, Louise W; Verlaan-de Vries, Matty

    2015-01-01

    with the DNA damaging agent methyl methanesulfonate (MMS). We have uncovered a dynamic set of 20 upregulated and 33 downregulated SUMO-2 conjugates, and 755 SUMO-2 sites, of which 362 were dynamic in response to MMS. In contrast to yeast, where a response is centered on homologous recombination, we identified......Small ubiquitin-like modifiers play critical roles in the DNA damage response (DDR). To increase our understanding of SUMOylation in the mammalian DDR, we employed a quantitative proteomics approach in order to identify dynamically regulated SUMO-2 conjugates and modification sites upon treatment...... dynamically SUMOylated interaction networks of chromatin modifiers, transcription factors, DNA repair factors, and nuclear body components. SUMOylated chromatin modifiers include JARID1B/KDM5B, JARID1C/KDM5C, p300, CBP, PARP1, SetDB1, and MBD1. Whereas SUMOylated JARID1B was ubiquitylated by the SUMO...

  10. Thermodynamic model of coherent island formation on vicinal substrate

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Xu, E-mail: zhangxubetter@gmail.com; Sun, Xiao-Hong [Henan Key Laboratory of Laser and Opto-electric Information Technology, School of Information Engineering, Zhengzhou University, Zhengzhou 450052 (China); Yu, Yanguang [School of Electrical, Computer and Telecommunications Engineering, University of Wollongong, Northfields Ave, Wollongong, New South Wales 2522 (Australia); Ren, Xiaomin [State Key Laboratory of Information Photonics and Optical Communications, Beijing University of Posts and Telecommunications, Beijing 100876 (China)

    2014-04-28

    A thermodynamic model has been proposed to address the formation of coherent island on the vicinal substrate. The morphological transition from square based island to elongated based one with various substrate misorientations is described. The initial stage of nucleation and growth process of islands in Stranski–Krastanow system is studied by taking into account the elastic deformations and the change of energy in the case of two-dimensional growth mode. The theoretical analysis shows the minimum nucleation barrier of island is on the decrease with increment of substrate misorientation, which means the nucleation of island on vicinal substrate is more favorable than that on flat substrate. By using the fitting data of experimental results done by Persichetti et al., [Phys. Rev. Lett. 104, 036104 (2010) and Phys. Rev. B 82, 121309(R) (2010)], we provide a meaningful explanation of the experimental observations.

  11. Mass Spectral Enhanced Detection of Ubls Using SWATH Acquisition: MEDUSA—Simultaneous Quantification of SUMO and Ubiquitin-Derived Isopeptides

    Science.gov (United States)

    Griffiths, John R.; Chicooree, Navin; Connolly, Yvonne; Neffling, Milla; Lane, Catherine S.; Knapman, Thomas; Smith, Duncan L.

    2014-05-01

    Protein modification by ubiquitination and SUMOylation occur throughout the cell and are responsible for numerous cellular functions such as apoptosis, DNA replication and repair, and gene transcription. Current methods for the identification of such modifications using mass spectrometry predominantly rely upon tryptic isopeptide tag generation followed by database searching with in vitro genetic mutation of SUMO routinely required. We have recently described a novel approach to ubiquitin and SUMO modification detection based upon the diagnostic a' and b' ions released from the isopeptide tags upon collision-induced dissociation of reductively methylated Ubl isopeptides (RUbI) using formaldehyde. Here, we significantly extend those studies by combining data-independent acquisition (DIA) with alternative labeling reagents to improve diagnostic ion coverage and enable relative quantification of modified peptides from both MS and MS/MS signals. Model synthetic ubiquitin and SUMO-derived isopeptides were labeled with mTRAQ reagents (Δ0, Δ4, and Δ8) and subjected to LC-MS/MS with SWATH acquisition. Novel diagnostic ions were generated upon CID, which facilitated the selective detection of these modified peptides. Simultaneous MS-based and MS/MS-based relative quantification was demonstrated for both Ub and SUMO-derived isopeptides across three channels in a background of mTRAQ-labeled Escherichia coli digest.

  12. The Ubiquitin-Like SUMO System and Heart Function: From Development to Disease.

    Science.gov (United States)

    Mendler, Luca; Braun, Thomas; Müller, Stefan

    2016-01-01

    SUMOylation is a ubiquitin-related transient posttranslational modification pathway catalyzing the conjugation of small ubiquitin-like modifier (SUMO) proteins (SUMO1, SUMO2, and SUMO3) to lysine residues of proteins. SUMOylation targets a wide variety of cellular regulators and thereby affects a multitude of different cellular processes. SUMO/sentrin-specific proteases are able to remove SUMOs from targets, contributing to a tight control of SUMOylated proteins. Genetic and cell biological experiments indicate a critical role of balanced SUMOylation/deSUMOylation for proper cardiac development, metabolism, and stress adaptation. Here, we review the current knowledge about SUMOylation/deSUMOylation in the heart and provide an integrated picture of cardiac functions of the SUMO system under physiologic or pathologic conditions. We also describe potential therapeutic approaches targeting the SUMO machinery to combat heart disease.

  13. Functional characterization of the SIZ/PIAS-type SUMO E3 ligases, OsSIZ1 and OsSIZ2 in rice

    KAUST Repository

    Park, Hyeongcheol

    2010-06-18

    Sumoylation is a post-translational regulatory process in diverse cellular processes in eukaryotes, involving conjugation/deconjugation of small ubiquitin-like modifier (SUMO) proteins to other proteins thus modifying their function. The PIAS [protein inhibitor of activated signal transducers and activators of transcription (STAT)] and SAP (scaffold attachment factor A/B/acinus/PIAS)/MIZ (SIZ) proteins exhibit SUMO E3-ligase activity that facilitates the conjugation of SUMO proteins to target substrates. Here, we report the isolation and molecular characterization of Oryza sativa SIZ1 (OsSIZ1) and SIZ2 (OsSIZ2), rice homologs of Arabidopsis SIZ1. The rice SIZ proteins are localized to the nucleus and showed sumoylation activities in a tobacco system. Our analysis showed increased amounts of SUMO conjugates associated with environmental stresses such as high and low temperature, NaCl and abscisic acid (ABA) in rice plants. The expression of OsSIZ1 and OsSIZ2 in siz1-2 Arabidopsis plants partially complemented the morphological mutant phenotype and enhanced levels of SUMO conjugates under heat shock conditions. In addition, ABA-hypersensitivity of siz1-2 seed germination was partially suppressed by OsSIZ1 and OsSIZ2. The results suggest that rice SIZ1 and SIZ2 are able to functionally complement Arabidopsis SIZ1 in the SUMO conjugation pathway. Their effects on the Arabidopsis mutant suggest a function for these genes related to stress responses and stress adaptation. © 2010 Blackwell Publishing Ltd.

  14. Global SUMO proteome responses guide gene regulation, mRNA biogenesis, and plant stress responses

    Directory of Open Access Journals (Sweden)

    Magdalena eMazur

    2012-09-01

    Full Text Available Small-ubiquitin-like MOdifier (SUMO is a key regulator of abiotic stress, disease resistance and development in plants. The identification of >350 plant SUMO targets has revealed many processes modulated by SUMO and potential consequences of SUMO on its targets. Importantly, highly related proteins are SUMO-modified in plants, yeast, and metazoans. Overlapping SUMO targets include heat-shock proteins, transcription regulators, histones, histone-modifying enzymes, proteins involved in DNA damage repair, but also proteins involved in mRNA biogenesis and nucleo-cytoplasmic transport. Proteomics studies indicate key roles for SUMO in gene repression by controlling histone (deacetylation activity at genomic loci. The responsible heavily sumoylated transcriptional repressor complexes are recruited by EAR (Ethylene-responsive element binding factor [ERF]-associated Amphiphilic Repression-motif containing transcription factors in plants. These transcription factors are not necessarily themselves a SUMO target. Conversely, SUMO acetylation prevents binding of downstream partners by preventing binding of SIMs (SUMO-interaction peptide motifs presents in these partners, while SUMO acetylation has emerged as mechanism to recruit specifically bromodomains; bromodomain are generally linked with gene activation. These findings strengthen the idea of a bidirectional sumo-/acetylation switch in gene regulation. Quantitative proteomics has highlighted that global sumoylation provides a dynamic response to protein damage involving SUMO chain-mediated protein degradation, but also SUMO E3 ligase-dependent transcription of HSP (Heat-shock protein genes. With these insights in SUMO function and novel technical advancements, we can now study SUMO dynamics in responses to (abiotic stress in plants.

  15. ATPase-Dependent Control of the Mms21 SUMO Ligase during DNA Repair

    NARCIS (Netherlands)

    M. Bermúdez-López (Marcelino); I. Pociño-Merino (Irene); H. Sanchez (Humberto); A. Bueno (Andrés); C. Guasch (Clàudia); S. Almedawar (Seba); S. Bru-Virgili (Sergi); E. Garí (Eloi); C. Wyman (Claire); D. Reverter (David); N. Colomina (Neus); J. Torres-Rosell (Jordi)

    2015-01-01

    textabstractModification of proteins by SUMO is essential for the maintenance of genome integrity. During DNA replication, the Mms21-branch of the SUMO pathway counteracts recombination intermediates at damaged replication forks, thus facilitating sister chromatid disjunction. The Mms21 SUMO ligase

  16. Modeling crawling cell movement on soft engineered substrates.

    Science.gov (United States)

    Löber, Jakob; Ziebert, Falko; Aranson, Igor S

    2014-03-07

    Self-propelled motion, emerging spontaneously or in response to external cues, is a hallmark of living organisms. Systems of self-propelled synthetic particles are also relevant for multiple applications, from targeted drug delivery to the design of self-healing materials. Self-propulsion relies on the force transfer to the surrounding. While self-propelled swimming in the bulk of liquids is fairly well characterized, many open questions remain in our understanding of self-propelled motion along substrates, such as in the case of crawling cells or related biomimetic objects. How is the force transfer organized and how does it interplay with the deformability of the moving object and the substrate? How do the spatially dependent traction distribution and adhesion dynamics give rise to complex cell behavior? How can we engineer a specific cell response on synthetic compliant substrates? Here we generalize our recently developed model for a crawling cell by incorporating locally resolved traction forces and substrate deformations. The model captures the generic structure of the traction force distribution and faithfully reproduces experimental observations, like the response of a cell on a gradient in substrate elasticity (durotaxis). It also exhibits complex modes of cell movement such as "bipedal" motion. Our work may guide experiments on cell traction force microscopy and substrate-based cell sorting and can be helpful for the design of biomimetic "crawlers" and active and reconfigurable self-healing materials.

  17. At the crossroads of SUMO and NF-κB

    Directory of Open Access Journals (Sweden)

    Schmidt Christian

    2003-11-01

    Full Text Available Abstract Background Recognition of pathogens by immune receptors leads to activation of macrophages, dendritic cells, and lymphocytes. Signals are communicated to enhance expression of target molecules such as cytokines and adhesion molecules, depending on activation of various inducible transcription factors, among which the family NF-κB transcription factors plays an evolutionarily conserved and critical role. Classical activation of NF-κB involves phosphorylation, polyubiquitination and subsequent degradation of the inhibitor molecules of NF-κB, referred to as IκB. Modification of IκBα, one of the mammalian IκB isoforms, with the small ubiquitin-like modifier (SUMO results its protection from degradation. Presentation of the hypothesis SUMO-IκBα localizes in the nucleus. The nuclear SUMO-IκBα pool may be dynamic. SUMO-IκBα functions as synergy control factor. Testing the hypothesis Immunoprecipitation from cellular fractions, 35S methionine pulse-chase, and FRET assays should reveal the localization of SUMO-IκBα and the dynamics of the pool. Expression of SUMOylation defective IκBα in an IκBα -/- background should yield insights into the function of SUMO-IκBα. Implication of the hypothesis IκBα contains the required SUMOylation motif but IκBβ does not. The suggested study would provide evidence whether or not IκBα and IκBβ can substitute each other. In addition, the suggested assays would reveal a possible redundancy in controlling transcriptional activity of NF-κB.

  18. Role of SUMO-specific protease 2 in reprogramming cellular glucose metabolism.

    Directory of Open Access Journals (Sweden)

    Shuang Tang

    Full Text Available Most cancer cells exhibit a shift in glucose metabolic strategy, displaying increased glycolysis even with adequate oxygen supply. SUMO-specific proteases (SENPs de-SUMOylate substrates including HIF1α and p53,two key regulators in cancer glucose metabolism, to regulate their activity, stability and subcellular localization. However, the role of SENPs in tumor glucose metabolism remains unclear. Here we report that SUMO-specific protease 2 (SENP2 negatively regulates aerobic glycolysis in MCF7 and MEF cells. Over-expression of SENP2 reduces the glucose uptake and lactate production, increasing the cellular ATP levels in MCF7 cells, while SENP2 knockout MEF cells show increased glucose uptake and lactate production along with the decreased ATP levels. Consistently, the MCF7 cells over-expressing SENP2 exhibit decreased expression levels of key glycolytic enzymes and an increased rate of glucose oxidation compared with control MCF7 cells, indicating inhibited glycolysis but enhanced oxidative mitochondrial respiration. Moreover, SENP2 over-expressing MCF7 cells demonstrated a reduced amount of phosphorylated AKT, whereas SENP2 knockout MEFs exhibit increased levels of phosphorylated AKT. Furthermore, inhibiting AKT phosphorylation by LY294002 rescued the phenotype induced by SENP2 deficiency in MEFs. In conclusion, SENP2 represses glycolysis and shifts glucose metabolic strategy, in part through inhibition of AKT phosphorylation. Our study reveals a novel function of SENP2 in regulating glucose metabolism.

  19. Development of a space universal modular architecture (SUMO)

    Science.gov (United States)

    Collins, Bernie F.

    This concept paper proposes that the space community should develop and implement a universal standard for spacecraft modularity - to improve interoperability of spacecraft components. Pursuing a global industry consensus standard for open and modular spacecraft architecture will encourage trade, remove standards-related market barriers, and in the long run increase both value provided to customers and profitability of the space industrial sector. This concept paper sets out: (1) the goals for a SUMO standard and how it will benefit the space community; (2) background on spacecraft modularity and existing related standards; (3) the proposed technical scope of the current standardization effort; and (4) an approach for creating a SUMO standard.

  20. Kant e o sumo bem comunitário

    Directory of Open Access Journals (Sweden)

    Letícia Machado Spinelli

    2012-08-01

    Full Text Available http://dx.doi.org/10.5007/1677-2954.2012v11n1p37 O que aqui nos propomos fazer é explicitar o conceito de sumo bem enquanto bem comunitário, em que se evidencia um desdobramento da formulação do conceito de sumo bem sob a perspectiva de um bem coletivo. Esse conceito foi apresentado por Kant na terceira parte de A religião nos limites da simples razão, contexto no qual se dedicou a tratar da noção de um progresso moral nos termos de uma comunidade ética.

  1. A model of cytoskeletal reorientation in response to substrate stretching

    Directory of Open Access Journals (Sweden)

    Lazopoulos K.A.

    2008-01-01

    Full Text Available Living adherent cells change their orientation in response to substrate stretching such that their cytoskeletal components reorganize in a new direction. To study this phenomenon, we model the cytoskeleton as a planar system of elastic cables and struts both pinned at their endpoints to a flat flexible substrate. Tensed (pre-strained cables represent acting stress fibers, whereas compression-bearing struts represent microtubules. We assume that in response to uniaxial substrate stretching the model reorients and deforms into a new configuration that minimizes its total potential energy. Using the Maxwell's global stability criterion, we find global minima configurations during static extension and compression of the substrate. Based on these results, we predict reorientation during cyclic stretching of the substrate. We find that in response to cyclic stretching cells either reorient transversely to the direction of stretching, or exhibit multiple configurations symmetrically distributed relative to the direction of stretching. These predictions are consistent with experimental data on living cells from the literature.

  2. Ubiquitin E3 ligase FIEL1 regulates fibrotic lung injury through SUMO-E3 ligase PIAS4.

    Science.gov (United States)

    Lear, Travis; McKelvey, Alison C; Rajbhandari, Shristi; Dunn, Sarah R; Coon, Tiffany A; Connelly, William; Zhao, Joe Y; Kass, Daniel J; Zhang, Yingze; Liu, Yuan; Chen, Bill B

    2016-05-30

    The E3 small ubiquitin-like modifier (SUMO) protein ligase protein inhibitor of activated STAT 4 (PIAS4) is a pivotal protein in regulating the TGFβ pathway. In this study, we discovered a new protein isoform encoded by KIAA0317, termed fibrosis-inducing E3 ligase 1 (FIEL1), which potently stimulates the TGFβ signaling pathway through the site-specific ubiquitination of PIAS4. FIEL1 targets PIAS4 using a double locking mechanism that is facilitated by the kinases PKCζ and GSK3β. Specifically, PKCζ phosphorylation of PIAS4 and GSK3β phosphorylation of FIEL1 are both essential for the degradation of PIAS4. FIEL1 protein is highly expressed in lung tissues from patients with idiopathic pulmonary fibrosis (IPF), whereas PIAS4 protein levels are significantly reduced. FIEL1 overexpression significantly increases fibrosis in a bleomycin murine model, whereas FIEL1 knockdown attenuates fibrotic conditions. Further, we developed a first-in-class small molecule inhibitor toward FIEL1 that is highly effective in ameliorating fibrosis in mice. This study provides a basis for IPF therapeutic intervention by modulating PIAS4 protein abundance.

  3. Construction of a mouse Aos1-Uba2 chimeric SUMO-E1 enzyme, mAU, and its expression in baculovirus-insect cells

    Science.gov (United States)

    Nakayama, Tomofumi; Yuasa, Eri; Kanemaru, Ayumi; Saito, Masayuki; Saitoh, Hisato

    2014-01-01

    Small ubiquitin-related modifier (SUMO) is a highly conserved protein that is covalently attached to target proteins. This posttranslational modification, designated SUMOylation, is a major protein-conjugation-driven strategy designed to regulate structure and function of cellular proteins. SUMOylation consists of an enzymatic cascade involving the E1-activating enzyme and the E2-conjugating enzyme. The SUMO-E1 enzyme consists of two subunits, a heterodimer of activation of Smt3p 1 (Aos1) and ubiquitin activating enzyme 2 (Uba2), which resembles the N- and C-terminal halves of ubiquitin E1 (Uba1). Herein, we describe the rational design of a single polypeptide version of SUMO-E1, a chimera of mouse Aos1 and Uba2 subunits, termed mAU, in which the functional domains appear to be arranged in a fashion similar to Uba1. We also describe the construction of a mAU plasmid for expression in a baculovirus-insect cell system and present an in situ SUMOylation assay using the recombinant mAU. Our results showed that mAU has SUMO-E1 activity, thereby indicating that mAU can be expressed in baculovirus-insect cells and represents a suitable source of SUMO-E1. PMID:24637489

  4. Modelling of Substrate Noise and Mitigation Schemes for UWB Systems

    DEFF Research Database (Denmark)

    Shen, Ming; Mikkelsen, Jan H.; Larsen, Torben

    2012-01-01

    The last chapter of this first part of the book, chapter seven, is devoted to Modeling of Substrate Noise and Mitigation Schemes for Ultrawideband (UWB) systems, and is written by Ming Shen, Jan H. Mikkelsen, and Torben Larsen from Aalborg University, Denmark. In highly integrated mixed-mode desi...

  5. The Sumo World through the gambling cases in 2010 The Sumo World as Seen Through the Gambling Cases of 2010

    Directory of Open Access Journals (Sweden)

    Keiko Morita

    2012-11-01

    Full Text Available In summer, 2010, gambling cases of sumo wrestlers and bosses and the connection with the gangsters were reported day after day through mass media. This article chases the summary of a chain of these cases and the correspondence of the Sumo Association for it. The article looks at the present conditions of a unique competition called sumo from these a series of cases. At first, it examines the summary of the illegal gambling case of Ozeki Kotomitsuki, the top of Japanese sumo wrestlers, and the correspondence of Sumo Association and other social organs. It takes up a problem about holding the tournament at Nagoya this summer next. And it relates to dark customs of sumo world that a series of cases are able to happen and tries to explain such customs through the tradition of sumo. While writing this article, the news to ask the self-purification capacity of the Association and to suggest relations with the sumo world and the gangsters are reported every day. The darkness of the sumo world seems to be wide and deep.This article is in Japanese. In summer 2010, a scandal in gambling on baseball matches implicating sumo wrestlers and their bosses was reported day after day by Japanese mass media. This scandal revealed the connections between the sumo world and criminal organizations and lead to unprecedented actions, such as the dismissal of the active Ozeki Kotomitsuki, one of the top Japanese sumo wrestlers, and the suspension of live television broadcasting of the sumo tournament. The reason why the Japanese public instigated mass public debates may relate to the notion that sumo has an intimate association with the gods, reinforced with its special position in Japanese tradition and culture. This article first presents a brief history of sumo and explains its particular characteristics. Then it provides a summary of the chain of events in the baseball gambling scandal and the responses from the Japan Sumo Association. In conjunction with

  6. Modeling crawling cell movement on soft engineered substrates

    Science.gov (United States)

    Aronson, Igor

    2014-03-01

    Self-propelled motion, emerging spontaneously or in response to external cues, is a hallmark of living organisms. Systems of self-propelled synthetic particles are also relevant for multiple applications, from targeted drug delivery to the design of self-healing materials. Self-propulsion relies on the force transfer to the surrounding. While self-propelled swimming in the bulk of liquids is fairly well characterized, many open questions remain in our understanding of self-propelled motion along substrates, such as in the case of crawling cells or related biomimetic objects. How is the force transfer organized and how does it interplay with the deformability of the moving object and the substrate? How do the spatially dependent traction distribution and adhesion dynamics give rise to complex cell behavior? How can we engineer a specific cell response on synthetic compliant substrates? Here we present a phase-field model for a crawling cell by incorporating locally resolved traction forces and substrate deformations. The model captures the generic structure of the traction force distribution and faithfully reproduces experimental observations, like the response of a cell on a gradient in substrate elasticity (durotaxis). It also exhibits complex modes of cell movement such as ``bipedal'' motion. Our work may guide experiments on cell traction force microscopy and substrate-based cell sorting and can be helpful for the design of biomimetic ``crawlers'' and active and reconfigurable self-healing materials. This work was supported by the U.S. Department of Energy, Office of Basic Energy Sciences, Division of Materials Science and Engineering, under Contract DE-AC02-06CH11357.

  7. Modeling the Substrate Skin Effects in Mutual RL Characteristics.,

    Directory of Open Access Journals (Sweden)

    D. de Roest

    2003-12-01

    Full Text Available The goal of this work was to model the influence of the substrateskin effects on the distributed mutual impedance per unit lengthparameters of multiple coupled on-chip interconnects. The proposedanalytic model is based on the frequency-dependent distribution of thecurrent in the silicon substrate and the closed form integrationapproach. It is shown that the calculated frequency-dependentdistributed mutual inductance and the associated mutual resistance arein good agreement with the results obtained from CAD-oriented circuitmodeling technique.

  8. Principles of ubiquitin and SUMO modifications in DNA repair

    NARCIS (Netherlands)

    Bergink, Steven; Jentsch, Stefan

    2009-01-01

    With the discovery in the late 1980s that the DNA-repair gene RAD6 encodes a ubiquitin-conjugating enzyme, it became clear that protein modification by ubiquitin conjugation has a much broader significance than had previously been assumed. Now, two decades later, ubiquitin and its cousin SUMO are im

  9. Binding properties of SUMO-interacting motifs (SIMs) in yeast.

    Science.gov (United States)

    Jardin, Christophe; Horn, Anselm H C; Sticht, Heinrich

    2015-03-01

    Small ubiquitin-like modifier (SUMO) conjugation and interaction play an essential role in many cellular processes. A large number of yeast proteins is known to interact non-covalently with SUMO via short SUMO-interacting motifs (SIMs), but the structural details of this interaction are yet poorly characterized. In the present work, sequence analysis of a large dataset of 148 yeast SIMs revealed the existence of a hydrophobic core binding motif and a preference for acidic residues either within or adjacent to the core motif. Thus the sequence properties of yeast SIMs are highly similar to those described for human. Molecular dynamics simulations were performed to investigate the binding preferences for four representative SIM peptides differing in the number and distribution of acidic residues. Furthermore, the relative stability of two previously observed alternative binding orientations (parallel, antiparallel) was assessed. For all SIMs investigated, the antiparallel binding mode remained stable in the simulations and the SIMs were tightly bound via their hydrophobic core residues supplemented by polar interactions of the acidic residues. In contrary, the stability of the parallel binding mode is more dependent on the sequence features of the SIM motif like the number and position of acidic residues or the presence of additional adjacent interaction motifs. This information should be helpful to enhance the prediction of SIMs and their binding properties in different organisms to facilitate the reconstruction of the SUMO interactome.

  10. Modeling universal dynamics of cell spreading on elastic substrates.

    Science.gov (United States)

    Fan, Houfu; Li, Shaofan

    2015-11-01

    A three-dimensional (3D) multiscale moving contact line model is combined with a soft matter cell model to study the universal dynamics of cell spreading over elastic substrates. We have studied both the early stage and the late stage cell spreading by taking into account the actin tension effect. In this work, the cell is modeled as an active nematic droplet, and the substrate is modeled as a St. Venant Kirchhoff elastic medium. A complete 3D simulation of cell spreading has been carried out. The simulation results show that the spreading area versus spreading time at different stages obeys specific power laws, which is in good agreement with experimental data and theoretical prediction reported in the literature. Moreover, the simulation results show that the substrate elasticity may affect force dipole distribution inside the cell. The advantage of this approach is that it combines the hydrodynamics of actin retrograde flow with moving contact line model so that it can naturally include actin tension effect resulting from actin polymerization and actomyosin contraction, and thus it might be capable of simulating complex cellular scale phenomenon, such as cell spreading or even crawling.

  11. High Glucose Induces Sumoylation of Smad4 via SUMO2/3 in Mesangial Cells

    Directory of Open Access Journals (Sweden)

    Xueqin Zhou

    2014-01-01

    Full Text Available Recent studies have shown that sumoylation is a posttranslational modification involved in regulation of the transforming growth factor-β (TGF-β signaling pathway, which plays a critical role in renal fibrosis in diabetic nephropathy (DN. However, the role of sumoylation in the regulation of TGF-β signaling in DN is still unclear. In the present study, we investigated the expression of SUMO (SUMO1 and SUMO2/3 and Smad4 and the interaction between SUMO and Smad4 in cultured rat mesangial cells induced by high glucose. We found that SUMO1 and SUMO2/3 expression was significantly increased in the high glucose groups compared to the normal group P<0.05. Smad4 and fibronectin (FN levels were also increased in the high glucose groups in a dose-dependent manner. Coimmunoprecipitation and confocal laser scanning revealed that Smad4 interacted and colocalized with SUMO2/3, but not with SUMO1 in mesangial cells. Sumoylation (SUMO2/3 of Smad4 under high glucose condition was strongly enhanced compared to normal control P<0.05. These results suggest that high glucose may activate TGF-β/Smad signaling through sumoylation of Samd4 by SUMO2/3 in mesangial cells.

  12. Priming and substrate quality interactions in soil organic matter models

    Directory of Open Access Journals (Sweden)

    T. Wutzler

    2012-12-01

    Full Text Available Interactions between different qualities of soil organic matter (SOM affecting their turnover are rarely represented in models. In this study we propose three mathematical strategies at different levels of abstraction for representing those interactions. Implementing these strategies into the Introductory Carbon Balance Model (ICBM and applying them to several scenarios of litter input show that the different levels of abstraction are applicable on different time scales. We present a simple one-parameter equation of substrate limitation applicable at decadal time scale that is straightforward to implement into other models of SOM dynamics. We show how substrate quality interactions can explain priming effects, acceleration of turnover times in FACE experiments, and the slowdown of decomposition in long-term bare fallow experiments as an effect of energy limitation of microbial biomass. The mechanisms of those interactions need to be further scrutinized empirically for a more complete understanding. Overall, substrate quality interactions offer a valuable way of understanding and quantitatively modelling SOM dynamics.

  13. QSAR Models for P-450 (2D6) Substrate Activity

    DEFF Research Database (Denmark)

    Ringsted, Tine; Nikolov, Nikolai Georgiev; Jensen, Gunde Egeskov;

    2009-01-01

    activity relationship (QSAR) modelling systems. They cross validated (leave-groups-out) with concordances of 71%, 81% and 82%, respectively. Discrete organic European Inventory of Existing Commercial Chemical Substances (EINECS) chemicals were screened to predict an approximate percentage of CYP 2D6...... substrates. These chemicals are potentially present in the environment. The biological importance of the CYP 2D6 and the use of the software mentioned above were discussed....

  14. A Critical SUMO1 Modification of LKB1 Regulates AMPK Activity during Energy Stress

    KAUST Repository

    Ritho, Joan

    2015-07-23

    SUMOylation has been implicated in cellular stress adaptation, but its role in regulating liver kinase B1 (LKB1), a major upstream kinase of the energy sensor AMP-activated protein kinase (AMPK), is unknown. Here, we show that energy stress triggers an increase in SUMO1 modification of LKB1, despite a global reduction in both SUMO1 and SUMO2/3 conjugates. During metabolic stress, SUMO1 modification of LKB1 lysine 178 is essential in promoting its interaction with AMPK via a SUMO-interacting motif (SIM) essential for AMPK activation. The LKB1 K178R SUMO mutant had defective AMPK signaling and mitochondrial function, inducing death in energy-deprived cells. These results provide additional insight into how LKB1-AMPK signaling is regulated during energy stress, and they highlight the critical role of SUMOylation in maintaining the cell’s energy equilibrium.

  15. Regulation of SUMO2 Target Proteins by the Proteasome in Human Cells Exposed to Replication Stress

    DEFF Research Database (Denmark)

    Bursomanno, Sara; McGouran, Joanna F; Kessler, Benedikt M

    2015-01-01

    In human cells, SUMO2 is predominantly conjugated to target proteins in response to cellular stress. Previous studies suggested that proteins conjugated to SUMO2, but not to SUMO1, could be regulated by the ubiquitin-mediated proteasome system. Hence, we set out to understand the role of the prot......In human cells, SUMO2 is predominantly conjugated to target proteins in response to cellular stress. Previous studies suggested that proteins conjugated to SUMO2, but not to SUMO1, could be regulated by the ubiquitin-mediated proteasome system. Hence, we set out to understand the role...... of genome instability, which is suggested to drive tumorigenesis and possibly aging, our data will facilitate future functional studies in the fields of DNA metabolism and cancer biology....

  16. Lava-substrate heat transfer: Laboratory experiments and thermodynamic modeling

    Science.gov (United States)

    Rumpf, M.; Fagents, S. A.; Hamilton, C. W.; Wright, R.; Crawford, I.

    2012-12-01

    We have performed laboratory experiments and numerical modeling to investigate the heat transfer from a lava flow into various substrate materials, focusing on the effects of the differing thermophysical properties of substrate materials. Initial motivation for this project developed from the desire to understand the loss of solar wind volatiles embedded in lunar regolith deposits that were subsequently covered by a lava flow. The Moon lacks a significant atmosphere and magnetosphere, leaving the surface regolith exposed to bombardment by solar flare and solar wind particles, and by the cosmogenic products of galactic cosmic rays. Preservation of particle-rich regolith deposits may have occurred by the emplacement of an active lava flow on top of the regolith layer, provided the embedded particles survive heating by the lava. During future expeditions to the lunar surface, ancient regolith deposits could be sampled through surface drilling to extract the extra-lunar particles, revealing a history of the solar activity and galactic events not available on the Earth. This project also has important implications for terrestrial lava flows, particularly in the prediction of lava flow hazards. Lava erupted on Earth may be emplaced on various substrates, including solid lava rock, volcanic tephra, sands, soils, etc. The composition, grain size, consolidation, moisture content, etc. of these materials will vary greatly and have different effects on the cooling of the flow. Accounting for specific properties of the substrate could be an important improvement in lava flow models We have performed laboratory experiments in collaboration with the Department of Art and Art History at the University of Hawaii at Manoa in which ~5-6 kg of basalt, collected at Kilauea Volcano, Hawaii, is melted to ~1200 °C. The lava is poured into a device constructed of calcium silicate sheeting that has been filled with a solid or particulate substrate material and embedded with thermocouples

  17. SUMO and SUMO-Conjugating Enzyme E2 UBC9 Are Involved in White Spot Syndrome Virus Infection in Fenneropenaeus chinensis.

    Directory of Open Access Journals (Sweden)

    Xiaoqian Tang

    Full Text Available In previous work, small ubiquitin-like modifier (SUMO in hemocytes of Chinese shrimp Fenneropenaeus chinensis was found to be up-regulated post-white spot syndrome virus (WSSV infection using proteomic approach. However, the role of SUMO in viral infection is still unclear. In the present work, full length cDNAs of SUMO (FcSUMO and SUMO-conjugating enzyme E2 UBC9 (FcUBC9 were cloned from F. chinensis using rapid amplification of cDNA ends approach. The open reading frame (ORF of FcSUMO encoded a 93 amino acids peptide with the predicted molecular weight (M.W of 10.55 kDa, and the UBC9 ORF encoded a 160 amino acids peptide with the predicted M.W of 18.35 kDa. By quantitative real-time RT-PCR, higher mRNA transcription levels of FcSUMO and FcUBC9 were detected in hemocytes and ovary of F. chinensis, and the two genes were significantly up-regulated post WSSV infection. Subsequently, the recombinant proteins of FcSUMO and FcUBC9 were expressed in Escherichia coli BL21 (DE3, and employed as immunogens for the production of polyclonal antibody (PAb. Indirect immunofluorescence assay revealed that the FcSUMO and UBC9 proteins were mainly located in the hemocytes nuclei. By western blotting, a 13.5 kDa protein and a 18.7 kDa protein in hemocytes were recognized by the PAb against SUMO or UBC9 respectively. Furthermore, gene silencing of FcSUMO and FcUBC9 were performed using RNA interference, and the results showed that the number of WSSV copies and the viral gene expressions were inhibited by knockdown of either SUMO or UBC9, and the mortalities of shrimp were also reduced. These results indicated that FcSUMO and FcUBC9 played important roles in WSSV infection.

  18. Model of spontaneous evaporating droplet on solid horizontal substrate

    Science.gov (United States)

    Dunin, S. Z.; Nagornov, O. V.; Trifonenkov, V. P.

    2017-01-01

    Free evaporation of sessile liquid non-isothermal drop on solid substrate is analyzed. Exact formulae for temperature and concentration fields are found out as functions of dimensionless parameters. The non-uniform temperature distribution at the drop surface creates the thermocapillar Marangonni forces that change their direction in the vicinity of stagnation points. Direction of the forces and disposition of the stagnation points are derived as function of contact angle and thermodynamic parameters of model. Conditions for the stagnation points to appear are found out. Moreover, maximal value of contact angle corresponding to presence of stagnation points in droplet is calculated as a function of the thermal conductivity ratio.

  19. Viral Mimicry to Usurp Ubiquitin and SUMO Host Pathways

    Directory of Open Access Journals (Sweden)

    Peter Wimmer

    2015-08-01

    Full Text Available Posttranslational modifications (PTMs of proteins include enzymatic changes by covalent addition of cellular regulatory determinants such as ubiquitin (Ub and small ubiquitin-like modifier (SUMO moieties. These modifications are widely used by eukaryotic cells to control the functional repertoire of proteins. Over the last decade, it became apparent that the repertoire of ubiquitiylation and SUMOylation regulating various biological functions is not restricted to eukaryotic cells, but is also a feature of human virus families, used to extensively exploit complex host-cell networks and homeostasis. Intriguingly, besides binding to host SUMO/Ub control proteins and interfering with the respective enzymatic cascade, many viral proteins mimic key regulatory factors to usurp this host machinery and promote efficient viral outcomes. Advanced detection methods and functional studies of ubiquitiylation and SUMOylation during virus-host interplay have revealed that human viruses have evolved a large arsenal of strategies to exploit these specific PTM processes. In this review, we highlight the known viral analogs orchestrating ubiquitin and SUMO conjugation events to subvert and utilize basic enzymatic pathways.

  20. Modeling Growth of Cellulomonas cellulans NRRL B 4567 under Substrate Inhibition During Cellulase Production

    OpenAIRE

    Agarwal, R; Mahanty, B.; Dasu, V. Venkata

    2009-01-01

    Cellulase production study was performed in shake flask and bioreactor system using Cellulomonas cellulans NRRL B 4567 for initial substrate concentration from γS0 = 2 to 12 g L–1. The growth, substrate uptake profile and enzyme activity at different initial substrate concentrations were measured. The results inferred the presence of substrate inhibition kinetics. Various substrate inhibition models were tested and parameters were estimated, using non-linear regression analysis. Han-Levenspie...

  1. Whole-genome duplications followed by tandem duplications drive diversification of the protein modifier SUMO in Angiosperms

    NARCIS (Netherlands)

    Hammoudi, V.; Vlachakis, G.; Schranz, M.E.; van den Burg, H.A.

    2016-01-01

    The ubiquitin-like modifier (UBL) SUMO (Small Ubiquitin-Like Modifier) regulates protein function. Structural rather than sequence homology typifies UBL families. However, individual UBL types, such as SUMO, show remarkable sequence conservation. Selection pressure also operates at the SUMO gene cop

  2. Whole-genome duplications followed by tandem duplications drive diversification of the protein modifier SUMO in Angiosperms

    NARCIS (Netherlands)

    Hammoudi, Valentin; Vlachakis, Georgios; Schranz, Eric; Burg, van den Harrold A.

    2016-01-01

    The ubiquitin-like modifier (UBL) SUMO (Small Ubiquitin-Like Modifier) regulates protein function. Structural rather than sequence homology typifies UBL families. However, individual UBL types, such as SUMO, show remarkable sequence conservation. Selection pressure also operates at the SUMO gene

  3. Whole-genome duplications followed by tandem duplications drive diversification of the protein modifier SUMO in Angiosperms

    NARCIS (Netherlands)

    Hammoudi, V.; Vlachakis, G.; Schranz, M.E.; van den Burg, H.A.

    2016-01-01

    The ubiquitin-like modifier (UBL) SUMO (Small Ubiquitin-Like Modifier) regulates protein function. Structural rather than sequence homology typifies UBL families. However, individual UBL types, such as SUMO, show remarkable sequence conservation. Selection pressure also operates at the SUMO gene cop

  4. Whole-genome duplications followed by tandem duplications drive diversification of the protein modifier SUMO in Angiosperms

    NARCIS (Netherlands)

    Hammoudi, Valentin; Vlachakis, Georgios; Schranz, Eric; Burg, van den Harrold A.

    2016-01-01

    The ubiquitin-like modifier (UBL) SUMO (Small Ubiquitin-Like Modifier) regulates protein function. Structural rather than sequence homology typifies UBL families. However, individual UBL types, such as SUMO, show remarkable sequence conservation. Selection pressure also operates at the SUMO gene

  5. SUMO fusion technology for enhanced protein production in prokaryotic and eukaryotic expression systems.

    Science.gov (United States)

    Panavas, Tadas; Sanders, Carsten; Butt, Tauseef R

    2009-01-01

    In eukaryotic cells, the reversible attachment of small ubiquitin-like modifier (SUMO) protein is a post-translational modification that has been demonstrated to play an important role in various cellular processes. Moreover, it has been found that SUMO as an N-terminal fusion partner enhances functional protein production in prokaryotic and eukaryotic expression systems, based upon significantly improved protein stability and solubility. Following the expression and purification of the fusion protein, the SUMO-tag can be cleaved by specific (SUMO) proteases via their endopeptidase activity in vitro to generate the desired N-terminus of the released protein partner. In addition to its physiological relevance in eukaryotes, SUMO can, thus, be used as a powerful biotechnological tool for protein expression in prokaryotic and eukaryotic cell systems.In this chapter, we will describe the construction of a fusion protein with the SUMO-tag, its expression in Escherichia coli, and its purification followed by the removal of the SUMO-tag by a SUMO-specific protease in vitro.

  6. Comprehensive Identification of SUMO2/3 Targets and Their Dynamics during Mitosis

    DEFF Research Database (Denmark)

    Schou, Julie; Kelstrup, Christian D; Hayward, Daniel G

    2014-01-01

    During mitosis large alterations in cellular structures occur rapidly, which to a large extent is regulated by post-translational modification of proteins. Modification of proteins with the small ubiquitin-related protein SUMO2/3 regulates mitotic progression, but few mitotic targets have been...... identified so far. To deepen our understanding of SUMO2/3 during this window of the cell cycle, we undertook a comprehensive proteomic characterization of SUMO2/3 modified proteins in mitosis and upon mitotic exit. We developed an efficient tandem affinity purification strategy of SUMO2/3 modified proteins...... from mitotic cells. Combining this purification strategy with cell synchronization procedures and quantitative mass spectrometry allowed for the mapping of numerous novel targets and their dynamics as cells progressed out of mitosis. This identified RhoGDIα as a major SUMO2/3 modified protein...

  7. SUMO-interacting motifs of human TRIM5α are important for antiviral activity.

    Directory of Open Access Journals (Sweden)

    Gloria Arriagada

    2011-04-01

    Full Text Available Human TRIM5α potently restricts particular strains of murine leukemia viruses (the so-called N-tropic strains but not others (the B- or NB-tropic strains during early stages of infection. We show that overexpression of SUMO-1 in human 293T cells, but not in mouse MDTF cells, profoundly blocks N-MLV infection. This block is dependent on the tropism of the incoming virus, as neither B-, NB-, nor the mutant R110E of N-MLV CA (a B-tropic switch are affected by SUMO-1 overexpression. The block occurred prior to reverse transcription and could be abrogated by large amounts of restricted virus. Knockdown of TRIM5α in 293T SUMO-1-overexpressing cells resulted in ablation of the SUMO-1 antiviral effects, and this loss of restriction could be restored by expression of a human TRIM5α shRNA-resistant plasmid. Amino acid sequence analysis of human TRIM5α revealed a consensus SUMO conjugation site at the N-terminus and three putative SUMO interacting motifs (SIMs in the B30.2 domain. Mutations of the TRIM5α consensus SUMO conjugation site did not affect the antiviral activity of TRIM5α in any of the cell types tested. Mutation of the SIM consensus sequences, however, abolished TRIM5α antiviral activity against N-MLV. Mutation of lysines at a potential site of SUMOylation in the CA region of the Gag gene reduced the SUMO-1 block and the TRIM5α restriction of N-MLV. Our data suggest a novel aspect of TRIM5α-mediated restriction, in which the presence of intact SIMs in TRIM5α, and also the SUMO conjugation of CA, are required for restriction. We propose that at least a portion of the antiviral activity of TRIM5α is mediated through the binding of its SIMs to SUMO-conjugated CA.

  8. Small signal modeling of high electron mobility transistors on silicon and silicon carbide substrate with consideration of substrate loss mechanism

    Science.gov (United States)

    Sahoo, A. K.; Subramani, N. K.; Nallatamby, J. C.; Sylvain, L.; Loyez, C.; Quere, R.; Medjdoub, F.

    2016-01-01

    In this paper, we present a comparative study on small-signal modeling of AlN/GaN/AlGaN double hetero-structure high electron mobility transistors (HEMTs) grown on silicon (Si) and silicon carbide (SiC) substrate. The traditional small signal equivalent circuit model is modified to take into account the transmission loss mechanism of coplanar waveguide (CPW) line which cannot be neglected at high frequencies. CPWs and HEMTs-on-AlN/GaN/AlGaN epitaxial layers are fabricated on both the Si and SiC substrates. S-parameter measurements at room temperature are performed over the frequency range from 0.5 GHz to 40 GHz. Transmission loss of CPW lines are modeled with a distributed transmission line (TL) network and an equivalent circuit model is included in the small-signal transistor model topology. Measurements and simulations are compared and found to be in good agreement.

  9. Proteome-wide analysis of SUMO2 targets in response to pathological DNA replication stress in human cells

    DEFF Research Database (Denmark)

    Bursomanno, Sara; Beli, Petra; Khan, Asif M;

    2015-01-01

    SUMOylation is a form of post-translational modification involving covalent attachment of SUMO (Small Ubiquitin-like Modifier) polypeptides to specific lysine residues in the target protein. In human cells, there are four SUMO proteins, SUMO1-4, with SUMO2 and SUMO3 forming a closely related...... subfamily. SUMO2/3, in contrast to SUMO1, are predominantly involved in the cellular response to certain stresses, including heat shock. Substantial evidence from studies in yeast has shown that SUMOylation plays an important role in the regulation of DNA replication and repair. Here, we report a proteomic...... repair. We have also shown that deficiency of POLD3 leads to an increase in RPA-bound ssDNA when cells are under replication stress, suggesting that POLD3 plays a role in the cellular response to DNA replication stress. Considering that DNA replication stress is a source of genome instability...

  10. Modeling of organic substrate transformation in the high-rate activated sludge process.

    Science.gov (United States)

    Nogaj, Thomas; Randall, Andrew; Jimenez, Jose; Takacs, Imre; Bott, Charles; Miller, Mark; Murthy, Sudhir; Wett, Bernhard

    2015-01-01

    This study describes the development of a modified activated sludge model No.1 framework to describe the organic substrate transformation in the high-rate activated sludge (HRAS) process. New process mechanisms for dual soluble substrate utilization, production of extracellular polymeric substances (EPS), absorption of soluble substrate (storage), and adsorption of colloidal substrate were included in the modified model. Data from two HRAS pilot plants were investigated to calibrate and to validate the proposed model for HRAS systems. A subdivision of readily biodegradable soluble substrate into a slow and fast fraction were included to allow accurate description of effluent soluble chemical oxygen demand (COD) in HRAS versus longer solids retention time (SRT) systems. The modified model incorporates production of EPS and storage polymers as part of the aerobic growth transformation process on the soluble substrate and transformation processes for flocculation of colloidal COD to particulate COD. The adsorbed organics are then converted through hydrolysis to the slowly biodegradable soluble fraction. Two soluble substrate models were evaluated during this study, i.e., the dual substrate and the diauxic models. Both models used two state variables for biodegradable soluble substrate (SBf and SBs) and a single biomass population. The A-stage pilot typically removed 63% of the soluble substrate (SB) at an SRT <0.13 d and 79% at SRT of 0.23 d. In comparison, the dual substrate model predicted 58% removal at the lower SRT and 78% at the higher SRT, with the diauxic model predicting 32% and 70% removals, respectively. Overall, the dual substrate model provided better results than the diauxic model and therefore it was adopted during this study. The dual substrate model successfully described the higher effluent soluble COD observed in the HRAS systems due to the partial removal of SBs, which is almost completely removed in higher SRT systems.

  11. The p150 subunit of CAF-1 causes association of SUMO2/3 with the DNA replication foci

    Energy Technology Data Exchange (ETDEWEB)

    Uwada, Junsuke [Department of Biological Sciences, Graduate School of Science and Technology, Kumamoto University, 2-39-1 Kurokami, Kumamoto 860-8555 (Japan); Global COE (Centers of Excellence) Program, Global Initiative Center for Pulsed Power Engineering, Kumamoto University, 2-39-1 Kurokami, Kumamoto 860-8555 (Japan); Tanaka, Niina; Yamaguchi, Yutaro; Uchimura, Yasuhiro [Department of Biological Sciences, Graduate School of Science and Technology, Kumamoto University, 2-39-1 Kurokami, Kumamoto 860-8555 (Japan); Shibahara, Kei-ichi [Department of Integrated Genetics, National Institute of Genetics, Research Organization of Information and Systems, Mishima (Japan); Nakao, Mitsuyoshi [Department of Medical Cell Biology, Institute of Molecular Embryology and Genetics, Kumamoto University, Kumamoto (Japan); Saitoh, Hisato, E-mail: hisa@kumamoto-u.ac.jp [Department of Biological Sciences, Graduate School of Science and Technology, Kumamoto University, 2-39-1 Kurokami, Kumamoto 860-8555 (Japan); Global COE (Centers of Excellence) Program, Global Initiative Center for Pulsed Power Engineering, Kumamoto University, 2-39-1 Kurokami, Kumamoto 860-8555 (Japan)

    2010-01-01

    The small ubiquitin-related modifier 2/3 (SUMO2/3) can be post-translationally conjugated to a wide variety of proteins constituting chromatin, the platform for genetic and epigenetic regulation. Nevertheless, it is unclear how SUMO2/3 and SUMO2/3-modified proteins are delivered to the chromatin fibers. Here we report that the largest subunit of chromatin assembly factor 1 (CAF-1), human p150, interacts directly and preferentially with SUMO2/3. Amino acid residue of 98-105 in p150 is essential and sufficient for SUMO2/3 interaction. p150-SUMO2/3 interaction coincided with regions that replicate chromatin fibers, because accumulation of the proliferating cell nuclear antigen (PCNA), and incorporation of bromodeoxyuridine (BrdU) were detected at foci co-localized with both p150 and SUMO2/3 during the S-phase in a cell line expressing epitope-tagged p150. Although inhibition of SUMO2/3 expression had only a small effect on p150 deposition on the replication sites, depletion of p150 led to delocalization of SUMO2/3 from the replication foci. Furthermore, p150 mutants deficient in SUMO2/3 interaction, caused a major reduction of SUMO2/3 at the replication foci. Thus, our findings suggest an expanded role of p150 as a SUMO2/3-interacting factor, and raise the intriguing possibility that p150 plays a role in promoting delivery of SUMO2/3 or SUMO2/3-modified proteins (or both) on chromatin fibers during replication.

  12. Comprehensive identification of SUMO2/3 targets and their dynamics during mitosis.

    Science.gov (United States)

    Schou, Julie; Kelstrup, Christian D; Hayward, Daniel G; Olsen, Jesper V; Nilsson, Jakob

    2014-01-01

    During mitosis large alterations in cellular structures occur rapidly, which to a large extent is regulated by post-translational modification of proteins. Modification of proteins with the small ubiquitin-related protein SUMO2/3 regulates mitotic progression, but few mitotic targets have been identified so far. To deepen our understanding of SUMO2/3 during this window of the cell cycle, we undertook a comprehensive proteomic characterization of SUMO2/3 modified proteins in mitosis and upon mitotic exit. We developed an efficient tandem affinity purification strategy of SUMO2/3 modified proteins from mitotic cells. Combining this purification strategy with cell synchronization procedures and quantitative mass spectrometry allowed for the mapping of numerous novel targets and their dynamics as cells progressed out of mitosis. This identified RhoGDIα as a major SUMO2/3 modified protein, specifically during mitosis, mediated by the SUMO ligases PIAS2 and PIAS3. Our data provide a rich resource for further exploring the role of SUMO2/3 modifications in mitosis and cell cycle regulation.

  13. Modeling of the substrate influence on multielement THz detector operation

    CERN Document Server

    Sakhno, M; Sizov, F

    2015-01-01

    The development of THz multielement uncooled imagers based on focal plane arrays (FPAs) requires an optimization of the system parameters to achieve a homogeneous sensitivity of the array elements. Results of numerical simulation of the eight-element linear array of planar antennas with detecting elements, on a substrate of finite dimensions are presented. We establish how the substrate thickness h and the relative permittivity epsilon influence antenna pattern and antenna-detector matching for each element. We show that the antenna pattern depends on the detector position more than the antenna-detector impedance matching. The gain of array elements, the antenna-detector matching, and the homogeneity of the detector sensitivity can be simultaneously optimized by the proper choice of the substrate thickness h and the relative permittivity epsilon. We show that multielement systems with large substrate thickness and high relative permittivity are not suitable for the imaging system implementation. To achieve un...

  14. Identifications of SUMO-1 cDNA and Its Expression Patterns in Pacific White Shrimp Litopeanaeus vannamei

    Directory of Open Access Journals (Sweden)

    Yanisa Laoong-u-thai, Baoping Zhao, Amornrat Phongdara, Harry Ako, Jinzeng Yang

    2009-01-01

    Full Text Available Small ubiquitin-like modifiers (SUMO work in a similar way as ubiquitin to alter the biological properties of a target protein by conjugation. A shrimp SUMO cDNA named LvSUMO-1 was identified in Litopenaeus vannamei. LvSUMO-1 cDNA contains a coding sequence of 282 nucleotides with untranslated regions of 37 bp at 5'-end and 347 bp at 3'-end, respectively. The deduced 93 amino acids exhibit 83% identity with the Western Honeybee SUMO-1, and more than 65% homologies with human and mouse SUMO-1. LvSUMO-1 mRNA is expressed in most L. vannamei tissues with the highest level in hepatopancrease. The mRNA expression of LvSUMO-1 over development stages in L. Vammamei is distinguished by a low level in nauplius stage and relatively high level in postlarva stage with continuous expression until juvenile stage. The LvSUMO-1 protein and its conjugated proteins are detected in both cytoplasm and nucleus in several tissues. Interestingly, LvSUMO-1 mRNA levels are high in abdominal muscle during the premolt stage, wherein it has significant activities of protein degradation, suggesting its possible role in the regulation of shrimp muscle protein degradation.

  15. A Lubrication Model of Coating Flows over a Curved Substrate in Space

    CERN Document Server

    Roy, R V; Simpson, M E

    1997-01-01

    Consider the three-dimensional flow of a viscous Newtonian fluid upon an abitrarily curved substrate when the fluid film is thin as occurs in many draining, coating and biological flows. We derive a model of the dynamics of the film, the model being expressed in terms of the film thickness and the curvature tensor of the substrate. The model accurately includes the effects of the curvature of the substrate, via a physical multiple-scale approach, and gravity and inertia, via more rigorous centre manifold techniques. Numerical simulations exhibit some generic features of the dynamics of such thin fluid films on substrates with complex curvature.

  16. Modeling of the Substrate Influence on Multielement THz Detector Operation

    Science.gov (United States)

    Sakhno, M.; Gumenjuk-Sichevska, J.; Sizov, F.

    2014-09-01

    The development of THz multielement uncooled imagers based on focal plane arrays (FPAs) requires an optimization of the system parameters to achieve a homogeneous sensitivity of the array elements. Results of numerical simulation of the eight-element linear array of planar antennas with detecting elements, on a substrate of finite dimensions are presented. We establish how the substrate thickness h and the relative permittivity ɛ r influence antenna pattern and antenna-detector matching for each element. We show that the antenna pattern depends on the detector position more than the antenna - detector impedance matching. The gain of array elements, the antenna-detector matching, and the homogeneity of the detector sensitivity can be simultaneously optimized by the proper choice of the substrate thickness h and the relative permittivity ɛ r . We show that multielement systems with large substrate thickness and high relative permittivity are not suitable for the imaging system implementation. To achieve uniform multielement system sensitivity, substrates with low permittivity ( ɛ r corruption.

  17. The SUMO Ship Detector Algorithm for Satellite Radar Images

    Directory of Open Access Journals (Sweden)

    Harm Greidanus

    2017-03-01

    Full Text Available Search for Unidentified Maritime Objects (SUMO is an algorithm for ship detection in satellite Synthetic Aperture Radar (SAR images. It has been developed over the course of more than 15 years, using a large amount of SAR images from almost all available SAR satellites operating in L-, C- and X-band. As validated by benchmark tests, it performs very well on a wide range of SAR image modes (from Spotlight to ScanSAR and resolutions (from 1–100 m and for all types and sizes of ships, within the physical limits imposed by the radar imaging. This paper describes, in detail, the algorithmic approach in all of the steps of the ship detection: land masking, clutter estimation, detection thresholding, target clustering, ship attribute estimation and false alarm suppression. SUMO is a pixel-based CFAR (Constant False Alarm Rate detector for multi-look radar images. It assumes a K distribution for the sea clutter, corrected however for deviations of the actual sea clutter from this distribution, implementing a fast and robust method for the clutter background estimation. The clustering of detected pixels into targets (ships uses several thresholds to deal with the typically irregular distribution of the radar backscatter over a ship. In a multi-polarization image, the different channels are fused. Azimuth ambiguities, a common source of false alarms in ship detection, are removed. A reliability indicator is computed for each target. In post-processing, using the results of a series of images, additional false alarms from recurrent (fixed targets including range ambiguities are also removed. SUMO can run in semi-automatic mode, where an operator can verify each detected target. It can also run in fully automatic mode, where batches of over 10,000 images have successfully been processed in less than two hours. The number of satellite SAR systems keeps increasing, as does their application to maritime surveillance. The open data policy of the EU

  18. The potential role of As-sumo-1 in the embryonic diapause process and early embryo development of Artemia sinica.

    Science.gov (United States)

    Chu, Bing; Yao, Feng; Cheng, Cheng; Wu, Yang; Mei, Yanli; Li, Xuejie; Liu, Yan; Wang, Peisheng; Hou, Lin; Zou, Xiangyang

    2014-01-01

    During embryonic development of Artemia sinica, environmental stresses induce the embryo diapause phenomenon, required to resist apoptosis and regulate cell cycle activity. The small ubiquitin-related modifier-1 (SUMO), a reversible post-translational protein modifier, plays an important role in embryo development. SUMO regulates multiple cellular processes, including development and other biological processes. The molecular mechanism of diapause, diapause termination and the role of As-sumo-1 in this processes and in early embryo development of Artemia sinica still remains unknown. In this study, the complete cDNA sequences of the sumo-1 homolog, sumo ligase homolog, caspase-1 homolog and cyclin B homolog from Artemia sinica were cloned. The mRNA expression patterns of As-sumo-1, sumo ligase, caspase-1, cyclin B and the location of As-sumo-1 were investigated. SUMO-1, p53, Mdm2, Caspase-1, Cyclin B and Cyclin E proteins were analyzed during different developmental stages of the embryo of A. sinica. Small interfering RNA (siRNA) was used to verify the function of sumo-1 in A. sinica. The full-length cDNA of As-sumo-1 was 476 bp, encoding a 92 amino acid protein. The As-caspases-1 cDNA was 966 bp, encoding a 245 amino-acid protein. The As-sumo ligase cDNA was 1556 bp encoding, a 343 amino acid protein, and the cyclin B cDNA was 739 bp, encoding a 133 amino acid protein. The expressions of As-sumo-1, As-caspase-1 and As-cyclin B were highest at the 10 h stage of embryonic development, and As-sumo ligase showed its highest expression at 0 h. The expression of As-SUMO-1 showed no tissue or organ specificity. Western blotting showed high expression of As-SUMO-1, p53, Mdm2, Caspase-1, Cyclin B and Cyclin E at the 10 h stage. The siRNA caused abnormal development of the embryo, with increased malformation and mortality. As-SUMO-1 is a crucial regulation and modification protein resumption of embryonic diapause and early embryo development of A. sinica.

  19. Modification of Akt by SUMO conjugation regulates alternative splicing and cell cycle

    OpenAIRE

    Risso, Guillermo; Pelisch, Federico; Pozzi, Berta; Mammi, Pablo; Blaustein, Matías; Colman-Lerner, Alejandro; Srebrow, Anabella

    2013-01-01

    Akt/PKB is a key signaling molecule in higher eukaryotes and a crucial protein kinase in human health and disease. Phosphorylation, acetylation, and ubiquitylation have been reported as important regulatory post-translational modifications of this kinase. We describe here that Akt is modified by SUMO conjugation, and show that lysine residues 276 and 301 are the major SUMO attachment sites within this protein. We found that phosphorylation and SUMOylation of Akt appear as independent events. ...

  20. Mathematical model of organic substrate degradation in solid waste windrow composting.

    Science.gov (United States)

    Seng, Bunrith; Kristanti, Risky Ayu; Hadibarata, Tony; Hirayama, Kimiaki; Katayama-Hirayama, Keiko; Kaneko, Hidehiro

    2016-01-01

    Organic solid waste composting is a complex process that involves many coupled physical, chemical and biological mechanisms. To understand this complexity and to ease in planning, design and management of the composting plant, mathematical model for simulation is usually applied. The aim of this paper is to develop a mathematical model of organic substrate degradation and its performance evaluation in solid waste windrow composting system. The present model is a biomass-dependent model, considering biological growth processes under the limitation of moisture, oxygen and substrate contents, and temperature. The main output of this model is substrate content which was divided into two categories: slowly and rapidly degradable substrates. To validate the model, it was applied to a laboratory scale windrow composting of a mixture of wood chips and dog food. The wastes were filled into a cylindrical reactor of 6 cm diameter and 1 m height. The simulation program was run for 3 weeks with 1 s stepwise. The simulated results were in reasonably good agreement with the experimental results. The MC and temperature of model simulation were found to be matched with those of experiment, but limited for rapidly degradable substrates. Under anaerobic zone, the degradation of rapidly degradable substrate needs to be incorporated into the model to achieve full simulation of a long period static pile composting. This model is a useful tool to estimate the changes of substrate content during composting period, and acts as a basic model for further development of a sophisticated model.

  1. SUMO meets meiosis: an encounter at the synaptonemal complex: SUMO chains and sumoylated proteins suggest that heterogeneous and complex interactions lie at the centre of the synaptonemal complex.

    Science.gov (United States)

    Watts, Felicity Z; Hoffmann, Eva

    2011-07-01

    Recent discoveries have identified the small ubiquitin-like modifier (SUMO) as the potential 'missing link' that could explain how the synaptonemal complex (SC) is formed during meiosis. The SC is important for a variety of chromosome interactions during meiosis and appears ladder-like. It is formed when 'axes' of the two homologous chromosomes become connected by the deposition of transverse filaments, forming the steps of the ladder. Although several components of axial and transverse elements have been identified, how the two are connected to form the SC has remained an enigma. Recent discoveries suggest that SUMO modification underlies protein-protein interactions within the SC of budding yeast. The versatility of SUMO in regulating protein-protein interactions adds an exciting new dimension to our understanding of the SC and suggests that SCs are not homogenous structures throughout the nucleus. We propose that this heterogeneity may allow differential regulation of chromosome structure and function.

  2. SUMO/FREND: vision system for autonomous satellite grapple

    Science.gov (United States)

    Obermark, Jerome; Creamer, Glenn; Kelm, Bernard E.; Wagner, William; Henshaw, C. Glen

    2007-04-01

    SUMO/FREND is a risk reduction program for an advanced servicing spacecraft sponsored by DARPA and executed by the Naval Center for Space Technology at the Naval Research Laboratory in Washington, DC. The overall program will demonstrate the integration of many techniques needed in order to autonomously rendezvous and capture customer satellites at geosynchronous orbits. A flight-qualifiable payload is currently under development to prove out challenging aspects of the mission. The grappling process presents computer vision challenges to properly identify and guide the final step in joining the pursuer craft to the customer. This paper will provide an overview of the current status of the project with an emphasis on the challenges, techniques, and directions of the machine vision processes to guide the grappling.

  3. Wrestling with Chromosomes: The Roles of SUMO During Meiosis.

    Science.gov (United States)

    Nottke, Amanda C; Kim, Hyun-Min; Colaiácovo, Monica P

    2017-01-01

    Meiosis is a specialized form of cell division required for the formation of haploid gametes and therefore is essential for successful sexual reproduction. Various steps are exquisitely coordinated to ensure accurate chromosome segregation during meiosis, thereby promoting the formation of haploid gametes from diploid cells. Recent studies are demonstrating that an important form of regulation during meiosis is exerted by the post-translational protein modification known as sumoylation. Here, we review and discuss the various critical steps of meiosis in which SUMO-mediated regulation has been implicated thus far. These include the maintenance of meiotic centromeric heterochromatin , meiotic DNA double-strand break repair and homologous recombination, centromeric coupling, and the assembly of a proteinaceous scaffold between homologous chromosomes known as the synaptonemal complex.

  4. VP6-SUMO Self-Assembly as Nanocarriers for Gastrointestinal Delivery

    Directory of Open Access Journals (Sweden)

    V. Palmieri

    2015-01-01

    Full Text Available High proteolytic degradation and poor absorption through epithelial barriers are major challenges to successful oral delivery of therapeutics. Nanoparticle platforms can enhance drug stability and extend the residence time in gastrointestinal (GI tract. However, drug delivery systems are often inactivated in acidic environment of stomach or suffer poor absorption from intestinal cells due to the mucus layer. To overcome these issues we developed a drug delivery system constituted by a protein construct made by a Rotavirus capsid protein (VP6 and the small ubiquitin-like modifier SUMO. This chimeric construct allows specificity towards intestinal cells, the Rotavirus natural target, combined by an enhanced stability given by the eukaryotic protein transporter SUMO. Furthermore SUMO can act as a molecular switch that facilitates import/export of its ligand to the nucleus, the hypersensitive subcellular site target of many cell killing therapies. In this paper we show that SUMO-VP6 constructs self-assembly into stable nanocarriers. SUMO-VP6 nanocarriers display ideal features for drug delivery: a small size and high monodispersity, a high stability in different pH conditions and a high uptake in the nuclear and cytoplasmic compartment of intestinal cells. These features make SUMO-VP6 nanocarriers a promising novel system for oral delivery of poorly soluble drugs.

  5. Cell cycle-dependent SUMO-1 conjugation to nuclear mitotic apparatus protein (NuMA)

    Energy Technology Data Exchange (ETDEWEB)

    Seo, Jae Sung; Kim, Ha Na; Kim, Sun-Jick; Bang, Jiyoung; Kim, Eun-A; Sung, Ki Sa [Department of Biological Sciences, Sungkyunkwan University, Suwon 440-746 (Korea, Republic of); Yoon, Hyun-Joo [TissueGene Inc. 9605 Medical Center Dr., Rockville, MD 20850 (United States); Yoo, Hae Yong [Department of Health Sciences and Technology, Samsung Advanced Institute for Health Sciences and Technology, Samsung Medical Center, Sungkyunkwan University, Seoul 135-710 (Korea, Republic of); Choi, Cheol Yong, E-mail: choicy@skku.ac.kr [Department of Biological Sciences, Sungkyunkwan University, Suwon 440-746 (Korea, Republic of)

    2014-01-03

    Highlights: •NuMA is modified by SUMO-1 in a cell cycle-dependent manner. •NuMA lysine 1766 is the primary target site for SUMOylation. •SUMOylation-deficient NuMA induces multiple spindle poles during mitosis. •SUMOylated NuMA induces microtubule bundling. -- Abstract: Covalent conjugation of proteins with small ubiquitin-like modifier 1 (SUMO-1) plays a critical role in a variety of cellular functions including cell cycle control, replication, and transcriptional regulation. Nuclear mitotic apparatus protein (NuMA) localizes to spindle poles during mitosis, and is an essential component in the formation and maintenance of mitotic spindle poles. Here we show that NuMA is a target for covalent conjugation to SUMO-1. We find that the lysine 1766 residue is the primary NuMA acceptor site for SUMO-1 conjugation. Interestingly, SUMO modification of endogenous NuMA occurs at the entry into mitosis and this modification is reversed after exiting from mitosis. Knockdown of Ubc9 or forced expression of SENP1 results in impairment of the localization of NuMA to mitotic spindle poles during mitosis. The SUMOylation-deficient NuMA mutant is defective in microtubule bundling, and multiple spindles are induced during mitosis. The mitosis-dependent dynamic SUMO-1 modification of NuMA might contribute to NuMA-mediated formation and maintenance of mitotic spindle poles during mitosis.

  6. KSHV latent protein LANA2 inhibits sumo2 modification of p53

    Science.gov (United States)

    Laura, Marcos-Villar; de la Cruz-Herrera, Carlos F; Ferreirós, Alba; Baz-Martínez, Maite; Lang, Valerie; Vidal, Anxo; Muñoz-Fontela, Cesar; Rodríguez, Manuel S; Collado, Manuel; Rivas, Carmen

    2015-01-01

    Tumor suppressor p53 plays a crucial antiviral role and targeting of p53 by viral proteins is a common mechanism involved in virus oncogenesis. The activity of p53 is tightly regulated at the post-translational levels through a myriad of modifications. Among them, modification of p53 by SUMO has been associated with the onset of cellular senescence. Kaposi´s sarcoma-associated herpesvirus (KSHV) expresses several proteins targeting p53, including the latent protein LANA2 that regulates polyubiquitylation and phosphorylation of p53. Here we show that LANA2 also inhibits the modification of p53 by SUMO2. Furthermore, we show that the reduction of p53-SUMO2 conjugation by LANA2, as well as the p53-LANA2 interaction, both require the SUMOylation of the viral protein and its interaction with SUMO or SUMOylated proteins in a non-covalent manner. Finally, we show that the control of p53-SUMO2 conjugation by LANA2 correlates with its ability to inhibit SUMO2- and type I interferon-induced senescence. These results highlight the importance of p53 SUMOylation in the control of virus infection and suggest that viral oncoproteins could contribute to viral infection and cell transformation by abrogating p53 SUMOylation. PMID:25607652

  7. A monoclonal antibody against the human SUMO-1 protein obtained by immunization with recombinant protein and CpG-DNA-liposome complex.

    Science.gov (United States)

    Kim, Dongbum; Lee, Joo Young; Song, Dae-Geun; Kwon, Sanghoon; Lee, Younghee; Pan, Cheol-Ho; Kwon, Hyung-Joo

    2013-10-01

    Post-translational modification regulated by conjugation of a small ubiquitin-like modifier (SUMO) is involved in various cellular processes. In this study, we expressed and purified recombinant human SUMO-1 (hSUMO-1). BALB/c mice were immunized with a complex of hSUMO-1 protein and Lipoplex(O) to produce hSUMO-1-specific antibodies. Using conventional hybridoma technology, we obtained four hybridoma clones derived from the mouse with the highest antibody titer against hSUMO-1. Based on Western blot analysis, our hSUMO-1 monoclonal antibody specifically recognizes hSUMO-1, but not other SUMO proteins. These results support that the anti-hSUMO-1 monoclonal antibody produced with the aid of Lipoplex(O) adjuvant is specific and that Lipoplex(O) is useful for development of monoclonal antibodies against recombinant protein. In addition, we analyzed human tissues to examine the distribution of hSUMO-1. Higher expression of hSUMO-1 was detected in normal adrenal gland, esophagus, pancreas, liver, stomach, kidney, and uterus than in corresponding cancer tissues, suggesting a tumor suppressive function of hSUMO-1.

  8. Adhesion of silver nanoparticles on the clay substrates; modeling and experiment

    Energy Technology Data Exchange (ETDEWEB)

    Tokarsky, Jonas, E-mail: jonas.tokarsky.fmmi@vsb.cz [Nanotechnology Centre, VSB-Technical University Ostrava, 17. listopadu 15/2172, 708 33 Ostrava (Czech Republic); Capkova, Pavla [Nanotechnology Centre, VSB-Technical University Ostrava, 17. listopadu 15/2172, 708 33 Ostrava (Czech Republic); Rafaja, David; Klemm, Volker [Institute of Materials Science, TU Bergakademie Freiberg, Gustav-Zeuner-Street 5, D-09599 Freiberg (Germany); Valaskova, Marta; Kukutschova, Jana; Tomasek, Vladimir [Nanotechnology Centre, VSB-Technical University Ostrava, 17. listopadu 15/2172, 708 33 Ostrava (Czech Republic)

    2010-02-15

    Adhesion of silver nanoparticles on the montmorillonite and kaolinite substrates has been investigated using molecular modeling (force field calculations) that enabled the estimation and comparison of adhesion energies for Ag/montmorillonite and Ag/kaolinite nanocomposites and revealed the preferred orientation of Ag nanoparticles on the silicate substrates. Results of the modeling have been confronted with experiment (X-ray fluorescence, high-resolution transmission electron microscopy). This confrontation has shown that the results of the modeling are consistent with the experimental data and illustrated the capability of the molecular modeling for prediction of the nanoparticles orientation, structure and stability of the nanoparticle/substrate nanocomposite.

  9. Association of SUMO4 M55V polymorphism with autoimmune diabetes in Latvian patients.

    Science.gov (United States)

    Sedimbi, Saikiran K; Shastry, Arun; Park, Yongsoo; Rumba, Ingrida; Sanjeevi, Carani B

    2006-10-01

    Small ubiquitin-related modifier (SUMO4), located in IDDM5, has been identified as a potential susceptibility gene for type 1 diabetes mellitus (T1DM). The novel polymorphism M55V, causing an amino acid change in the evolutionarily conserved met55 residue has been shown to activate the nuclear factor kappaB (NF-kappaB), hence the suspected role of SUMO4 in the pathogenicity of T1DM. The M55V polymorphism has been shown to be associated with susceptibility to T1DM in Asians, but not in Caucasians. Latent autoimmune diabetes in adults (LADA) is a slowly progressive form of T1DM and SUMO4 M55V has not been studied in LADA to date. The current study aims to test whether Latvians are similar to Caucasians in susceptibility to autoimmune diabetes (T1DM and LADA), with respect to SUMO4 M55V. We studied, age- and sex-matched, Latvian T1DM patients (n = 100) and healthy controls (n = 90) and LADA patients (n = 45) and healthy controls (n = 95). SUMO4 M55V polymorphism was analyzed using polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP). The allelic frequencies of the A and G alleles were compared with HLA DR3-DR4-positive and HLA DR3-DR4-negative patients to identify any potential relation between HLA DR3-DR4 and SUMO4 M55V. We found no significant association between SUMO4 M55V and T1DM susceptibility in Latvians, the results being in concurrence with the previous studies in Caucasians of British and Canadian origin. Comparison of the A and G alleles with HLA DR3-DR4 did not result in any significant P values. No significant association was found between SUMO4 M55V and LADA. SUMO4 M55V is not associated with susceptibility to T1DM and LADA in Latvians, and Latvians exhibit similarity to other Caucasians with respect to association of SUMO4 M55V with autoimmune diabetes.

  10. A mesoscopic stochastic model for the specific consumption rate in substrate-limited microbial growth

    Science.gov (United States)

    2017-01-01

    The specific consumption rate of substrate, as well as the associated specific growth rate, is an essential parameter in the mathematical description of substrate-limited microbial growth. In this paper we develop a completely new kinetic model of substrate transport, based on recent knowledge on the structural biology of transport proteins, which correctly describes very accurate experimental results at near-zero substrate concentration values found in the literature, where the widespread Michaelis-Menten model fails. Additionally, our model converges asymptotically to Michaelis-Menten predictions as substrate concentration increases. Instead of the single active site enzymatic reaction of Michaelis-Menten type, the proposed model assumes a multi-site kinetics, simplified as an apparent all-or-none mechanism for the transport, which is controlled by means of the local substrate concentration in the close vicinity of the transport protein. Besides, the model also assumes that this local concentration is not equal to the mean substrate concentration experimentally determined in the culture medium. Instead, we propose that it fluctuates with a mostly exponential distribution of Weibull type. PMID:28187189

  11. PIAS1 is a GATA4 SUMO ligase that regulates GATA4-dependent intestinal promoters independent of SUMO ligase activity and GATA4 sumoylation.

    Directory of Open Access Journals (Sweden)

    Narasimhaswamy S Belaguli

    Full Text Available GATA4 confers cell type-specific gene expression on genes expressed in cardiovascular, gastro-intestinal, endocrine and neuronal tissues by interacting with various ubiquitous and cell-type-restricted transcriptional regulators. By using yeast two-hybrid screening approach, we have identified PIAS1 as an intestine-expressed GATA4 interacting protein. The physical interaction between GATA4 and PIAS1 was confirmed in mammalian cells by coimmunoprecipitation and two-hybrid analysis. The interacting domains were mapped to the second zinc finger and the adjacent C-terminal basic region of GATA4 and the RING finger and the adjoining C-terminal 60 amino acids of PIAS1. PIAS1 and GATA4 synergistically activated IFABP and SI promoters but not LPH promoters suggesting that PIAS1 differentially activates GATA4 targeted promoters. In primary murine enterocytes PIAS1 was recruited to the GATA4-regulated IFABP promoter. PIAS1 promoted SUMO-1 modification of GATA4 on lysine 366. However, sumoylation was not required for the nuclear localization and stability of GATA4. Further, neither GATA4 sumoylation nor the SUMO ligase activity of PIAS1 was required for coactivation of IFABP promoter by GATA4 and PIAS1. Together, our results demonstrate that PIAS1 is a SUMO ligase for GATA4 that differentially regulates GATA4 transcriptional activity independent of SUMO ligase activity and GATA4 sumoylation.

  12. A phenomenological model of coating/substrate adhesion and interfacial bimetallic peeling stress in composite mirrors

    Science.gov (United States)

    Mcelroy, Paul M.; Lawson, Daniel D.

    1990-01-01

    Adhesion and interfacial stress between metal films and structural composite material substrates is discussed. A theoretical and conceptual basis for selecting coating materials for composite mirror substrates is described. A phenomenological model that interrelates cohesive tensile strength of thin film coatings and interfacial peeling stresses is presented. The model serves as a basis in determining gradiated materials response and compatibility of composite substrate and coating combinations. Parametric evaluation of material properties and geometrical factors such as coating thickness are used to determine the threshold stress levels for maintaining adhesion at the different interfaces.

  13. Mathematical modeling of the growth and development of the mussel Mytilus galloprovincialis on artificial substrates

    Science.gov (United States)

    Vasechkina, E. F.; Kazankova, I. I.

    2014-11-01

    A mathematical model simulating the growth and development of the mussel Mytilus galloprovincialis Lam. on artificial substrates has been constructed. The model is based on experimental data and contains mathematical descriptions of the filtration, respiration, excretion, spawning, and growth of an individual during its ontogenesis from the moment it attaches to a solid substrate to the attainment of a marketable size. The test computations have been compared to the available observation data for mussel farms.

  14. Identification of SUMO Targets by a Novel Proteomic Approach in Plants

    Institute of Scientific and Technical Information of China (English)

    Gema López-Torrejón; Davide Guerra; Rafael Catalá; Julio Salinas; Juan C.del Pozo

    2013-01-01

    Post-translational modifications (PTMs) chemically and physically alter the properties of proteins,including their folding,subcellular localization,stability,activity,and consequently their function.In spite of their relevance,studies on PTMs in plants are still limited.Small Ubiquitin-like Modifier (SUMO) modification regulates several biological processes by affecting protein-protein interactions,or changing the subcellular localizations of the target proteins.Here,we describe a novel proteomic approach to identify SUMO targets that combines 2-D liquid chromatography,immunodetection,and mass spectrometry (MS) analyses.We have applied this approach to identify nuclear SUMO targets in response to heat shock.Using a bacterial SUMOylation system,we validated that some of the targets identified here are,in fact,labeled with SUMO1.Interestingly,we found that GIGANTEA (GI),a photoperiodic-pathway protein,is modified with SUMO in response to heat shock both in vitro and in vivo.

  15. SUMO-1 Enhancing the p53-induced HepG2 Cell Apoptosis

    Institute of Scientific and Technical Information of China (English)

    LU Xingrong; YI Jilin

    2005-01-01

    Summary: In order to investigate the effect of small ubiquitin-like modifier-1 (SUMO-1) on the p53-induced HepG2 cell apoptosis, HepG2 cells were transfected by recombinant plasmids as pwtp53, pMDM2 and pSUMO-1 respectively. Western blot was employed to detect the protein expression of the transfected recombinant plasmids and the rate of apoptosis was measured by flow cytometry. The results showed that in cells transfected with pwtp53 and pwtp53+pSUMO-1, the apoptosis rate was (16.79±1.62) % and (18.15±1.36) % respectively, while transfected with pwtp53+pMDM2, the rate was decreased to (5.17±1.23) %. The apoptosis rate was (14.06±1.84) % in the cells transfected with pwtp53+pMDM2+pSUMO-1, significantly higher than that in the cells Transfected with pwtp53+pMDM2 (P<0.01). The apoptosis rates in the cells were all less than 2 % and had no significant difference among the groups. It was suggested that in the HepG2 cells, SUMO-1 can increase the apoptosis induced by wild-type p53 through binding to p53 protein, post-translational modification and inhibiting the p53 degradation by MDM2.

  16. Lamin A reassembly at the end of mitosis is regulated by its SUMO-interacting motif.

    Science.gov (United States)

    Moriuchi, Takanobu; Kuroda, Masaki; Kusumoto, Fumiya; Osumi, Takashi; Hirose, Fumiko

    2016-03-01

    Modification of proteins with small ubiquitin-related modifier (SUMO; SUMOylation) is involved in the regulation of various biological processes. Recent studies have demonstrated that noncovalent associations between SUMOylated proteins and co-operative proteins containing SUMO-interacting motifs (SIMs) are important for the spatiotemporal organization of many protein complexes. In this study, we demonstrate that interactions between lamin A, a major component of the nuclear lamina, and SUMO isoforms are dependent on one of the four SIMs (SIM3) resided in lamin A polypeptide in vitro. Live cell imaging and immunofluorescence staining showed that SIM3 is required for accumulation of lamin A on the chromosomes during telophase, and subsequent evaluation of a panel of deletion mutants determined that a 156-amino acid region spanning the carboxyl-terminal Ig-fold domain of lamin A is sufficient for this accumulation. Notably, mutation of SIM3 abrogated the dephosphorylation of mitosis-specific phosphorylation at Ser-22 of lamin A, which normally occurs during telophase, and the subsequent nuclear lamina reorganization. Furthermore, expression of a conjugation-defective SUMO2 mutant, which was previously shown to inhibit endogenous SUMOylation in a dominant-negative manner, also impaired the accumulation of wild type lamin A on telophase chromosomes. These findings suggest that interactions between SIM3 of lamin A and a putative SUMO2-modified protein plays an important role in the reorganization of the nuclear lamina at the end of mitosis.

  17. Global analysis of SUMO chain function reveals multiple roles in chromatin regulation.

    Science.gov (United States)

    Srikumar, Tharan; Lewicki, Megan C; Costanzo, Michael; Tkach, Johnny M; van Bakel, Harm; Tsui, Kyle; Johnson, Erica S; Brown, Grant W; Andrews, Brenda J; Boone, Charles; Giaever, Guri; Nislow, Corey; Raught, Brian

    2013-04-01

    Like ubiquitin, the small ubiquitin-related modifier (SUMO) proteins can form oligomeric "chains," but the biological functions of these superstructures are not well understood. Here, we created mutant yeast strains unable to synthesize SUMO chains (smt3(allR)) and subjected them to high-content microscopic screening, synthetic genetic array (SGA) analysis, and high-density transcript profiling to perform the first global analysis of SUMO chain function. This comprehensive assessment identified 144 proteins with altered localization or intensity in smt3(allR) cells, 149 synthetic genetic interactions, and 225 mRNA transcripts (primarily consisting of stress- and nutrient-response genes) that displayed a >1.5-fold increase in expression levels. This information-rich resource strongly implicates SUMO chains in the regulation of chromatin. Indeed, using several different approaches, we demonstrate that SUMO chains are required for the maintenance of normal higher-order chromatin structure and transcriptional repression of environmental stress response genes in budding yeast.

  18. Production of FMDV virus-like particles by a SUMO fusion protein approach in Escherichia coli

    Directory of Open Access Journals (Sweden)

    Liang Shu-Mei

    2009-08-01

    Full Text Available Abstract Virus-like particles (VLPs are formed by the self-assembly of envelope and/or capsid proteins from many viruses. Some VLPs have been proven successful as vaccines, and others have recently found applications as carriers for foreign antigens or as scaffolds in nanoparticle biotechnology. However, production of VLP was usually impeded due to low water-solubility of recombinant virus capsid proteins. Previous studies revealed that virus capsid and envelope proteins were often posttranslationally modified by SUMO in vivo, leading into a hypothesis that SUMO modification might be a common mechanism for virus proteins to retain water-solubility or prevent improper self-aggregation before virus assembly. We then propose a simple approach to produce VLPs of viruses, e.g., foot-and-mouth disease virus (FMDV. An improved SUMO fusion protein system we developed recently was applied to the simultaneous expression of three capsid proteins of FMDV in E. coli. The three SUMO fusion proteins formed a stable heterotrimeric complex. Proteolytic removal of SUMO moieties from the ternary complexes resulted in VLPs with size and shape resembling the authentic FMDV. The method described here can also apply to produce capsid/envelope protein complexes or VLPs of other disease-causing viruses.

  19. Characterization of a novel posttranslational modification in polypyrimidine tract-binding proteins by SUMO1.

    Science.gov (United States)

    Han, Wei; Wang, Lin; Yin, Bin; Peng, Xiaozhong

    2014-04-01

    Polypyrimidine tract-binding protein 1 (PTBP1) and its brainspecific homologue, PTBP2, are associated with pre-mRNAs and influence pre-mRNA processing, as well as mRNA metabolism and transport. They play important roles in neural differentiation and glioma development. In our study, we detected the expression of the two proteins in glioma cells and predicted that they may be sumoylated using SUMOplot analyses. We confirmed that PTBP1 and PTBP2 can be modified by SUMO1 with co-immunoprecipitation experiments using 293ET cells transiently co-expressing SUMO1 and either PTBP1 or PTBP2. We also found that SUMO1 modification of PTBP2 was enhanced by Ubc9 (E2). The mutation of the sumoylation site (Lys137) of PTBP2 markedly inhibited its modification by SUMO1. Interestingly, in T98G glioma cells, the level of sumoylated PTBP2 was reduced compared to that of normal brain cells. Overall, this study shows that PTBP2 is posttranslationally modified by SUMO1.

  20. Enhanced protein expression in the baculovirus/insect cell system using engineered SUMO fusions.

    Science.gov (United States)

    Liu, Li; Spurrier, Joshua; Butt, Tauseef R; Strickler, James E

    2008-11-01

    Recombinant protein expression in insect cells varies greatly from protein to protein. A fusion tag that is not only a tool for detection and purification, but also enhances expression and/or solubility would greatly facilitate both structure/function studies and therapeutic protein production. We have shown that fusion of SUMO (small ubiquitin-related modifier) to several test proteins leads to enhanced expression levels in Escherichia coli. In eukaryotic expression systems, however, the SUMO tag could be cleaved by endogenous desumoylase. In order to adapt SUMO-fusion technology to these systems, we have developed an alternative SUMO-derived tag, designated SUMOstar, which is not processed by native SUMO proteases. In the present study, we tested the SUMOstar tag in a baculovirus/insect cell system with several proteins, i.e. mouse UBP43, human tryptase beta II, USP4, USP15, and GFP. Our results demonstrate that fusion to SUMOstar enhanced protein expression levels at least 4-fold compared to either the native or His(6)-tagged proteins. We isolated active SUMOstar tagged UBP43, USP4, USP15, and GFP. Tryptase was active following cleavage with a SUMOstar specific protease. The SUMOstar system will make significant impact in difficult-to-express proteins and especially to those proteins that require the native N-terminal residue for function.

  1. Cerebral ischemia/stroke and small ubiquitin-like modifier (SUMO) conjugation--a new target for therapeutic intervention?

    Science.gov (United States)

    Yang, Wei; Sheng, Huaxin; Homi, H Mayumi; Warner, David S; Paschen, Wulf

    2008-08-01

    Transient cerebral ischemia/stroke activates various post-translational protein modifications such as phosphorylation and ubiquitin conjugation that are believed to play a major role in the pathological process triggered by an interruption of blood supply and culminating in cell death. A new system of post-translational protein modification has been identified, termed as small ubiquitin-like modifier (SUMO) conjugation. Like ubiquitin, SUMO is conjugated to the lysine residue of target proteins in a complex process. This review summarizes observations from recent experiments focusing on the effect of cerebral ischemia on SUMO conjugation. Transient global and focal cerebral ischemia both induced a rapid, dramatic and long-lasting rise in levels of SUMO2/3 conjugation. After transient focal cerebral ischemia, SUMO conjugation was particularly prominent in neurons located at the border of the ischemic territory where SUMO-conjugated proteins translocated to the nucleus. Many SUMO conjugation target proteins are transcription factors and sumoylation has been shown to have a major impact on the activity, stability, and cellular localization of target proteins. The rise in levels of SUMO-conjugated proteins is therefore likely to have a major effect on the fate of post-ischemic neurons. The sumoylation process could provide an exciting new target for therapeutic intervention.

  2. The peeling behavior of nanowires and carbon nanotubes from a substrate using continuum modeling

    Science.gov (United States)

    Li, Yue; Xiong, Yan; Zhou, Zhikang; Tang, Bingxian; Yang, Zhaoyao; Zhao, Junhua

    2017-02-01

    The peeling behavior of different nanowires or single-walled/multi-walled carbon nanotubes (CNTs) from a substrate is studied by using the Kendall model of the continuum mechanics, where a basic assumption is that the deformation of the part of the nanowire/nanotube attached to the substrate under peeling force is ignored. The cohesive energy between a nanowire (or a CNT) and a substrate is obtained through continuum modeling of the van der Waals interaction, which has high accuracy by comparison of our molecular dynamics simulations. Our analytical results show that the peeling behavior strongly depends on the peeling angle, the pre-tension, the separation distance toward the substrate, the radius, and the Young's modulus of the nanowire (or the CNT). In particular, the peeling forces with a generalized peeling model in the steady-state stage are compared with those of the classical Kendall model. In the generalized peeling model, the effect of the bending stiffness and cohesive energy between the bending nanowire and the substrate on peeling forces is considered. The obtained analytical solution should be of great help for understanding the interaction between the nanostructures and the substrates, and designing nanoelectromechanical systems.

  3. Modeling the effect of substrate stoichiometry on microbial carbon use efficiency and soil C cycling

    Science.gov (United States)

    Abramoff, R. Z.; Tang, J.; Georgiou, K.; Brodie, E.; Torn, M. S.; Riley, W. J.

    2015-12-01

    Microorganisms degrade soil organic matter (SOM) and apportion newly acquired substrates into enzyme production, biomass growth, and respiration. The fraction of acquired substrate that is released into the atmosphere as heterotrophic respiration is determined by the microbial carbon use efficiency (CUE), commonly defined as the fraction of carbon uptake that is allocated to microbial growth and enzyme production. Despite recent demonstrations that changes in CUE can greatly affect predictions of global soil C stocks, most models do not incorporate process-level representation of CUE or how it varies with substrate stoichiometry. Here we introduce coupled C and N cycling into a prognostic CUE model that uses the dynamic energy budget theory to predict CUE at each time step. We solve this model over a range of substrate C:N to simulate the effects of N addition on CUE, and test the model against previously published measurements of CUE after nutrient enrichment with a range of substrates. We find that CUE declines with microbial N limitation due to C overflow and acquisition strategies that favor N immobilization. We also demonstrate that including an intracellular reserve pool in the model alleviates decreases in CUE by allowing excess C to be stored during periods of N limitation. Consistent with previous studies, we find that predictions of soil C stocks are highly sensitive to CUE. Furthermore, we show that interactive effects between substrate inputs and temperature result in a wide range of possible CUE values under global change scenarios.

  4. A combined BEM/FEM method for IC substrate modeling

    NARCIS (Netherlands)

    Schrik, E.

    2006-01-01

    The research presented in this thesis was done in the context of the modeling of parasitic physical effects, like field couplings and crosstalk, that may adversely affect the functional performance of Integrated Circuits (ICs). The modeling of parasitic effects, through simulations with the resultin

  5. The effect of substrate concentration on biohydrogen production by using kinetic models

    Institute of Scientific and Technical Information of China (English)

    WANG JianLong; WAN Wei

    2008-01-01

    The effect of substrate concentration ranging from 0 to 300 g/L on fermentative hydrogen production by mixed cultures was investigated in batch tests using glucose as substrate. The experimental results showed that, at 35℃ and initial pH 7.0, during the fermentative hydrogen production, the hydrogen production potential and hydrogen production rate increased with increasing substrate concentration from 0 to 25 g/L. The maximal hydrogen production potential of 426.8 mL and maximal hydrogen pro-duction rate of 15.1 mlJh were obtained at the substrate concentration of 25 g/L. The maximal hydrogen yield and the maximal substrate degradation efficiency were respectively 384.3 mL/g glucose and 97.6%, at the substrate concentration of 2 g/L. The modified Logistic model could be used to describe the progress of cumulative hydrogen production in this study successfully. The Han-Levenspiel model could be used to describe the effect of substrate concentration on fermentative hydrogen production rate.

  6. The effect of substrate concentration on biohydrogen production by using kinetic models

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    The effect of substrate concentration ranging from 0 to 300 g/L on fermentative hydrogen production by mixed cultures was investigated in batch tests using glucose as substrate. The experimental results showed that, at 35℃ and initial pH 7.0, during the fermentative hydrogen production, the hydrogen production potential and hydrogen production rate increased with increasing substrate concentration from 0 to 25 g/L. The maximal hydrogen production potential of 426.8 mL and maximal hydrogen pro-duction rate of 15.1 mL/h were obtained at the substrate concentration of 25 g/L. The maximal hydrogen yield and the maximal substrate degradation efficiency were respectively 384.3 mL/g glucose and 97.6%, at the substrate concentration of 2 g/L. The modified Logistic model could be used to describe the progress of cumulative hydrogen production in this study successfully. The Han-Levenspiel model could be used to describe the effect of substrate concentration on fermentative hydrogen production rate.

  7. Proteome-wide analysis of SUMO2 targets in response to pathological DNA replication stress in human cells.

    Science.gov (United States)

    Bursomanno, Sara; Beli, Petra; Khan, Asif M; Minocherhomji, Sheroy; Wagner, Sebastian A; Bekker-Jensen, Simon; Mailand, Niels; Choudhary, Chunaram; Hickson, Ian D; Liu, Ying

    2015-01-01

    SUMOylation is a form of post-translational modification involving covalent attachment of SUMO (Small Ubiquitin-like Modifier) polypeptides to specific lysine residues in the target protein. In human cells, there are four SUMO proteins, SUMO1-4, with SUMO2 and SUMO3 forming a closely related subfamily. SUMO2/3, in contrast to SUMO1, are predominantly involved in the cellular response to certain stresses, including heat shock. Substantial evidence from studies in yeast has shown that SUMOylation plays an important role in the regulation of DNA replication and repair. Here, we report a proteomic analysis of proteins modified by SUMO2 in response to DNA replication stress in S phase in human cells. We have identified a panel of 22 SUMO2 targets with increased SUMOylation during DNA replication stress, many of which play key functions within the DNA replication machinery and/or in the cellular response to DNA damage. Interestingly, POLD3 was found modified most significantly in response to a low dose aphidicolin treatment protocol that promotes common fragile site (CFS) breakage. POLD3 is the human ortholog of POL32 in budding yeast, and has been shown to act during break-induced recombinational repair. We have also shown that deficiency of POLD3 leads to an increase in RPA-bound ssDNA when cells are under replication stress, suggesting that POLD3 plays a role in the cellular response to DNA replication stress. Considering that DNA replication stress is a source of genome instability, and that excessive replication stress is a hallmark of pre-neoplastic and tumor cells, our characterization of SUMO2 targets during a perturbed S-phase should provide a valuable resource for future functional studies in the fields of DNA metabolism and cancer biology.

  8. Kinetic models of cell growth, substrate utilization and bio ...

    African Journals Online (AJOL)

    STORAGESEVER

    2008-05-02

    May 2, 2008 ... A simple model was proposed using the Logistic Equation for the growth,. Leudeking-Piret ... (melanoidin) which may create many problems and also .... Where, the constant µ is defined as the specific growth rate. Equation 1 ...

  9. The identification of new substrates of human DHRS7 by molecular modeling and in vitro testing.

    Science.gov (United States)

    Zemanová, Lucie; Kirubakaran, Palani; Pato, Ignacio Hernando; Štambergová, Hana; Vondrášek, Jiří

    2017-07-04

    Human DHRS7 (SDR34C1) is one of insufficiently described enzymes of the short-chain dehydrogenase/reductase superfamily. The members of this superfamily often play an important pato/physiological role in the human body, participating in the metabolism of diverse substrates (e.g. retinoids, steroids, xenobiotics). A systematic approach to the identification of novel, physiological substrates of DHRS7 based on a combination of homology modeling, structure-based virtual screening and experimental evaluation has been used. Three novel substrates of DHRS7 (dihydrotestosterone, benzil and 4,4'-dimetylbenzil) have been described. Copyright © 2017 Elsevier B.V. All rights reserved.

  10. Characterisation of the SUMO-like domains of Schizosaccharomyces pombe Rad60.

    Directory of Open Access Journals (Sweden)

    Lara K Boyd

    Full Text Available The S. pombe Rad60 protein is required for the repair of DNA double strand breaks, recovery from replication arrest, and is essential for cell viability. It has two SUMO-like domains (SLDs at its C-terminus, an SXS motif and three sequences that have been proposed to be SUMO-binding motifs (SBMs. SMB1 is located in the middle of the protein, SBM2 is in SLD1 and SBM3 is at the C-terminus of SLD2. We have probed the functions of the two SUMO-like domains, SLD1 and SLD2, and the putative SBMs. SLD1 is essential for viability, while SLD2 is not. rad60-SLD2Δ cells are sensitive to DNA damaging agents and hydroxyurea. Neither ubiquitin nor SUMO can replace SLD1 or SLD2. Cells in which either SBM1 or SBM2 has been mutated are viable and are wild type for response to MMS and HU. In contrast mutation of SBM3 results in significant sensitivity to MMS and HU. These results indicate that the lethality resulting from deletion of SLD1 is not due to loss of SBM2, but that mutation of SBM3 produces a more severe phenotype than does deletion of SLD2. Using chemical denaturation studies, FPLC and dynamic light scattering we show this is likely due to the destabilisation of SLD2. Thus we propose that the region corresponding to the putative SBM3 forms part of the hydrophobic core of SLD2 and is not a SUMO-interacting motif. Over-expression of Hus5, which is the SUMO conjugating enzyme and known to interact with Rad60, does not rescue rad60-SLD2Δ, implying that as well as having a role in the sumoylation process as previously described, Rad60 has a Hus5-independent function.

  11. Enhanced in vitro refolding of fibroblast growth factor 15 with the assistance of SUMO fusion partner.

    Directory of Open Access Journals (Sweden)

    Bo Kong

    Full Text Available Fibroblast growth factor 15 (Fgf15 is the mouse orthologue of human FGF19. Fgf15 is highly expressed in the ileum and functions as an endocrine signal to regulate liver function, including bile acid synthesis, hepatocyte proliferation and insulin sensitivity. In order to fully understand the function of Fgf15, methods are needed to produce pure Fgf15 protein in the prokaryotic system. However, when expressed in Escherichia coli (E. coli, the recombinant Fgf15 protein was insoluble and found only in inclusion bodies. In the current study, we report a method to produce recombinant Fgf15 protein in E. coli through the use of small ubiquitin-related modifier (SUMO fusion tag. Even though the SUMO has been shown to strongly improve protein solubility and expression levels, our studies suggest that the SUMO does not improve Fgf15 protein solubility. Instead, proper refolding of Fgf15 protein was achieved when Fgf15 was expressed as a partner protein of the fusion tag SUMO, followed by in vitro dialysis refolding. After refolding, the N-terminal SUMO tag was cleaved from the recombinant Fgf15 fusion protein by ScUlp1 (Ubiquitin-Like Protein-Specific Protease 1 from S. cerevisiae. With or without the SUMO tag, the refolded Fgf15 protein was biologically active, as revealed by its ability to reduce hepatic Cyp7a1 mRNA levels in mice. In addition, recombinant Fgf15 protein suppressed Cyp7a1 mRNA levels in a dose-dependent manner. In summary, we have developed a successful method to express functional Fgf15 protein in prokaryotic cells.

  12. Kinetics Modelling of the Biodegradation of Benzene, Toluene and Phenol as Single Substrate and Mixed Substrate by Using Pseudomonas putida

    OpenAIRE

    Mathur, A K; Majumder, C. B.

    2010-01-01

    In the present work, kinetics of the biodegradation of benzene, toluene and phenol by using a pure culture of Pseudomonas putida (MTCC 1194) was determined by measuring the specific growth rate and degradation rate with substrate concentration as a function of time in a batch reactor. In general, the degradation rate of benzene, toluene and phenol increased with the increase in the initial substrate concentration and then decreased after reaching a maximum, showing substrate inhibition kineti...

  13. Anterior mediastinal abscess diagnosed in a young sumo wrestler after closed blunt chest trauma

    Institute of Scientific and Technical Information of China (English)

    Tatsuro Sassa; Ken-ichiro Kobayashi; Masayuki Ota; Takuya Washino; Mayu Hikone; Naoya Sakamoto; Sentaro Iwabuchi

    2015-01-01

    Most mediastinal abscesses result from infections after thoracotomy, esophageal perforation or penetrating chest trauma.This disease is rarely caused by closed blunt chest trauma.All previously reported such cases after closed blunt chest trauma presented with hematoma and sternal osteomyelitis resulting from sternal fracture.Here we report a 15-year-old sumo wrestler who presented with an anterior mediastinal abscess without any mediastinal fracture.The mediastinal abscess resulted from the hematogenous spread of Staphylococcus aureus to a hematoma that might have been caused by a closed blunt chest trauma incurred during sumo wrestling exercises.

  14. Hard Spherocylinders of Two Different Lengths as a Model System of a Nematic Liquid Crystal on an Anisotropic Substrate

    Science.gov (United States)

    Koda, Tomonori; Hyodo, Yosuke; Momoi, Yuichi; Kwak, Musun; Kang, Dongwoo; Choi, Youngseok; Nishioka, Akihiro; Haba, Osamu; Yonetake, Koichiro

    2016-02-01

    In this article, we describe the effects of an anisotropic substrate on the alignment of a nematic liquid crystal. We examine how the substrate affects the alignment of a nematic liquid crystal by Monte Carlo simulation. The liquid crystal on a substrate was described by the phase separation of liquid crystal molecules and substrate molecules, both of which were modeled by hard particles. We used hard rods to represent both the liquid crystal and the substrate. The length of the hard rods representing the substrate was adjusted to represent the degree of substrate anisotropy. The results show that the nematic alignment could either be reinforced or weakened, depending on the length of the substrate rods. Mean field theory is used to analyze the simulation results. We confirmed that the distance over which the substrate affects the bulk liquid crystal is about 3 nm for the present hard-rod-based model.

  15. Substrate turnover at low carbon concentrations in a model drinking water distribution system

    DEFF Research Database (Denmark)

    Boe-Hansen, Rasmus; Albrechtsen, Hans-Jørgen; Arvin, Erik;

    2002-01-01

    utilisation and bacterial growth at low nutrient conditions in a model distribution system. The model system consisted of two loops in series, where flow rate and retention time were controlled independently. Spiking the drinking water of the model system with two different environmentally realistic......Water quality changes caused by microbial activity in the distribution network can cause serious problems. Reducing the amount of microbial available substrate may be an effective way to control bacterial aftergrowth. The purpose of the present study was to study the kinetics of substrate...

  16. Modeling of the Bacillus subtilis Bacterial Biofilm Growing on an Agar Substrate

    Directory of Open Access Journals (Sweden)

    Xiaoling Wang

    2015-01-01

    Full Text Available Bacterial biofilms are organized communities composed of millions of microorganisms that accumulate on almost any kinds of surfaces. In this paper, a biofilm growth model on an agar substrate is developed based on mass conservation principles, Fick’s first law, and Monod’s kinetic reaction, by considering nutrient diffusion between biofilm and agar substrate. Our results show biofilm growth evolution characteristics such as biofilm thickness, active biomass, and nutrient concentration in the agar substrate. We quantitatively obtain biofilm growth dependence on different parameters. We provide an alternative mathematical method to describe other kinds of biofilm growth such as multiple bacterial species biofilm and also biofilm growth on various complex substrates.

  17. Capturing a Dynamic Chaperone-Substrate Interaction Using NMR-Informed Molecular Modeling.

    Science.gov (United States)

    Salmon, Loïc; Ahlstrom, Logan S; Horowitz, Scott; Dickson, Alex; Brooks, Charles L; Bardwell, James C A

    2016-08-10

    Chaperones maintain a healthy proteome by preventing aggregation and by aiding in protein folding. Precisely how chaperones influence the conformational properties of their substrates, however, remains unclear. To achieve a detailed description of dynamic chaperone-substrate interactions, we fused site-specific NMR information with coarse-grained simulations. Our model system is the binding and folding of a chaperone substrate, immunity protein 7 (Im7), with the chaperone Spy. We first used an automated procedure in which NMR chemical shifts inform the construction of system-specific force fields that describe each partner individually. The models of the two binding partners are then combined to perform simulations on the chaperone-substrate complex. The binding simulations show excellent agreement with experimental data from multiple biophysical measurements. Upon binding, Im7 interacts with a mixture of hydrophobic and hydrophilic residues on Spy's surface, causing conformational exchange within Im7 to slow down as Im7 folds. Meanwhile, the motion of Spy's flexible loop region increases, allowing for better interaction with different substrate conformations, and helping offset losses in Im7 conformational dynamics that occur upon binding and folding. Spy then preferentially releases Im7 into a well-folded state. Our strategy has enabled a residue-level description of a dynamic chaperone-substrate interaction, improving our understanding of how chaperones facilitate substrate folding. More broadly, we validate our approach using two other binding partners, showing that this approach provides a general platform from which to investigate other flexible biomolecular complexes through the integration of NMR data with efficient computational models.

  18. Modeling of air-droplet interaction, substrate melting and coating buildup in thermal spraying

    Science.gov (United States)

    Wei, Guanghua

    Among the many surface coating techniques now available, thermal spray is known to offer the most advantages. It can meet a wide range of technical and engineering requirements in a relatively inexpensive and easily controllable way with the capability of producing repeatable results. In the last few decades a lot of important strides have been made in the field of measurements and modelling of thermal spraying. However, due to the complex of the process and the lack of basic materials-based knowledge about the particle melting, spreading and deposition, the relationship between the process parameters and the coating properties still remains unclear. In thermal spraying, a particle is melted to form a droplet with morphology and thermal- and kinetic-energy status change by the interaction with the plasma/flame. In order to produce higher-quality coatings and expand the use of this versatile family of technologies, modelling of the particle behaviors during in-flight, spreading and deposition is essential. This thesis investigates the connections between particle characteristics and coating properties. Momentum, heat and mass transfer phenomena related to particle in-flight, droplet impacting, spreading, and splat layering are studied. Numerical models are developed to establish the quantitative relationships between spray parameters, particle and substrate properties and deposition characteristics. Most existing theoretical studies of in-flight particle assume that the particle is in a spherical shape without voids inside. The behavior of porous particles in thermal spray has not been well understood. However, the presence of voids in the feedstock powders may have a great impact on particle in-flight behaviors such as particle acceleration, melting and oxidation because a hollowed particle is also lighter than a densed one and this will affect the particle trajectory. The particle shape also needs to be taken into account because it influences the drag force and

  19. Redox regulation of SUMO enzymes is required for ATM activity and survival in oxidative stress.

    Science.gov (United States)

    Stankovic-Valentin, Nicolas; Drzewicka, Katarzyna; König, Cornelia; Schiebel, Elmar; Melchior, Frauke

    2016-06-15

    To sense and defend against oxidative stress, cells depend on signal transduction cascades involving redox-sensitive proteins. We previously identified SUMO (small ubiquitin-related modifier) enzymes as downstream effectors of reactive oxygen species (ROS). Hydrogen peroxide transiently inactivates SUMO E1 and E2 enzymes by inducing a disulfide bond between their catalytic cysteines. How important their oxidation is in light of many other redox-regulated proteins has however been unclear. To selectively disrupt this redox switch, we identified a catalytically fully active SUMO E2 enzyme variant (Ubc9 D100A) with strongly reduced propensity to maintain a disulfide with the E1 enzyme in vitro and in cells. Replacement of Ubc9 by this variant impairs cell survival both under acute and mild chronic oxidative stresses. Intriguingly, Ubc9 D100A cells fail to maintain activity of the ATM-Chk2 DNA damage response pathway that is induced by hydrogen peroxide. In line with this, these cells are also more sensitive to the ROS-producing chemotherapeutic drugs etoposide/Vp16 and Ara-C. These findings reveal that SUMO E1~E2 oxidation is an essential redox switch in oxidative stress.

  20. An Arabidopsis SUMO E3 Ligase, SIZ1, Negatively Regulates Photomorphogenesis by Promoting COP1 Activity

    KAUST Repository

    Lin, Xiao-Li

    2016-04-29

    COP1 (CONSTITUTIVE PHOTOMORPHOGENIC 1), a ubiquitin E3 ligase, is a central negative regulator of photomorphogenesis. However, how COP1 activity is regulated by post-translational modifications remains largely unknown. Here we show that SUMO (small ubiquitin-like modifier) modification enhances COP1 activity. Loss-of-function siz1 mutant seedlings exhibit a weak constitutive photomorphogenic phenotype. SIZ1 physically interacts with COP1 and mediates the sumoylation of COP1. A K193R substitution in COP1 blocks its SUMO modification and reduces COP1 activity in vitro and in planta. Consistently, COP1 activity is reduced in siz1 and the level of HY5, a COP1 target protein, is increased in siz1. Sumoylated COP1 may exhibits higher transubiquitination activity than does non-sumoylated COP1, but SIZ1-mediated SUMO modification does not affect COP1 dimerization, COP1-HY5 interaction, and nuclear accumulation of COP1. Interestingly, prolonged light exposure reduces the sumoylation level of COP1, and COP1 mediates the ubiquitination and degradation of SIZ1. These regulatory mechanisms may maintain the homeostasis of COP1 activity, ensuing proper photomorphogenic development in changing light environment. Our genetic and biochemical studies identify a function for SIZ1 in photomorphogenesis and reveal a novel SUMO-regulated ubiquitin ligase, COP1, in plants.

  1. Analysis of Changes in SUMO-2/3 Modification during Breast Cancer Progression and Metastasis

    DEFF Research Database (Denmark)

    Subramonian, Divya; Raghunayakula, Sarita; Olsen, Jesper V;

    2014-01-01

    SUMOylation is an essential posttranslational modification and regulates many cellular processes. Dysregulation of SUMOylation plays a critical role in metastasis, yet how its perturbation affects this lethal process of cancer is not well understood. We found that SUMO-2/3 modification is greatly...

  2. Functional expression and purification of recombinant Hepcidin25 production in Escherichia coli using SUMO fusion technology.

    Science.gov (United States)

    Sadr, Vahideh; Saffar, Behnaz; Emamzadeh, Rahman

    2017-04-30

    Hepcidin25 is a small cysteine-rich peptide hormone known as a new class of antimicrobial peptides. The purpose of the present study was to express, purify and investigate the antibacterial properties of recombinant human hepcidin25 protein production in Escherichia coli. Human hepcidin25 gene was optimized and fused to a small ubiquitin-related modifier (SUMO) gene for higher expression. Then SUMO-hepcidin25 was cloned into the pET-32a (+) vector and expressed in E. coli Origami. The fusion protein with a molecular weight of approximately 35kDa was analyzed on SDS-PAGE gel. The highest expression was observed after 6h induction and the fusion protein consisted approximately 47% of the total cellular protein. The purified SUMO-hepcidin25 purity was determined to be higher than 95%, with a final yield of 3.9mgl(-)(1) of media. The recombinant hepcidin25 showed antibacterial activity against both Gram negative (Klebsiella pneumonia) and Gram positive (Staphylococcus aureus and Bacillus cereus) bacteria with minimum inhibitory concentrations (MICs) of 150μgml(-1), 18.7μg/ml(-1) and 37.5μg/ml(-1), respectively. These results indicated that thioredoxin and SUMO dual fusion system is an efficient production system for synthesis functional human hepcidin25.

  3. Study of MDM2 and SUMO-1 expression in actinic cheilitis and lip cancer.

    Science.gov (United States)

    Oliveira Alves, Mônica Ghislaine; da Mota Delgado, Adriana; Balducci, Ivan; Carvalho, Yasmin Rodarte; Cavalcante, Ana Sueli Rodrigues; Almeida, Janete Dias

    2014-11-01

    Actinic cheilitis exhibits a potential of malignant transformation in 10-20 % of cases. The objective of this study was to compare the expression of MDM2 and SUMO-1 proteins between actinic cheilitis (AC) and squamous cell carcinoma (SCC) of the lip. The sample consisted of lower lip mucosa specimens obtained from cases with a clinical and histopathological diagnosis of AC (n = 26) and SCC (n = 25) and specimens of labial semi-mucosa (n = 15) without clinical alterations or inflammation. The tissue samples were stained with hematoxylin-eosin and anti-MDM2 and anti-SUMO-1 antibodies. Data were analyzed by the Kruskal-Wallis and Dunn's tests (5 %). The median expression of MDM2 (kW = 36.8565; df = 3-1 = 2; p = 0.0001) and SUMO-1 (kW = 32.7080; df = 3-1 = 2; p = 0.0001) was similar in cases of AC and SCC of the lip, but differed significantly from that observed for normal labial semi-mucosa. Despite the limitations of the present study, immunohistochemistry demonstrated the overexpression of important proteins (MDM2 and SUMO-1) related to regulatory mechanisms of apoptosis in AC and SCC of the lip, but further studies are needed.

  4. Modeling of the rough spherical nanoparticles manipulation on a substrate based on the AFM nanorobot

    Science.gov (United States)

    Zakeri, M.; Faraji, J.

    2014-12-01

    In this paper, dynamic behavior of the rough spherical micro/nanoparticles during pulling/pushing on the flat substrate has been investigated and analyzed. For this purpose, at first, two hexagonal roughness models (George and Cooper) were studied and then evaluations for adhesion force were determined for rough particle manipulation on flat substrate. These two models were then changed by using of the Rabinovich theory. Evaluations were determined for contact adhesion force between rough particle and flat substrate; depth of penetration evaluations were determined by the Johnson-Kendall-Roberts contact mechanic theory and the Schwartz method and according to Cooper and George roughness models. Then, the novel contact theory was used to determine a dynamic model for rough micro/nanoparticle manipulation on flat substrate. Finally, simulation of particle dynamic behavior was implemented during pushing of rough spherical gold particles with radii of 50, 150, 400, 600, and 1,000 nm. Results derived from simulations of particles with several rates of roughness on flat substrate indicated that compared to results for flat particles, inherent roughness on particles might reduce the rate of critical force needed for sliding and rolling given particles. Given a fixed radius for roughness value and increased roughness height, evaluations for sliding and rolling critical forces showed greater reduction. Alternately, the rate of critical force was shown to reduce relative to an increased roughness radius. With respect to both models, based on the George roughness model, the predicted rate of adhesion force was greater than that determined in the Cooper roughness model, and as a result, the predicted rate of critical force based on the George roughness model was closer to the critical force value of flat particle.

  5. Experimental Results and Integrated Modeling of Bacterial Growth on an Insoluble Hydrophobic Substrate (Phenanthrene)

    DEFF Research Database (Denmark)

    Adam, Iris K. U.; Rein, Arno; Miltner, Anja

    2014-01-01

    was fitted to the test results for the rates of dissolution, metabolism, and growth. The strains showed similar efficiency, with v(max) values of 12-18 g dw g(-1) d(-1), yields of 0.21 g g(-1), maximum growth rates of 2.5-3.8 d(-1), and decay rates of 0.04-0.05 d(-1). Sensitivity analysis with the model......Metabolism of a low-solubility substrate is limited by dissolution and availability and can hardly be determined. We developed a numerical model for simultaneously calculating dissolution kinetics of such substrates and their metabolism and microbial growth (Monod kinetics with decay) and tested...... shows that (i) retention in crystals or NAPLs or by sequestration competes with biodegradation, (ii) bacterial growth conditions (dissolution flux and resulting chemical activity of substrate) are more relevant for the final state of the system than the initial biomass, and (iii) the desorption flux...

  6. Two-dimensional modeling of water and heat fluxes in green roof substrates

    Science.gov (United States)

    Suarez, F. I.; Sandoval, V. P.

    2016-12-01

    Due to public concern towards sustainable development, greenhouse gas emissions and energy efficiency, green roofs have become popular in the last years. Green roofs integrate vegetation into infrastructures to reach additional benefits that minimize negative impacts of the urbanization. A properly designed green roof can reduce environmental pollution, noise levels, energetic requirements or surface runoff. The correct performance of green roofs depends on site-specific conditions and on each component of the roof. The substrate and the vegetation layers strongly influence water and heat fluxes on a green roof. The substrate is an artificial media that has an improved performance compared to natural soils as it provides critical resources for vegetation survival: water, nutrients, and a growing media. Hence, it is important to study the effects of substrate properties on green roof performance. The objective of this work is to investigate how the thermal and hydraulic properties affect the behavior of a green roof through numerical modeling. The substrates that were investigated are composed by: crushed bricks and organic soil (S1); peat with perlite (S2); crushed bricks (S3); mineral soil with tree leaves (S4); and a mixture of topsoil and mineral soil (S5). The numerical model utilizes summer-arid meteorological information to evaluate the performance of each substrate. Results show that the area below the water retention curve helps to define the substrate that retains more water. In addition, the non-linearity of the water retention curve can increment the water needed to irrigate the roof. The heat propagation through the roof depends strongly on the hydraulic behavior, meaning that a combination of a substrate with low thermal conductivity and more porosity can reduce the heat fluxes across the roof. Therefore, it can minimize the energy consumed of an air-conditioner system.

  7. Advantages of geographically weighted regression for modeling benthic substrate in two Greater Yellowstone Ecosystem streams

    Science.gov (United States)

    Sheehan, Kenneth R.; Strager, Michael P.; Welsh, Stuart

    2013-01-01

    Stream habitat assessments are commonplace in fish management, and often involve nonspatial analysis methods for quantifying or predicting habitat, such as ordinary least squares regression (OLS). Spatial relationships, however, often exist among stream habitat variables. For example, water depth, water velocity, and benthic substrate sizes within streams are often spatially correlated and may exhibit spatial nonstationarity or inconsistency in geographic space. Thus, analysis methods should address spatial relationships within habitat datasets. In this study, OLS and a recently developed method, geographically weighted regression (GWR), were used to model benthic substrate from water depth and water velocity data at two stream sites within the Greater Yellowstone Ecosystem. For data collection, each site was represented by a grid of 0.1 m2 cells, where actual values of water depth, water velocity, and benthic substrate class were measured for each cell. Accuracies of regressed substrate class data by OLS and GWR methods were calculated by comparing maps, parameter estimates, and determination coefficient r 2. For analysis of data from both sites, Akaike’s Information Criterion corrected for sample size indicated the best approximating model for the data resulted from GWR and not from OLS. Adjusted r 2 values also supported GWR as a better approach than OLS for prediction of substrate. This study supports GWR (a spatial analysis approach) over nonspatial OLS methods for prediction of habitat for stream habitat assessments.

  8. El sumo bien kantiano: el objeto construído de la ley moral

    Directory of Open Access Journals (Sweden)

    Alonso Villarán

    2016-01-01

    Full Text Available El presente artículo tiene tres objetivos: a. compilar las críticas planteadas en contra del sumo bien kantiano (un mundo donde la virtud es premiada con felicidad; b. elaborar una revisión crítica de las respectivas defensas; y c. desarrollar una nueva defensa, con énfasis en el problema de derivación (según el cual el sumo bien no se deriva de la ley moral. Respecto al primero punto, el artículo muestra que son al menos cinco los problemas que amenazan la doctrina: heteronomía, derivación, imposibilidad, irrelevancia y dualismo. Respecto al segundo, el artículo revela que las múltiples defensas elaboradas en nombre del sumo bien son fuente, a su vez, de otros problemas, como los de incompletitud, ilusión, injusticia, deslealtad y escisión. Respecto al tercer y principal punto, y empezando por la derivación, el artículo propone ver al sumo bien como el objeto construido de la ley moral. Resuelto el problema de derivación, el artículo ofrece breves respuestas a los otros problemas en juego. El artículo también traduce por primera vez, del inglés al español, fragmentos de importantes publicaciones sobre el sumo bien.

  9. How can cells sense the elasticity of a substrate? An analysis using a cell tensegrity model

    Directory of Open Access Journals (Sweden)

    G De Santis

    2011-10-01

    Full Text Available A eukaryotic cell attaches and spreads on substrates, whether it is the extracellular matrix naturally produced by the cell itself, or artificial materials, such as tissue-engineered scaffolds. Attachment and spreading require the cell to apply forces in the nN range to the substrate via adhesion sites, and these forces are balanced by the elastic response of the substrate. This mechanical interaction is one determinant of cell morphology and, ultimately, cell phenotype. In this paper we use a finite element model of a cell, with a tensegrity structure to model the cytoskeleton of actin filaments and microtubules, to explore the way cells sense the stiffness of the substrate and thereby adapt to it. To support the computational results, an analytical 1D model is developed for comparison. We find that (i the tensegrity hypothesis of the cytoskeleton is sufficient to explain the matrix-elasticity sensing, (ii cell sensitivity is not constant but has a bell-shaped distribution over the physiological matrix-elasticity range, and (iii the position of the sensitivity peak over the matrix-elasticity range depends on the cytoskeletal structure and in particular on the F-actin organisation. Our model suggests that F-actin reorganisation observed in mesenchymal stem cells (MSCs in response to change of matrix elasticity is a structural-remodelling process that shifts the sensitivity peak towards the new value of matrix elasticity. This finding discloses a potential regulatory role of scaffold stiffness for cell differentiation.

  10. Small- and large-signal modeling of InP HBTs in transferred-substrate technology

    DEFF Research Database (Denmark)

    Johansen, Tom Keinicke; Rudolph, Matthias; Jensen, Thomas

    2014-01-01

    In this paper, the small- and large-signal modeling of InP heterojunction bipolar transistors (HBTs) in transferred substrate (TS) technology is investigated. The small-signal equivalent circuit parameters for TS-HBTs in two-terminal and three-terminal configurations are determined by employing...

  11. A two-substrate Michaelis-Menten model for the growth of self-replicating polymers.

    Science.gov (United States)

    Ferreira, R

    1987-10-07

    A two-substrate Michaelis-Menten model is proposed for the growth of autocatalytic self-replicating polymers. Selective growth depends on the existence of two complementary pairs of monomers. Discrimination among sequences results from different products of binding constants, KCGnKAUm. The results support an earlier renormalization group treatment (Ferreira & Tsallis, 1985).

  12. Modeling of InP HBTs in Transferred-Substrate Technology for Millimeter-Wave Applications

    DEFF Research Database (Denmark)

    Johansen, Tom Keinicke; Rudolph, Matthias; Jensen, Thomas;

    2013-01-01

    In this paper, the modeling of InP heterojunction bipolar transistors (HBTs) in transferred substrate (TS) technology is investigated. At first, a direct parameter extraction methodology dedicated to III-V based HBTs is employed to determine the small-signal equivalent circuit parameters from...

  13. Comparative modeling of the human monoamine transporters: similarities in substrate binding.

    Science.gov (United States)

    Koldsø, Heidi; Christiansen, Anja B; Sinning, Steffen; Schiøtt, Birgit

    2013-02-20

    The amino acid compositions of the substrate binding pockets of the three human monoamine transporters are compared as is the orientation of the endogenous substrates, serotonin, dopamine, and norepinephrine, bound in these. Through a combination of homology modeling, induced fit dockings, molecular dynamics simulations, and uptake experiments in mutant transporters, we propose a common binding mode for the three substrates. The longitudinal axis of the substrates is similarly oriented with these, forming an ionic interaction between the ammonium group and a highly conserved aspartate, Asp98 (serotonin transporter, hSERT), Asp79 (dopamine transporter, hDAT), and Asp75 (norepinephrine transporter, hNET). The 6-position of serotonin and the para-hydroxyl groups of dopamine and norepinephrine were found to face Ala173 in hSERT, Gly153 in hDAT, and Gly149 in hNET. Three rotations of the substrates around the longitudinal axis were identified. In each mode, an aromatic hydroxyl group of the substrates occupied equivalent volumes of the three binding pockets, where small changes in amino acid composition explains the differences in selectivity. Uptake experiments support that the 5-hydroxyl group of serotonin and the meta-hydroxyl group norepinephrine and dopamine are placed in the hydrophilic pocket around Ala173, Ser438, and Thr439 in hSERT corresponding to Gly149, Ser419, Ser420 in hNET and Gly153 Ser422 and Ala423 in hDAT. Furthermore, hDAT was found to possess an additional hydrophilic pocket around Ser149 to accommodate the para-hydroxyl group. Understanding these subtle differences between the binding site compositions of the three transporters is imperative for understanding the substrate selectivity, which could eventually aid in developing future selective medicines.

  14. Global (Q)SAR models on substrates for human Cytochrome P450 3A4

    DEFF Research Database (Denmark)

    Ringsted, Tine; Nikolov, Nikolai Georgiev; Wedebye, Eva Bay;

    The Cytochrome P450 (CYP) is a superfamily of enzymes which catalyze the metabolism of a wide range of endobiotics and xenobiotics. The latter category comprises drugs and about 75% of marketed drugs are metabolised by CYP enzymes. Besides drugs, CYP enzymes detoxify environmental compounds...... domain. Domain coverage of EINECS chemicals and number of predicted substrates are discussed. Reference: C.W. Yap and Y.Z. Chen, Prediction of cytochrome p450 3A4, 2D6, and 2C9 inhibitors and substrates by using support vector machines, J. Chem. Inf. Model. 45 (2005), pp. 982–992....... but paradoxically they also have the ability to form reactive intermediates which can damage DNA, lipids and proteins. It is therefore important to gain knowledge on which substrates that can potentially be metabolised by CYP. The CYP 3A4 isoenzyme plays a dominant role by the metabolic elimination of up to 35...

  15. Modeling and Design Guidelines for P⁺ Guard Rings in Lightly Doped CMOS Substrates

    DEFF Research Database (Denmark)

    Shen, Ming; Mikkelsen, Jan H.; Zhang, Ke;

    2013-01-01

    This paper presents a compact model for ${rm P}^{+}$ guard rings in lightly doped CMOS substrates featuring a P-well layer. Simple expressions for the impedances in the model are derived based on a conformal mapping approach. The model can be used to predict the noise suppression performance...... of ${rm P}^{+}$ guard rings in terms of S-parameters, which is useful for substrate noise mitigation in mixed-signal system-on-chips. Validation of the model has been done by both electromagnetic simulation and experimental results from guard rings implemented using a standard 0.18-$mu{rm m}$ CMOS process....... In addition, design guidelines have been drawn for minimizing the guard ring size while maintaining the noise suppression performance....

  16. Molecular Modeling of Peroxidase and Polyphenol Oxidase: Substrate Specificity and Active Site Comparison

    Directory of Open Access Journals (Sweden)

    Lalida Shank

    2010-09-01

    Full Text Available Peroxidases (POD and polyphenol oxidase (PPO are enzymes that are well known to be involved in the enzymatic browning reaction of fruits and vegetables with different catalytic mechanisms. Both enzymes have some common substrates, but each also has its specific substrates. In our computational study, the amino acid sequence of grape peroxidase (ABX was used for the construction of models employing homology modeling method based on the X-ray structure of cytosolic ascorbate peroxidase from pea (PDB ID:1APX, whereas the model of grape polyphenol oxidase was obtained directly from the available X-ray structure (PDB ID:2P3X. Molecular docking of common substrates of these two enzymes was subsequently studied. It was found that epicatechin and catechin exhibited high affinity with both enzymes, even though POD and PPO have different binding pockets regarding the size and the key amino acids involved in binding. Predicted binding modes of substrates with both enzymes were also compared. The calculated docking interaction energy of trihydroxybenzoic acid related compounds shows high affinity, suggesting specificity and potential use as common inhibitor to grape ascorbate peroxidase and polyphenol oxidase.

  17. Modeling the microbial growth of two Escherichia coli strains in a multi-substrate environment

    OpenAIRE

    Poccia,M. E.; Beccaria, A. J.; R. G. Dondo

    2014-01-01

    The microbial growth in multi-substrate environments may be viewed as an optimal resources allocation problem. The optimization aims at maximizing some biological objective like the biomass growth. The models developed using this hypothesis are called “cybernetic” and they represent the complex cell structure as an optimizing function that regulates the intracellular enzymatic machinery. In this work, a cybernetic model was developed to represent the growth of two E. coli strains (JM 109 and ...

  18. SUMO-1 Promotes Ishikawa Cell Proliferation and Apoptosis in Endometrial Cancer by Increasing Sumoylation of Histone H4.

    Science.gov (United States)

    Zheng, Jindan; Liu, Lili; Wang, Shanfeng; Huang, Xin

    2015-10-01

    To investigate the functional role of SUMO-1 on cell proliferation and apoptosis in endometrial cancer cells (Ishikawa line) and to explore the underlying regulatory mechanisms. Different concentrations of estradiol (E2) and small interfering RNA (siRNA) targeting the SUMO-1 (siCo) were treated in Ishikawa cells, and then quantitative polymerase chain reaction was used to examine the expression of progesterone receptor (PR) expression in Ishikawa cells. Western blots were applied to validate histone H4 sumoylation. CCK8 assay and flow cytometry were performed to investigate cell proliferation and apoptosis in Ishikawa cells. Estradiol up-regulated the expression of PR messenger RNA, most obviously at 100 nM. SUMO-1 siRNA decreased the PR expression. Estradiol up-regulated H4 sumoylation and caused the increase of Ishikawa cell proliferation, whereas SUMO-1 siRNA down-regulated H4 sumoylation, inhibited the cell proliferation, and induced apoptosis. Furthermore, SUMO-1 siRNA-transfected cells were arrested in the S and G2/M phases and E2 increased the S and G2/M phases of Ishikawa cells. SUMO-1 regulates the Ishikawa cell proliferation and apoptosis by the sumoylation of histone H4.

  19. Slx8 removes Pli1-dependent protein-SUMO conjugates including SUMOylated topoisomerase I to promote genome stability.

    Directory of Open Access Journals (Sweden)

    Roland Steinacher

    Full Text Available The SUMO-dependent ubiquitin ligase Slx8 plays key roles in promoting genome stability, including the processing of trapped Topoisomerase I (Top1 cleavage complexes and removal of toxic SUMO conjugates. We show that it is the latter function that constitutes Slx8's primary role in fission yeast. The SUMO conjugates in question are formed by the SUMO ligase Pli1, which is necessary for limiting spontaneous homologous recombination when Top1 is present. Surprisingly there is no requirement for Pli1 to limit recombination in the vicinity of a replication fork blocked at the programmed barrier RTS1. Notably, once committed to Pli1-mediated SUMOylation Slx8 becomes essential for genotoxin resistance, limiting both spontaneous and RTS1 induced recombination, and promoting normal chromosome segregation. We show that Slx8 removes Pli1-dependent Top1-SUMO conjugates and in doing so helps to constrain recombination at RTS1. Overall our data highlight how SUMOylation and SUMO-dependent ubiquitylation by the Pli1-Slx8 axis contribute in different ways to maintain genome stability.

  20. Slx8 Removes Pli1-Dependent Protein-SUMO Conjugates Including SUMOylated Topoisomerase I to Promote Genome Stability

    Science.gov (United States)

    Steinacher, Roland; Osman, Fekret; Lorenz, Alexander; Bryer, Claire; Whitby, Matthew C.

    2013-01-01

    The SUMO-dependent ubiquitin ligase Slx8 plays key roles in promoting genome stability, including the processing of trapped Topoisomerase I (Top1) cleavage complexes and removal of toxic SUMO conjugates. We show that it is the latter function that constitutes Slx8's primary role in fission yeast. The SUMO conjugates in question are formed by the SUMO ligase Pli1, which is necessary for limiting spontaneous homologous recombination when Top1 is present. Surprisingly there is no requirement for Pli1 to limit recombination in the vicinity of a replication fork blocked at the programmed barrier RTS1. Notably, once committed to Pli1-mediated SUMOylation Slx8 becomes essential for genotoxin resistance, limiting both spontaneous and RTS1 induced recombination, and promoting normal chromosome segregation. We show that Slx8 removes Pli1-dependent Top1-SUMO conjugates and in doing so helps to constrain recombination at RTS1. Overall our data highlight how SUMOylation and SUMO-dependent ubiquitylation by the Pli1-Slx8 axis contribute in different ways to maintain genome stability. PMID:23936535

  1. SIZ1-Dependent Post-Translational Modification by SUMO Modulates Sugar Signaling and Metabolism in Arabidopsis thaliana.

    Science.gov (United States)

    Castro, Pedro Humberto; Verde, Nuno; Lourenço, Tiago; Magalhães, Alexandre Papadopoulos; Tavares, Rui Manuel; Bejarano, Eduardo Rodríguez; Azevedo, Herlânder

    2015-12-01

    Post-translational modification mechanisms function as switches that mediate the balance between optimum growth and the response to environmental stimuli, by regulating the activity of key proteins. SUMO (small ubiquitin-like modifier) attachment, or sumoylation, is a post-translational modification that is essential for the plant stress response, also modulating hormonal circuits to co-ordinate developmental processes. The Arabidopsis SUMO E3 ligase SAP and Miz 1 (SIZ1) is the major SUMO conjugation enhancer in response to stress, and is implicated in several aspects of plant development. Here we report that known SUMO targets are over-represented in multiple carbohydrate-related proteins, suggesting a functional link between sumoylation and sugar metabolism and signaling in plants. We subsequently observed that SUMO-conjugated proteins accumulate in response to high doses of sugar in a SIZ1-dependent manner, and that the null siz1 mutant displays increased expression of sucrose and starch catabolic genes and shows reduced starch levels. We demonstrated that SIZ1 controls germination time and post-germination growth via osmotic and sugar-dependent signaling, respectively. Glucose was specifically linked to SUMO-sugar interplay, with high levels inducing root growth inhibition and aberrant root hair morphology in siz1. The use of sugar analogs and sugar marker gene expression analysis allowed us to implicate SIZ1 in a signaling pathway dependent on glucose metabolism, probably involving modulation of SNF1-related kinase 1 (SnRK1) activity.

  2. Development of Conformation Independent Computational Models for the Early Recognition of Breast Cancer Resistance Protein Substrates

    Directory of Open Access Journals (Sweden)

    Melisa Edith Gantner

    2013-01-01

    Full Text Available ABC efflux transporters are polyspecific members of the ABC superfamily that, acting as drug and metabolite carriers, provide a biochemical barrier against drug penetration and contribute to detoxification. Their overexpression is linked to multidrug resistance issues in a diversity of diseases. Breast cancer resistance protein (BCRP is the most expressed ABC efflux transporter throughout the intestine and the blood-brain barrier, limiting oral absorption and brain bioavailability of its substrates. Early recognition of BCRP substrates is thus essential to optimize oral drug absorption, design of novel therapeutics for central nervous system conditions, and overcome BCRP-mediated cross-resistance issues. We present the development of an ensemble of ligand-based machine learning algorithms for the early recognition of BCRP substrates, from a database of 262 substrates and nonsubstrates compiled from the literature. Such dataset was rationally partitioned into training and test sets by application of a 2-step clustering procedure. The models were developed through application of linear discriminant analysis to random subsamples of Dragon molecular descriptors. Simple data fusion and statistical comparison of partial areas under the curve of ROC curves were applied to obtain the best 2-model combination, which presented 82% and 74.5% of overall accuracy in the training and test set, respectively.

  3. Substrate Creep on The Fatigue Life of A Model Dental Multilayer Structure

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, J; Huang, M; Niu, X; soboyejo, W

    2006-10-09

    In this paper, we investigated the effects of substrate creep on the fatigue behavior of a model dental multilayer structure, in which a top glass layer was bonded to a polycarbonate substrate through a dental adhesive. The top glass layers were ground using 120 grit or 600 grit sand papers before bonding to create different sub-surface crack sizes and morphologies. The multilayer structures were tested under cyclic Hertzian contact loading to study crack growth and obtain fatigue life curves. The experiment results showed that the fatigue lives of the multilayer structures were impaired by increasing crack sizes in the sub-surfaces. They were also significantly reduced by the substrate creep when tested at relatively low load levels i.e. P{sub m} < 60 N (Pm is the maximum magnitude of cyclic load). But at relatively high load levels i.e. P{sub m} > 65 N, slow crack growth (SCG) was the major failure mechanisms. A modeling study was then carried out to explore the possible failure mechanisms over a range of load levels. It is found that fatigue life at relatively low load levels can be better estimated by considering the substrate creep effect (SCE).

  4. Towards the modeling of nanoindentation of virus shells: Do substrate adhesion and geometry matter?

    Science.gov (United States)

    Bousquet, Arthur; Dragnea, Bogdan; Tayachi, Manel; Temam, Roger

    2016-12-01

    Soft nanoparticles adsorbing at surfaces undergo deformation and buildup of elastic strain as a consequence of interfacial adhesion of similar magnitude with constitutive interactions. An example is the adsorption of virus particles at surfaces, a phenomenon of central importance for experiments in virus nanoindentation and for understanding of virus entry. The influence of adhesion forces and substrate corrugation on the mechanical response to indentation has not been studied. This is somewhat surprising considering that many single-stranded RNA icosahedral viruses are organized by soft intermolecular interactions while relatively strong adhesion forces are required for virus immobilization for nanoindentation. This article presents numerical simulations via finite elements discretization investigating the deformation of a thick shell in the context of slow evolution linear elasticity and in presence of adhesion interactions with the substrate. We study the influence of the adhesion forces in the deformation of the virus model under axial compression on a flat substrate by comparing the force-displacement curves for a shell having elastic constants relevant to virus capsids with and without adhesion forces derived from the Lennard-Jones potential. Finally, we study the influence of the geometry of the substrate in two-dimensions by comparing deformation of the virus model adsorbed at the cusp between two cylinders with that on a flat surface.

  5. SUMO modification regulates BLM and RAD51 interaction at damaged replication forks.

    Directory of Open Access Journals (Sweden)

    Karen J Ouyang

    2009-12-01

    Full Text Available The gene mutated in Bloom's syndrome, BLM, is important in the repair of damaged replication forks, and it has both pro- and anti-recombinogenic roles in homologous recombination (HR. At damaged forks, BLM interacts with RAD51 recombinase, the essential enzyme in HR that catalyzes homology-dependent strand invasion. We have previously shown that defects in BLM modification by the small ubiquitin-related modifier (SUMO cause increased gamma-H2AX foci. Because the increased gamma-H2AX could result from defective repair of spontaneous DNA damage, we hypothesized that SUMO modification regulates BLM's function in HR repair at damaged forks. To test this hypothesis, we treated cells that stably expressed a normal BLM (BLM+ or a SUMO-mutant BLM (SM-BLM with hydroxyurea (HU and examined the effects of stalled replication forks on RAD51 and its DNA repair functions. HU treatment generated excess gamma-H2AX in SM-BLM compared to BLM+ cells, consistent with a defect in replication-fork repair. SM-BLM cells accumulated increased numbers of DNA breaks and were hypersensitive to DNA damage. Importantly, HU treatment failed to induce sister-chromatid exchanges in SM-BLM cells compared to BLM+ cells, indicating a specific defect in HR repair and suggesting that RAD51 function could be compromised. Consistent with this hypothesis, RAD51 localization to HU-induced repair foci was impaired in SM-BLM cells. These data suggested that RAD51 might interact noncovalently with SUMO. We found that in vitro RAD51 interacts noncovalently with SUMO and that it interacts more efficiently with SUMO-modified BLM compared to unmodified BLM. These data suggest that SUMOylation controls the switch between BLM's pro- and anti-recombinogenic roles in HR. In the absence of BLM SUMOylation, BLM perturbs RAD51 localization at damaged replication forks and inhibits fork repair by HR. Conversely, BLM SUMOylation relieves its inhibitory effects on HR, and it promotes RAD51 function.

  6. Modelling anaerobic co-digestion in Benchmark Simulation Model No. 2: Parameter estimation, substrate characterisation and plant-wide integration.

    Science.gov (United States)

    Arnell, Magnus; Astals, Sergi; Åmand, Linda; Batstone, Damien J; Jensen, Paul D; Jeppsson, Ulf

    2016-07-01

    Anaerobic co-digestion is an emerging practice at wastewater treatment plants (WWTPs) to improve the energy balance and integrate waste management. Modelling of co-digestion in a plant-wide WWTP model is a powerful tool to assess the impact of co-substrate selection and dose strategy on digester performance and plant-wide effects. A feasible procedure to characterise and fractionate co-substrates COD for the Benchmark Simulation Model No. 2 (BSM2) was developed. This procedure is also applicable for the Anaerobic Digestion Model No. 1 (ADM1). Long chain fatty acid inhibition was included in the ADM1 model to allow for realistic modelling of lipid rich co-substrates. Sensitivity analysis revealed that, apart from the biodegradable fraction of COD, protein and lipid fractions are the most important fractions for methane production and digester stability, with at least two major failure modes identified through principal component analysis (PCA). The model and procedure were tested on bio-methane potential (BMP) tests on three substrates, each rich on carbohydrates, proteins or lipids with good predictive capability in all three cases. This model was then applied to a plant-wide simulation study which confirmed the positive effects of co-digestion on methane production and total operational cost. Simulations also revealed the importance of limiting the protein load to the anaerobic digester to avoid ammonia inhibition in the digester and overloading of the nitrogen removal processes in the water train. In contrast, the digester can treat relatively high loads of lipid rich substrates without prolonged disturbances.

  7. Dynamics of cell shape and forces on micropatterned substrates predicted by a cellular Potts model.

    Science.gov (United States)

    Albert, Philipp J; Schwarz, Ulrich S

    2014-06-03

    Micropatterned substrates are often used to standardize cell experiments and to quantitatively study the relation between cell shape and function. Moreover, they are increasingly used in combination with traction force microscopy on soft elastic substrates. To predict the dynamics and steady states of cell shape and forces without any a priori knowledge of how the cell will spread on a given micropattern, here we extend earlier formulations of the two-dimensional cellular Potts model. The third dimension is treated as an area reservoir for spreading. To account for local contour reinforcement by peripheral bundles, we augment the cellular Potts model by elements of the tension-elasticity model. We first parameterize our model and show that it accounts for momentum conservation. We then demonstrate that it is in good agreement with experimental data for shape, spreading dynamics, and traction force patterns of cells on micropatterned substrates. We finally predict shapes and forces for micropatterns that have not yet been experimentally studied. Copyright © 2014 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  8. Electromagnetic Modelling of Raman Enhancement from Nanoscale Structures as a Means to Predict the Efficacy of SERS Substrates

    Directory of Open Access Journals (Sweden)

    Richard J. C. Brown

    2007-01-01

    Full Text Available The requirement to optimise the balance between signal enhancement and reproducibility in surface enhanced Raman spectroscopy (SERS is stimulating the development of novel substrates for enhancing Raman signals. This paper describes the application of finite element electromagnetic modelling to predict the Raman enhancement produced from a variety of SERS substrates with differently sized, spaced and shaped morphologies with nanometre dimensions. For the first time, a theoretical comparison between four major generic types of SERS substrate (including metal nanoparticles, structured surfaces, and sharp tips has been performed and the results are presented and discussed. The results of the modelling are consistent with published experimental data from similar substrates.

  9. Gold Incorporated Mesoporous Silica Thin Film Model Surface as a Robust SERS and Catalytically Active Substrate

    Directory of Open Access Journals (Sweden)

    Anandakumari Chandrasekharan Sunil Sekhar

    2016-05-01

    Full Text Available Ultra-small gold nanoparticles incorporated in mesoporous silica thin films with accessible pore channels perpendicular to the substrate are prepared by a modified sol-gel method. The simple and easy spin coating technique is applied here to make homogeneous thin films. The surface characterization using FESEM shows crack-free films with a perpendicular pore arrangement. The applicability of these thin films as catalysts as well as a robust SERS active substrate for model catalysis study is tested. Compared to bare silica film our gold incorporated silica, GSM-23F gave an enhancement factor of 103 for RhB with a laser source 633 nm. The reduction reaction of p-nitrophenol with sodium borohydride from our thin films shows a decrease in peak intensity corresponding to –NO2 group as time proceeds, confirming the catalytic activity. Such model surfaces can potentially bridge the material gap between a real catalytic system and surface science studies.

  10. A comprehensive model of anaerobic bioconversion of complex substrates to biogas

    DEFF Research Database (Denmark)

    Angelidaki, Irini; Ellegaard, Lars; Ahring, Birgitte Kiær

    1999-01-01

    .e., carbohydrates, lipids, and proteins, the concentration of intermediates such as volatile fatty acids and long-chain fatty acids, and important inorganic components, i.e., ammonia, phosphate, cations, and anions. This allows dynamic changes of the process during a shift of substrate composition to be simulated......) constitute the primary modulating factors in the model. The model was rested with success in lab-scale reactors codigesting manure with glycerol trioleate or manure with gelatin. Finally, the model was validated using results from a full-scale biogas plant codigesting manure together with a proteinous...

  11. Defining Human Tyrosine Kinase Phosphorylation Networks Using Yeast as an In Vivo Model Substrate.

    Science.gov (United States)

    Corwin, Thomas; Woodsmith, Jonathan; Apelt, Federico; Fontaine, Jean-Fred; Meierhofer, David; Helmuth, Johannes; Grossmann, Arndt; Andrade-Navarro, Miguel A; Ballif, Bryan A; Stelzl, Ulrich

    2017-08-23

    Systematic assessment of tyrosine kinase-substrate relationships is fundamental to a better understanding of cellular signaling and its profound alterations in human diseases such as cancer. In human cells, such assessments are confounded by complex signaling networks, feedback loops, conditional activity, and intra-kinase redundancy. Here we address this challenge by exploiting the yeast proteome as an in vivo model substrate. We individually expressed 16 human non-receptor tyrosine kinases (NRTKs) in Saccharomyces cerevisiae and identified 3,279 kinase-substrate relationships involving 1,351 yeast phosphotyrosine (pY) sites. Based on the yeast data without prior information, we generated a set of linear kinase motifs and assigned ∼1,300 known human pY sites to specific NRTKs. Furthermore, experimentally defined pY sites for each individual kinase were shown to cluster within the yeast interactome network irrespective of linear motif information. We therefore applied a network inference approach to predict kinase-substrate relationships for more than 3,500 human proteins, providing a resource to advance our understanding of kinase biology. Copyright © 2017 Elsevier Inc. All rights reserved.

  12. At the interface: Biotic-abiotic interactions between substrates and a model epithlium

    Science.gov (United States)

    Covell, Alan D.

    The need for determining the fundamental mechanisms that define the interaction of biological systems with underlying materials, both natural and synthetic, is important as humanity endeavors to improve the quality of life of individuals through technology. Recently, much work has focused on the role of material properties on the behavior of cells. Most of these studies have concentrated their efforts on fibroblastic cell lines and more recently different kinds of stems cells. While these cells represent an important subset of cells in complex organisms, they do not manifest cell-cell interactions, a feature of epithelial cells, the most abundant cell type. Epithelial cells represent the largest cell type in the body and introduce an intrinsic complexity when researching the interaction of biological systems with materials. Adherens junctions (AJ) play a significant role in many signaling pathways, and therefore there is need to investigate how physical interactions with underlying substrates affect cell-cell interactions, such as the adhesion properties between cells, as well as how cell-substrate interactions influence the morphology and growth of epithelial cells. In this work I seek to determine the effects and identify mechanisms that epithelial cells use to "read" their environment. To do this I examined changes in cell behavior (growth, morphological, adhesion) of a model epithelium on substrates that have similar composition but significant differences in surface organization. In such a manner, I probed the limitations at which the nanoscale differences in substrate topography affect cellular behavior.

  13. USP7/HAUSP: A SUMO deubiquitinase at the heart of DNA replication.

    Science.gov (United States)

    Smits, Veronique A J; Freire, Raimundo

    2016-09-01

    DNA replication is both highly conserved and controlled. Problematic DNA replication can lead to genomic instability and therefore carcinogenesis. Numerous mechanisms work together to achieve this tight control and increasing evidence suggests that post-translational modifications (phosphorylation, ubiquitination, SUMOylation) of DNA replication proteins play a pivotal role in this process. Here we discuss such modifications in the light of a recent article that describes a novel role for the deubiquitinase (DUB) USP7/HAUSP in the control of DNA replication. USP7 achieves this function by an unusual and novel mechanism, namely deubiquitination of SUMOylated proteins at the replication fork, making USP7 also a SUMO DUB (SDUB). This work extends previous observations of increased levels of SUMO and low levels of ubiquitin at the on-going replication fork. Here, we discuss this novel study, its contribution to the DNA replication and genomic stability field and what questions arise from this work.

  14. Modeling of stable and metastable structures of Co, Cr, or Fe deposited on Ag(100) substrates

    Energy Technology Data Exchange (ETDEWEB)

    Canzian, A. [Grupo de Caracterizacion y Modelacion de Materiales, UTN, FRGP, H. Yrigoyen 288, (B1617FRP) Gral. Pacheco (Argentina); Bozzolo, G., E-mail: guille_bozzolo@yahoo.co [Loyola University of Maryland, 4501 N. Charles St, Baltimore, MD 21210 (United States); Mosca, H.O. [Gerencia de Investigacion y Aplicaciones, Comision Nacional de Energia Atomica, Av. Gral. Paz 1499 (B1650KNA), San Martin (Argentina)

    2011-01-31

    Atomistic modeling of the deposition of Co, Cr, or Fe on a Ag(100) substrate is performed using the Bozzolo-Ferrante-Smith method for alloys, in order to describe the similarities and differences between the three cases. An atom-by-atom description of the deposition process explains the growth patterns from an early stage, establishing a criterion for the determination of the ensuing growth modes.

  15. Covalent conjugation of the equine infectious anemia virus Gag with SUMO.

    Science.gov (United States)

    Wang, Jinzhong; Wen, Shuping; Zhao, Rui; Qi, Jing; Liu, Zhao; Li, Weiwei; An, Jing; Wood, Charles; Wang, Ying

    2017-05-06

    The conjugation of small ubiquitin-like modifier (SUMO) to the target protein, namely, SUMOylation, is involved in the regulation of many important biological events including host-pathogen interaction. Some viruses have evolved to exploit the host SUMOylation machinery to modify their own protein. Retroviral Gag protein plays critical roles in the viral life cycle. The HIV-1 p6 and the Moloney murine leukemia virus CA have been reported to be conjugated with SUMO. In this study, we report for the first time, to our knowledge, the covalent conjugation of equine infectious anemia virus (EIAV) Gag with SUMO. The C-terminal p9 domain of Gag is a main target for SUMOylation and SUMO is attached to multiple sites of p9, including K30 whose mutation abolished p9 SUMOylation completely. The SUMOylation of p9, but not the p9-K30 mutant, was also detected in equine fibroblastic cells ATCC(®) CCL-57™. Ubc9 and its C93 residue are indispensable for the SUMOylation of p9. Using confocal microscopy, it is found that EIAV Gag localizes primarily, if not exclusively, in the cytoplasm of the cell and the co-localization of EIAV Gag with Ubc9 was observed. Our findings that EIAV Gag is SUMOylated at p9-K30, together with previous findings on the defects of p9-K30 mutant in viral DNA translocation from cytoplasm to the nucleus, suggests that SUMOylation of Gag may be involved in such functions. Copyright © 2017 Elsevier Inc. All rights reserved.

  16. Scavenger receptors mediate the role of SUMO and Ftz-f1 in Drosophila steroidogenesis.

    Directory of Open Access Journals (Sweden)

    Ana Talamillo

    2013-04-01

    Full Text Available SUMOylation participates in ecdysteroid biosynthesis at the onset of metamorphosis in Drosophila melanogaster. Silencing the Drosophila SUMO homologue smt3 in the prothoracic gland leads to reduced lipid content, low ecdysone titers, and a block in the larval-pupal transition. Here we show that the SR-BI family of Scavenger Receptors mediates SUMO functions. Reduced levels of Snmp1 compromise lipid uptake in the prothoracic gland. In addition, overexpression of Snmp1 is able to recover lipid droplet levels in the smt3 knockdown prothoracic gland cells. Snmp1 expression depends on Ftz-f1 (an NR5A-type orphan nuclear receptor, the expression of which, in turn, depends on SUMO. Furthermore, we show by in vitro and in vivo experiments that Ftz-f1 is SUMOylated. RNAi-mediated knockdown of ftz-f1 phenocopies that of smt3 at the larval to pupal transition, thus Ftz-f1 is an interesting candidate to mediate some of the functions of SUMO at the onset of metamorphosis. Additionally, we demonstrate that the role of SUMOylation, Ftz-f1, and the Scavenger Receptors in lipid capture and mobilization is conserved in other steroidogenic tissues such as the follicle cells of the ovary. smt3 knockdown, as well as ftz-f1 or Scavenger knockdown, depleted the lipid content of the follicle cells, which could be rescued by Snmp1 overexpression. Therefore, our data provide new insights into the regulation of metamorphosis via lipid homeostasis, showing that Drosophila Smt3, Ftz-f1, and SR-BIs are part of a general mechanism for uptake of lipids such as cholesterol, required during development in steroidogenic tissues.

  17. Arabidopsis SUMO protease ASP1 positively regulates flowering time partially through regulating FLC stability 

    KAUST Repository

    Kong, Xiangxiong

    2016-12-07

    The initiation of flowering is tightly regulated by the endogenous and environment signals, which is crucial for the reproductive success of flowering plants. It is well known that autonomous and vernalization pathways repress transcription of FLOWERING LOCUS C (FLC), a focal floral repressor, but how its protein stability is regulated remains largely unknown. Here, we found that mutations in a novel Arabidopsis SUMO protease 1 (ASP1) resulted in a strong late-flowering phenotype under long-days, but to a lesser extent under short-days. ASP1 localizes in the nucleus and exhibited a SUMO protease activity in vitro and in vivo. The conserved Cys-577 in ASP1 is critical for its enzymatic activity, as well as its physiological function in the regulation of flowering time. Genetic and gene expression analyses demonstrated that ASP1 promotes transcription of positive regulators of flowering, such as FT, SOC1 and FD, and may function in both CO-dependent photoperiod pathway and FLC-dependent pathways. Although the transcription level of FLC was not affected in the loss-of-function asp1 mutant, the protein stability of FLC was increased in the asp1 mutant. Taken together, this study identified a novel bona fide SUMO protease, ASP1, which positively regulates transition to flowering at least partly by repressing FLC protein stability.

  18. Regulation of mitotic spindle asymmetry by SUMO and the spindle-assembly checkpoint in yeast.

    Science.gov (United States)

    Leisner, Christian; Kammerer, Daniel; Denoth, Annina; Britschi, Mirjam; Barral, Yves; Liakopoulos, Dimitris

    2008-08-26

    During mitosis, the kinetochore microtubules capture and segregate chromosomes, and the astral microtubules position the spindle within the cell. Although the spindle is symmetric, proper positioning of the spindle in asymmetrically dividing cells generally correlates with the formation of morphologically and structurally distinct asters [1]. In budding yeast, the spindle-orientation proteins Kar9 and dynein decorate only one aster of the metaphase spindle and direct it toward the bud [2, 3]. The mechanisms controlling the distribution of Kar9 and dynein remain unclear. Here, we show that SUMO regulates astral-microtubule function in at least two ways. First, Kar9 was sumoylated in vivo. Sumoylation and Cdk1-dependent phosphorylation of Kar9 independently promoted Kar9 asymmetry on the spindle. Second, proper regulation of kinetochore function by SUMO was also required for Kar9 asymmetry. Indeed, activation of the spindle-assembly checkpoint (SAC) due to SUMO and kinetochore defects promoted symmetric redistribution of Kar9 in a Mad2-dependent manner. The control of Kar9 distribution by the SAC was independent of Kar9 sumoylation and phosphorylation. Together, our data reveal that three independent mechanisms contribute to Kar9 asymmetry: Cdk1-dependent phosphorylation, sumoylation, and SAC signaling. Hence, the two seemingly independent spindle domains, kinetochores and astral microtubules, function in a tightly coordinated fashion.

  19. Tandem SUMO fusion vectors for improving soluble protein expression and purification.

    Science.gov (United States)

    Guerrero, Fernando; Ciragan, Annika; Iwaï, Hideo

    2015-12-01

    Availability of highly purified proteins in quantity is crucial for detailed biochemical and structural investigations. Fusion tags are versatile tools to facilitate efficient protein purification and to improve soluble overexpression of proteins. Various purification and fusion tags have been widely used for overexpression in Escherichia coli. However, these tags might interfere with biological functions and/or structural investigations of the protein of interest. Therefore, an additional purification step to remove fusion tags by proteolytic digestion might be required. Here, we describe a set of new vectors in which yeast SUMO (SMT3) was used as the highly specific recognition sequence of ubiquitin-like protease 1, together with other commonly used solubility enhancing proteins, such as glutathione S-transferase, maltose binding protein, thioredoxin and trigger factor for optimizing soluble expression of protein of interest. This tandem SUMO (T-SUMO) fusion system was tested for soluble expression of the C-terminal domain of TonB from different organisms and for the antiviral protein scytovirin.

  20. Dynamic Modelling Reveals 'Hotspots' on the Pathway to Enzyme-Substrate Complex Formation.

    Directory of Open Access Journals (Sweden)

    Shane E Gordon

    2016-03-01

    Full Text Available Dihydrodipicolinate synthase (DHDPS catalyzes the first committed step in the diaminopimelate pathway of bacteria, yielding amino acids required for cell wall and protein biosyntheses. The essentiality of the enzyme to bacteria, coupled with its absence in humans, validates DHDPS as an antibacterial drug target. Conventional drug design efforts have thus far been unsuccessful in identifying potent DHDPS inhibitors. Here, we make use of contemporary molecular dynamics simulation and Markov state models to explore the interactions between DHDPS from the human pathogen Staphylococcus aureus and its cognate substrate, pyruvate. Our simulations recover the crystallographic DHDPS-pyruvate complex without a priori knowledge of the final bound structure. The highly conserved residue Arg140 was found to have a pivotal role in coordinating the entry of pyruvate into the active site from bulk solvent, consistent with previous kinetic reports, indicating an indirect role for the residue in DHDPS catalysis. A metastable binding intermediate characterized by multiple points of intermolecular interaction between pyruvate and key DHDPS residue Arg140 was found to be a highly conserved feature of the binding trajectory when comparing alternative binding pathways. By means of umbrella sampling we show that these binding intermediates are thermodynamically metastable, consistent with both the available experimental data and the substrate binding model presented in this study. Our results provide insight into an important enzyme-substrate interaction in atomistic detail that offers the potential to be exploited for the discovery of more effective DHDPS inhibitors and, in a broader sense, dynamic protein-drug interactions.

  1. Swimming near the substrate: a simple robotic model of stingray locomotion.

    Science.gov (United States)

    Blevins, Erin; Lauder, George V

    2013-03-01

    Studies of aquatic locomotion typically assume that organisms move through unbounded fluid. However, benthic fishes swim close to the substrate and will experience significant ground effects, which will be greatest for fishes with wide spans such as benthic batoids and flatfishes. Ground effects on fixed-wing flight are well understood, but these models are insufficient to describe the dynamic interactions between substrates and undulating, oscillating fish. Live fish alter their swimming behavior in ground effect, complicating comparisons of near-ground and freestream swimming performance. In this study, a simple, stingray-inspired physical model offers insights into ground effects on undulatory swimmers, contrasting the self-propelled swimming speed, power requirements, and hydrodynamics of fins swimming with fixed kinematics near and far from a solid boundary. Contrary to findings for gliding birds and other fixed-wing fliers, ground effect does not necessarily enhance the performance of undulating fins. Under most kinematic conditions, fins do not swim faster in ground effect, power requirements increase, and the cost of transport can increase by up to 10%. The influence of ground effect varies with kinematics, suggesting that benthic fish might modulate their swimming behavior to minimize locomotor penalties and incur benefits from swimming near a substrate.

  2. Modal response of a computational vocal fold model with a substrate layer of adipose tissue.

    Science.gov (United States)

    Jones, Cameron L; Achuthan, Ajit; Erath, Byron D

    2015-02-01

    This study demonstrates the effect of a substrate layer of adipose tissue on the modal response of the vocal folds, and hence, on the mechanics of voice production. Modal analysis is performed on the vocal fold structure with a lateral layer of adipose tissue. A finite element model is employed, and the first six mode shapes and modal frequencies are studied. The results show significant changes in modal frequencies and substantial variation in mode shapes depending on the strain rate of the adipose tissue. These findings highlight the importance of considering adipose tissue in computational vocal fold modeling.

  3. Benzophenone-based photochemical micropatterning of biomolecules to create model substrates and instructive biomaterials.

    Science.gov (United States)

    Turgeon, Aurora J; Harley, Brendan A; Bailey, Ryan C

    2014-01-01

    The extracellular matrix (ECM) is a dynamic and heterogeneous environment that controls many aspects of cell behavior. Not surprisingly, many different approaches have focused on creating model substrates that recapitulate the biomolecular, topographical, and mechanical properties of the ECM for in vitro studies of cell behavior. This chapter details a general, versatile method for the spatially controlled deposition of multiple biomolecules onto both planar and topographically complex support structures with micrometer resolution. This approach is based upon the well-understood photochemical UV crosslinking of benzophenone (BP) to solution-phase biomolecules. This is a molecularly general strategy that can be utilized to immobilize biomolecules onto any surface prefunctionalized with BP. Examples described herein include modification of planar and corrugated glass substrates as well as collagen-glycosaminoglycan biomaterials configured either as highly porous scaffolds or nonporous membranes with a variety of biomolecular targets, including proteins, glycoproteins, and carbohydrates.

  4. Computational modelling of multi-cell migration in a multi-signalling substrate

    Science.gov (United States)

    Jamaleddin Mousavi, Seyed; Doblaré, Manuel; Hamdy Doweidar, Mohamed

    2014-04-01

    Cell migration is a vital process in many biological phenomena ranging from wound healing to tissue regeneration. Over the past few years, it has been proven that in addition to cell-cell and cell-substrate mechanical interactions (mechanotaxis), cells can be driven by thermal, chemical and/or electrical stimuli. A numerical model was recently presented by the authors to analyse single cell migration in a multi-signalling substrate. That work is here extended to include multi-cell migration due to cell-cell interaction in a multi-signalling substrate under different conditions. This model is based on balancing the forces that act on the cell population in the presence of different guiding cues. Several numerical experiments are presented to illustrate the effect of different stimuli on the trajectory and final location of the cell population within a 3D heterogeneous multi-signalling substrate. Our findings indicate that although multi-cell migration is relatively similar to single cell migration in some aspects, the associated behaviour is very different. For instance, cell-cell interaction may delay single cell migration towards effective cues while increasing the magnitude of the average net cell traction force as well as the local velocity. Besides, the random movement of a cell within a cell population is slightly greater than that of single cell migration. Moreover, higher electrical field strength causes the cell slug to flatten near the cathode. On the other hand, as with single cell migration, the existence of electrotaxis dominates mechanotaxis, moving the cells to the cathode or anode pole located at the free surface. The numerical results here obtained are qualitatively consistent with related experimental works.

  5. A homology model of Xyloglucan Xylosyltransferase 2 reveals critical amino acids involved in substrate binding.

    Science.gov (United States)

    Culbertson, Alan T; Tietze, Alesia A; Tietze, Daniel; Chou, Yi-Hsiang; Smith, Adrienne L; Young, Zachary T; Zabotina, Olga A

    2016-09-01

    In dicotyledonous plants, xyloglucan (XyG) is the most abundant hemicellulose of the primary cell wall. The enzymes involved in XyG biosynthesis have been identified through reverse-genetics and activity was characterized by heterologous expression. Currently, there is no information on the atomic structures or amino acids involved in activity or substrate binding of any of the Golgi-localized XyG biosynthetic enzymes. A homology model of the xyloglucan xylosyltransferase 2 (XXT2) catalytic domain was built on the basis of the crystal structure of A64Rp. Molecular dynamics simulations revealed that the homology model retains the glycosyltransferase (GT)-A fold of the template structure used to build the homology model indicating that XXT2 likely has a GT-A fold. According to the XXT2 homology model, six amino acids (Phe204, Lys207, Asp228, Ser229, Asp230, His378) were selected and their contribution in catalytic activity was investigated. Site-directed mutagenesis studies show that Asp228, Asp230 and His378 are critical for XXT2 activity and are predicted to be involved in coordination of manganese ion. Lys207 was also found to be critical for protein activity and the homology model indicates a critical role in substrate binding. Additionally, Phe204 mutants have less of an impact on XXT2 activity with the largest effect when replaced with a polar residue. This is the first study that investigates the amino acids involved in substrate binding of the XyG-synthesizing xylosyltransferases and contributes to the understanding of the mechanisms of polysaccharide-synthesizing GTs and XyG biosynthesis. © The Author 2016. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  6. The Key Regulator for Language and Speech Development, FOXP2, is a Novel Substrate for SUMOylation.

    Science.gov (United States)

    Meredith, Leslie J; Wang, Chiung-Min; Nascimento, Leticia; Liu, Runhua; Wang, Lizhong; Yang, Wei-Hsiung

    2016-02-01

    Transcription factor forkhead box protein P2 (FOXP2) plays an essential role in the development of language and speech. However, the transcriptional activity of FOXP2 regulated by the post-translational modifications remains unknown. Here, we demonstrated that FOXP2 is clearly defined as a SUMO target protein at the cellular levels as FOXP2 is covalently modified by both SUMO1 and SUMO3. Furthermore, SUMOylation of FOXP2 was significantly decreased by SENP2 (a specific SUMOylation protease). We further showed that FOXP2 is selectively SUMOylated in vivo on a phylogenetically conserved lysine 674 but the SUMOylation does not alter subcellular localization and stability of FOXP2. Interestingly, we observed that human etiological FOXP2 R553H mutation robustly reduces its SUMOylation potential as compared to wild-type FOXP2. In addition, the acidic residues downstream the core SUMO motif on FOXP2 are required for its full SUMOylation capacity. Finally, our functional analysis using reporter gene assays showed that SUMOylation may modulate transcriptional activity of FOXP2 in regulating downstream target genes (DISC1, SRPX2, and MiR200c). Altogether, we provide the first evidence that FOXP2 is a substrate for SUMOylation and SUMOylation of FOXP2 plays a functional role in regulating its transcriptional activity.

  7. Stability of small ubiquitin-like modifier (SUMO) proteases OVERLY TOLERANT TO SALT1 and -2 modulates salicylic acid signalling and SUMO1/2 conjugation in Arabidopsis thaliana.

    Science.gov (United States)

    Bailey, Mark; Srivastava, Anjil; Conti, Lucio; Nelis, Stuart; Zhang, Cunjin; Florance, Hannah; Love, Andrew; Milner, Joel; Napier, Richard; Grant, Murray; Sadanandom, Ari

    2016-01-01

    Small ubiquitin-like modifier proteases 1 and 2 (SUMO1/2) have been linked to the regulation of salicylic acid (SA)-mediated defence signalling in Arabidopsis thaliana. In order to define the role of the SUMO proteases OVERLY TOLERANT TO SALT1 and -2 (OTS1/2) in defence and to provide insight into SUMO1/2-mediated regulation of SA signalling, we examined the status of SA-mediated defences in ots1/2 mutants. The ots1 ots2 double mutant displayed enhanced resistance to virulent Pseudomonas syringae and higher levels of SA compared with wild-type (WT) plants. Furthermore, ots1 ots2 mutants exhibited upregulated expression of the SA biosynthesis gene ICS1 in addition to enhanced SA-responsive ICS1 expression beyond that of WT. SA stimulated OTS1/2 degradation and promoted accumulation of SUMO1/2 conjugates. These results indicate that OTS1 and -2 act in a feedback loop in SA signalling and that de novo OTS1/2 synthesis works antagonistically to SA-promoted degradation, adjusting the abundance of OTS1/2 to moderate SA signalling. Accumulation of SUMO1/2 conjugates coincides with SA-promoted OTS degradation and may play a positive role in SA-mediated signalling in addition to its repressive roles reported elsewhere.

  8. Substrate specificity and molecular modelling of the feline herpesvirus-1 thymidine kinase.

    Science.gov (United States)

    Hussein, Islam T M; Miguel, Ricardo Núñez; Tiley, Laurence S; Field, Hugh J

    2008-01-01

    Feline herpesvirus-1 (FHV-1) causes a severe upper respiratory and ocular disease in cats. An effective antiviral compound is required for treating FHV-1 infections. The virus-encoded thymidine kinase (TK) is the molecular basis for selective activation of commonly used antiviral nucleoside analogue drugs, e.g. acyclovir (ACV), penciclovir (PCV) and ganciclovir (GCV). The substrate specificity of a recombinant FHV-1 TK, expressed in Escherichia coli, was studied. FHV-1 TK efficiently phosphorylated its natural substrate deoxythymidine. However, it exhibited relatively lower affinity for the guanosine analogue substrates. PCV was most efficiently phosphorylated, followed by GCV, with approximately twofold reduction in the phosphorylation rate. The lowest phosphorylation rate was recorded for ACV. To correlate these biochemical data with structural features of the FHV-1 TK, a three-dimensional (3D) model of this enzyme was constructed based on sequence homology with two other herpesviral TKs, encoded by equine herpesvirus-4 (EHV-4) and herpes simplex-1 (HSV-1). Mutational analysis of the amino acids forming the FHV-1 TK active site identified two residues (Y29 and F144) as being critical for the differential ability of this enzyme to phosphorylate nucleoside analogues. A double substitution of Y29H/F144Y resulted in a threefold increase in the ACV phosphorylation rate.

  9. Modified kinetic-hydraulic UASB reactor model for treatment of wastewater containing biodegradable organic substrates.

    Science.gov (United States)

    El-Seddik, Mostafa M; Galal, Mona M; Radwan, A G; Abdel-Halim, Hisham S

    2016-01-01

    This paper addresses a modified kinetic-hydraulic model for up-flow anaerobic sludge blanket (UASB) reactor aimed to treat wastewater of biodegradable organic substrates as acetic acid based on Van der Meer model incorporated with biological granules inclusion. This dynamic model illustrates the biomass kinetic reaction rate for both direct and indirect growth of microorganisms coupled with the amount of biogas produced by methanogenic bacteria in bed and blanket zones of reactor. Moreover, the pH value required for substrate degradation at the peak specific growth rate of bacteria is discussed for Andrews' kinetics. The sensitivity analyses of biomass concentration with respect to fraction of volume of reactor occupied by granules and up-flow velocity are also demonstrated. Furthermore, the modified mass balance equations of reactor are applied during steady state using Newton Raphson technique to obtain a suitable degree of freedom for the modified model matching with the measured results of UASB Sanhour wastewater treatment plant in Fayoum, Egypt.

  10. Allosteric activation of SENP1 by SUMO1 β-grasp domain involves a dock-and-coalesce mechanism.

    Science.gov (United States)

    Guo, Jingjing; Zhou, Huan-Xiang

    2016-08-31

    Small ubiquitin-related modifiers (SUMOs) are conjugated to proteins to regulate a variety of cellular processes. SENPs are cysteine proteases with a catalytic center located within a channel between two subdomains that catalyzes SUMO C-terminal cleavage for processing of SUMO precursors and de-SUMOylation of target proteins. The β-grasp domain of SUMOs binds to an exosite cleft, and allosterically activates SENPs via an unknown mechanism. Our molecular dynamics simulations showed that binding of the β-grasp domain induces significant conformational and dynamic changes in SENP1, including widening of the exosite cleft and quenching of nanosecond dynamics in all but a distal region. A dock-and-coalesce mechanism emerges for SENP-catalyzed SUMO cleavage: the wedging of the β-grasp domain enables the docking of the proximal portion of the C-terminus and the strengthened cross-channel motional coupling initiates inter-subdomain correlated motions to allow for the distal portion to coalesce around the catalytic center.

  11. Theory of dielectric loss in Graphene-on-substrate: A tight- binding model study

    Science.gov (United States)

    Sahu, Sivabrata; Panda, S. K.; Rout, G. C.

    2016-09-01

    Graphene-on-substrate exhibits interesting dielectric behaviour due to screening of coulomb interaction induced by many body effects. In this communication we attempt to study the dielectric loss property of graphene within tight-binding model approach. The Hamiltonian consisting of electron hopping upto third-nearest-neighbour's with impurities in two in equivalent sub-lattices. The graphene-on-substrate raises the energy +Δ at one sub lattice and reduces energy -Δ at other sub lattice. Further we introduced coulomb interaction between π - electrons at the two sub lattices separately with the same effective coulomb interaction. We calculate polarization function Π(q, ω) which is a two particle Green's function arising due to charge-charge correlation by using Zubarev's Green's function technique. Finally we calculate dielectric function of graphene i.e. ε(q, ω) =1+Π(q,ω) at arbitrary wave vector q and frequency ra. The dielectric loss in graphene calculated from the imaginary part of dielectric function which is a measure of absorption spectrum. Only a few Fragmentary theoretical attempts have been made to utilize the full frequency and wave vector dependent dielectric function. We compute numerically the frequency dependent dielectric loss function for 100x100 momentum grid points. We observe a low energy Plasmon resonance peak and a high energy flat peak arising due to absorption of optical energy at substrate induced gap. With increase of small Plasmon wave vector, both low and high energy peaks approach each other. The dielectric loss at low energies exhibits a parabolic curve, but it exhibit a clear peak on introduction of higher order electron hopping's. The Coulomb interaction suppresses induced gap in graphene and decreases the optical energy absorption spectra. The increase of substrate induced gap shifts the high energy flat peak to higher energies and enhances the dielectric loss throughout the frequency range. Finally the effect of doping on

  12. Characterization of the SUMO-binding activity of the myeloproliferative and mental retardation (MYM-type zinc fingers in ZNF261 and ZNF198.

    Directory of Open Access Journals (Sweden)

    Catherine M Guzzo

    Full Text Available SUMO-binding proteins interact with SUMO modified proteins to mediate a wide range of functional consequences. Here, we report the identification of a new SUMO-binding protein, ZNF261. Four human proteins including ZNF261, ZNF198, ZNF262, and ZNF258 contain a stretch of tandem zinc fingers called myeloproliferative and mental retardation (MYM-type zinc fingers. We demonstrated that MYM-type zinc fingers from ZNF261 and ZNF198 are necessary and sufficient for SUMO-binding and that individual MYM-type zinc fingers function as SUMO-interacting motifs (SIMs. Our binding studies revealed that the MYM-type zinc fingers from ZNF261 and ZNF198 interact with the same surface on SUMO-2 recognized by the archetypal consensus SIM. We also present evidence that MYM-type zinc fingers in ZNF261 contain zinc, but that zinc is not required for SUMO-binding. Immunofluorescence microscopy studies using truncated fragments of ZNF198 revealed that MYM-type zinc fingers of ZNF198 are necessary for localization to PML-nuclear bodies (PML-NBs. In summary, our studies have identified and characterized the SUMO-binding activity of the MYM-type zinc fingers in ZNF261 and ZNF198.

  13. Intense Resistance Exercise Promotes the Acute and Transient Nuclear Translocation of Small Ubiquitin-Related Modifier (SUMO-1 in Human Myofibres

    Directory of Open Access Journals (Sweden)

    Sebastian Gehlert

    2016-04-01

    Full Text Available Protein sumoylation is a posttranslational modification triggered by cellular stress. Because general information concerning the role of small ubiquitin-related modifier (SUMO proteins in adult skeletal muscle is sparse, we investigated whether SUMO-1 proteins will be subjected to time-dependent changes in their subcellular localization in sarcoplasmic and nuclear compartments of human type I and II skeletal muscle fibers in response to acute stimulation by resistance exercise (RE. Skeletal muscle biopsies were taken at baseline (PRE, 15, 30, 60, 240 min and 24 h post RE from 6 male subjects subjected to a single bout of one-legged knee extensions. SUMO-1 localization was determined via immunohistochemistry and confocal laser microscopy. At baseline SUMO-1 was localized in perinuclear regions of myonuclei. Within 15 and up to 60 min post exercise, nuclear SUMO-1 localization was significantly increased (p < 0.01, declining towards baseline levels within 240 min post exercise. Sarcoplasmic SUMO-1 localization was increased at 15 min post exercise in type I and up to 30 min post RE in type II myofibres. The changing localization of SUMO-1 proteins acutely after intense muscle contractions points to a role for SUMO proteins in the acute regulation of the skeletal muscle proteome after exercise.

  14. A Proposal for a Structural Model of the Feline Calicivirus Protease Bound to the Substrate Peptide under Physiological Conditions

    Directory of Open Access Journals (Sweden)

    Masaru Yokoyama

    2017-07-01

    Full Text Available Feline calicivirus (FCV protease functions to cleave viral precursor proteins during productive infection. Previous studies have mapped a protease-coding region and six cleavage sites in viral precursor proteins. However, how the FCV protease interacts with its substrates remains unknown. To gain insights into the interactions, we constructed a molecular model of the FCV protease bound with the octapeptide containing a cleavage site of the capsid precursor protein by homology modeling and docking simulation. The complex model was used to screen for the substrate mimic from a chemical library by pharmacophore-based in silico screening. With this structure-based approach, we identified a compound that has physicochemical features and arrangement of the P3 and P4 sites of the substrate in the protease, is predicted to bind to FCV proteases in a mode similar to that of the authentic substrate, and has the ability to inhibit viral protease activity in vitro and in the cells, and to suppress viral replication in FCV-infected cells. The complex model was further subjected to molecular dynamics simulation to refine the enzyme-substrate interactions in solution. The simulation along with a variation study predicted that the authentic substrate and anti-FCV compound share a highly conserved binding site. These results suggest the validity of our in silico model for elucidating protease-substrate interactions during FCV replication and for developing antivirals.

  15. A Proposal for a Structural Model of the Feline Calicivirus Protease Bound to the Substrate Peptide under Physiological Conditions.

    Science.gov (United States)

    Yokoyama, Masaru; Oka, Tomoichiro; Takagi, Hirotaka; Kojima, Hirotatsu; Okabe, Takayoshi; Nagano, Tetsuo; Tohya, Yukinobu; Sato, Hironori

    2017-01-01

    Feline calicivirus (FCV) protease functions to cleave viral precursor proteins during productive infection. Previous studies have mapped a protease-coding region and six cleavage sites in viral precursor proteins. However, how the FCV protease interacts with its substrates remains unknown. To gain insights into the interactions, we constructed a molecular model of the FCV protease bound with the octapeptide containing a cleavage site of the capsid precursor protein by homology modeling and docking simulation. The complex model was used to screen for the substrate mimic from a chemical library by pharmacophore-based in silico screening. With this structure-based approach, we identified a compound that has physicochemical features and arrangement of the P3 and P4 sites of the substrate in the protease, is predicted to bind to FCV proteases in a mode similar to that of the authentic substrate, and has the ability to inhibit viral protease activity in vitro and in the cells, and to suppress viral replication in FCV-infected cells. The complex model was further subjected to molecular dynamics simulation to refine the enzyme-substrate interactions in solution. The simulation along with a variation study predicted that the authentic substrate and anti-FCV compound share a highly conserved binding site. These results suggest the validity of our in silico model for elucidating protease-substrate interactions during FCV replication and for developing antivirals.

  16. Langmuir adsorption study of the interaction of CdSe/ZnS quantum dots with model substrates: influence of substrate surface chemistry and pH.

    Science.gov (United States)

    Park, Jung Jin; Lacerda, Silvia H De Paoli; Stanley, Scott K; Vogel, Brandon M; Kim, Sangcheol; Douglas, Jack F; Raghavan, Dharmaraj; Karim, Alamgir

    2009-01-06

    We investigate the utility of Langmuir adsorption measurements for characterizing nanoparticle-substrate interactions. Spherical CdSe/ZnS core-shell nanoparticles were chosen as representative particles because of their widespread use in biological labeling measurements and their relatively monodisperse dimensions. In particular, the quantum dots were functionalized with 11-mercaptoundecanoic acid, and we utilized an amine-terminated self-assembled monolayer (SAM) as a model substrate. SAMs with different end-groups (-CH(3) and -COOH) were also considered to contrast with the adsorption behavior on the amine-terminated SAM substrates. We followed the kinetics of nanoparticle adsorption on the aminosilane layer by quartz crystal microgravimetry (QCM) over a range of particle concentrations and determined the corresponding Langmuir adsorption isotherms. Analysis of both equilibrium adsorption and kinetic adsorption data allowed us to determine a consistent value of the Langmuir adsorption equilibrium constant for the amine-terminated SAM at room temperature (K(L) approximately 2.7 (micromol/L)(-1)), providing a useful characterization of the nanoparticle-substrate interaction. The effect of varying solution pH on Langmuir adsorption was also investigated in order to gain insight into the role of electrostatic interactions on nanoparticle adsorption. The equilibrium extent of adsorption was found to be maximum at about pH 7. These changes of nanoparticle adsorption were further quantified and validated by X-ray photoelectron spectroscopy (XPS) and confocal fluorescence microscopy measurements. We conclude that Langmuir adsorption measurements provide a promising approach for quantifying nanoparticle-substrate interactions.

  17. Age-related changes of protein SUMOylation balance in the AβPP Tg2576 mouse model of Alzheimer's disease

    Directory of Open Access Journals (Sweden)

    Robert eNisticò

    2014-04-01

    Full Text Available Alzheimer's disease (AD is a complex disorder that affects the central nervous system causing a severe neurodegeneration. This pathology affects an increasing number of people worldwide due to the overall aging of the human population. In recent years SUMO protein modification has emerged as a possible cellular mechanism involved in AD. Some of the proteins engaged in the physiopathological process of AD, like BACE1, GSK3-β tau, AβPP and JNK, are in fact subject to protein SUMO modifications or interactions. Here, we have investigated the SUMO/deSUMOylation balance and SUMO-related proteins during the onset and progression of the pathology in the Tg2576 mouse model of AD. We examined four age-stages (1.5; 3; 6; 17 months old and observed shows an increase in SUMO-1 protein conjugation at 3 and 6 months in transgenic mice with respect to WT in both cortex and hippocampus. Interestingly this is paralleled by increased expression levels of Ubc9 and SENP1 in both brain regions. At 6 months of age also the SUMO-1 mRNA resulted augmented. SUMO-2-ylation was surprisingly decreased in old transgenic mice and was unaltered in the other time windows. The fact that alterations in SUMO/deSUMOylation equilibrium occur from the early phases of AD suggests that global posttranslational modifications may play an important role in the mechanisms underlying disease pathogenesis, thus providing potential targets for pharmacological interventions.

  18. Towards a modeling synthesis of two or three-dimensional circuits through substrate coupling and interconnections

    CERN Document Server

    Gontrand, Christian

    2014-01-01

    The number of transistors in integrated circuits doubles every two years, as stipulated by Moore's law, and this has been the driving force for the huge development of the microelectronics industry in the past 50 years - currently advanced to the nanometric scale.This e-book is dedicated to electronic noises and parasites, accounting for issues involving substrate coupling and interconnections, in the perspective of the 3D integration: a second track for enhancing integration, also compatible with Moore's law. This reference explains the modeling of 3D circuits without delving into the latest

  19. Multi-dimensional modeling of atmospheric copper-sulfidation corrosion on non-planar substrates.

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Ken Shuang

    2004-11-01

    This report documents the author's efforts in the deterministic modeling of copper-sulfidation corrosion on non-planar substrates such as diodes and electrical connectors. A new framework based on Goma was developed for multi-dimensional modeling of atmospheric copper-sulfidation corrosion on non-planar substrates. In this framework, the moving sulfidation front is explicitly tracked by treating the finite-element mesh as a pseudo solid with an arbitrary Lagrangian-Eulerian formulation and repeatedly performing re-meshing using CUBIT and re-mapping using MAPVAR. Three one-dimensional studies were performed for verifying the framework in asymptotic regimes. Limited model validation was also carried out by comparing computed copper-sulfide thickness with experimental data. The framework was first demonstrated in modeling one-dimensional copper sulfidation with charge separation. It was found that both the thickness of the space-charge layers and the electrical potential at the sulfidation surface decrease rapidly as the Cu{sub 2}S layer thickens initially but eventually reach equilibrium values as Cu{sub 2}S layer becomes sufficiently thick; it was also found that electroneutrality is a reasonable approximation and that the electro-migration flux may be estimated by using the equilibrium potential difference between the sulfidation and annihilation surfaces when the Cu{sub 2}S layer is sufficiently thick. The framework was then employed to model copper sulfidation in the solid-state-diffusion controlled regime (i.e. stage II sulfidation) on a prototypical diode until a continuous Cu{sub 2}S film was formed on the diode surface. The framework was also applied to model copper sulfidation on an intermittent electrical contact between a gold-plated copper pin and gold-plated copper pad; the presence of Cu{sub 2}S was found to raise the effective electrical resistance drastically. Lastly, future research needs in modeling atmospheric copper sulfidation are discussed.

  20. Dynamic wetting model for the isotropic-to-nematic transition over a flat substrate.

    Science.gov (United States)

    Rey, Alejandro D; Herrera-Valencia, E E

    2014-03-14

    Phase ordering over solid substrates is a ubiquitous and important soft material transformation process whose description incorporates wetting, anchoring and phase transition kinetics. In this paper the kinetics of the isotropic-to-nematic isothermal phase transition over a flat solid surface in a growing spherical drop is analyzed based on the Landau-de Gennes Q-tensor order parameter equations. The model, based on a previously derived interface force balance and a newly derived contact line force balance, is shown to be consistent with the generic model of conservative interface and contact line motions. The advancing dynamic contact angle equation is extracted from kinematic compatibility between the moving isotropic-nematic interface and contact line. A tractable surface phase transition kinetic model obtained by focusing on the dominant phase transition and wetting driving forces yields: (i) the constant advancing dynamic contact angle θ, and (ii) the contact line speed as a function of undercooling ΔT. It is shown that as undercooling increases, the surface phase transition mode approaches the bulk phase transition mode, such that θ approaches π. The elastic and wetting parameters that control the phase transformation process are identified and experiments for their determination are defined. These dynamic wetting and surface phase transition results significantly expand existing characterization methods of LC-substrate interfaces based on static phase transition droplet methods.

  1. Cellulose hydrolysis in evolving substrate morphologies I: A general modeling formalism.

    Science.gov (United States)

    Zhou, Wen; Schüttler, Heinz-Bernd; Hao, Zhiqian; Xu, Ying

    2009-10-01

    We develop a general framework for a realistic rate equation modeling of cellulose hydrolysis using non-complexed cellulase. Our proposed formalism, for the first time, takes into account explicitly the time evolution of the random substrate morphology resulting from the hydrolytic cellulose chain fragmentation and solubilization. This is achieved by integrating novel geometrical concepts to quantitatively capture the time-dependent random morphology, together with the enzymatic chain fragmentation, into a coupled morphology-plus-kinetics rate equation approach. In addition, an innovative site number representation, based on tracking available numbers of beta(1,4) glucosidic bonds, of different "site" types, exposed to attacks by different enzyme types, is presented. This site number representation results in an ordinary differential equation (ODE) system, with a substantially reduced ODE system size, compared to earlier chain fragmentation kinetics approaches. This formalism enables us to quantitatively simulate both the hydrolytically evolving random substrate morphology and the profound, and heretofore neglected, morphology effects on the hydrolysis kinetics. By incorporating the evolving morphology on an equal footing with the hydrolytic chain fragmentation, our formalism provides a framework for the realistic modeling of the entire solubilization process, beyond the short-time limit and through near-complete hydrolytic conversion. As part I of two companion papers, the present paper focuses on the development of the general modelling formalism. Results and testable experimental predictions from detailed numerical simulations are presented in part II.

  2. Structural differences of matrix metalloproteinases. Homology modeling and energy minimization of enzyme-substrate complexes

    DEFF Research Database (Denmark)

    Terp, G E; Christensen, I T; Jørgensen, Flemming Steen

    2000-01-01

    Matrix metalloproteinases are extracellular enzymes taking part in the remodeling of extracellular matrix. The structures of the catalytic domain of MMP1, MMP3, MMP7 and MMP8 are known, but structures of enzymes belonging to this family still remain to be determined. A general approach...... to the homology modeling of matrix metalloproteinases, exemplified by the modeling of MMP2, MMP9, MMP12 and MMP14 is described. The models were refined using an energy minimization procedure developed for matrix metalloproteinases. This procedure includes incorporation of parameters for zinc and calcium ions...... in the AMBER 4.1 force field, applying a non-bonded approach and a full ion charge representation. Energy minimization of the apoenzymes yielded structures with distorted active sites, while reliable three-dimensional structures of the enzymes containing a substrate in active site were obtained. The structural...

  3. DNA damage-inducible SUMOylation of HERC2 promotes RNF8 binding via a novel SUMO-binding Zinc finger

    DEFF Research Database (Denmark)

    Danielsen, Jannie Michaela Rendtlew; Povlsen, Lou Klitgaard; Villumsen, Bine Hare

    2012-01-01

    Nonproteolytic ubiquitylation of chromatin surrounding deoxyribonucleic acid (DNA) double-strand breaks (DSBs) by the RNF8/RNF168/HERC2 ubiquitin ligases facilitates restoration of genome integrity by licensing chromatin to concentrate genome caretaker proteins near the lesions. In parallel......, SUMOylation of so-far elusive upstream DSB regulators is also required for execution of this ubiquitin-dependent chromatin response. We show that HERC2 and RNF168 are novel DNA damage-dependent SUMOylation targets in human cells. In response to DSBs, both HERC2 and RNF168 were specifically modified with SUMO1...... at DSB sites in a manner dependent on the SUMO E3 ligase PIAS4. SUMOylation of HERC2 was required for its DSB-induced association with RNF8 and for stabilizing the RNF8-Ubc13 complex. We also demonstrate that the ZZ Zinc finger in HERC2 defined a novel SUMO-specific binding module, which together...

  4. Proteins with two SUMO-like domains in chromatin-associated complexes: The RENi (Rad60-Esc2-NIP45 family

    Directory of Open Access Journals (Sweden)

    Eisenhaber Birgit

    2005-02-01

    Full Text Available Abstract Background Post-translational modification by Small Ubiquitin-like Modifiers (SUMO has been implicated in protein targeting, in the maintenance of genomic integrity and in transcriptional control. But the specific molecular effects of SUMO modification on many target proteins remain to be elucidated. Recent findings point at the importance of SUMO-mediated histone NAD-dependent deacetylase (HDAC recruitment in transcriptional regulation. Results We describe the RENi family of SUMO-like domain proteins (SDP with the unique feature of typically containing two carboxy-terminal SUMO-like domains. Using sequence analytic evidence, we collect family members from animals, fungi and plants, most prominent being yeast Rad60, Esc2 and mouse NIP45 http://mendel.imp.univie.ac.at/SEQUENCES/reni/. Different proteins of the novel family are known to interact directly with histone NAD-dependent deacetylases (HDACs, structural maintenance of chromosomes (SMC proteins, and transcription factors. In particular, the highly non-trivial designation of the first of the two successive SUMO-domains in non-plant RENi provides a rationale for previously published functionally impaired mutant variants. Conclusions Till now, SUMO-like proteins have been studied exclusively in the context of their covalent conjugation to target proteins. Here, we present the exciting possibility that SUMO domain proteins, similarly to ubiquitin modifiers, have also evolved in a second line – namely as multi-domain proteins that are non-covalently attached to their target proteins. We suggest that the SUMO stable fusion proteins of the RENi family, which we introduce in this work, might mimic SUMO and share its interaction motifs (in analogy to the way that ubiquitin-like domains mimic ubiquitin. This presumption is supported by parallels in the spectrum of modified or bound proteins e.g. transcription factors and chromatin-associated proteins and in the recruitment of HDAC-activity.

  5. Functional Crosstalk between the PP2A and SUMO Pathways Revealed by Analysis of STUbL Suppressor, razor 1-1.

    Directory of Open Access Journals (Sweden)

    Minghua Nie

    2016-07-01

    Full Text Available Posttranslational modifications (PTMs provide dynamic regulation of the cellular proteome, which is critical for both normal cell growth and for orchestrating rapid responses to environmental stresses, e.g. genotoxins. Key PTMs include ubiquitin, the Small Ubiquitin-like MOdifier SUMO, and phosphorylation. Recently, SUMO-targeted ubiquitin ligases (STUbLs were found to integrate signaling through the SUMO and ubiquitin pathways. In general, STUbLs are recruited to target proteins decorated with poly-SUMO chains to ubiquitinate them and drive either their extraction from protein complexes, and/or their degradation at the proteasome. In fission yeast, reducing or preventing the formation of SUMO chains can circumvent the essential and DNA damage response functions of STUbL. This result indicates that whilst some STUbL "targets" have been identified, the crucial function of STUbL is to antagonize SUMO chain formation. Herein, by screening for additional STUbL suppressors, we reveal crosstalk between the serine/threonine phosphatase PP2A-Pab1B55 and the SUMO pathway. A hypomorphic Pab1B55 mutant not only suppresses STUbL dysfunction, but also mitigates the phenotypes associated with deletion of the SUMO protease Ulp2, or mutation of the STUbL cofactor Rad60. Together, our results reveal a novel role for PP2A-Pab1B55 in modulating SUMO pathway output, acting in parallel to known critical regulators of SUMOylation homeostasis. Given the broad evolutionary functional conservation of the PP2A and SUMO pathways, our results could be relevant to the ongoing attempts to therapeutically target these factors.

  6. Three-Dimensional Numerical Model of Cell Morphology during Migration in Multi-Signaling Substrates

    Science.gov (United States)

    Mousavi, Seyed Jamaleddin; Hamdy Doweidar, Mohamed

    2015-01-01

    Cell Migration associated with cell shape changes are of central importance in many biological processes ranging from morphogenesis to metastatic cancer cells. Cell movement is a result of cyclic changes of cell morphology due to effective forces on cell body, leading to periodic fluctuations of the cell length and cell membrane area. It is well-known that the cell can be guided by different effective stimuli such as mechanotaxis, thermotaxis, chemotaxis and/or electrotaxis. Regulation of intracellular mechanics and cell’s physical interaction with its substrate rely on control of cell shape during cell migration. In this notion, it is essential to understand how each natural or external stimulus may affect the cell behavior. Therefore, a three-dimensional (3D) computational model is here developed to analyze a free mode of cell shape changes during migration in a multi-signaling micro-environment. This model is based on previous models that are presented by the same authors to study cell migration with a constant spherical cell shape in a multi-signaling substrates and mechanotaxis effect on cell morphology. Using the finite element discrete methodology, the cell is represented by a group of finite elements. The cell motion is modeled by equilibrium of effective forces on cell body such as traction, protrusion, electrostatic and drag forces, where the cell traction force is a function of the cell internal deformations. To study cell behavior in the presence of different stimuli, the model has been employed in different numerical cases. Our findings, which are qualitatively consistent with well-known related experimental observations, indicate that adding a new stimulus to the cell substrate pushes the cell to migrate more directionally in more elongated form towards the more effective stimuli. For instance, the presence of thermotaxis, chemotaxis and electrotaxis can further move the cell centroid towards the corresponding stimulus, respectively, diminishing the

  7. Three-dimensional numerical model of cell morphology during migration in multi-signaling substrates.

    Directory of Open Access Journals (Sweden)

    Seyed Jamaleddin Mousavi

    Full Text Available Cell Migration associated with cell shape changes are of central importance in many biological processes ranging from morphogenesis to metastatic cancer cells. Cell movement is a result of cyclic changes of cell morphology due to effective forces on cell body, leading to periodic fluctuations of the cell length and cell membrane area. It is well-known that the cell can be guided by different effective stimuli such as mechanotaxis, thermotaxis, chemotaxis and/or electrotaxis. Regulation of intracellular mechanics and cell's physical interaction with its substrate rely on control of cell shape during cell migration. In this notion, it is essential to understand how each natural or external stimulus may affect the cell behavior. Therefore, a three-dimensional (3D computational model is here developed to analyze a free mode of cell shape changes during migration in a multi-signaling micro-environment. This model is based on previous models that are presented by the same authors to study cell migration with a constant spherical cell shape in a multi-signaling substrates and mechanotaxis effect on cell morphology. Using the finite element discrete methodology, the cell is represented by a group of finite elements. The cell motion is modeled by equilibrium of effective forces on cell body such as traction, protrusion, electrostatic and drag forces, where the cell traction force is a function of the cell internal deformations. To study cell behavior in the presence of different stimuli, the model has been employed in different numerical cases. Our findings, which are qualitatively consistent with well-known related experimental observations, indicate that adding a new stimulus to the cell substrate pushes the cell to migrate more directionally in more elongated form towards the more effective stimuli. For instance, the presence of thermotaxis, chemotaxis and electrotaxis can further move the cell centroid towards the corresponding stimulus, respectively

  8. Modeling of an aerobic biofilm reactor with double-limiting substrate kinetics: bifurcational and dynamical analysis.

    Science.gov (United States)

    Olivieri, Giuseppe; Russo, Maria Elena; Marzocchella, Antonio; Salatino, Piero

    2011-01-01

    A mathematical model of an aerobic biofilm reactor is presented to investigate the bifurcational patterns and the dynamical behavior of the reactor as a function of different key operating parameters. Suspended cells and biofilm are assumed to grow according to double limiting kinetics with phenol inhibition (carbon source) and oxygen limitation. The model presented by Russo et al. is extended to embody key features of the phenomenology of the granular-supported biofilm: biofilm growth and detachment, gas-liquid oxygen transport, phenol, and oxygen uptake by both suspended and immobilized cells, and substrate diffusion into the biofilm. Steady-state conditions and stability, and local dynamic behavior have been characterized. The multiplicity of steady states and their stability depend on key operating parameter values (dilution rate, gas-liquid mass transfer coefficient, biofilm detachment rate, and inlet substrate concentration). Small changes in the operating conditions may be coupled with a drastic change of the steady-state scenario with transcritical and saddle-node bifurcations. The relevance of concentration profiles establishing within the biofilm is also addressed. When the oxygen level in the liquid phase is <10% of the saturation level, the biofilm undergoes oxygen starvation and the active biofilm fraction becomes independent of the dilution rate. © 2011 American Institute of Chemical Engineers Biotechnol. Prog., 2011.

  9. [Performance and substrate inhibition kinetics model of nitritation process in inverse turbulent bed reactor].

    Science.gov (United States)

    Jin, Ren-Cun; Yang, Guang-Feng; Ma, Chun; Zheng, Ping

    2011-01-01

    The performance of a nitritation inverse turbulent bed (ITB) reactor was tested and the substrate inhibition kinetics characteristics of the reactor were analyzed. The results showed that a rapid reactor startup could be realized within 20 d with a strategy that combined the biofilm attachment method named "precoating carrier treatment" and "rapid suspending sludge discharge", with the feeding strategy named "low strength, high load". When operated at a hydraulic retention time of 3 h and influent NH4(+) -N of 700 mg x L(-1), corresponding to a nitrogen loading rate of 5.60 kg x (m3 x d)(-1), a maximum NH4(+) -N removal rate of 4.25 kg x (m3 x d)(-1) was observed. The maximum NO2(-) -N production rate was as high as 3.70 kg x (m3 x d)(-1). Four inhibition kinetic models (Haldane, Edwards, Aiba and Luong) were analyzed through non-linear regression to represent the inhibitions caused by substrate of nitritation process and the parameters of models were gained, which were r(max) of 1.84 kg x (m3 x d)(-1), K(IH) of 97.4 mg x L(-1) and K(m) of 0.188 mg x L(-1) for Haldane model, and r(max) of 1.83 kg x (m3 x d)(-1) and K(IA) of 114 mg x L(-1) for Aiba model. It was proposed that Haldane and Aiba models well fitted the process data harvested in the ITB reactor.

  10. SUMO E3 Ligase AtMMS21 Regulates Drought Tolerance in Arabidopsis thaliana

    Institute of Scientific and Technical Information of China (English)

    Shengchun Zhang; Yanli Qi; Ming Liu; ChengweiYang

    2013-01-01

    Post-translational modifications of proteins by small ubiquitin-like modifiers (SUMOs) play crucial roles in plant growth and development,and in stress responses.The MMS21 is a newly-identified Arabidopsis thaliana L.SUMO E3 ligase gene aside from the SIZ1,and its function requires further elucidation.Here,we show that MMS21 deficient plants display improved drought tolerance,and constitutive expression of MMS21 reduces drought tolerance.The expression of MMS21 was reduced by abscisic acid (ABA),polyethylene glycol (PEG) or drought stress.Under drought conditions,mms21 mutants showed the highest survival rate and the slowest water loss,and accumulated a higher level of free proline compared to wild-type (WT) and MMS21 over-expression plants.Stomatal aperture,seed germination and cotyledon greening analysis indicated that mms21 was hypersensitive to ABA.Molecular genetic analysis revealed that MMS21 deficiency led to elevated expression of a series of ABA-mediated stress-responsive genes,including COR15A,RD22,and P5CS1 The ABA and drought-induced stress-responsive genes,including RAB18,RD29A and RD29B,were inhibited by constitutive expression of MMS21.Moreover,ABA-induced accumulation of SUMO-protein conjugates was blocked in the mms21 mutant.We thus conclude that MMS21 plays a role in the drought stress response,likely through regulation of gene expression in an ABA-dependent pathway.

  11. Proof of concept for turbulence measurements with the RPAS SUMO during the BLLAST campaign

    Science.gov (United States)

    Båserud, Line; Reuder, Joachim; Jonassen, Marius O.; Kral, Stephan T.; Paskyabi, Mostafa B.; Lothon, Marie

    2016-10-01

    The micro-RPAS (remotely piloted aircraft system) SUMO (Small Unmanned Meteorological Observer) equipped with a five-hole-probe (5HP) system for turbulent flow measurements was operated in 49 flight missions during the BLLAST (Boundary-Layer Late Afternoon and Sunset Turbulence) field campaign in 2011. Based on data sets from these flights, we investigate the potential and limitations of airborne velocity variance and TKE (turbulent kinetic energy) estimations by an RPAS with a take-off weight below 1 kg. The integration of the turbulence probe in the SUMO system was still in an early prototype stage during this campaign, and therefore extensive post-processing of the data was required. In order to be able to calculate the three-dimensional wind vector, flow probe measurements were first synchronized with the autopilot's attitude and velocity data. Clearly visible oscillations were detected in the resulting vertical velocity, w, even after correcting for the aircraft motion. The oscillations in w were identified as the result of an internal time shift between the inertial measurement unit (IMU) and the GPS sensors, leading to insufficient motion correction, especially for the vertical wind component, causing large values of σw. Shifting the IMU 1-1.5 s forward in time with respect to the GPS yields a minimum for σw and maximum covariance between the IMU pitch angle and the GPS climb angle. The SUMO data show a good agreement to sonic anemometer data from a 60 m tower for σu, but show slightly higher values for σv and σw. Vertical TKE profiles, obtained from consecutive flight legs at different altitudes, show reasonable results, both with respect to the overall TKE level and the temporal variation. A thorough discussion of the methods used and the identified uncertainties and limitations of the system for turbulence measurements is included and should help the developers and users of other systems with similar problems.

  12. Parameter passing between molecular dynamics and continuum models for droplets on solid substrates: the static case.

    Science.gov (United States)

    Tretyakov, Nikita; Müller, Marcus; Todorova, Desislava; Thiele, Uwe

    2013-02-14

    We study equilibrium properties of polymer films and droplets on a solid substrate employing particle-based simulation techniques (molecular dynamics) and a continuum description. Parameter-passing techniques are explored that facilitate a detailed comparison of the two models. In particular, the liquid-vapor, solid-liquid, and solid-vapor interface tensions, and the Derjaguin or disjoining pressure are determined by molecular dynamics simulations. This information is then introduced into continuum descriptions accounting for (i) the full curvature and (ii) a long-wave approximation of the curvature (thin film model). A comparison of the dependence of the contact angle on droplet size indicates that the theories agree well if the contact angles are defined in a compatible manner.

  13. An Immunosensing System Using Stilbene Glycoside as a Fluorogenic Substrate for an Enzymatic Reaction Model

    Directory of Open Access Journals (Sweden)

    Ya-Fei Tan

    2008-09-01

    Full Text Available A natural product, stilbene glycoside (2,3,5,4’-tetrahydroxydiphenylethylene-2-O-glucoside, TBG, has been evaluated for the first time as a potential substrate for horseradish peroxidase (HRP-catalyzed fluorogenic reactions. The properties of TBG as a fluorogenic substrate for HRP and its application in a fluorometric enzyme-linked immunosensing system were compared with commercially available substrates such as p-hydroxyphenylpropionic acid (pHPPA, chavicol and Amplex red using Brucella melitensis antibody (BrAb as a model analyte. The immunosensing body based on HRP-BrAb was constructed by dispersing graphite, BrAg and paraffin wax at room temperature. In a competitive immunoassay procedure, the BrAb competed with HRP-BrAb to react with the immobilized BrAg. In the enzymatic reaction, the binding HRP-BrAb on the sensing body surface can catalyze the polymerization reaction of TBG by H2O2 forming fluorescent dimers and causing an increase in fluorescence intensity. TBG showed comparable ability for HRP detection and its enzyme-linked immunosensing reaction system, in a linear detection ranging of 3.5´10-8~7.6´10-6g/L and with a detection limit of 1.7´10-9 g/L. The immobilized biocomposite surface could be regenerated with excellent reproducibility (RSD=3.8% by simply polishing with an alumina paper. The proposed immunosensing system has been used to determine the BrAb in rabbit serum samples with satisfactory results.

  14. PERFORMANCE ANALYSIS OF COMPOSITE LEAF SPRING IN A DEFENCE SUMO VEHICLE

    OpenAIRE

    M. Venkatesan; V. C. SATHISH GANDHI; E. JANARTHAN

    2015-01-01

    The composite material has taking place a major role in an automobiles industries. The leaf spring, which is considered for this study is a specially designed leaf spring used in SUMO design by the ordinance factory. The leaf spring which is an automotive component used to absorb vibrations induced during the motion of vehicle. It also acts as a structure to support vertical loading due to the weight of the vehicle and payload. In this study the Finite element method is used for analysing ...

  15. Applying multibeam sonar and mathematical modeling for mapping seabed substrate and biota of offshore shallows

    Science.gov (United States)

    Herkül, Kristjan; Peterson, Anneliis; Paekivi, Sander

    2017-06-01

    Both basic science and marine spatial planning are in a need of high resolution spatially continuous data on seabed habitats and biota. As conventional point-wise sampling is unable to cover large spatial extents in high detail, it must be supplemented with remote sensing and modeling in order to fulfill the scientific and management needs. The combined use of in situ sampling, sonar scanning, and mathematical modeling is becoming the main method for mapping both abiotic and biotic seabed features. Further development and testing of the methods in varying locations and environmental settings is essential for moving towards unified and generally accepted methodology. To fill the relevant research gap in the Baltic Sea, we used multibeam sonar and mathematical modeling methods - generalized additive models (GAM) and random forest (RF) - together with underwater video to map seabed substrate and epibenthos of offshore shallows. In addition to testing the general applicability of the proposed complex of techniques, the predictive power of different sonar-based variables and modeling algorithms were tested. Mean depth, followed by mean backscatter, were the most influential variables in most of the models. Generally, mean values of sonar-based variables had higher predictive power than their standard deviations. The predictive accuracy of RF was higher than that of GAM. To conclude, we found the method to be feasible and with predictive accuracy similar to previous studies of sonar-based mapping.

  16. Diamond-like carbon films deposited on three-dimensional shape substrate model by liquid electrochemical technique

    Energy Technology Data Exchange (ETDEWEB)

    He, Y.Y. [Institute of Nano-photonics, School of Physics and Materials Engineering, Dalian Nationalities University, 116600 Dalian (China); Zhang, G.F. [School of Materials Science and Engineering, Dalian University of Technology, 116024, Dalian China (China); Zhao, Y.; Liu, D.D. [Institute of Nano-photonics, School of Physics and Materials Engineering, Dalian Nationalities University, 116600 Dalian (China); Cong, Y., E-mail: congyan@ciomp.ac.cn [Institute of Nano-photonics, School of Physics and Materials Engineering, Dalian Nationalities University, 116600 Dalian (China); Buck, V. [Thin Film Technology Group, Faculty of Physics, University Duisburg-Essen and CeNIDE, 47057 Duisburg (Germany)

    2015-09-01

    Diamond-like carbon (DLC) films were deposited on three-dimensional (3D) shape substrate model by electrolysis of 2-propanol solution at low temperature (60 °C). This 3D shape model was composed of a horizontally aligned stainless steel wafer and vertically aligned stainless steel rods. Morphology and microstructure of the films were analyzed by scanning electron microscopy (SEM), atomic force microscopy (AFM) and Raman spectroscopy, respectively. The results suggested there were only differences in film uniformity and thickness for two kinds of samples. The hydrogenated amorphous carbon films deposited on horizontally aligned substrate were smooth and homogeneous. And the film thickness of DLC films gained on the vertical substrates decreased along vertical direction. It is believed that bubble formation could enhance nucleation on the wetted capillary area. This experiment shows that deposition of DLC films by liquid phase deposition on 3D shape conductive substrates is possible. - Highlights: • DLC film is expected to be deposited on complex surface/shape substrate. • DLC film is deposited on 3D shape substrate by liquid electrochemical method. • Horizontal substrate is covered by smooth and homogeneous DLC films. • Film thickness decreases along vertical direction due to boiling effect.

  17. Mechanistic modeling of enzymatic hydrolysis of cellulose integrating substrate morphology and cocktail composition.

    Science.gov (United States)

    Huron, Maïté; Hudebine, Damien; Lopes Ferreira, Nicolas; Lachenal, Dominique

    2016-05-01

    A mechanistic model of enzymatic hydrolysis taking into account the morphology of the cellulosic particles and its evolution with time was developed. The individual behavior of the main enzymes involved in the reaction (cellobiohydrolases, endoglucanases, and β-glucosidases), as well as synergy effects, were also included. A large panel of experimental tests was done to fit and validate the model. This database included different enzymes mixtures and operating conditions and allowed to determine and compare with accuracy the adsorption and kinetic parameters of the different enzymes. Model predictions on short hydrolysis times were very satisfactory. On longer times, a deactivation constant was added to represent the hydrolysis slowdown. The model also allowed to predict the impact of enzymes ratios and initial substrate parameters (chain length distribution, polymerization degree) on hydrolysis, and to follow the evolution of these parameters with time. This model revealed general trends on the impact of cellulose morphology on hydrolysis. It is a useful tool to better understand the mechanisms involved in enzymatic hydrolysis of cellulose and to determine optimal cellulolytic cocktails for process design. © 2015 Wiley Periodicals, Inc.

  18. Modeling the behavioral substrates of associate learning and memory - Adaptive neural models

    Science.gov (United States)

    Lee, Chuen-Chien

    1991-01-01

    Three adaptive single-neuron models based on neural analogies of behavior modification episodes are proposed, which attempt to bridge the gap between psychology and neurophysiology. The proposed models capture the predictive nature of Pavlovian conditioning, which is essential to the theory of adaptive/learning systems. The models learn to anticipate the occurrence of a conditioned response before the presence of a reinforcing stimulus when training is complete. Furthermore, each model can find the most nonredundant and earliest predictor of reinforcement. The behavior of the models accounts for several aspects of basic animal learning phenomena in Pavlovian conditioning beyond previous related models. Computer simulations show how well the models fit empirical data from various animal learning paradigms.

  19. Structural Analysis of a Complex between Small Ubiquitin-like Modifier 1 (SUMO1) and the ZZ Domain of CREB-binding Protein (CBP/p300) Reveals a New Interaction Surface on SUMO

    DEFF Research Database (Denmark)

    Diehl, Carl; Akke, Mikael; Bekker-Jensen, Simon;

    2016-01-01

    )N Carr-Purcell-Meiboom-Gill (CPMG) relaxation dispersions, which define the off rates for the ZZ domain and SIM motif and show that the dynamic binding process has different characteristics for the two cases. Furthermore, in the absence of bound ligands SUMO1 transiently samples a high energy...

  20. Model-based predictions of anaerobic digestion of agricultural substrates for biogas production.

    Science.gov (United States)

    Zhou, Haidong; Löffler, Daniel; Kranert, Martin

    2011-12-01

    A modified Anaerobic Digestion Model No. 1 (ADM1), calibrated on a laboratory digester with a feeding mix of 30% weight of cow manure and 70% weight of corn silage, was implemented, showing its performances of simulation as a decision-making and planning-supporting tool for the anaerobic digestion of agricultural substrates. The virtual fermenter obtained was used to conduct simulations with different feeding compositions and loading rates of cow manure, corn silage, grass silage and rape oil. All simulations were started at the same initial state which was represented by a steady state with an organic loading rate of 2.5 kg ODM/(mdigester3∗d). The effects of the different feeding combinations on biogas composition and biogas yield were predicted reasonably, and partly verified with the available literature data. Results demonstrated that the simulations could be helpful for taking decisions on agricultural biogas plant operation or experimental set-ups, if used advisedly.

  1. Molecular modelling studies of substrate binding to the lipase from Rhizomucor miehei

    Science.gov (United States)

    Yagnik, Asutosh T.; Littlechild, Jennifer A.; Turner, Nicholas J.

    1997-05-01

    Lipase enzymes have found increasingly widespread use, especially in biotransformation reactions in organic synthesis. Due to their efficiency and high enantioselectivity, they can be employed in a variety of reactions to carry out asymmetric hydrolyses, esterifications and transesterifications. However, the reasons for their stereospecificity have not been fully correlated with the enzyme structure. Employing molecular modelling techniques and existing experimental data, a transesterification reaction using Rhizomucor miehei lipase was studied. The results indicate that the major controlling factor for this reaction is hydrophobic in nature, providing support for previous literature hypotheses. In addition, computational experiments suggest that the origin of enantioselectivity is the formation of essential hydrogen bonds in and around the catalytic triad of active site residues. Only one enantiomer of the substrate is able to form these hydrogen bonds during the formation of the first tetrahedral transition state.

  2. Structural modelling of substrate binding and inhibition in penicillin V acylase from Pectobacterium atrosepticum.

    Science.gov (United States)

    Avinash, V S; Panigrahi, Priyabrata; Suresh, C G; Pundle, Archana V; Ramasamy, Sureshkumar

    2013-08-09

    Penicillin V acylases (PVAs) and bile salt hydrolases (BSHs) have considerable sequence and structural similarity; however, they vary significantly in their substrate specificity. We have identified a PVA from a Gram-negative organism, Pectobacterium atrosepticum (PaPVA) that turned out to be a remote homolog of the PVAs and BSHs reported earlier. Even though the active site residues were conserved in PaPVA it showed high specificity towards penV and interestingly the penV acylase activity was inhibited by bile salts. Comparative modelling and docking studies were carried out to understand the structural differences of the binding site that confer this characteristic property. We show that PaPVA exhibits significant differences in structure, which are in contrast to those of known PVAs and such enzymes from Gram-negative bacteria require further investigation.

  3. Escherichia coli removal from model substrates: Underlying mechanism based on nanofluid structural forces.

    Science.gov (United States)

    Shim, Jiyoung; Nikolov, Alex; Wasan, Darsh

    2017-07-15

    Understanding the interactions between bacteria and solid surfaces that result in bacterial adhesion and removal is of immense importance for reducing foodborne illness outbreaks. A nanofluid formulation comprised of a sodium dodecyl sulfate (SDS) micellar aqueous solution in the presence of an organic acid (as a pH controller) was used to test the E. coli K12 removal from two substrates, polyvinylchloride (PVC) and partially hydrophobic glass. We investigated the bacterial removal efficacy based on the combined effect of the nanofluid's structural forces and bacterial isoelectric point. To quantify the bacteria-PVC coverage, we used fluorescence microscope. The Langmuir isotherm at the low volume fraction was applied to estimate the adsorption energy of E. coli K12. We obtained a value of about 2.5±0.2kT. This value compared favorably with the value of 2.1kT reported previously for E. coli NCTC 9002 (Vanloosdrecht et al., 1989). We applied the dynamic light scattering method to estimate the radius of the gyration of E. coli K12. The radius of the gyration was used to estimate the limit of surface area covered by the bacterium and compared it to the surface area measured from the image taken with fluorescence microscope. We found that they are in good agreement with each other. We modeled the nanofluid oscillatory structural energy against the E. coli K12 adsorption energy by applying the statistical mechanics approach. Based on the model prediction, the oscillatory interaction energy was estimated at the vertex between a bacterium and the substrate (i.e., the wedge film's interaction energy at one particle layer). The evaluated film's repulsive energy due to the oscillatory structural forces (OSF) was about 15.6±4.4kT of the 0.02M SMNF (the SDS micellar nanofluid formulation) and several times higher than the bacterial adsorption energy, 2.5±0.2kT. The OSF of the 0.06M SMNF was measured by AFM (the oscillatory decay force curve). The period and number of

  4. A finite element formulation for modeling dynamic wetting on flexible substrates and in deformable porous media.

    Energy Technology Data Exchange (ETDEWEB)

    Schunk, Peter Randall; Cairncross, Richard A. (Drexel University, Philadelphia, PA); Madasu, S. (Drexel University, Philadelphia, PA)

    2004-03-01

    This report summarizes research advances pursued with award funding issued by the DOE to Drexel University through the Presidential Early Career Award (PECASE) program. Professor Rich Cairncross was the recipient of this award in 1997. With it he pursued two related research topics under Sandia's guidance that address the outstanding issue of fluid-structural interactions of liquids with deformable solid materials, focusing mainly on the ubiquitous dynamic wetting problem. The project focus in the first four years was aimed at deriving a predictive numerical modeling approach for the motion of the dynamic contact line on a deformable substrate. A formulation of physical model equations was derived in the context of the Galerkin finite element method in an arbitrary Lagrangian/Eulerian (ALE) frame of reference. The formulation was successfully integrated in Sandia's Goma finite element code and tested on several technologically important thin-film coating problems. The model equations, the finite-element implementation, and results from several applications are given in this report. In the last year of the five-year project the same physical concepts were extended towards the problem of capillary imbibition in deformable porous media. A synopsis of this preliminary modeling and experimental effort is also discussed.

  5. SUMO-conjugating enzyme E2 UBC9 mediates viral immediate-early protein SUMOylation in crayfish to facilitate reproduction of white spot syndrome virus.

    Science.gov (United States)

    Chen, An-Jing; Gao, Lu; Wang, Xian-Wei; Zhao, Xiao-Fan; Wang, Jin-Xing

    2013-01-01

    Successful viruses have evolved superior strategies to escape host defenses or exploit host biological pathways. Most of the viral immediate-early (ie) genes are essential for viral infection and depend solely on host proteins; however, the molecular mechanisms are poorly understood. In this study, we focused on the modification of viral IE proteins by the crayfish small ubiquitin-related modifier (SUMO) and investigated the role of SUMOylation during the viral life cycle. SUMO and SUMO ubiquitin-conjugating enzyme 9 (UBC9) involved in SUMOylation were identified in red swamp crayfish (Procambarus clarkii). Both SUMO and UBC9 were upregulated in crayfish challenged with white spot syndrome virus (WSSV). Replication of WSSV genes increased in crayfish injected with recombinant SUMO or UBC9, but injection of mutant SUMO or UBC9 protein had no effect. Subsequently, we analyzed the mechanism by which crayfish SUMOylation facilitates WSSV replication. Crayfish UBC9 bound to all three WSSV IE proteins tested, and one of these IE proteins (WSV051) was covalently modified by SUMO in vitro. The expression of viral ie genes was affected and that of late genes was significantly inhibited in UBC9-silenced or SUMO-silenced crayfish, and the inhibition effect was rescued by injection of recombinant SUMO or UBC9. The results of this study demonstrate that viral IE proteins can be modified by crayfish SUMOylation, prompt the expression of viral genes, and ultimately benefit WSSV replication. Understanding of the mechanisms by which viruses exploit host components will greatly improve our knowledge of the virus-host "arms race" and contribute to the development of novel methods against virulent viruses.

  6. Elg1, the major subunit of an alternative RFC complex, interacts with SUMO-processing proteins.

    Science.gov (United States)

    Parnas, Oren; Amishay, Rona; Liefshitz, Batia; Zipin-Roitman, Adi; Kupiec, Martin

    2011-09-01

    PCNA is a homotrimeric ring with important roles in DNA replication and repair. PCNA is loaded and unloaded by the RFC complex, which is composed of five subunits (Rfc1-5). Three additional complexes that share with RFC the small subunits (Rfc2-5) and contain alternative large subunits were found in yeast and other eukaryotes. We have recently reported that one of these, the Elg1-RFC complex, interacts with SUMOylated PCNA and may play a role in its unloading during DNA repair. Here we report that a yeast-two-hybrid screen with the N terminus of Elg1(which interacts with SUMOylated PCNA) uncovered interactions with proteins that belong to the SUMO pathway, including Slx5 and Slx8, which form an E3 ubiquitin ligase that ubiquitinates SUMOylated proteins. Mutations in SLX5 result in a genomic instability phenotype similar to that of elg1 mutants. The physical interaction between the N terminus of Elg1 and Slx5 is mediated by poly-SUMO chains but not by PCNA modifications, and requires Siz2, but not Siz1, activity. Thus our results highlight the many important roles played by Elg1, some of which are PCNA-dependent and some PCNA-independent.

  7. Antagonistic roles of ubiquitin ligase HEI10 and SUMO ligase RNF212 regulate meiotic recombination.

    Science.gov (United States)

    Qiao, Huanyu; Prasada Rao, H B D; Yang, Ye; Fong, Jared H; Cloutier, Jeffrey M; Deacon, Dekker C; Nagel, Kathryn E; Swartz, Rebecca K; Strong, Edward; Holloway, J Kim; Cohen, Paula E; Schimenti, John; Ward, Jeremy; Hunter, Neil

    2014-02-01

    Crossover recombination facilitates the accurate segregation of homologous chromosomes during meiosis. In mammals, poorly characterized regulatory processes ensure that every pair of chromosomes obtains at least one crossover, even though most recombination sites yield non-crossovers. Designation of crossovers involves selective localization of the SUMO ligase RNF212 to a minority of recombination sites, where it stabilizes pertinent factors such as MutSγ (ref. 4). Here we show that the ubiquitin ligase HEI10 (also called CCNB1IP1) is essential for this crossover/non-crossover differentiation process. In HEI10-deficient mice, RNF212 localizes to most recombination sites, and dissociation of both RNF212 and MutSγ from chromosomes is blocked. Consequently, recombination is impeded, and crossing over fails. In wild-type mice, HEI10 accumulates at designated crossover sites, suggesting that it also has a late role in implementing crossing over. As with RNF212, dosage sensitivity for HEI10 indicates that it is a limiting factor for crossing over. We suggest that SUMO and ubiquitin have antagonistic roles during meiotic recombination that are balanced to effect differential stabilization of recombination factors at crossover and non-crossover sites.

  8. PERFORMANCE ANALYSIS OF COMPOSITE LEAF SPRING IN A DEFENCE SUMO VEHICLE

    Directory of Open Access Journals (Sweden)

    M. VENKATESAN

    2015-05-01

    Full Text Available The composite material has taking place a major role in an automobiles industries. The leaf spring, which is considered for this study is a specially designed leaf spring used in SUMO design by the ordinance factory. The leaf spring which is an automotive component used to absorb vibrations induced during the motion of vehicle. It also acts as a structure to support vertical loading due to the weight of the vehicle and payload. In this study the Finite element method is used for analysing the composite spring for different parameters such us stress, deformation and mode frequencies for three different ratios of epoxy and E fiberglass materials. The composite specimen has been made in the three different ratios of material combination by hand layout moulding technique. The three different samples are 40% epoxy and 60% Efiberglass, 60% epoxy and 40% E-fiberglass, 70% epoxy and 30% E-fiberglass. The experiments were carried out for different test like tensile test, flexural test and hardness test. The experimental results are well within the simulation results and identified that the 40% epoxy and 60% E-fiberg as composite leaf spring is suitable for designing a spring in SUMO vehicle.

  9. Centromere binding and a conserved role in chromosome stability for SUMO-dependent ubiquitin ligases.

    Directory of Open Access Journals (Sweden)

    Loes A L van de Pasch

    Full Text Available The Saccharomyces cerevisiae Slx5/8 complex is the founding member of a recently defined class of SUMO-targeted ubiquitin ligases (STUbLs. Slx5/8 has been implicated in genome stability and transcription, but the precise contribution is unclear. To characterise Slx5/8 function, we determined genome-wide changes in gene expression upon loss of either subunit. The majority of mRNA changes are part of a general stress response, also exhibited by mutants of other genome integrity pathways and therefore indicative of an indirect effect on transcription. Genome-wide binding analysis reveals a uniquely centromeric location for Slx5. Detailed phenotype analyses of slx5Δ and slx8Δ mutants show severe mitotic defects that include aneuploidy, spindle mispositioning, fish hooks and aberrant spindle kinetics. This is associated with accumulation of the PP2A regulatory subunit Rts1 at centromeres prior to entry into anaphase. Knockdown of the human STUbL orthologue RNF4 also results in chromosome segregation errors due to chromosome bridges. The study shows that STUbLs have a conserved role in maintenance of chromosome stability and links SUMO-dependent ubiquitination to a centromere-specific function during mitosis.

  10. Negative regulation of TLR inflammatory signaling by the SUMO-deconjugating enzyme SENP6.

    Directory of Open Access Journals (Sweden)

    Xing Liu

    Full Text Available The signaling of Toll-like receptors (TLRs induces host defense against microbial invasion. Protein posttranslational modifications dynamically shape the strength and duration of the signaling pathways. It is intriguing to explore whether de-SUMOylation could modulate the TLR signaling. Here we identified SUMO-specific protease 6 (SENP6 as an intrinsic attenuator of the TLR-triggered inflammation. Depletion of SENP6 significantly potentiated the NF-κB-mediated induction of the proinflammatory genes. Consistently, SENP6-knockdown mice were more susceptible to endotoxin-induced sepsis. Mechanistically, the small ubiquitin-like modifier 2/3 (SUMO-2/3 is conjugated onto the Lysine residue 277 of NF-κB essential modifier (NEMO/IKKγ, and this impairs the deubiquitinase CYLD to bind NEMO, thus strengthening the inhibitor of κB kinase (IKK activation. SENP6 reverses this process by catalyzing the de-SUMOylation of NEMO. Our study highlights the essential function of the SENP family in dampening TLR signaling and inflammation.

  11. Structural analysis and evolution of specificity of the SUMO UFD E1-E2 interactions

    Science.gov (United States)

    Liu, Bing; Lois, L. Maria; Reverter, David

    2017-01-01

    SUMO belongs to the ubiquitin-like family (UbL) of protein modifiers. SUMO is conserved among eukaryotes and is essential for the regulation of processes such as DNA damage repair, transcription, DNA replication and mitosis. UbL modification of proteins occurs via a specific enzymatic cascade formed by the crosstalk between the E1-activating enzyme, the E2-conjugating enzyme and the E3-ligase. An essential discrimination step in all UbL modifiers corresponds to the interaction between E1 and E2 enzymes, which is mediated by the recruitment of the E2 to the UFD domain (Ubiquitin-Fold Domain) of the E1 enzyme. To gain insights in the properties of this interface, we have compared the structures of the complexes between E1 UFD domain and E2 in human and yeast, revealing two alternative UFD platforms that interact with a conserved E2. Comparative sequence analysis of the E1 UFD domain indicates that the E2 binding region has been conserved across phylogenetic closely related species, in which higher sequence conservation can be found in the E2 binding region than in the entire UFD domain. These distinctive strategies for E1-E2 interactions through the UFD domain might be the consequence of a high selective pressure to ensure specificity of each modifier conjugation system. PMID:28165030

  12. Uncovering SUMOylation Dynamics during Cell-Cycle Progression Reveals FoxM1 as a Key Mitotic SUMO Target Protein

    DEFF Research Database (Denmark)

    Schimmel, Joost; Eifler, Karolin; Sigurdsson, Jón Otti;

    2014-01-01

    Loss of small ubiquitin-like modification (SUMOylation) in mice causes genomic instability due to the missegregation of chromosomes. Currently, little is known about the identity of relevant SUMO target proteins that are involved in this process and about global SUMOylation dynamics during cell-c...

  13. The Smc5-Smc6 complex and SUMO modification of Rad52 regulates recombinational repair at the ribosomal gene locus

    DEFF Research Database (Denmark)

    Torres-Rosell, Jordi; Sunjevaric, Ivana; De Piccoli, Giacomo;

    2007-01-01

    at an extranucleolar site. The nucleolar exclusion of Rad52 recombination foci entails Mre11 and Smc5-Smc6 complexes and depends on Rad52 SUMO (small ubiquitin-related modifier) modification. Remarkably, mutations that abrogate these activities result in the formation of Rad52 foci within the nucleolus and cause r...

  14. Cleavage of model substrates by archaeal RNase P: role of protein cofactors in cleavage-site selection.

    Science.gov (United States)

    Sinapah, Sylvie; Wu, Shiying; Chen, Yu; Pettersson, B M Fredrik; Gopalan, Venkat; Kirsebom, Leif A

    2011-02-01

    RNase P is a catalytic ribonucleoprotein primarily involved in tRNA biogenesis. Archaeal RNase P comprises a catalytic RNase P RNA (RPR) and at least four protein cofactors (RPPs), which function as two binary complexes (POP5•RPP30 and RPP21• RPP29). Exploiting the ability to assemble a functional Pyrococcus furiosus (Pfu) RNase P in vitro, we examined the role of RPPs in influencing substrate recognition by the RPR. We first demonstrate that Pfu RPR, like its bacterial and eukaryal counterparts, cleaves model hairpin loop substrates albeit at rates 90- to 200-fold lower when compared with cleavage by bacterial RPR, highlighting the functionally comparable catalytic cores in bacterial and archaeal RPRs. By investigating cleavage-site selection exhibited by Pfu RPR (±RPPs) with various model substrates missing consensus-recognition elements, we determined substrate features whose recognition is facilitated by either POP5•RPP30 or RPP21•RPP29 (directly or indirectly via the RPR). Our results also revealed that Pfu RPR + RPP21•RPP29 displays substrate-recognition properties coinciding with those of the bacterial RPR-alone reaction rather than the Pfu RPR, and that this behaviour is attributable to structural differences in the substrate-specificity domains of bacterial and archaeal RPRs. Moreover, our data reveal a hierarchy in recognition elements that dictates cleavage-site selection by archaeal RNase P.

  15. Cancer cell mechanics with altered cytoskeletal behavior and substrate effects: A 3D finite element modeling study.

    Science.gov (United States)

    Katti, Dinesh R; Katti, Kalpana S

    2017-05-23

    A robust computational model of a cancer cell is presented using finite element modeling. The model accurately captures nuances of the various components of the cellular substructure. The role of degradation of cytoskeleton on overall elastic properties of the cancer cell is reported. The motivation for degraded cancer cellular substructure, the cytoskeleton is the observation that the innate mechanics of cytoskeleton is disrupted by various anti-cancer drugs as therapeutic treatments for the destruction of the cancer tumors. We report a significant influence on the degradation of the cytoskeleton on the mechanics of cancer cell. Further, a simulations based study is reported where we evaluate mechanical properties of the cancer cell attached to a variety of substrates. The loading of the cancer cell is less influenced by nature of the substrate, but low modulus substrates such as osteoblasts and hydrogels indicate a significant change in unloading behavior and also the plastic deformation. Overall, softer substrates such as osteoblasts and other bone cells result in a much altered unloading response as well as significant plastic deformation. These substrates are relevant to metastasis wherein certain type of cancers such as prostate and breast cancer cells migrate to the bone and colonize through mesenchymal to epithelial transition. The modeling study presented here is an important first step in the development of strong predictive methodologies for cancer progression. Copyright © 2017 Elsevier Ltd. All rights reserved.

  16. Substrate reduction augments the efficacy of enzyme therapy in a mouse model of Fabry disease.

    Directory of Open Access Journals (Sweden)

    John Marshall

    Full Text Available Fabry disease is an X-linked glycosphingolipid storage disorder caused by a deficiency in the activity of the lysosomal hydrolase α-galactosidase A (α-gal. This deficiency results in accumulation of the glycosphingolipid globotriaosylceramide (GL-3 in lysosomes. Endothelial cell storage of GL-3 frequently leads to kidney dysfunction, cardiac and cerebrovascular disease. The current treatment for Fabry disease is through infusions of recombinant α-gal (enzyme-replacement therapy; ERT. Although ERT can markedly reduce the lysosomal burden of GL-3 in endothelial cells, variability is seen in the clearance from several other cell types. This suggests that alternative and adjuvant therapies may be desirable. Use of glucosylceramide synthase inhibitors to abate the biosynthesis of glycosphingolipids (substrate reduction therapy, SRT has been shown to be effective at reducing substrate levels in the related glycosphingolipidosis, Gaucher disease. Here, we show that such an inhibitor (eliglustat tartrate, Genz-112638 was effective at lowering GL-3 accumulation in a mouse model of Fabry disease. Relative efficacy of SRT and ERT at reducing GL-3 levels in Fabry mouse tissues differed with SRT being more effective in the kidney, and ERT more efficacious in the heart and liver. Combination therapy with ERT and SRT provided the most complete clearance of GL-3 from all the tissues. Furthermore, treatment normalized urine volume and uromodulin levels and significantly delayed the loss of a nociceptive response. The differential efficacies of SRT and ERT in the different tissues indicate that the combination approach is both additive and complementary suggesting the possibility of an improved therapeutic paradigm in the management of Fabry disease.

  17. Modeling of metal nanocluster growth on patterned substrates and surface pattern formation under ion bombardment

    Energy Technology Data Exchange (ETDEWEB)

    Numazawa, Satoshi

    2012-11-01

    This work addresses the metal nanocluster growth process on prepatterned substrates, the development of atomistic simulation method with respect to an acceleration of the atomistic transition states, and the continuum model of the ion-beam inducing semiconductor surface pattern formation mechanism. Experimentally, highly ordered Ag nanocluster structures have been grown on pre-patterned amorphous SiO{sub 2} surfaces by oblique angle physical vapor deposition at room temperature. Despite the small undulation of the rippled surface, the stripe-like Ag nanoclusters are very pronounced, reproducible and well-separated. The first topic is the investigation of this growth process with a continuum theoretical approach to the surface gas condensation as well as an atomistic cluster growth model. The atomistic simulation model is a lattice-based kinetic Monte-Carlo (KMC) method using a combination of a simplified inter-atomic potential and experimental transition barriers taken from the literature. An effective transition event classification method is introduced which allows a boost factor of several thousand compared to a traditional KMC approach, thus allowing experimental time scales to be modeled. The simulation predicts a low sticking probability for the arriving atoms, millisecond order lifetimes for single Ag monomers and {approx}1 nm square surface migration ranges of Ag monomers. The simulations give excellent reproduction of the experimentally observed nanocluster growth patterns. The second topic specifies the acceleration scheme utilized in the metallic cluster growth model. Concerning the atomistic movements, a classical harmonic transition state theory is considered and applied in discrete lattice cells with hierarchical transition levels. The model results in an effective reduction of KMC simulation steps by utilizing a classification scheme of transition levels for thermally activated atomistic diffusion processes. Thermally activated atomistic movements

  18. Fibrinogen adsorption mechanisms at the gold substrate revealed by QCM-D measurements and RSA modeling.

    Science.gov (United States)

    Kubiak, Katarzyna; Adamczyk, Zbigniew; Cieśla, Michał

    2016-03-01

    Adsorption kinetics of fibrinogen at a gold substrate at various pHs was thoroughly studied using the QCM-D method. The experimental were interpreted in terms of theoretical calculations performed according to the random sequential adsorption model (RSA). In this way, the hydration functions and water factors of fibrinogen monolayers were quantitatively evaluated at various pHs. It was revealed that for the lower range of fibrinogen coverage the hydration function were considerably lower than previously obtained for the silica sensor [33]. The lower hydration of fibrinogen monolayers on the gold sensor was attributed to its higher roughness. However, for higher fibrinogen coverage the hydration functions for both sensors became identical exhibiting an universal behavior. By using the hydration functions, the fibrinogen adsorption/desorption runs derived from QCM-D measurements were converted to the Γd vs. the time relationships. This allowed to precisely determine the maximum coverage that varied between 1.6mgm(-2) at pH 3.5 and 4.5mgm(-2) at pH 7.4 (for ionic strength of 0.15M). These results agree with theoretical eRSA modeling and previous experimental data derived by using ellipsometry, OWLS and TIRF. Various fibrinogen adsorption mechanisms were revealed by exploiting the maximum coverage data. These results allow one to develop a method for preparing fibrinogen monolayers of well-controlled coverage and molecule orientation. Copyright © 2015 Elsevier B.V. All rights reserved.

  19. Modeling fungicides mobility in undisturbed vineyard soil cores unamended and amended with spent mushroom substrates.

    Science.gov (United States)

    Marín-Benito, Jesús María; Rodríguez-Cruz, María Sonia; Sánchez-Martín, María Jesús; Mamy, Laure

    2015-09-01

    The performance of the pesticide fate model PRZM to predict the fate of two fungicides, penconazole and metalaxyl, and the major metabolite of metalaxyl (CGA-62826), in amended and unamended vineyard soils was tested from undisturbed soils columns experiments. Three different treatments were tested in two soils: control soil (unamended), and soil amended with fresh or composted spent mushroom substrates, which correspond to common agricultural practices in Spain. Leaching experiments were performed under non-saturated flow conditions. The model was parameterized with laboratory and literature data, and using pedotransfer functions. It was first calibrated for water flow against chloride breakthrough curves. The key parameter was the hydrodynamic dispersion coefficient (DISP). No leaching of penconazole, the most hydrophobic fungicide, was observed. It remained in the top 0-8 cm of the column. In any case, simulations were highly correlated to the experimental results. On the contrary, metalaxyl and its metabolite were consistently found in the leachates. A calibration step of the Kd of metalaxyl and CGA-62826 and of DISP for CGA-62826 was necessary to obtain good prediction of the leaching of both compounds. PRZM generally simulated acceptable metalaxyl vertical distribution in the soil profiles although results were overestimated for its metabolite. Nevertheless, PRZM can be reasonably used to assess the leaching (through breakthrough curves) and vertical distribution of fungicides in amended soils, knowing their DISP values. Copyright © 2015 Elsevier Ltd. All rights reserved.

  20. Homology modeling of Homo sapiens lipoic acid synthase: Substrate docking and insights on its binding mode.

    Science.gov (United States)

    Krishnamoorthy, Ezhilarasi; Hassan, Sameer; Hanna, Luke Elizabeth; Padmalayam, Indira; Rajaram, Rama; Viswanathan, Vijay

    2017-05-07

    Lipoic acid synthase (LIAS) is an iron-sulfur cluster mitochondrial enzyme which catalyzes the final step in the de novo pathway for the biosynthesis of lipoic acid, a potent antioxidant. Recently there has been significant interest in its role in metabolic diseases and its deficiency in LIAS expression has been linked to conditions such as diabetes, atherosclerosis and neonatal-onset epilepsy, suggesting a strong inverse correlation between LIAS reduction and disease status. In this study we use a bioinformatics approach to predict its structure, which would be helpful to understanding its role. A homology model for LIAS protein was generated using X-ray crystallographic structure of Thermosynechococcus elongatus BP-1 (PDB ID: 4U0P). The predicted structure has 93% of the residues in the most favour region of Ramachandran plot. The active site of LIAS protein was mapped and docked with S-Adenosyl Methionine (SAM) using GOLD software. The LIAS-SAM complex was further refined using molecular dynamics simulation within the subsite 1 and subsite 3 of the active site. To the best of our knowledge, this is the first study to report a reliable homology model of LIAS protein. This study will facilitate a better understanding mode of action of the enzyme-substrate complex for future studies in designing drugs that can target LIAS protein. Copyright © 2017 Elsevier Ltd. All rights reserved.

  1. Genetic Polymorphism of SUMO-Specific Cysteine Proteases - SENP1 and SENP2 in Breast Cancer.

    Science.gov (United States)

    Mirecka, Alicja; Morawiec, Zbigniew; Wozniak, Katarzyna

    2016-10-01

    SENP proteases take part in post-translational modification of proteins known as sumoylation. They catalyze three distinct processes during sumoylation: processing of SUMO protein, deconjugation of SUMO from the target protein, and chain editing which mentions to the dismantling of SUMO chain. Many proteins that are involved in the basic processes of cells, such as regulation of transcription, DNA repair or cell cycle control, are sumoylated. The aim of these studies was to investigate an association between polymorphic variants (SNPs) of the SENP1 gene (c.1691 + 36C > T, rs12297820) and SENP2 gene (c.902C > A, p.Thr301Lys, rs6762208) and a risk of breast cancer occurrence. We performed a case-control study in 324 breast cancer cases and 335 controls using PCR-RLFP. In the case of the SENP1 gene polymorphism we did not find any association between this polymorphism and breast cancer risk. In the case of SENP2 gene polymorphism we observed higher risk of breast cancer for carriers of the A allele (OR =1.33; 95 % CI 1.04-1.69). Our analysis also showed the genotype C/C (OR =0.67, 95 % CI 0.48-0.93) and the allele C (OR =0.75, 95 % CI 0.59-0.69) of this polymorphism decrease a risk of breast cancer. We also checked the distribution of genotypes and frequency of alleles of the SENP1 and SENP2 genes polymorphisms in groups of patients with different hormone receptor status, patients with positive and negative lymph node status and patients with different tumor grade. Odds ratio analysis showed a higher risk of metastases in women with the genotype C/C (OR =2.07, 95 % CI 1.06-4.05) and allele C (OR =2.10 95 % CI 1.10-4.01) of the c.1691 + 36C > T SENP1 gene polymorphism. Moreover, we observed reduced risk in women with the allele T (OR =0.48, 95 % CI 0.25-0.91) in this polymorphic site. In the case of SENP2 gene polymorphism we observed that the A/A genotype correlated with the lack of estrogen receptor (OR =1.94, 95 % CI 1.04-3.62). Our results suggest

  2. The SUMOylation of Kif18A and its regulatory mechanism%Kif18A 的 SUMO 化修饰及其调控机制

    Institute of Scientific and Technical Information of China (English)

    李艳; 夏南松; 左勇

    2015-01-01

    Objective To investigate the SUMOylation of Kif18A and its regulatory mechanism.Methods Plas-mids Flag -Kif18A and HA -SUMO1 or His -SUMO1 were co -transfected into HEK293T cells.Flag -Kif18A proteins were pulled down by Flag M2 beads and its SUMOylation was detected by immunoblotting with anti -HA.His -SUMO1 conjugated proteins were also precipitated by TALON Resin and the level of SUMOylated Kif18A was detected by immno-blotting with Flag antibody.The potential sites of Kif18A SUMOlation were predicted by SUMOsp 2.0 software and its coding sequences were mutated from lysine to arginine on the Flag -Kif18A plasmid.Kif18A wide and mutant types were separately expressed in HEK293T cells.The change in the quantity of SUMOylated Kif18A was adopted to determine the site of SUMOylation in Flag -Kif18A.Furthermore, plasmids Flag -Kif18A, HA -SUMO1 and SENP1 WT or mutant were co -transfected into HEK293T cells.Then the amount of SUMOylated Kif18A was detected by immunoprecipitation to determine whether Kif18A can be de -SUMOylated by SENP1.Finally, HeLa cells were synchronized in G2 /M phase by Nocodazole treatment.The SUMOylation of Kif18A in HeLa cells in G2 /M phase were detected by immunoprecipitati-on and immunobloting.Results Kif18A could be SUMOylated by SUMO1 or SUMO2.K47 and K148 were two impor-tant sites for Kif18A SUMOylation.Besides, Kif18A could be de -SUMOylated by SENP1.The SUMOylation of Kif18A was decreased in HeLa cells arrested in G2 /M phase.Conclusion Kif18A can be SUMOylated and its SUMOylation is regulated by mitosis progression.%目的:探讨 Kif18A 的 SUMO 化修饰及其调控机制。方法在 HEK293T 细胞中共转染 Flag -Kif18A、HA -SUMO1或 His -SUMO1质粒,免疫沉淀富集 Flag -Kif18A,免疫印迹检测 HA -SUMO;用 TALON 树脂富集His -SUMO1蛋白,免疫印迹检测 Flag -Kif18A 考察 Kif18A 的 SUMO 化水平。利用 SUMOsp 2.0软件预测Kif18A 可能发生 SUMO 化的潜在位点,将 Flag -Kif18A

  3. Establishment of a P-glycoprotein substrate screening model and its preliminary application

    Institute of Scientific and Technical Information of China (English)

    Yi Wang; Jiang Cao; Su Zeng

    2004-01-01

    studying drug intestinal absorption mechanism,predicting the drug permeability characteristics and screening new multi-drug resistance reversing agents. With this model,quercetin can be found to be transported by P-gp, and it is a P-gp substrate.

  4. Molecular metal sulfide cluster model for substrate binding to oil-refinery hydrodesulfurization catalysts.

    Science.gov (United States)

    Herbst, Konrad; Monari, Magda; Brorson, Michael

    2002-03-25

    Reaction between [(eta5-Cp')3Mo3S4]+ and [Ni(1,5-cod)2] (Cp' = methylcyclopentadienyl; 1,5-cod = 1,5-cyclooctadiene) in THF at ambient temperature yielded a coordinatively unsaturated cubane-like cluster cation, [(eta5-Cp')3Mo3S4Ni]+. The ligand sphere at the Ni atom could be saturated by coordinating dimethyl sulfide, diethyl sulfide, di(tert-butyl) sulfide, tetrahydrothiophene, thiochroman-4-ol, 1,4-dithiane, pyridine, quinoline, or 4,4'-bipyridine. The products structurally model a mode of substrate coordination on proposed binding sites of heterogeneous MoNi sulfide hydrotreating catalysts. No stable coordination compounds could be isolated for thiophene derivatives. X-ray crystal structures are reported for the ligand-bridged dicluster compounds [[(eta5-Cp')3Mo3S4Ni]2(mu-C4H4S2)][pts]2 (C4H8S2 = 1,4-dithiane) and [[(eta5-Cp')3Mo3S4Ni]2(mu-bipy)][pts]2 (bipy = 4,4'-bipyridine).

  5. Active-site models for complexes of quinolinate synthase with substrates and intermediates

    Energy Technology Data Exchange (ETDEWEB)

    Soriano, Erika V.; Zhang, Yang; Colabroy, Keri L.; Sanders, Jennie M.; Settembre, Ethan C.; Dorrestein, Pieter C.; Begley, Tadhg P.; Ealick, Steven E., E-mail: see3@cornell.edu [Cornell University, Ithaca, NY 14853-1301 (United States)

    2013-09-01

    Structural studies of quinolinate synthase suggest a model for the enzyme–substrate complex and an enzyme–intermediate complex with a [4Fe–4S] cluster. Quinolinate synthase (QS) catalyzes the condensation of iminoaspartate and dihydroxyacetone phosphate to form quinolinate, the universal precursor for the de novo biosynthesis of nicotinamide adenine dinucleotide. QS has been difficult to characterize owing either to instability or lack of activity when it is overexpressed and purified. Here, the structure of QS from Pyrococcus furiosus has been determined at 2.8 Å resolution. The structure is a homodimer consisting of three domains per protomer. Each domain shows the same topology with a four-stranded parallel β-sheet flanked by four α-helices, suggesting that the domains are the result of gene triplication. Biochemical studies of QS indicate that the enzyme requires a [4Fe–4S] cluster, which is lacking in this crystal structure, for full activity. The organization of domains in the protomer is distinctly different from that of a monomeric structure of QS from P. horikoshii [Sakuraba et al. (2005 ▶), J. Biol. Chem.280, 26645–26648]. The domain arrangement in P. furiosus QS may be related to protection of cysteine side chains, which are required to chelate the [4Fe–4S] cluster, prior to cluster assembly.

  6. Influence of respiratory substrate in carbon steel corrosion by a Sulphate Reducing Prokaryote model organism.

    Science.gov (United States)

    Dall'agnol, Leonardo T; Cordas, Cristina M; Moura, José J G

    2014-06-01

    Sulphate Reducing Prokaryotes (SRP) are an important group of microorganisms involved in biocorrosion processes. Sulphide production is recognized as a fundamental cause of corrosion and nitrate is often used as treatment. The present work analyses the influence of respiratory substrates in the metal, from off-shore installations, SRP influenced corrosion, using Desulfovibrio desulfuricans ATTC 27774 as model organism, since this can switch from sulphate to nitrate. Open Circuit Potential over 6days in different conditions was measured, showing an increase around 200 and 90mV for the different media. Tafel plots were constructed allowing Ecorr and jcorr calculations. For SRP in sulphate and nitrate media Ecorr values of -824 and -728mV, and jcorr values of 2.5 and 3.7μAcm(-2), respectively, were attained indicating that in nitrate, the resultant corrosion rate is larger than in sulphate. Also, it is shown that the equilibrium of sulphide in the solution/gas phases is a key factor to the evolution of corrosion Nitrate prevents pitting but promotes general corrosion and increases the corrosion potential and iron dissolution 40 times when compared to sulphate. Our results demonstrate that nitrate injection strategy in oil fields has to be considered carefully as option to reduce souring and localized corrosion.

  7. The ubiquitin- and SUMO-dependent signaling response to DNA double-strand breaks

    DEFF Research Database (Denmark)

    Bekker-Jensen, Simon; Mailand, Niels

    2011-01-01

    DNA double-strand breaks (DSBs) represent the most destructive type of chromosomal lesion and trigger rapid chromatin restructuring accompanied by accumulation of proteins in the vicinity of the DSB. Non-proteolytic ubiquitylation of chromatin surrounding DSBs, mediated by the RNF8/RNF168 ubiquitin...... ligase cascade, has emerged as a key mechanism for restoration of genome integrity by licensing the DSB-modified chromatin to concentrate genome caretaker proteins such as 53BP1 and BRCA1 near the lesions. In parallel, SUMOylation of upstream DSB regulators is also required for execution...... of this ubiquitin-dependent chromatin response, but its molecular basis is currently unclear. Here, we discuss recent insights into how ubiquitin- and SUMO-dependent signaling processes cooperate to orchestrate protein interactions with sites of DNA damage to facilitate DSB repair....

  8. Measurement and thermal modeling of sapphire substrate temperature at III-Nitride MOVPE conditions

    Science.gov (United States)

    Creighton, J. Randall; Coltrin, Michael E.; Figiel, Jeffrey J.

    2017-04-01

    Growth rates and alloy composition of AlGaN grown by MOVPE is often very temperature dependent due to the presence of gas-phase parasitic chemical processes. These processes make wafer temperature measurement highly important, but in fact such measurements are very difficult because of substrate transparency in the near-IR ( 900 nm) where conventional pyrometers detect radiation. The transparency problem can be solved by using a mid-IR pyrometer operating at a wavelength ( 7500 nm) where sapphire is opaque. We employ a mid-IR pyrometer to measure the sapphire wafer temperature and simultaneously a near-IR pyrometer to measure wafer pocket temperature, while varying reactor pressure in both a N2 and H2 ambient. Near 1300 °C, as the reactor pressure is lowered from 300 Torr to 10 Torr the wafer temperature drops dramatically, and the ∆T between the pocket and wafer increases from 20 °C to 250 °C. Without the mid-IR pyrometer the large wafer temperature change with pressure would not have been noted. In order to explain this behavior we have developed a quasi-2D thermal model that includes a proper accounting of the pressure-dependent thermal contact resistance, and also accounts for sapphire optical transmission. The model and experimental results demonstrate that at most growth conditions the majority of the heat is transported from the wafer pocket to the wafer via gas conduction, in the free molecular flow limit. In this limit gas conductivity is independent of gap size but first order in pressure, and can quantitatively explain results from 20 to 300 Torr. Further analysis yields a measure of the thermal accommodation coefficients; α(H2) =0.23, α(N2) =0.50, which are in the range typically measured.

  9. A novel high-voltage device structure with an N+ ring in substrate and the breakdown voltage model

    Institute of Scientific and Technical Information of China (English)

    Li Qi; Zhu Jinluan; Wang Weidong; Yue Hongwei; Jin Liangnian

    2011-01-01

    A novel high-voltage device structure with a floating heavily doped N+ ring embedded in the substrate is reported,which is called FR LDMOS.When the N+ ring is introduced in the device substrate,the electric field peak of the main junction is reduced due to the transfer of the voltage from the main junction to the N+ ring junction,and the vertical breakdown characteristic is improved significantly.Based on the Poisson equation of cylindrical coordinates,a breakdown voltage model is developed.The numerical results indicate that the breakdown voltage of the proposed device is increased by 56% in comparison to conventional LDMOS.

  10. [Macrokinetic basis for the model of microbial growth in a limited volume under constant conditions with a single leading substrate].

    Science.gov (United States)

    Gendugov, V M; Glazunov, G P

    2013-01-01

    Within the framework of the macrokinetic approach and continuum and chemical/biochemical gross reaction conceptions, an equation describing the complete dynamics of microbial growth and decline as function of a variable concentration of the leading substrate was deduced. This equation allows us to distinguish quantitatively and qualitatively the stages of microbial growth and the intervals of microbial tolerance to the initial concentration of the leading substrate. Adequacy of the model was confirmed by comparison with experimental dynamics of aerobic microorganisms in the samples of groundwater collected from a region polluted with uranium.

  11. COMPARISON OF NORMALIZED MAXIMUM AEROBIC CAPACITY AND BODY COMPOSITION OF SUMO WRESTLERS TO ATHLETES IN COMBAT AND OTHER SPORTS

    Directory of Open Access Journals (Sweden)

    Matthew D. Beekley

    2006-07-01

    Full Text Available Sumo wrestling is unique in combat sport, and in all of sport. We examined the maximum aerobic capacity and body composition of sumo wrestlers and compared them to untrained controls. We also compared "aerobic muscle quality", meaning VO2max normalized to predicted skeletal muscle mass (SMM (VO2max /SMM, between sumo wrestlers and controls and among previously published data for male athletes from combat, aerobic, and power sports. Sumo wrestlers, compared to untrained controls, had greater (p < 0.05 body mass (mean ± SD; 117.0 ± 4.9 vs. 56.1 ± 9.8 kg, percent fat (24.0 ± 1.4 vs. 13.3 ± 4.5, fat-free mass (88.9 ± 4.2 vs. 48.4 �� 6.8 kg, predicted SMM (48.2 ± 2.9 vs. 20.6 ± 4.7 kg and absolute VO2max (3.6 ± 1.3 vs. 2.5 ± 0.7 L·min-1. Mean VO2max /SMM (ml·kg SMM-1·min-1 was significantly different (p < 0.05 among aerobic athletes (164.8 ± 18.3, combat athletes (which was not different from untrained controls; 131.4 ± 9.3 and 128.6 ± 13.6, respectively, power athletes (96.5 ± 5.3, and sumo wrestlers (71.4 ± 5.3. There was a strong negative correlation (r = - 0.75 between percent body fat and VO2max /SMM (p < 0.05. We conclude that sumo wrestlers have some of the largest percent body fat and fat-free mass and the lowest "aerobic muscle quality" (VO2max /SMM, both in combat sport and compared to aerobic and power sport athletes. Additionally, it appears from analysis of the relationship between SMM and absolute VO2max for all sports that there is a "ceiling" at which increases in SMM do not result in additional increases in absolute VO2max

  12. Technical Note: A generic law-of-the-minimum flux limiter for simulating substrate limitation in biogeochemical models

    Directory of Open Access Journals (Sweden)

    J. Y. Tang

    2015-08-01

    Full Text Available We present a generic flux limiter to account for mass limitations from an arbitrary number of substrates in a biogeochemical reaction network. The flux limiter is based on the observation that substrate (e.g., nitrogen, phosphorus limitation in biogeochemical models can be represented as to ensure mass conservative and non-negative numerical solutions to the governing ordinary differential equations. Application of the flux limiter includes two steps: (1 formulate the biogeochemical processes with a matrix of stoichiometric coefficients and (2 apply Liebig's law of the minimum using the dynamic stoichiometric relationship of the reactants. This approach contrasts with the ad hoc down-regulation approaches that are implemented in many existing models (such as CLM4.5 and the ACME (Accelerated Climate Modeling for Energy Land Model (ALM of carbon and nutrient interactions, which are error prone when adding new processes, even for experienced modelers. Through an example implementation with a Century-like decomposition model that includes carbon, nitrogen, and phosphorus, we show that our approach (1 produced almost identical results to that from the ad hoc down-regulation approaches under non-limiting nutrient conditions; and (2 properly resolved the negative solutions under substrate-limited conditions where the simple clipping approach failed; and (3 successfully avoided the potential conceptual ambiguities that are implied by those ad hoc down-regulation approaches. We expect our approach will make future biogeochemical models easier to improve and more robust.

  13. Modeling of Axially Loaded Nanowires Embedded in Elastic Substrate Media with Inclusion of Nonlocal and Surface Effects

    Directory of Open Access Journals (Sweden)

    Suchart Limkatanyu

    2013-01-01

    Full Text Available Nonlocal and surface effects are incorporated into a bar-elastic substrate element to account for small-scale and size-dependent effects on axial responses of nanowires embedded in elastic substrate media. The virtual displacement principle, employed to consistently derive the governing differential equation as well as the boundary conditions, forms the core of the displacement-based finite element formulation of the nanowire-elastic substrate element. The element displacement shape functions, analytically derived based on homogeneous solution to the governing differential equilibrium equation of the problem, result in the exact element stiffness matrix and equivalent load vector. Two numerical simulations employing the proposed model are performed to study characteristics and behavior of the nanowire-substrate system. The first simulation involves investigation of responses of the wire embedded in elastic substrate. The second examines influences of several system parameters on the contact stiffness and reveals the size-dependent effect on the effective Young's modulus of the system.

  14. Substrate stiffness and oxygen as regulators of stem cell differentiation during skeletal tissue regeneration: a mechanobiological model.

    Directory of Open Access Journals (Sweden)

    Darren Paul Burke

    Full Text Available Extrinsic mechanical signals have been implicated as key regulators of mesenchymal stem cell (MSC differentiation. It has been possible to test different hypotheses for mechano-regulated MSC differentiation by attempting to simulate regenerative events such as bone fracture repair, where repeatable spatial and temporal patterns of tissue differentiation occur. More recently, in vitro studies have identified other environmental cues such as substrate stiffness and oxygen tension as key regulators of MSC differentiation; however it remains unclear if and how such cues determine stem cell fate in vivo. As part of this study, a computational model was developed to test the hypothesis that substrate stiffness and oxygen tension regulate stem cell differentiation during fracture healing. Rather than assuming mechanical signals act directly on stem cells to determine their differentiation pathway, it is postulated that they act indirectly to regulate angiogenesis and hence partially determine the local oxygen environment within a regenerating tissue. Chondrogenesis of MSCs was hypothesized to occur in low oxygen regions, while in well vascularised regions of the regenerating tissue a soft local substrate was hypothesised to facilitate adipogenesis while a stiff substrate facilitated osteogenesis. Predictions from the model were compared to both experimental data and to predictions of a well established computational mechanobiological model where tissue differentiation is assumed to be regulated directly by the local mechanical environment. The model predicted all the major events of fracture repair, including cartilaginous bridging, endosteal and periosteal bony bridging and bone remodelling. It therefore provides support for the hypothesis that substrate stiffness and oxygen play a key role in regulating MSC fate during regenerative events such as fracture healing.

  15. Modelling the anaerobic digestion of solid organic waste – Substrate characterisation method for ADM1 using a combined biochemical and kinetic parameter estimation approach

    OpenAIRE

    Poggio, D.; Walker, M.; Nimmo, W; Ma, L; Pourkashanian, M.

    2016-01-01

    This work proposes a novel and rigorous substrate characterisation methodology to be used with ADM1\\ud to simulate the anaerobic digestion of solid organic waste. The proposed method uses data from both\\ud direct substrate analysis and the methane production from laboratory scale anaerobic digestion experiments\\ud and involves assessment of four substrate fractionation models. The models partition the organic\\ud matter into a mixture of particulate and soluble fractions with the decision on t...

  16. Comparing kinetic Monte Carlo and thin-film modeling of transversal instabilities of ridges on patterned substrates

    Science.gov (United States)

    Tewes, Walter; Buller, Oleg; Heuer, Andreas; Thiele, Uwe; Gurevich, Svetlana V.

    2017-03-01

    We employ kinetic Monte Carlo (KMC) simulations and a thin-film continuum model to comparatively study the transversal (i.e., Plateau-Rayleigh) instability of ridges formed by molecules on pre-patterned substrates. It is demonstrated that the evolution of the occurring instability qualitatively agrees between the two models for a single ridge as well as for two weakly interacting ridges. In particular, it is shown for both models that the instability occurs on well defined length and time scales which are, for the KMC model, significantly larger than the intrinsic scales of thermodynamic fluctuations. This is further evidenced by the similarity of dispersion relations characterizing the linear instability modes.

  17. Structural diversity in the dandelion (Taraxacum officinale polyphenol oxidase family results in different responses to model substrates.

    Directory of Open Access Journals (Sweden)

    Mareike E Dirks-Hofmeister

    Full Text Available Polyphenol oxidases (PPOs are ubiquitous type-3 copper enzymes that catalyze the oxygen-dependent conversion of o-diphenols to the corresponding quinones. In most plants, PPOs are present as multiple isoenzymes that probably serve distinct functions, although the precise relationship between sequence, structure and function has not been addressed in detail. We therefore compared the characteristics and activities of recombinant dandelion PPOs to gain insight into the structure-function relationships within the plant PPO family. Phylogenetic analysis resolved the 11 isoenzymes of dandelion into two evolutionary groups. More detailed in silico and in vitro analyses of four representative PPOs covering both phylogenetic groups were performed. Molecular modeling and docking predicted differences in enzyme-substrate interactions, providing a structure-based explanation for grouping. One amino acid side chain positioned at the entrance to the active site (position HB2+1 potentially acts as a "selector" for substrate binding. In vitro activity measurements with the recombinant, purified enzymes also revealed group-specific differences in kinetic parameters when the selected PPOs were presented with five model substrates. The combination of our enzyme kinetic measurements and the in silico docking studies therefore indicate that the physiological functions of individual PPOs might be defined by their specific interactions with different natural substrates.

  18. Structural diversity in the dandelion (Taraxacum officinale) polyphenol oxidase family results in different responses to model substrates.

    Science.gov (United States)

    Dirks-Hofmeister, Mareike E; Singh, Ratna; Leufken, Christine M; Inlow, Jennifer K; Moerschbacher, Bruno M

    2014-01-01

    Polyphenol oxidases (PPOs) are ubiquitous type-3 copper enzymes that catalyze the oxygen-dependent conversion of o-diphenols to the corresponding quinones. In most plants, PPOs are present as multiple isoenzymes that probably serve distinct functions, although the precise relationship between sequence, structure and function has not been addressed in detail. We therefore compared the characteristics and activities of recombinant dandelion PPOs to gain insight into the structure-function relationships within the plant PPO family. Phylogenetic analysis resolved the 11 isoenzymes of dandelion into two evolutionary groups. More detailed in silico and in vitro analyses of four representative PPOs covering both phylogenetic groups were performed. Molecular modeling and docking predicted differences in enzyme-substrate interactions, providing a structure-based explanation for grouping. One amino acid side chain positioned at the entrance to the active site (position HB2+1) potentially acts as a "selector" for substrate binding. In vitro activity measurements with the recombinant, purified enzymes also revealed group-specific differences in kinetic parameters when the selected PPOs were presented with five model substrates. The combination of our enzyme kinetic measurements and the in silico docking studies therefore indicate that the physiological functions of individual PPOs might be defined by their specific interactions with different natural substrates.

  19. Mechanism of the Quorum-Quenching Lactonase (AiiA) from Bacillus thuringiensis. 2. Substrate Modeling and Active Site Mutations

    Energy Technology Data Exchange (ETDEWEB)

    Momb, Jessica; Wang, Canhui; Liu, Dali; Thomas, Pei W.; Petsko, Gregory A.; Guo, Hua; Ringe, Dagmar; Fast, Walter (UNM); (Brandeis); (Texas)

    2008-12-02

    The N-acyl-l-homoserine lactone hydrolases (AHL lactonases) have attracted considerable attention because of their ability to quench AHL-mediated quorum-sensing pathways in Gram-negative bacteria and because of their relation to other enzymes in the metallo-{beta}-lactamase superfamily. To elucidate the detailed catalytic mechanism of AHL lactonase, mutations are made on residues that presumably contribute to substrate binding and catalysis. Steady-state kinetic studies are carried out on both the wild-type and mutant enzymes using a spectrum of substrates. Two mutations, Y194F and D108N, present significant effects on the overall catalysis. On the basis of a high-resolution structural model of the enzyme-product complex, a hybrid quantum mechanical/molecular mechanical method is used to model the substrate binding orientation and to probe the effect of the Y194F mutation. Combining all experimental and computational results, we propose a detailed mechanism for the ring-opening hydrolysis of AHL substrates as catalyzed by the AHL lactonase from Bacillus thuringiensis. Several features of the mechanism that are also found in related enzymes are discussed and may help to define an evolutionary thread that connects the hydrolytic enzymes of this mechanistically diverse superfamily.

  20. Modeling gravity effects on water retention and gas transport characteristics in plant growth substrates

    DEFF Research Database (Denmark)

    Deepagoda Thuduwe Kankanamge Kelum, Chamindu; Jones, Scott B.; Tuller, Markus

    2014-01-01

    Growing plants to facilitate life in outer space, for example on the International Space Station (ISS) or at planned deep-space human outposts on the Moon or Mars, has received much attention with regard to NASA’s advanced life support system research. With the objective of in situ resource...... utilization to conserve energy and to limit transport costs, native materials mined on Moon or Mars are of primary interest for plant growth media in a future outpost, while terrestrial porous substrates with optimal growth media characteristics will be useful for onboard plant growth during space missions...... permeability characteristics of six plant growth substrates for potential applications in space, including two terrestrial analogs for lunar and Martian soils and four particulate substrates widely used in reduced gravity experiments. To simulate reduced gravity water characteristics, the predictions...

  1. Dielectric constant of graphene-on-polarized substrate: A tight-binding model study

    Indian Academy of Sciences (India)

    SIVABRATA SAHU; S K S PARASHAR; G C ROUT

    2017-07-01

    We report here a microscopic tight-binding theoretical study of the dynamic dielectric response of graphene-on-polarizable substrate with impurity. The Hamiltonian consists of first, second and third nearest neighbour electron hopping interactions besides doping and substrate-induced effects on graphene. We have introduced electron–electron correlation effect at A and B sublattices of graphene which is considered within Hartree–Fock mean-field approximation. The electron occupancies at both sublattices are calculated and solvedself-consistently and numerically for both up- and down-spin orientations. The polarization function appearing in the dielectric function is a two-particle Green’s function which is calculated by using Zubarev’s Green’s function technique. The temperature and optical frequency-dependent dielectric function is evaluated and compared with experimental data by varying Coulomb correlation energy, substrate-induced gap and impurity concentrations.

  2. High-level expression and purification of soluble recombinant FGF21 protein by SUMO fusion in Escherichia coli

    Directory of Open Access Journals (Sweden)

    Huang Yadong

    2010-02-01

    Full Text Available Abstract Background Fibroblast growth factor 21 (FGF21 is a promising drug candidate to combat metabolic diseases. However, high-level expression and purification of recombinant FGF21 (rFGF21 in Escherichia coli (E. coli is difficult because rFGF21 forms inclusion bodies in the bacteria making it difficult to purify and obtain high concentrations of bioactive rFGF21. To overcome this problem, we fused the FGF21 with SUMO (Small ubiquitin-related modifier by polymerase chain reaction (PCR, and expressed the fused gene in E. coli BL21(DE3. Results By inducing with IPTG, SUMO-FGF21 was expressed at a high level. Its concentration reached 30% of total protein, and exceeded 95% of all soluble proteins. The fused protein was purified by DEAE sepharose FF and Ni-NTA affinity chromatography. Once cleaved by the SUMO protease, the purity of rFGF21 by high performance liquid chromatography (HPLC was shown to be higher than 96% with low endotoxin level (in vivo animal experiments showed that rFGF21 produced by using this method, could decrease the concentration of plasma glucose in diabetic rats by streptozotocin (STZ injection. Conclusions This study demonstrated that SUMO, when fused with FGF21, was able to promote its soluble expression of the latter in E. coli, making it more convenient to purify rFGF21 than previously. This may be a better method to produce rFGF21 for pharmaceutical research and development.

  3. Modeling neural differentiation on micropatterned substrates coated with neural matrix components

    Directory of Open Access Journals (Sweden)

    Patricia eGarcía-Parra

    2012-03-01

    Full Text Available Topographical and biochemical characteristics of the substrate are critical for neuronal differentiation including axonal outgrowth and regeneration of neural circuits in vivo. Contact stimuli and signaling molecules allow neurons to develop and stabilize synaptic contacts. Here we present the development, characterization and functional validation of a new polymeric support able to induce neuronal differentiation in both PC12 cell line and adult primary skin-derived precursor cells in vitro. By combining a photolithographic technique with use of neural extracellular matrix as a substrate, a biocompatible and efficient microenvironment for neuronal differentiation was developed.

  4. Real Estate in the DNA Damage Response: Ubiquitin and SUMO Ligases Home in on DNA Double-Strand Breaks.

    Science.gov (United States)

    Dantuma, Nico P; Pfeiffer, Annika

    2016-01-01

    Ubiquitin and the ubiquitin-like modifier SUMO are intimately connected with the cellular response to various types of DNA damage. A striking feature is the local accumulation of these proteinaceous post-translational modifications in the direct vicinity to DNA double-strand breaks, which plays a critical role in the formation of ionizing radiation-induced foci. The functional significance of these modifications is the coordinated recruitment and removal of proteins involved in DNA damage signaling and repair in a timely manner. The central orchestrators of these processes are the ubiquitin and SUMO ligases that are responsible for accurately tagging a broad array of chromatin and chromatin-associated proteins thereby changing their behavior or destination. Despite many differences in the mode of action of these enzymes, they share some striking features that are of direct relevance for their function in the DNA damage response. In this review, we outline the molecular mechanisms that are responsible for the recruitment of ubiquitin and SUMO ligases and discuss the importance of chromatin proximity in this process.

  5. DNA Damage Response and SUMO Modification%SUMO化修饰在DNA损伤响应中的作用

    Institute of Scientific and Technical Information of China (English)

    蒋华东; 李钰

    2012-01-01

    物理或化学等多种因素均可以引起DNA损伤.为维持机体基因组的稳定性,机体形成了精确完整的机制来修复损伤的DNA. SUMO(small ubiquitin-related modifier,SUMO)化修饰与其他蛋白翻译后修饰一样,具有多种生物学功能.近年来的研究表明,其在DNA损伤修复中也具有非常重要的作用.该文就DNA损伤修复、SUMO化修饰系统及其二者关系的最新研究进展作了较为全面的介绍和总结.%DNA damage can be produced widely in our body by physical and/or chemical factors. Therefore, in order to maintain the genomic integrity, DNA must be protected from DNA damage and repaired correctly by forming accurate mechanism if DNA damage happened. SUMOylation has multiple biological functions like other post-translation modification. Emerging evidence has showed that SUMOs play vital roles in DNA damage repair. This review focuses on the newly research about DNA damage, SUMOylation and the relationship between them and provides a complete introducing and summarization.

  6. Post-Translational Modifications of Kaposi’s Sarcoma-Associated Herpesvirus (KSHV Regulatory Proteins-SUMO and KSHV

    Directory of Open Access Journals (Sweden)

    Mel eCampbell

    2012-02-01

    Full Text Available Reactivation from a latent state is an important feature of infection and disease caused by many herpesviruses. KSHV latency can be envisioned as an outcome that is balanced between factors that promote viral gene expression and lytic replication against those that facilitate gene silencing and establish or maintain latency. A large body of work has focused on the activities of the key viral regulatory proteins involved in KSHV latent or lytic states. Moreover, recent studies have also begun to document the importance of epigenetic landscape evolution of the KSHV viral genome during latency and reactivation. However, one area of KSHV molecular virology that remains largely unanswered is the precise role of post-translational modifications on the activities of viral factors that function during latency and reactivation. In this review, we will summarize the post-translational modifications associated with three viral factors whose activities contribute to the viral state. The viral proteins discussed are the two major KSHV encoded transcription factors, K-Rta and K-bZIP (KSHV basic leucine zipper and the viral latency-associated nuclear antigen (LANA. A special emphasis will be placed on the role of the sumoylation pathway in the modulation of the KSHV lifecycle. Newly uncovered SUMO-dependent properties of LANA and K-Rta will also be presented, namely LANA histone-targeting SUMO E3 ligase activity and K-Rta SUMO-targeted ubiquitin ligase function.

  7. Homology modeling of mosquito cytochrome P450 enzymes involved in pyrethroid metabolism: insights into differences in substrate selectivity

    Directory of Open Access Journals (Sweden)

    Rongnoparut Pornpimol

    2011-09-01

    Full Text Available Abstract Background Cytochrome P450 enzymes (P450s have been implicated in insecticide resistance. Anopheles minumus mosquito P450 isoforms CYP6AA3 and CYP6P7 are capable of metabolizing pyrethroid insecticides, however CYP6P8 lacks activity against this class of compounds. Findings Homology models of the three An. minimus P450 enzymes were constructed using the multiple template alignment method. The predicted enzyme model structures were compared and used for molecular docking with insecticides and compared with results of in vitro enzymatic assays. The three model structures comprise common P450 folds but differences in geometry of their active-site cavities and substrate access channels are prominent. The CYP6AA3 model has a large active site allowing it to accommodate multiple conformations of pyrethroids. The predicted CYP6P7 active site is more constrained and less accessible to binding of pyrethroids. Moreover the predicted hydrophobic interface in the active-site cavities of CYP6AA3 and CYP6P7 may contribute to their substrate selectivity. The absence of CYP6P8 activity toward pyrethroids appears to be due to its small substrate access channel and the presence of R114 and R216 that may prevent access of pyrethroids to the enzyme heme center. Conclusions Differences in active site topologies among CYPAA3, CYP6P7, and CYP6P8 enzymes may impact substrate binding and selectivity. Information obtained using homology models has the potential to enhance the understanding of pyrethroid metabolism and detoxification mediated by P450 enzymes.

  8. Modeling of penconazole and metalaxyl mobility in undisturbed vineyard soil cores, unamended and amended with spent mushroom substrate

    Science.gov (United States)

    Marin-Benito, Jesus Maria; Mamy, Laure; Rodriguez-Cruz, Maria Sonia; Sanchez-Martin, Maria Jesus

    2013-04-01

    In Spain, one of the main producers of mushrooms in the world, a huge amount of the substrates used for the growth of mushrooms have to be eliminated after harvest. However, this substrate represents a promising amendment because of its high organic matter content and, in particular, it could be used in vineyard soils because they generally are poor in organic matter. But the effect of this amendment on the fate in soils of fungicides that are massively used in vineyards is unknown. Therefore, the objectives of this work were to model the mobility of two fungicides, penconazole and metalaxyl, in undisturbed vineyard soil columns using the PRZM3 (Pesticide Root Zone Model) parameterized with laboratory data, and to compare the simulations with the experimental results obtained in mobility studies. Soil cores (40 cm x 9 cm d.i.) were collected from experimental plots in three different vineyard soils of La Rioja (Spain). Three different treatments were tested in each soil: natural (control) soil, soil amended with fresh spent mushroom substrate, and soil amended with composted spent mushroom substrate. The leaching of fungicides was studied in non-incubated and incubated (outdoors for 77 days) soil cores under unsaturated flow conditions. In general, the addition of mushroom substrates decreased the leaching of fungicides compared to control soils. For the most hydrophobic fungicide, penconazole, the predictions obtained by the model were highly correlated (r > 0.88) with the experimental results. Penconazole was never observed in the leachates, its vertical distribution was similar within all soil profiles, and retention of almost all the fungicide was into the topsoil (0-8cm). For the less hydrophobic fungicide, metalaxyl, and the CGA 62826 metabolite generated from its degradation during the experimental period, PRZM3 was not able to reproduce the observations and it was necessary to calibrate the model. After calibration, the correlation between model predictions

  9. Non-native Conformers of Cystic Fibrosis Transmembrane Conductance Regulator NBD1 Are Recognized by Hsp27 and Conjugated to SUMO-2 for Degradation.

    Science.gov (United States)

    Gong, Xiaoyan; Ahner, Annette; Roldan, Ariel; Lukacs, Gergely L; Thibodeau, Patrick H; Frizzell, Raymond A

    2016-01-22

    A newly identified pathway for selective degradation of the common mutant of the cystic fibrosis transmembrane conductance regulator (CFTR), F508del, is initiated by binding of the small heat shock protein, Hsp27. Hsp27 collaborates with Ubc9, the E2 enzyme for protein SUMOylation, to selectively degrade F508del CFTR via the SUMO-targeted ubiquitin E3 ligase, RNF4 (RING finger protein 4) (1). Here, we ask what properties of CFTR are sensed by the Hsp27-Ubc9 pathway by examining the ability of NBD1 (locus of the F508del mutation) to mimic the disposal of full-length (FL) CFTR. Similar to FL CFTR, F508del NBD1 expression was reduced 50-60% by Hsp27; it interacted preferentially with the mutant and was modified primarily by SUMO-2. Mutation of the consensus SUMOylation site, Lys(447), obviated Hsp27-mediated F508del NBD1 SUMOylation and degradation. As for FL CFTR and NBD1 in vivo, SUMO modification using purified components in vitro was greater for F508del NBD1 versus WT and for the SUMO-2 paralog. Several findings indicated that Hsp27-Ubc9 targets the SUMOylation of a transitional, non-native conformation of F508del NBD1: (a) its modification decreased as [ATP] increased, reflecting stabilization of the nucleotide-binding domain by ligand binding; (b) a temperature-induced increase in intrinsic fluorescence, which reflects formation of a transitional NBD1 conformation, was followed by its SUMO modification; and (c) introduction of solubilizing or revertant mutations to stabilize F508del NBD1 reduced its SUMO modification. These findings indicate that the Hsp27-Ubc9 pathway recognizes a non-native conformation of mutant NBD1, which leads to its SUMO-2 conjugation and degradation by the ubiquitin-proteasome system.

  10. Continuum Plate Theory and Atomistic Modeling to Find the Flexural Rigidity of a Graphene Sheet Interacting with a Substrate

    Directory of Open Access Journals (Sweden)

    M. W. Roberts

    2010-01-01

    Full Text Available Using a combination of continuum modeling, atomistic simulations, and numerical optimization, we estimate the flexural rigidity of a graphene sheet. We consider a rectangular sheet that is initially parallel to a rigid substrate. The sheet interacts with the substrate by van der Waals forces and deflects in response to loading on a pair of opposite edges. To estimate the flexural rigidity, we model the graphene sheet as a continuum and numerically solve an appropriate differential equation for the transverse deflection. This solution depends on the flexural rigidity. We then use an optimization procedure to find the value of the flexural rigidity that minimizes the difference between the numerical solutions and the deflections predicted by atomistic simulations. This procedure predicts a flexural rigidity of 0.26 nN nm=1.62 eV.

  11. Finite dipole model for extreme near-field thermal radiation between a tip and planar SiC substrate

    Science.gov (United States)

    Jarzembski, Amun; Park, Keunhan

    2017-04-01

    Recent experimental studies have measured the infrared (IR) spectrum of tip-scattered near-field thermal radiation for a SiC substrate and observed up to a 50cm-1 redshift of the surface phonon polariton (SPhP) resonance peak [1,2]. However, the observed spectral redshift cannot be explained by the conventional near-field thermal radiation model based on the point dipole approximation. In the present work, a heated tip is modeled as randomly fluctuating point charges (or fluctuating finite dipoles) aligned along the primary axis of a prolate spheroid, and quasistatic tip-substrate charge interactions are considered to formulate the effective polarizability and self-interaction Green's function. The finite dipole model (FDM), combined with fluctuational electrodynamics, allows the computation of tip-plane thermal radiation in the extreme near-field (i.e., H / R ≲ 1 , where H is the tip-substrate gap distance and R is the tip radius), which cannot be calculated with the point dipole approximation. The FDM provides the underlying physics on the spectral redshift of tip-scattered near-field thermal radiation as observed in experiments. In addition, the SPhP peak in the near-field thermal radiation spectrum may split into two peaks as the gap distance decreases into the extreme near-field regime. This observation suggests that scattering-type spectroscopic measurements may not convey the full spectral features of tip-plane extreme near-field thermal radiation.

  12. Exploring the potential of the RPA system SUMO for multipurpose boundary-layer missions during the BLLAST campaign

    Science.gov (United States)

    Reuder, Joachim; Båserud, Line; Jonassen, Marius O.; Kral, Stephan T.; Müller, Martin

    2016-06-01

    In June and July 2011 the RPAS (Remotely Piloted Aircraft System) SUMO (Small Unmanned Meteorological Observer) performed a total number of 299 scientific flights during the BLLAST (Boundary Layer Late Afternoon and Sunset Turbulence) campaign in southern France. Three different types of missions were performed: vertical profiling of the mean meteorological parameters (temperature, humidity and wind), horizontal surveys of the surface temperature and horizontal transects for the estimation of turbulence. The manuscript provides an introduction to the corresponding SUMO operations, including regulatory issues and the coordination of manned and unmanned airborne operations for boundary-layer research that have been pioneered during the BLLAST campaign. The main purpose of the SUMO flight strategy was atmospheric profiling at high temporal resolution. A total of 168 profile flights were performed during the campaign with typically more than 10 flights per Intensive Observational Period (IOP) day. The collected data allow for a detailed study of boundary-layer structure and dynamics and will be used for further analysis, e.g. the determination of profiles of sensible and latent heat fluxes. First, tests of a corresponding method have shown very promising results and have provided surface-flux values in close agreement with those from ground-based eddy-covariance measurements. In addition, 74 horizontal surveys of the IR emission of the surface were performed at altitudes of around 65 m. Each of those surveys covers a typical area of around 1 km2 and allows for an estimation of the surface-temperature variability, important information for the assessment of the heterogeneity of the surface forcing as a function of soil and vegetation properties. The comparison with other surface-temperature measurements shows that the raw data of the airborne and ground observations can differ considerably, but that even a very simple multiple regression method can reduce those

  13. SUMO: solar ultraviolet monitor and ozone nanosatellite for spectral irradiance, ozone and Earth radiative budget simultaneous evaluation

    Science.gov (United States)

    Damé, Luc

    SUMO is an innovative proof-of-concept nano-satellite which aims to measure on the same platform the different components of the Earth radiation budget, the solar energy input and the energy reemitted at the top of the Earth atmosphere, with a particular focus on the UV part of the spectrum and on the ozone layer, which are the most sensitive to the solar variability. The far UV (FUV) is the only wavelength band with energy absorbed in the high atmosphere (stratosphere), in the ozone (Herzberg continuum, 200-220 nm) and oxygen bands, and its high variability is most probably at the origin of a climate influence (UV affects stratospheric dynamics and temperatures, altering interplanetary waves and weather patterns both poleward and downward to the lower stratosphere and tropopause regions). Recent measurements at the time of the last solar minimum suggest that variations in the UV may be larger than previously assumed what implies a very different response in both stratospheric ozone and temperature. A simultaneous observation of the incoming FUV and of the ozone (O _{3}) production, would bring an invaluable information on this process of solar-climat forcing. Space instruments have already measured the different components of the Earth radiative budget but this is, to our knowledge, the first time that all instruments are operated on the same platform. This characteristic guarantees by itself obtaining original scientific results. SUMO is a 10x10x30 cm (3) nanosatellite (``3U"), the payload occupying ``1U", i.e. a cube of 10x10x10 cm (3) for 1 kg and 1 W of power. Orbit is polar since a further challenge in understanding the relation between solar UV variability and stratospheric ozone on arctic and antarctic regions. SUMO definition has been completed (platform and payload assembly integration and tests are possible in 24 months) and it is now intended to be proposed to CNES for a flight in 2017. Mission is expected to last up to 1 year. Follow-up is 2 fold: on

  14. Structural differences of matrix metalloproteinases. Homology modeling and energy minimization of enzyme-substrate complexes

    DEFF Research Database (Denmark)

    Terp, G E; Christensen, I T; Jørgensen, Flemming Steen

    2000-01-01

    in the AMBER 4.1 force field, applying a non-bonded approach and a full ion charge representation. Energy minimization of the apoenzymes yielded structures with distorted active sites, while reliable three-dimensional structures of the enzymes containing a substrate in active site were obtained. The structural...... differences between the eight enzyme-substrate complexes were studied with particular emphasis on the active site, and possible sites for obtaining selectivity among the MMP's are discussed. Differences in the P1' pocket are well-documented and have been extensively exploited in inhibitor design. The present......Matrix metalloproteinases are extracellular enzymes taking part in the remodeling of extracellular matrix. The structures of the catalytic domain of MMP1, MMP3, MMP7 and MMP8 are known, but structures of enzymes belonging to this family still remain to be determined. A general approach...

  15. Myxococcus xanthus gliding motors are elastically coupled to the substrate as predicted by the focal adhesion model of gliding motility

    CERN Document Server

    Balagam, Rajesh; Czerwinski, Fabian; Sun, Mingzhai; Kaplan, Heidi B; Shaevitz, Joshua W; Igoshin, Oleg A

    2014-01-01

    Myxococcus xanthus is a model organism for studying bacterial social behaviors due to its ability to form complex multi-cellular structures. Knowledge of M. xanthus surface gliding motility and the mechanisms that coordinate it are critically important to our understanding of collective cell behaviors. Although the mechanism of gliding motility is still under investigation, recent experiments suggest that there are two possible mechanisms underlying force production for cell motility: the focal adhesion mechanism and the helical rotor mechanism which differ in the biophysics of the cell-substrate interactions. Whereas the focal adhesion model predicts an elastic coupling, the helical rotor model predicts a viscous coupling. Using a combination of computational modeling, imaging, and force microscopy, we find evidence for elastic coupling in support of the focal adhesion model. Using a biophysical model of the M. xanthus cell, we investigated how the mechanical interactions between cells are affected by intera...

  16. A Cross-Shore Model of Barrier Island Migration over a Compressible Substrate

    Science.gov (United States)

    2010-01-01

    substrate Marine Geology 271 (2010) 1–16 ⁎ Corresponding author. Tel.: +1 251 694 3719; fax: +1 251 690 3464. E-mail address: Julie.D.Rosati...ScienceDirect Marine Geology j ourna l homepage: www.e lsev ie r.com/ locate /margeo Report Documentation Page Form ApprovedOMB No. 0704-0188 Public reporting...Egypt (Stanley and Warne, 1998), the Ganges–Brahmaputra Rivers, Bangladesh , India (Allison, 1998); and the Yangtze River, China (Xiqing, 1998

  17. Expression and characterization of SUMO-conjugated metal-responsive transcription factor 1: SIM-dependent cross-interaction and distinct DNA binding activity.

    Science.gov (United States)

    Lin, Chang-Yi; Liu, Ya-Chuan; Lin, Meng-Chieh; Thi Nguyen, Thao; Tam, Ming F; Chein, Chih-Yuan; Lin, Meng-Ting; Lin, Lih-Yuan

    2013-04-01

    Metal-responsive transcription factor 1 (MTF-1) regulates a variety of genes involving in metal homeostasis and oxidative stress. We have shown that MTF-1 can be conjugated by small ubiquitin-like modifier (SUMO) and forms complexes with cellular factor(s) in a SUMO-interacting motif (SIM)-dependent manner. To investigate whether the interaction of MTF-1 and its SUMO conjugate occurs, we expressed and isolated MTF-1 and sumoylated MTF-1 (S-MTF-1) for functional studies. Various conditions were examined to optimize the expressions of MTF-1 and S-MTF-1. Results from affinity column chromatography demonstrated that the unmodified MTF-1 consistently co-eluted with the S-MTF-1. Mutations at the SIM did not reduce the level of MTF-1 sumoylation but the sumoylated product can then be purified to homogeneity. The presence of MTF-1 cross-interaction was further supported by in vitro pull-down assays. The ability of the purified proteins in binding metal-responsive element (MRE) was assessed with electrophoretic mobility shift assay. Noticeably, MTF-1 required the presence of cell extracts to render the binding activity. However, S-MTF-1 binds MRE in void of other cellular factors. The same characteristic was found for MTF-1 with SUMO fusion at the carboxyl terminus. These results indicate that the presence of SUMO moiety allows the protein to interact directly with MRE.

  18. [A simulation model for predicting the dry matter allocation in cut lily plants under effects of substrate water potential].

    Science.gov (United States)

    Dong, Yong-Yi; Li, Gang; An, Dong-Sheng; Luo, Wei-Hong

    2012-04-01

    Dry matter allocation and translocation is the base of the formation of appearance quality of ornamental plants, and strongly affected by water supply. Taking cut lily cultivar 'Sorbonne' as test material, a culture experiment of different planting dates and water supply levels was conducted in a multi-span greenhouse in Nanjing from March 2009 to January 2010 to quantitatively analyze the seasonal changes of the dry matter allocation and translocation in 'Sorbonne' plants and the effects of substrate water potential on the dry matter allocation indices for different organs (flower, stem, leaf, bulb, and root), aimed to define the critical substrate water potential for the normal growth of the cultivar, and establish a simulation model for predicting the dry matter allocation in cut lily plants under effects of substrate water potential. The model established in this study gave a good prediction on the dry mass of plant organs, with the coefficient of determination and the relative root mean square error between the simulated and measured values of the cultivar' s flower dry mass, stem dry mass, leaf dry mass, bulb dry mass, and root dry mass being 0.96 and 19.2%, 0.95 and 12.4%, 0.86 and 19.4%, 0.95 and 12.2%, and 0.85 and 31.7%, respectively. The critical water potential for the water management of cut lily could be -15 kPa.

  19. Salinosporamides A and B Inhibit Proteasome Activity and Delay the Degradation of N-end Rule Model Substrates

    Energy Technology Data Exchange (ETDEWEB)

    Shin, Seungkyun; Bang, Daein; Choi, Wonhoon; Lee, Minjae [Kyung Hee Univ., Yongin (Korea, Republic of); Kim, Seonghwan; Oh, Dongchan [Seoul National Univ., Seoul (Korea, Republic of)

    2013-05-15

    The proteasome, which is highly evolutionarily conserved, is responsible for the degradation of most short-lived proteins in cells. Small-molecule inhibitors targeting the proteasome's degradative activity have been extensively developed as lead compounds for various human diseases. An exemplified molecule is bortezomib, which was approved by FDA in 2003 for the treatment of multiple myeloma. Here, using transiently and stably expressed N-end rule model substrates in mammalian cells, we evaluated and identified that salinosporamide A and salinosporamide B effectively inhibited the proteasomal degradation. Considering that a variety of proteasome substrates are implicated in the pathogenesis of many diseases, they have the potential to be clinically applicable as therapeutic agents.

  20. A finite element model for the analysis of buckling driven delaminations of thin films on rigid substrates

    Science.gov (United States)

    Gruttmann, F.; Pham, V. D.

    2008-02-01

    The delamination process of thin films on rigid substrates is investigated. Such systems are typically subject to high residual compression and modest adhesion causing them to buckling driven blisters. In certain cases buckles with the shape of telephone cords are observed. A finite element model for quasi-static delamination growth is developed. Applying a Reissner-Mindlin shell kinematic for the film allows C 0- continuous shape functions. The traction vector at the film-substrate interface is obtained from the derivative of a cohesive free energy. Incorporation of loading and unloading conditions is considered for the irreversible process. The equilibrium state is computed iteratively in dependence of the compressive residual stresses. The computed telephone cord delaminations are stable asymmetric configurations whereas the symmetric configurations are unstable.

  1. The influence of metal ions on the substrate binding pocket of human alcohol dehydrogenase β 2β 2 by molecular modeling

    Science.gov (United States)

    Liu, Hsuan-Liang; Ho, Yih; Hsu, Chia-Ming

    2003-04-01

    Based on theoretical molecular modeling performed in this study, both structural and catalytic zinc ions, Zn s and Zn a, respectively, were shown to influence the structural integrity of the substrate binding pocket of human alcohol dehydrogenase β 2β 2 in the middle and outer regions. The replacement of both Zn s and Zn a with different metal ions restricts the access of bulky substrates to the bottom of the active site by narrowing the bottleneck formed between L116 and V294, whereas it does not affect substrate binding affinity since the accessible surface area of the substrate binding pocket remains more than 80% of the wild-type.

  2. Complex SUMO-1 regulation of cardiac transcription factor Nkx2-5.

    Directory of Open Access Journals (Sweden)

    Mauro W Costa

    Full Text Available Reversible post-translational protein modifications such as SUMOylation add complexity to cardiac transcriptional regulation. The homeodomain transcription factor Nkx2-5/Csx is essential for heart specification and morphogenesis. It has been previously suggested that SUMOylation of lysine 51 (K51 of Nkx2-5 is essential for its DNA binding and transcriptional activation. Here, we confirm that SUMOylation strongly enhances Nkx2-5 transcriptional activity and that residue K51 of Nkx2-5 is a SUMOylation target. However, in a range of cultured cell lines we find that a point mutation of K51 to arginine (K51R does not affect Nkx2-5 activity or DNA binding, suggesting the existence of additional Nkx2-5 SUMOylated residues. Using biochemical assays, we demonstrate that Nkx2-5 is SUMOylated on at least one additional site, and this is the predominant site in cardiac cells. The second site is either non-canonical or a "shifting" site, as mutation of predicted consensus sites and indeed every individual lysine in the context of the K51R mutation failed to impair Nkx2-5 transcriptional synergism with SUMO, or its nuclear localization and DNA binding. We also observe SUMOylation of Nkx2-5 cofactors, which may be critical to Nkx2-5 regulation. Our data reveal highly complex regulatory mechanisms driven by SUMOylation to modulate Nkx2-5 activity.

  3. The modeling of ethanol production by Kluyveromyces marxianus using whey as substrate in continuous A-Stat bioreactors.

    Science.gov (United States)

    Gabardo, Sabrina; Pereira, Gabriela Feix; Rech, Rosane; Ayub, Marco Antônio Záchia

    2015-09-01

    We investigated the kinetics of whey bioconversion into ethanol by Kluyveromyces marxianus in continuous bioreactors using the "accelerostat technique" (A-stat). Cultivations using free and Ca-alginate immobilized cells were evaluated using two different acceleration rates (a). The kinetic profiles of these systems were modeled using four different unstructured models, differing in the expressions for the specific growth (μ) and substrate consumption rates (r s), taking into account substrate limitation and product inhibition. Experimental data showed that the dilution rate (D) directly affected cell physiology and metabolism. The specific growth rate followed the dilution rate (μ≈D) for the lowest acceleration rate (a = 0.0015 h(-2)), condition in which the highest ethanol yield (0.52 g g(-1)) was obtained. The highest acceleration rate (a = 0.00667 h(-2)) led to a lower ethanol yield (0.40 g g(-1)) in the system where free cells were used, whereas with immobilized cells ethanol yields increased by 23 % (0.49 g g(-1)). Among the evaluated models, Monod and Levenspiel combined with Ghose and Tyagi models were found to be more appropriate for describing the kinetics of whey bioconversion into ethanol. These results may be useful in scaling up the process for ethanol production from whey.

  4. The use of the radius of gyration in a WLC polymer model of cancer cell adhesion to glycosaminoglycans substrates

    Science.gov (United States)

    Peramo, Antonio; Matthews, Garrett

    2009-03-01

    Glycosaminoglycans (GAG) are a group of polysaccharides involved in several biological functions, including cell adhesion. Most of their biological properties are derived from the interactions of the chains with their environment, hence the interest in developing physical models that could describe their interactions with whole cells. As linear biopolymers with low polydispersity, GAG can be described using polymer models of Gaussian chain distributions, like the WLC (worm-like chain) model. We found that the adhesion of whole cancer cells to glass substrates coated with GAG appear to be dependent on the charge per dimer and degree of sulfation of the GAG chain. We have hypothesized that the adhesion of whole cancer cells to GAG substrates can be described as a function of polysaccharide radius of gyration and used the WLC model describing the global structure of the GAGs to analyze this relationship. We will show that the adhesion of the cancer cells has a linear response with the radius of gyration and is essentially controlled by the charge per dimer. This dominating mechanism is not eliminated when the cells are resuspended in media with heparin. We then propose how these physical properties could be used to predict the preferred molecular structures of compounds for use as anti-metastatic or anti-inflammatory agents.

  5. Atomistic Modelling of Confined Polypropylene Chains between Ferric Oxide Substrates at Melt Temperature

    Directory of Open Access Journals (Sweden)

    Ali Gooneie

    2016-10-01

    Full Text Available The interactions and conformational characteristics of confined molten polypropylene (PP chains between ferric oxide (Fe2O3 substrates were investigated by molecular dynamics (MD simulations. A comparative analysis of the adsorbed amount shows strong adsorption of the chains on the high-energy surface of Fe2O3. Local structures formed in the polymer film were studied utilizing density profiles, orientation of bonds, and end-to-end distance of chains. At interfacial regions, the backbone carbon-carbon bonds of the chains preferably orient in the direction parallel to the surface while the carbon-carbon bonds with the side groups show a slight tendency to orient normal to the surface. Based on the conformation tensor data, the chains are compressed in the normal direction to the substrates in the interfacial regions while they tend to flatten in parallel planes with respect to the surfaces. The orientation of the bonds as well as the overall flattening of the chains in planes parallel to the solid surfaces are almost identical to that of the unconfined PP chains. Also, the local pressure tensor is anisotropic closer to the solid surfaces of Fe2O3 indicating the influence of the confinement on the buildup imbalance of normal and tangential pressures.

  6. Droplet evaporation on a horizontal substrate under gravity field by mesoscopic modeling.

    Science.gov (United States)

    Xie, Chiyu; Zhang, Jianying; Bertola, Volfango; Wang, Moran

    2016-02-01

    The evaporation of water drop deposited on a horizontal substrate is investigated using a lattice Boltzmann method (LBM) for multiphase flows with a large-density ratio. To account for the variation of evaporation flux distribution along the drop interface, a novel evaporation scheme is introduced into the LBM framework, and validated by comparison with experimental data. We aim at discovering the effect of gravity on the evaporating drop in detail, and various evaporation conditions are considered as well as different wetting properties of the substrates. An effective diameter is introduced as an indicator of the critical drop size under which gravity is negligible. Our results show that such critical diameter is much smaller than the capillary length, which has been widely accepted as the critical size in previous and current works. The critical diameter is found to be almost independent of the evaporation conditions and the surface wettability. A correlation between this critical diameter and the capillary length is also proposed for easy use in applications.

  7. Modelling the growth kinetics of Kocuria marina DAGII as a function of single and binary substrate during batch production of β-Cryptoxanthin.

    Science.gov (United States)

    Mitra, Ruchira; Chaudhuri, Surabhi; Dutta, Debjani

    2017-01-01

    In the present investigation, growth kinetics of Kocuria marina DAGII during batch production of β-Cryptoxanthin (β-CRX) was studied by considering the effect of glucose and maltose as a single and binary substrate. The importance of mixed substrate over single substrate has been emphasised in the present study. Different mathematical models namely, the Logistic model for cell growth, the Logistic mass balance equation for substrate consumption and the Luedeking-Piret model for β-CRX production were successfully implemented. Model-based analyses for the single substrate experiments suggested that the concentrations of glucose and maltose higher than 7.5 and 10.0 g/L, respectively, inhibited the growth and β-CRX production by K. marina DAGII. The Han and Levenspiel model and the Luong product inhibition model accurately described the cell growth in glucose and maltose substrate systems with a R (2) value of 0.9989 and 0.9998, respectively. The effect of glucose and maltose as binary substrate was further investigated. The binary substrate kinetics was well described using the sum-kinetics with interaction parameters model. The results of production kinetics revealed that the presence of binary substrate in the cultivation medium increased the biomass and β-CRX yield significantly. This study is a first time detailed investigation on kinetic behaviours of K. marina DAGII during β-CRX production. The parameters obtained in the study might be helpful for developing strategies for commercial production of β-CRX by K. marina DAGII.

  8. pkaPS: prediction of protein kinase A phosphorylation sites with the simplified kinase-substrate binding model

    Directory of Open Access Journals (Sweden)

    Schneider Georg

    2007-01-01

    Full Text Available Abstract Background Protein kinase A (cAMP-dependent kinase, PKA is a serine/threonine kinase, for which ca. 150 substrate proteins are known. Based on a refinement of the recognition motif using the available experimental data, we wished to apply the simplified substrate protein binding model for accurate prediction of PKA phosphorylation sites, an approach that was previously successful for the prediction of lipid posttranslational modifications and of the PTS1 peroxisomal translocation signal. Results Approximately 20 sequence positions flanking the phosphorylated residue on both sides have been found to be restricted in their sequence variability (region -18...+23 with the site at position 0. The conserved physical pattern can be rationalized in terms of a qualitative binding model with the catalytic cleft of the protein kinase A. Positions -6...+4 surrounding the phosphorylation site are influenced by direct interaction with the kinase in a varying degree. This sequence stretch is embedded in an intrinsically disordered region composed preferentially of hydrophilic residues with flexible backbone and small side chain. This knowledge has been incorporated into a simplified analytical model of productive binding of substrate proteins with PKA. Conclusion The scoring function of the pkaPS predictor can confidently discriminate PKA phosphorylation sites from serines/threonines with non-permissive sequence environments (sensitivity of ~96% at a specificity of ~94%. The tool "pkaPS" has been applied on the whole human proteome. Among new predicted PKA targets, there are entirely uncharacterized protein groups as well as apparently well-known families such as those of the ribosomal proteins L21e, L22 and L6. Availability The supplementary data as well as the prediction tool as WWW server are available at http://mendel.imp.univie.ac.at/sat/pkaPS. Reviewers Erik van Nimwegen (Biozentrum, University of Basel, Switzerland, Sandor Pongor (International

  9. STELLA software as a tool for modelling phosphorus removal in a constructed wetland employing dewatered alum sludge as main substrate.

    Science.gov (United States)

    Kumar, J L G; Wang, Z Y; Zhao, Y Q; Babatunde, A O; Zhao, X H; Jørgensen, S E

    2011-01-01

    A dynamic simulation model was developed for the removal of soluble reactive phosphorus (SRP) from the vertical flow constructed wetlands (VFCW) using a dynamic software program called STELLA (structural thinking, experiential learning laboratory with animation) 9.1.3 to aid in simulating the environmental nature and succession of relationship between interdependent components and processes in the VFCW system. In particular, the VFCW employed dewatered alum sludge as its main substrate to enhance phosphorus (P) immobilization. Although computer modelling of P in treatment wetland has been well studied especially in recent years, there is still a need to develop simple and realistic models that can be used for investigating the dynamics of SRP in VFCWs. The state variables included in the model are dissolved phosphorus (DISP), plant phosphorus (PLAP), detritus phosphorus (DETP), plant biomass (PLBI) and adsorbed phosphorus (ADSP). The major P transformation processes considered in this study were adsorption, plant and microbial uptake and decomposition. The forcing functions which were considered in the model are temperature, radiation, volume of wastewater, P concentration, contact time, flow rate and the adsorbent (i.e., alum sludge). The model results revealed that up to 72% of the SRP can be removed through adsorption process whereas the uptake by plants is about 20% and the remaining processes such as microbial P utilization and decomposition, accounted for 7% SRP removal based on the mass balance calculations. The results obtained indicate that the model can be used to simulate outflow SRP concentration, and it can also be used to estimate the amount of P removed by individual processes in the VFCW using alum-sludge as a substrate.

  10. Modeling of Thermal Distributions around a Barrier at the Interface of Coating and Substrate

    Directory of Open Access Journals (Sweden)

    Ali Sahin

    2013-01-01

    Full Text Available Due to constant heat flux, the thermal distribution around an insulated barrier at the interface of substrate and functionally graded material (FGM which are essentially two-phase particulate composites is examined in such a way that the volume fractions of the constituents vary continuously in the thickness direction. Using integral transform method, two-dimensional steady-state diffusion equation with variable conductivity is turned into constant coefficient differential equation. Reducing that equation to a singular integral equation with Cauchy type, the temperature distribution around the barrier is obtained by defining an unknown function, which is called density function, as a series expansion of orthogonal polynomials. Results are shown for different thickness and nonhomogeneity parameters of FGM.

  11. Multi-scale modeling for prediction of distributed cellular properties in response to substrate spatial gradients in a continuously run microreactor

    DEFF Research Database (Denmark)

    Lencastre Fernandes, Rita; Krühne, Ulrich; Nopens, Ingmar;

    2012-01-01

    microbioreactor is simulated. A multiscale model consisting of the coupling of a population balance model, a kinetic model and a flow model was developed in order to predict simultaneously local concentrations of substrate (glucose), product (ethanol) and biomass, as well as the local cell size distributions....

  12. Intranuclear targeting and nuclear export of the adenovirus E1B-55K protein are regulated by SUMO1 conjugation

    OpenAIRE

    Kindsmüller, Kathrin; Groitl, Peter; Härtl, Barbara; Blanchette, Paola; Hauber, Joachim; Dobner, Thomas

    2007-01-01

    We have investigated the requirements for CRM1-mediated nuclear export and SUMO1 conjugation of the adenovirus E1B-55K protein during productive infection. Our data show that CRM1 is the major export receptor for E1B-55K in infected cells. Functional inactivation of the E1B-55K CRM1-dependent nuclear export signal (NES) or leptomycin B treatment causes an almost complete redistribution of the viral protein from the cytoplasm to the nucleus and its accumulation at the periphery of the viral re...

  13. 8051 Mikrodenetleyicili Bir Sumo Robot Tasarımı ve Uygulaması

    Directory of Open Access Journals (Sweden)

    Mehmet ALBAYRAK

    2009-04-01

    Full Text Available Bu çalısmada, 8051 mikrodenetleyici kontrollü bir sumo robotun mekanik tasarımı, kontrol kartı ve yazılımı gerçeklestirilmistir. Robot, yarısma ve egitim amaçlı tasarlandıgı için hücum amaçlı strateji içeren yazılım gelistirilerek yüklenmistir. 8051 mikrodenetleyici içindeki kontrol yazılımı PC 'nin seri portuna baglı mikrodenetleyici programlama kartına takılıp, kolaylıkla güncellenebildigi için yazılım stratejisi her yarısma için degistirilebilir niteliktedir. Microdenetleyici yazılımı μVision Keil derleyicisinde C kodu kullanılarak hazırlanmıstır. Mekanik tasarım asamasında robot dıs yüzeyinin darbelere dayanıklı malzemeden seçilmesine ve tekerleklerin sürtünmesinin fazla (kaymaz olmasına özen gösterilmistir. Robotun gücünü belirlenen agırlık sınırları içinde en fazla hale getirebilmek için 4 ayrı disli DC motor ve 4 amperlik akü kullanılmıstır. Rakip algılama mesafe sensörü ve kontrast sensörü (beyaz algılama seçiminde cevap verme süresi çok az olan sensörler tercih edilmistir.

  14. An experimental model of COD abatement in MBBR based on biofilm growth dynamic and on substrates' removal kinetics.

    Science.gov (United States)

    Siciliano, Alessio; De Rosa, Salvatore

    2016-08-01

    In this study, the performance of a lab-scale Moving Bed Biofilm Reactor (MBBR) under different operating conditions was analysed. Moreover, the dependence of the reaction rates both from the concentration and biodegradability of substrates and from the biofilm surface density, by means of several batch kinetic tests, was investigated. The reactor controls exhibited an increasing COD (Chemical Oxygen Demand) removal, reaching maximum yields (close to 90%) for influent loadings of up to12.5 gCOD/m(2)d. From this value, the pilot plant performance decreased to yields of only about 55% for influent loadings greater than 16 gCOD/m(2)d. In response to the influent loading increase, the biofilm surface density exhibited a logistic growing trend until reaching a maximum amount of total attached solids of about 9.5 g/m(2). The kinetic test results indicated that the COD removal rates for rapidly biodegradable, rapidly hydrolysable and slowly biodegradable substrates were not affected by the organic matter concentrations. Instead, first-order kinetics were detected with respect to biofilm surface density. The experimental results permitted the formulation of a mathematical model to predict the MBBR organic matter removal efficiency. The validity of the model was successfully tested in the lab-scale plant.

  15. Molecular modeling and docking of novel laccase from multiple serotype of Yersinia enterocolitica suggests differential and multiple substrate binding.

    Science.gov (United States)

    Singh, Deepti; Sharma, Krishna Kant; Dhar, Mahesh Shanker; Virdi, Jugsharan Singh

    2014-06-20

    Multi-copper oxidases (MCOs) are widely distributed in bacteria, where they are responsible for metal homeostasis, acquisition and oxidation. Using specific primers, yacK coding for MCO was amplified from different serotypes of Yersinia enterocolitica biovar 1A. Homology modeling of the protein followed by docking with five well-known substrates for different MCO's (viz., 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulphonic acid [ABTS], syringaldazine, L-tyrosine, ammonium ferrous sulfate and guaiacol), lignin monomers (Coniferyl alcohol, p-coumaryl alcohol and sinapyl alcohol) and two inhibitors i.e., kojic acid and N-hydroxyglycine was done. The docking gave maximum GoldScore i.e., 91.93 and 72.64 with ammonium ferrous sulfate and ABTS, respectively. Similarly, docking with ICM gave -82.10 and -83.61 docking score, confirming the protein to be true laccase with ferroxidase activity. Further, validation with ammonium ferrous sulfate as substrate gave laccase activity of 0.36Units/L/min. Guaiacol, L-tyrosine, and lignin monomers showed good binding affinity with protein models with GoldScores of 35.89, 41.82, 40.41, 41.12 and 43.10, respectively. The sequence study of all the cloned Yack genes showed serotype specific clade in dendrogram. There was distinct discrimination in the ligand binding affinity of Y. enterocolitica laccase, among strains of same clonal groups, suggesting it as a tool for phylogenetic studies.

  16. Influence of dormancy on microbial competition under intermittent substrate supply: insights from model simulations.

    Science.gov (United States)

    Stolpovsky, Konstantin; Fetzer, Ingo; Van Cappellen, Philippe; Thullner, Martin

    2016-06-01

    Most natural environments are characterized by frequent changes of their abiotic conditions. Microorganisms can respond to such changes by switching their physiological state between activity and dormancy allowing them to endure periods of unfavorable abiotic conditions. As a consequence, the competitiveness of microbial species is not simply determined by their growth performance under favorable conditions but also by their ability and readiness to respond to periods of unfavorable environmental conditions. The present study investigates the relevance of factors controlling the abundance and activity of individual bacterial species competing for an intermittently supplied substrate. For this purpose, numerical experiments were performed addressing the response of microbial systems to regularly applied feeding pulses. Simulation results show that community dynamics may exhibit a non-trivial link to the frequency of the external constraints and that for a certain combination of these environmental conditions coexistence of species is possible. The ecological implication of our results is that even non-dominant, neglected species can have a strong influence on realized species composition of dominant key species, due to their invisible presence enable the coexistence between important key species and by this affecting provided function of the system.

  17. Modeling of triangular-shaped substrates for light trapping in microcrystalline silicon solar cells

    Science.gov (United States)

    Zi, Wei; Hu, Jian; Ren, Xiaodong; Ren, Xianpei; Wei, QingBo; Liu, Shengzhong (Frank)

    2017-01-01

    The influence of triangular grating used as a light trapping structure on the optical wave propagation within thin-film microcrystalline silicon (μc-Si:H) solar cells is investigated. A finite difference time domain (FDTD) approach is used to rigorously solve the Maxwell's equations in three dimensions. We apply two parameters of mean surface roughness (Sa) and slope (k) to define triangular structure and study their influence on the absorption of μc-Si:H. When Sa and k are set to 400 nm and 1, respectively, a largest enhancement of absorption is achieved. The optimum short circuit photocurrent (Jsc) of a 1-μm thick μc-Si:H solar cell made on such a textured substrate can reach 27.0 mA/cm2. The carrier generation rate in the μc-Si:H material is also rigorously analyzed. Finally, we identify some key optical losses in μc-Si:H solar cells and propose for further optimizing the device design.

  18. The copper spoil heap Knappenberg, Austria, as a model for metal habitats – Vegetation, substrate and contamination

    Energy Technology Data Exchange (ETDEWEB)

    Adlassnig, Wolfram; Weiss, Yasmin S. [University of Vienna, Core Facility Cell Imaging and Ultrastructure Research, Althanstraße 14, A-1090 Vienna (Austria); Sassmann, Stefan [University of Vienna, Core Facility Cell Imaging and Ultrastructure Research, Althanstraße 14, A-1090 Vienna (Austria); University of Exeter, College of Life and Environmental Sciences, Biosciences, Stocker Road, Exeter EX4 4QD (United Kingdom); Steinhauser, Georg [Leibniz University Hannover, Institute of Radioecology and Radiation Protection, Herrenhäuser Straße 2, D30419 Hannover (Germany); Hofhansl, Florian [University of Vienna, Department of Microbiology and Ecosystem Science, Althanstraße 14, A-1090 Vienna (Austria); Instituto Nacional de Pesquisas da Amazônia, Coordenação de Dinâmica Ambiental, Manaus (Brazil); Baumann, Nils [Helmholtz-Zentrum Dresden-Rossendorf, Division of Biogeochemistry, Bautzner Landstraße 400, D-01328 Dresden (Germany); Lichtscheidl, Irene K. [University of Vienna, Core Facility Cell Imaging and Ultrastructure Research, Althanstraße 14, A-1090 Vienna (Austria); Lang, Ingeborg, E-mail: ingeborg.lang@univie.ac.at [University of Vienna, Core Facility Cell Imaging and Ultrastructure Research, Althanstraße 14, A-1090 Vienna (Austria)

    2016-09-01

    Historic mining in the Eastern Alps has left us with a legacy of numerous spoil heaps hosting specific, metal tolerant vegetation. Such habitats are characterized by elevated concentrations of toxic elements but also by high irradiation, a poorly developed substrate or extreme pH of the soil. This study investigates the distribution of vascular plants, mosses and lichens on a copper spoil heap on the ore bearing Knappenberg formed by Prebichl Layers and Werfener Schist in Lower Austria. It serves as a model for discriminating between various ecological traits and their effects on vegetation. Five distinct clusters were distinguished: (1) The bare, metal rich Central Spoil Heap was only colonised by highly resistant specialists. (2) The Northern and (3) Southern Peripheries contained less copper; the contrasting vegetation was best explained by the different microclimate. (4) A forest over acidic bedrock hosted a vegetation overlapping with the periphery of the spoil heap. (5) A forest over calcareous bedrock was similar to the spoil heap with regard to pH and humus content but hosted a vegetation differing strongly to all other habitats. Among the multiple toxic elements at the spoil heap, only Cu seems to exert a crucial influence on the vegetation pattern. Besides metal concentrations, irradiation, humidity, humus, pH and grain size distribution are important for the establishment of a metal tolerant vegetation. The difference between the species poor Northern and the diverse Southern Periphery can be explained by the microclimate rather than by the substrate. All plant species penetrating from the forest into the periphery of the spoil heap originate from the acidic but not from the calcareous bedrock. - Highlights: • Strong impact on plant diversity by isolation and extreme abiotic conditions • Both, microclimate and substrate explain species distribution. • Increased cellular metal tolerance of plants from the Central Spoil Heap • Among toxic elements

  19. Application of Thermal Network Model to Transient Thermal Analysis of Power Electronic Package Substrate

    National Research Council Canada - National Science Library

    Ishizuka, Masaru; Hatakeyama, Tomoyuki; Funawatashi, Yuichi; Koizumi, katsuhiro

    2011-01-01

    .... This paper describes an application of the thermal network method to the transient thermal analysis of multichip modules and proposes a simple model for the thermal analysis of multichip modules as a preliminary thermal design tool. On the basis of the result of transient thermal analysis, the validity of the thermal network method and the simple thermal analysis model is confirmed.

  20. A Mathematical Model for Dynamic Simulation of Anaerobic Digestion of Complex Substrates

    DEFF Research Database (Denmark)

    Angelidaki, Irini; Ellegaard, L.; Ahring, Birgitte Kiær

    1993-01-01

    of pH and temperature characteristics in order to accurately simulate free ammonia concentration. Free ammonia and acetate constitute the primary modulating factors in the model. The model has been applied for the simulation of digestion of cattle manure in continuously stirred tank reactors (CSTRs...

  1. Analytical and Numerical Modelling of FRP Debonding from Concrete Substrate under Pure Shearing

    Institute of Scientific and Technical Information of China (English)

    PAN Jinlong; XU Zhun; C K Y Leung; LI Zongjin

    2012-01-01

    External bonding of fiber reinforced polymer (FRP) composites on the concrete structures has been proved to be an effective and efficient way to strengthen concrete structures.For a FRP strengthened concrete beam,it is usually observed that the failure occurs in the concrete and a thin layer of concrete is attached on the surface of the debonded FRP plate.To study the debond behavior between concrete and FRP composites,an analytical model based on the three-parameter model is developed to study the debonding behavior for the FRP-to-concrete joint under pure shearing.Then,nonlinear FEM analysis is conducted to verify the proposed analytical model.The FEM results shows good agreement with the results from the model.Finally,with the analytical model,sensitivity analyses are performed to study the effect of the interfacial parameters or the geometric parameters on the debonding behavior.

  2. A model describing Debaryomyces hansenii growth and substrate consumption during a smear soft cheese deacidification and ripening.

    Science.gov (United States)

    Riahi, M H; Trelea, I C; Picque, D; Leclercq-Perlat, M-N; Hélias, A; Corrieu, G

    2007-05-01

    A mechanistic model for Debaryomyces hansenii growth and substrate consumption, lactose conversion into lactate by lactic acid bacteria, as well as lactose and lactate transfer from the core toward the rind was established. The model described the first step (14 d) of the ripening of a smear soft cheese and included the effects of temperature and relative humidity of the ripening chamber on the kinetic parameters. Experimental data were collected from experiments carried out in an aseptic pilot scale ripening chamber under 9 different combinations of temperature (8, 12, and 16 degrees C) and relative humidity (85, 93, and 99%) according to a complete experimental design. The model considered the cheese as a system with 2 compartments (rind and core) and included 5 state evolution equations and 16 parameters. The model succeeded in predicting D. hansenii growth and lactose and lactate concentrations during the first step of ripening (curd deacidification) in core and rind. The nonlinear data-fitting method allowed the determination of tight confidence intervals for the model parameters. The residual standard error (RSE) between model predictions and experimental data was close to the experimental standard deviation between repeated experiments.

  3. A whole-body model for glycogen regulation reveals a critical role for substrate cycling in maintaining blood glucose homeostasis.

    Directory of Open Access Journals (Sweden)

    Ke Xu

    2011-12-01

    Full Text Available Timely, and sometimes rapid, metabolic adaptation to changes in food supply is critical for survival as an organism moves from the fasted to the fed state, and vice versa. These transitions necessitate major metabolic changes to maintain energy homeostasis as the source of blood glucose moves away from ingested carbohydrates, through hepatic glycogen stores, towards gluconeogenesis. The integration of hepatic glycogen regulation with extra-hepatic energetics is a key aspect of these adaptive mechanisms. Here we use computational modeling to explore hepatic glycogen regulation under fed and fasting conditions in the context of a whole-body model. The model was validated against previous experimental results concerning glycogen phosphorylase a (active and glycogen synthase a dynamics. The model qualitatively reproduced physiological changes that occur during transition from the fed to the fasted state. Analysis of the model reveals a critical role for the inhibition of glycogen synthase phosphatase by glycogen phosphorylase a. This negative regulation leads to high levels of glycogen synthase activity during fasting conditions, which in turn increases substrate (futile cycling, priming the system for a rapid response once an external source of glucose is restored. This work demonstrates that a mechanistic understanding of the design principles used by metabolic control circuits to maintain homeostasis can benefit from the incorporation of mathematical descriptions of these networks into "whole-body" contextual models that mimic in vivo conditions.

  4. Offshore Substrate

    Data.gov (United States)

    California Department of Resources — This shapefile displays the distribution of substrate types from Pt. Arena to Pt. Sal in central/northern California. Originally this data consisted of seven paper...

  5. SUMO modification of Stra13 is required for repression of cyclin D1 expression and cellular growth arrest.

    Directory of Open Access Journals (Sweden)

    Yaju Wang

    Full Text Available Stra13, a basic helix-loop-helix (bHLH transcription factor is involved in myriad biological functions including cellular growth arrest, differentiation and senescence. However, the mechanisms by which its transcriptional activity and function are regulated remain unclear. In this study, we provide evidence that post-translational modification of Stra13 by Small Ubiquitin-like Modifier (SUMO dramatically potentiates its ability to transcriptionally repress cyclin D1 and mediate G(1 cell cycle arrest in fibroblast cells. Mutation of SUMO acceptor lysines 159 and 279 located in the C-terminal repression domain has no impact on nuclear localization; however, it abrogates association with the co-repressor histone deacetylase 1 (HDAC1, attenuates repression of cyclin D1, and prevents Stra13-mediated growth suppression. HDAC1, which promotes cellular proliferation and cell cycle progression, antagonizes Stra13 sumoylation-dependent growth arrest. Our results uncover an unidentified regulatory axis between Stra13 and HDAC1 in progression through the G(1/S phase of the cell cycle, and provide new mechanistic insights into regulation of Stra13-mediated transcriptional repression by sumoylation.

  6. SUMO ligase PIAS1 functions as a target gene selective androgen receptor coregulator on prostate cancer cell chromatin.

    Science.gov (United States)

    Toropainen, Sari; Malinen, Marjo; Kaikkonen, Sanna; Rytinki, Miia; Jääskeläinen, Tiina; Sahu, Biswajyoti; Jänne, Olli A; Palvimo, Jorma J

    2015-01-01

    Androgen receptor (AR) is a ligand-activated transcription factor that plays a central role in the development and growth of prostate carcinoma. PIAS1 is an AR- and SUMO-interacting protein and a putative transcriptional coregulator overexpressed in prostate cancer. To study the importance of PIAS1 for the androgen-regulated transcriptome of VCaP prostate cancer cells, we silenced its expression by RNAi. Transcriptome analyses revealed that a subset of the AR-regulated genes is significantly influenced, either activated or repressed, by PIAS1 depletion. Interestingly, PIAS1 depletion also exposed a new set of genes to androgen regulation, suggesting that PIAS1 can mask distinct genomic loci from AR access. In keeping with gene expression data, silencing of PIAS1 attenuated VCaP cell proliferation. ChIP-seq analyses showed that PIAS1 interacts with AR at chromatin sites harboring also SUMO2/3 and surrounded by H3K4me2; androgen exposure increased the number of PIAS1-occupying sites, resulting in nearly complete overlap with AR chromatin binding events. PIAS1 interacted also with the pioneer factor FOXA1. Of note, PIAS1 depletion affected AR chromatin occupancy at binding sites enriched for HOXD13 and GATA motifs. Taken together, PIAS1 is a genuine chromatin-bound AR coregulator that functions in a target gene selective fashion to regulate prostate cancer cell growth. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

  7. Modelling the anaerobic digestion of solid organic waste - Substrate characterisation method for ADM1 using a combined biochemical and kinetic parameter estimation approach.

    Science.gov (United States)

    Poggio, D; Walker, M; Nimmo, W; Ma, L; Pourkashanian, M

    2016-07-01

    This work proposes a novel and rigorous substrate characterisation methodology to be used with ADM1 to simulate the anaerobic digestion of solid organic waste. The proposed method uses data from both direct substrate analysis and the methane production from laboratory scale anaerobic digestion experiments and involves assessment of four substrate fractionation models. The models partition the organic matter into a mixture of particulate and soluble fractions with the decision on the most suitable model being made on quality of fit between experimental and simulated data and the uncertainty of the calibrated parameters. The method was tested using samples of domestic green and food waste and using experimental data from both short batch tests and longer semi-continuous trials. The results showed that in general an increased fractionation model complexity led to better fit but with increased uncertainty. When using batch test data the most suitable model for green waste included one particulate and one soluble fraction, whereas for food waste two particulate fractions were needed. With richer semi-continuous datasets, the parameter estimation resulted in less uncertainty therefore allowing the description of the substrate with a more complex model. The resulting substrate characterisations and fractionation models obtained from batch test data, for both waste samples, were used to validate the method using semi-continuous experimental data and showed good prediction of methane production, biogas composition, total and volatile solids, ammonia and alkalinity.

  8. MOLECULAR MODELLING OF HUMAN ALDEHYDE OXIDASE AND IDENTIFICATION OF THE KEY INTERACTIONS IN THE ENZYME-SUBSTRATE COMPLEX

    Directory of Open Access Journals (Sweden)

    Siavoush Dastmalchi

    2005-05-01

    Full Text Available Aldehyde oxidase (EC 1.2.3.1, a cytosolic enzyme containing FAD, molybdenum and iron-sulphur cluster, is a member of non-cytochrome P-450 enzymes called molybdenum hydroxylases which is involved in the metabolism of a wide range of endogenous compounds and many drug substances. Drug metabolism is one of the important characteristics which influences many aspects of a therapeutic agent such as routes of administration, drug interaction and toxicity and therefore, characterisation of the key interactions between enzymes and substrates is very important from drug development point of view. The aim of this study was to generate a three-dimensional model of human aldehyde oxidase (AO in order to assist us to identify the mode of interaction between enzyme and a set of phethalazine/quinazoline derivatives. Both sequence-based (BLAST and inverse protein fold recognition methods (THREADER were used to identify the crystal structure of bovine xanthine dehydrogenase (pdb code of 1FO4 as the suitable template for comparative modelling of human AO. Model structure was generated by aligning and then threading the sequence of human AO onto the template structure, incorporating the associated cofactors, and molecular dynamics simulations and energy minimization using GROMACS program. Different criteria which were measured by the PROCHECK, QPACK, VERIFY-3D were indicative of a proper fold for the predicted structural model of human AO. For example, 97.9 percentages of phi and psi angles were in the favoured and most favoured regions in the ramachandran plot, and all residues in the model are assigned environmentally positive compatibility scores. Further evaluation on the model quality was performed by investigation of AO-mediated oxidation of a set of phthalazine/quinazoline derivatives to develop QSAR model capable of describing the extent of the oxidation. Substrates were aligned by docking onto the active site of the enzyme using GOLD technology and then

  9. Establishment and experimental verification of the photoresist model considering interface slip between photoresist and concave spherical substrate

    Directory of Open Access Journals (Sweden)

    S. Yang

    2015-07-01

    Full Text Available A thickness distribution model of photoresist spin-coating on concave spherical substrate (CSS has been developed via both theoretical studies and experimental verification. The stress of photoresist on rotating CSS is analyzed and the boundary conditions of hydrodynamic equation are presented under the non-lubricating condition. Moreover, a multivariable polynomial equation of photoresist-layer thickness distribution is derived by analyzing and deducing the flow equation where the evaporation rate, substrate topography, interface slip between liquid and CSS, and the variation of rotational speed and photoresist parameters are considered in detail. Importantly, the photoresist-layer thickness at various CSS rotational speeds and liquid concentrations can be obtained according to the theoretical equation. The required photoresist viscosity and concentration parameters of different photoresist coating thickness under a certain coating speeds can be also solved through this equation. It is noted that the calculated theoretical values are well consistent with the experimental results which were measured with various CSS rotational speeds and liquid concentrations at steady state. Therefore, both our experimental results and theoretical analysis provide the guidance for photoresist dilution and pave the way for potential improvements and microfabrication applications in the future.

  10. SUMO proteases ULP1c and ULP1d are required for development and osmotic stress responses in Arabidopsis thaliana

    DEFF Research Database (Denmark)

    Castro, Pedro Humberto Araújo R F; Couto, Daniel; Freitas, Sara

    2016-01-01

    diminished in vitro root growth under low water potential and higher stomatal aperture, yet leaf transpirational water loss and whole drought tolerance were not significantly altered. Generation of a triple siz1 ulp1c ulp1d mutant suggests that ULP1c/d and the SUMO E3 ligase SIZ1 may display separate...

  11. Interferon-induced antiviral Mx1 GTPase is associated with components of the SUMO-1 system and promyelocytic leukemia protein nuclear bodies.

    Science.gov (United States)

    Engelhardt, O G; Ullrich, E; Kochs, G; Haller, O

    2001-12-10

    Mx proteins are interferon-induced large GTPases, some of which have antiviral activity against a variety of viruses. The murine Mx1 protein accumulates in the nucleus of interferon-treated cells and is active against members of the Orthomyxoviridae family, such as the influenza viruses and Thogoto virus. The mechanism by which Mx1 exerts its antiviral action is still unclear, but an involvement of undefined nuclear factors has been postulated. Using the yeast two-hybrid system, we identified cellular proteins that interact with Mx1 protein. The Mx1 interactors were mainly nuclear proteins. They included Sp100, Daxx, and Bloom's syndrome protein (BLM), all of which are known to localize to specific subnuclear domains called promyelocytic leukemia protein nuclear bodies (PML NBs). In addition, components of the SUMO-1 protein modification system were identified as Mx1-interacting proteins, namely the small ubiquitin-like modifier SUMO-1 and SAE2, which represents subunit 2 of the SUMO-1 activating enzyme. Analysis of the subcellular localization of Mx1 and some of these interacting proteins by confocal microscopy revealed a close spatial association of Mx1 with PML NBs. This suggests a role of PML NBs and SUMO-1 in the antiviral action of Mx1 and may allow us to discover novel functions of this large GTPase.

  12. A multiphase mixture model for substrate concentration distribution characteristics and photo-hydrogen production performance of the entrapped-cell photobioreactor.

    Science.gov (United States)

    Guo, Cheng-Long; Cao, Hong-Xia; Pei, Hong-Shan; Guo, Fei-Qiang; Liu, Da-Meng

    2015-04-01

    A multiphase mixture model was developed for revealing the interaction mechanism between biochemical reactions and transfer processes in the entrapped-cell photobioreactor packed with gel granules containing Rhodopseudomonas palustris CQK 01. The effects of difference operation parameters, including operation temperature, influent medium pH value and porosity of packed bed, on substrate concentration distribution characteristics and photo-hydrogen production performance were investigated. The results showed that the model predictions were in good agreement with the experimental data reported. Moreover, the operation temperature of 30 °C and the influent medium pH value of 7 were the most suitable conditions for photo-hydrogen production by biodegrading substrate. In addition, the lower porosity of packed bed was beneficial to enhance photo-hydrogen production performance owing to the improvement on the amount of substrate transferred into gel granules caused by the increased specific area for substrate transfer in the elemental volume.

  13. Application of Thermal Network Model to Transient Thermal Analysis of Power Electronic Package Substrate

    Directory of Open Access Journals (Sweden)

    Masaru Ishizuka

    2011-01-01

    Full Text Available In recent years, there is a growing demand to have smaller and lighter electronic circuits which have greater complexity, multifunctionality, and reliability. High-density multichip packaging technology has been used in order to meet these requirements. The higher the density scale is, the larger the power dissipation per unit area becomes. Therefore, in the designing process, it has become very important to carry out the thermal analysis. However, the heat transport model in multichip modules is very complex, and its treatment is tedious and time consuming. This paper describes an application of the thermal network method to the transient thermal analysis of multichip modules and proposes a simple model for the thermal analysis of multichip modules as a preliminary thermal design tool. On the basis of the result of transient thermal analysis, the validity of the thermal network method and the simple thermal analysis model is confirmed.

  14. Growth and lipid production of Umbelopsis isabellina on a solid substrate - Mechanistic modeling and validation

    NARCIS (Netherlands)

    Meeuwse, P.; Klok, A.J.; Haemers, S.; Tramper, J.; Rinzema, A.

    2012-01-01

    Microbial lipids are an interesting feedstock for biodiesel. Their production from agricultural waste streams by fungi cultivated in solid-state fermentation may be attractive, but the yield of this process is still quite low. In this article, a mechanistic model is presented that describes growth,

  15. The effect of roxatidine acetate and cimetidine on hepatic drug clearance assessed by simultaneous administration of three model substrates.

    Science.gov (United States)

    Tanaka, E; Nakamura, K

    1989-08-01

    The effect of pretreatment for 7 days with either roxatidine acetate 75 mg twice daily or cimetidine 200 mg four times daily on the kinetics of antipyrine (AP), trimethadione (TMO) and indocyanine green (ICG) was studied in seven healthy, male, nonsmoking subjects. After pretreatment with cimetidine, the plasma clearances (CL) of AP and TMO were significantly lower and the elimination half-life (t1/2) of AP was significantly increased. The volumes of distribution (V) of AP and TMO were not affected. After roxatidine acetate, the pharmacokinetics of AP and TMO were unchanged. The cumulative renal excretion (% dose) and formation clearance of 3-hydroxymethyl-3-nor-antipyrine (NORA) were lowered by cimetidine treatment, but not following the administration of roxatidine acetate. ICG clearance was not changed significantly by either pretreatment. The results of this study show that roxatidine acetate does not impair the metabolism of three model substrates used to assess hepatic drug clearance.

  16. Deposition, diffusion, and aggregation on small-world networks: a model for nanostructure growth on the defective substrate

    Energy Technology Data Exchange (ETDEWEB)

    Shao Zhigang [Department of Physics, Wuhan University, Wuhan 430072 (China); Zou Xianwu [Department of Physics, Wuhan University, Wuhan 430072 (China)]. E-mail: xwzou@whu.edu.cn; Tan Zhijie [Department of Physics, Wuhan University, Wuhan 430072 (China); Huang Shengyou [Department of Physics, Wuhan University, Wuhan 430072 (China); Jin Zhunzhi [Department of Physics, Wuhan University, Wuhan 430072 (China)

    2004-10-11

    Deposition, diffusion, and aggregation (DDA) on a two-dimensional small-world network have been investigated by computer simulations. This model is characterized by two parameters: the clustering exponent {alpha} and long-range connection rate {phi}. The results show that as {alpha} and {phi} vary there exists a continuous crossover in the fractal dimension Df from 1.65 to 2, which corresponds to the crossover from the DDA pattern to dense one. The change of the aggregation pattern results from the long-range connection in the network, which reduces the effect of screening during the aggregation. With primitive analysis, we obtain the expressions of the fractal dimension Df and the crossover point {alpha}c. These results may be useful to describe the vapor deposition, molecular-beam epitaxy, and similar experiments on defective substrates.

  17. Study of the role of functional variants of SLC22A4, RUNX1 and SUMO4 in systemic lupus erythematosus

    Science.gov (United States)

    Orozco, G; Sánchez, E; Gómez, L M; González‐Gay, M A; López‐Nevot, M A; Torres, B; Ortego‐Centeno, N; Jiménez‐Alonso, J; de Ramón, E; Román, J Sánchez; Anaya, J M; Sturfelt, G; Gunnarsson, I; Svennungsson, E; Alarcón‐Riquelme, M; González‐Escribano, M F; Martín, J

    2006-01-01

    Background Functional polymorphisms of the solute carrier family 22, member 4 (SLC22A4), runt related transcription factor 1 (RUNX1) and small ubiquitin‐like modifier 4 (SUMO4) genes have been shown to be associated with several autoimmune diseases. Objective To test the possible role of these variants in susceptibility to or severity of systemic lupus erythematosus (SLE), on the basis that common genetic bases are shared by autoimmune disorders. Methods 597 SLE patients and 987 healthy controls of white Spanish origin were studied. Two additional cohorts of 228 SLE patients from Sweden and 122 SLE patients from Colombia were included. A case–control association study was carried out with six single nucleotide polymorphisms (SNP) spanning the SLC22A4 gene, one SNP in RUNX1 gene, and one additional SNP in SUM04 gene. Results No significant differences were observed between SLE patients and healthy controls when comparing the distribution of the genotypes or alleles of any of the SLC22A4, RUNX1, or SUMO4 polymorphisms tested. Significant differences were found in the distribution of the SUMO4 genotypes and alleles among SLE patients with and without nephritis, but after multiple testing correction, the significance of the association was lost. The association of SUMO4 with nephritis could not be verified in two independent SLE cohorts from Sweden and Colombia. Conclusions These results suggest that the SLC22A4, RUNX1, and SUMO4 polymorphisms analysed do not play a role in the susceptibility to or severity of SLE. PMID:16249223

  18. Toward a biaxial model of "bipolar" affective disorders: further exploration of genetic, molecular and cellular substrates.

    Science.gov (United States)

    Askland, Kathleen

    2006-08-01

    Current epidemiologic and genetic evidence strongly supports the heritability of bipolar disease. Inconsistencies across linkage and association analyses have been primarily interpreted as suggesting polygenic, nonMendelian and variably-penetrant inheritance (i.e., in terms of interacting disease models). An equally-likely explanation for this genetic complexity is that trait, locus and allelic heterogeneities (i.e., a heterogeneous disease model) are primarily responsible for observed variability at the population level. The two models of genetic complexity are not mutually-exclusive, and are in fact likely to co-exist both in trait determination and disease expression. However, the current model proposes that, while both types of complex genetics are likely central to observable affective trait spectra, inheritance patterns, gross phenotypic categories and treatment-responsiveness in affective disease (as well as the widespread inconsistencies across such studies) may be primarily explained in terms of a heterogeneous disease model. Gene-gene, gene-protein and protein-protein interactions, then, are most likely to serve as trait determinants and 'phenotypic modifiers' rather than as primary pathogenic determinants. Moreover, while locus heterogeneity indicates the presence of multiple susceptibility genes at the population level, it does not necessitate polygenic inheritance at the individual or pedigree level. Rather, it is compatible with the possibility of mono- or bigenic determination of disease susceptibility within individuals/pedigrees. More specifically, the biaxial model proposes that integration of specific findings from genetic linkage and association studies, ion channels research as well as pharmacologic mechanism, phenotypic specificity and effectiveness studies suggests that each gene of potential etiologic significance in primary affective illness might be categorized into one of two classes, according to their primary role in neuronal

  19. Thermal behavior analysis of GaN based epi-material on different substrates by means of a physical-thermal model

    Energy Technology Data Exchange (ETDEWEB)

    Tang, Xiao; Rousseau, Michel; Defrance, Nicolas; Hoel, Virginie; Soltani, Ali; De Jaeger, Jean-Claude [Institut d' Electronique de Microelectronique et de Nanotechnologie, UMR CNRS 8520, Lille University, Avenue Poincare, BP 60069, 59652 Villeneuve d' Ascq (France); Langer, Robert [PICOGIGA International, Place Marcel Rebuffat, Parc de Villejust, 91971 Courtaboeuf Cedex (France)

    2010-08-15

    For gallium nitride (GaN) based microwave power devices, the thermal behavior due to the self-heating effect constitutes a main limitation because the power dissipation is very high. For this study, a predictive physical-thermal model has been developed to analyze the physical and thermal phenomena observed in experiment. In this paper, the thermal performances of AlGaN/GaN epitaxies grown on different substrates are determined. It is found that compared with Si substrate, composite substrates: SopSiC (mono-Si/poly-SiC) and SiCopSiC (mono-SiC/poly-SiC) substrates (V. Hoel et al., Electron. Lett. 44, 238 (2008) 1 and T.J. Anderson et al., J. Vac. Sci. Technol. B 24, 2302 (2006) 2) present better thermal resistances especially at high dissipated power densities with an improvement of 18% for SopSiC substrate and 25% for SiCopSiC substrate at 12 W/mm. Furthermore, polycrystalline diamond is the most promising substrate with a thermal resistance of 5.4 W/m.K at 12 W/mm. The simulation results, such as lattice temperature and thermal resistance, are in good agreement with our measurements. (Abstract Copyright [2010], Wiley Periodicals, Inc.)

  20. Experimental Models of Oral Biofilms Developed on Inert Substrates: A Review of the Literature

    Science.gov (United States)

    Darrene, Lopez-Nguyen

    2016-01-01

    The oral ecosystem is a very complex environment where more than 700 different bacterial species can be found. Most of them are organized in biofilm on dental and mucosal surfaces. Studying this community is important because a rupture in stability can lead to the preeminence of pathogenic microorganisms, causing dental decay, gingivitis, or periodontitis. The multitude of species complicates biofilm analysis so its reproduction, collection, and counting are very delicate. The development of experimental models of dental biofilms was therefore essential and multiple in vitro designs have emerged, each of them especially adapted to observing biofilm formation of specific bacteria within specific environments. The aim of this review is to analyze oral biofilm models. PMID:27699173

  1. Experimental Models of Oral Biofilms Developed on Inert Substrates: A Review of the Literature

    Directory of Open Access Journals (Sweden)

    Lopez-Nguyen Darrene

    2016-01-01

    Full Text Available The oral ecosystem is a very complex environment where more than 700 different bacterial species can be found. Most of them are organized in biofilm on dental and mucosal surfaces. Studying this community is important because a rupture in stability can lead to the preeminence of pathogenic microorganisms, causing dental decay, gingivitis, or periodontitis. The multitude of species complicates biofilm analysis so its reproduction, collection, and counting are very delicate. The development of experimental models of dental biofilms was therefore essential and multiple in vitro designs have emerged, each of them especially adapted to observing biofilm formation of specific bacteria within specific environments. The aim of this review is to analyze oral biofilm models.

  2. Modeling the pulsed light inactivation of microorganisms naturally occurring on vegetable substrates.

    Science.gov (United States)

    Izquier, Adriana; Gómez-López, Vicente M

    2011-09-01

    Pulsed light (PL) is a fast non-thermal method for microbial inactivation. This research studied the kinetics of PL inactivation of microorganisms naturally occurring in some vegetables. Iceberg lettuce, white cabbage and Julienne-style cut carrots were subjected to increasing PL fluences up to 12J/cm(2) in order to study its effect on aerobic mesophilic bacteria determined by plate count. Also, sample temperature increase was determined by infrared thermometry. Survivors' curves were adjusted to several models. No shoulder but tail was observed. The Weibull model showed good fitting performance of data. Results for lettuce were: goodness-of-fit parameter RMSE=0.2289, fluence for the first decimal reduction δ=0.98±0.80J/cm(2) and concavity parameter p=0.33±0.08. Results for cabbage were: RMSE=0.0725, δ=0.81±0.23J/cm(2) and p=0.30±0.02; and for carrot: RMSE=0.1235, δ=0.39±0.24J/cm(2) and p=0.23±0.03. For lettuce, a log-linear and tail model was also suitable. Validation of the Weibull model produced determination coefficients of 0.88-0.96 and slopes of 0.78-0.99. Heating was too low to contribute to inactivation. A single low-energy pulse was enough to achieve one log reduction, with an ultrafast treatment time of 0.5ms. While PL efficacy was found to be limited to high residual counts, the achievable inactivation level may be considered useful for shelf-life extension.

  3. A lattice Boltzmann model for substrates with regularly structured surface roughness

    Science.gov (United States)

    Yagub, A.; Farhat, H.; Kondaraju, S.; Singh, T.

    2015-11-01

    Superhydrophobic surface characteristics are important in many industrial applications, ranging from the textile to the military. It was observed that surfaces fabricated with nano/micro roughness can manipulate the droplet contact angle, thus providing an opportunity to control the droplet wetting characteristics. The Shan and Chen (SC) lattice Boltzmann model (LBM) is a good numerical tool, which holds strong potentials to qualify for simulating droplets wettability. This is due to its realistic nature of droplet contact angle (CA) prediction on flat smooth surfaces. But SC-LBM was not able to replicate the CA on rough surfaces because it lacks a real representation of the physics at work under these conditions. By using a correction factor to influence the interfacial tension within the asperities, the physical forces acting on the droplet at its contact lines were mimicked. This approach allowed the model to replicate some experimentally confirmed Wenzel and Cassie wetting cases. Regular roughness structures with different spacing were used to validate the study using the classical Wenzel and Cassie equations. The present work highlights the strength and weakness of the SC model and attempts to qualitatively conform it to the fundamental physics, which causes a change in the droplet apparent contact angle, when placed on nano/micro structured surfaces.

  4. Imaging a cognitive model of apraxia: the neural substrate of gesture-specific cognitive processes.

    Science.gov (United States)

    Peigneux, Philippe; Van der Linden, Martial; Garraux, Gaetan; Laureys, Steven; Degueldre, Christian; Aerts, Joel; Del Fiore, Guy; Moonen, Gustave; Luxen, Andre; Salmon, Eric

    2004-03-01

    The present study aimed to ascertain the neuroanatomical basis of an influential neuropsychological model for upper limb apraxia [Rothi LJ, et al. The Neuropsychology of Action. 1997. Hove, UK: Psychology Press]. Regional cerebral blood flow was measured in healthy volunteers using H2 15O PET during performance of four tasks commonly used for testing upper limb apraxia, i.e., pantomime of familiar gestures on verbal command, imitation of familiar gestures, imitation of novel gestures, and an action-semantic task that consisted in matching objects for functional use. We also re-analysed data from a previous PET study in which we investigated the neural basis of the visual analysis of gestures. First, we found that two sets of discrete brain areas are predominantly engaged in the imitation of familiar and novel gestures, respectively. Segregated brain activation for novel gesture imitation concur with neuropsychological reports to support the hypothesis that knowledge about the organization of the human body mediates the transition from visual perception to motor execution when imitating novel gestures [Goldenberg Neuropsychologia 1995;33:63-72]. Second, conjunction analyses revealed distinctive neural bases for most of the gesture-specific cognitive processes proposed in this cognitive model of upper limb apraxia. However, a functional analysis of brain imaging data suggested that one single memory store may be used for "to-be-perceived" and "to-be-produced" gestural representations, departing from Rothi et al.'s proposal. Based on the above considerations, we suggest and discuss a revised model for upper limb apraxia that might best account for both brain imaging findings and neuropsychological dissociations reported in the apraxia literature. Copyright 2004 Wiley-Liss, Inc.

  5. Mathematical modeling of the "plant community -soil-like substrate -gas exchange with the human" closed ecosystem

    Science.gov (United States)

    Barkhatov, Yuri; Gubanov, Vladimir; Tikhomirov, Alexander A.; Degermendzhy, Andrey G.

    A mathematical model of the "plant community -soil-like substrate -gas exchange with the human" experimental biological life support system (BLSS) has been constructed to predict its functioning and estimate feasibility of controlling it. The mathematical model consists of three compartments -two `phytotron' models (with wheat and radish) and the `mycotron' model (for mushrooms). The following components are included in the model: edible mushrooms (mushroom fruit bodies and mycelium); wheat; radish; straw (processed by mycelium); dead organic matter in the phytotron (separately for the wheat unit and for the radish unit); worms; worms' coprolites; vermicompost used as a soil-like substrate (SLS); bacterial microflora; min-eral nitrogen, phosphorus and iron; products of the system intended for humans (wheat grains, radish roots and mushroom fruit bodies); oxygen and carbon dioxide. Under continuous gas exchange, the mass exchange between the compartments occurs at the harvesting time. The conveyor character of the closed ecosystem functioning has been taken into account -the num-ber of culture age groups can be regulated (in experiments -4 and 8 age groups). The conveyor cycle duration can be regulated as well. The module is designed for the food and gas exchange requirements of 1/30 of a virtually present human. Aim of model analysis is determination of investigation direction in real experimental BLSS. The model allows doing dynamic calcu-lations of closure coefficient based on the main elements taken into account in the model and evaluating all dynamic components of the system under different conditions and modes of its operation, especially under the conditions that can hardly be created experimentally. One of the sustainability conditions can be long-duration functioning of the system under the light-ing that is far from the optimum. The mathematical model of the system can demonstrate variants of its sustainable functioning or ruin under various critical

  6. Correlation between enzyme activity and substrate storage in a cell culture model system for Gaucher disease.

    Science.gov (United States)

    Schueler, U H; Kolter, T; Kaneski, C R; Zirzow, G C; Sandhoff, K; Brady, R O

    2004-01-01

    Gaucher disease, the most common sphingolipidosis, is caused by a decreased activity of glucosylceramide beta-glucosidase, resulting in the accumulation of glucosylceramide in macrophage-derived cells known as Gaucher cells. Much of the storage material is thought to originate from the turnover of cell membranes, such as phagocytosed red and white blood cells. In this study, an in vitro model of Gaucher disease was developed by treating the murine macrophage cell line J774 with a specific inhibitor of glucosylceramide beta-glucosidase, conduritol B-epoxide, and feeding red blood cell ghosts, in order to mimic the disease state. It was found in this model system that glucosylceramide beta-glucosidase activity could be reduced to about 11-15% of the normal control level before increased storage of glucosylceramide occurred. This in vitro system allows insight into the correlation between enzyme activity and lipid storage as predicted by the theory of residual enzyme activity that was proposed by Conzelmann and Sandhoff.

  7. Nernst-ping-pong model for evaluating the effects of the substrate concentration and anode potential on the kinetic characteristics of bioanode.

    Science.gov (United States)

    Peng, Sikan; Liang, Da-Wei; Diao, Peng; Liu, Yanyan; Lan, Fei; Yang, Yuhan; Lu, Shanfu; Xiang, Yan

    2013-05-01

    Understanding the electron-transfer mechanism and kinetic characteristics of bioanodes is greatly significant to enhance the electron-generating efficiencies in bioelectrochemical systems (BESs). A Nernst-ping-pong model is proposed here to investigate the kinetics and biochemical processes of bioanodes in a microbial electrolysis cell. This model can accurately describe the effects of the substrate (including substrate inhibition) and the anode potential on the current of bioanodes. Results show that the half-wave potential positively shifts as the substrate concentration increases, indicating that the rate-determining steps of anodic processes change from substrate oxidation to intracellular electron transport reaction. The anode potential has negligible effects on the enzymatic catalysis of anodic microbes in the range of -0.25 V to +0.1 V vs. a saturated calomel electrode. It turns out that to reduce the anodic energy loss caused by overpotential, higher substrate concentrations are preferred, if the substrate do not significantly and adversely affect the output current. Copyright © 2013 Elsevier Ltd. All rights reserved.

  8. Date seed characterisation, substrate extraction and process modelling for the production of polyhydroxybutyrate by Cupriavidus necator.

    Science.gov (United States)

    Yousuf, R G; Winterburn, J B

    2016-12-01

    Poly-3-hydroxybutrate (PHB) is a biodegradable polymer synthesised via bacterial fermentation as a means of storing carbon and energy under unbalanced growth conditions. The production cost of petroleum-based plastics is currently lower than that for biopolymers, and the carbon source is the most significant contributor to biopolymer production cost. A feasibility study to assess the suitability of using a date seed derived media as an alternative for PHB production under various stress conditions was investigated. Results include fructose extraction from date seeds and a mass transfer model to describe the process, demonstrating that the high nutrient content of date seeds makes them a promising raw material for microbial growth and that a meaningful amount of PHB can be produced without supplementation. Maximum dry cell weight and PHB concentrations were 6.3g/l and 4.6g/l respectively, giving a PHB content of 73%, when an initial fructose concentration of 10.8g/l was used. Copyright © 2016 Elsevier Ltd. All rights reserved.

  9. Determinants of brain cell metabolic phenotypes and energy substrate utilization unraveled with a modeling approach.

    Directory of Open Access Journals (Sweden)

    Aitana Neves

    Full Text Available Although all brain cells bear in principle a comparable potential in terms of energetics, in reality they exhibit different metabolic profiles. The specific biochemical characteristics explaining such disparities and their relative importance are largely unknown. Using a modeling approach, we show that modifying the kinetic parameters of pyruvate dehydrogenase and mitochondrial NADH shuttling within a realistic interval can yield a striking switch in lactate flux direction. In this context, cells having essentially an oxidative profile exhibit pronounced extracellular lactate uptake and consumption. However, they can be turned into cells with prominent aerobic glycolysis by selectively reducing the aforementioned parameters. In the case of primarily oxidative cells, we also examined the role of glycolysis and lactate transport in providing pyruvate to mitochondria in order to sustain oxidative phosphorylation. The results show that changes in lactate transport capacity and extracellular lactate concentration within the range described experimentally can sustain enhanced oxidative metabolism upon activation. Such a demonstration provides key elements to understand why certain brain cell types constitutively adopt a particular metabolic profile and how specific features can be altered under different physiological and pathological conditions in order to face evolving energy demands.

  10. SUMO4基因多态性与NF-κB、糖尿病及冠心病的关系%Role of SUMO4 gene polymorphism in NF-κB signalling pathway and its research advances in genetic susceptibility to DM and CAD

    Institute of Scientific and Technical Information of China (English)

    蒲连美; 南楠; 杨泽; 金泽宁; 刘铭

    2012-01-01

    Diabetes mellitus ( DM ) and coronary artery disease ( CAD ) are both complex chronic diseases which are caused by interactions of multiple genes with environmental factors.Nuclear factor kappa- B ( NF-κB ) is a family of critical transcription factors modulating the expression of diverse genes involved in the pathogenesis of several inflammatory diseases,including DM and CAD. IκBα is one of the most important inhibitory proteins for NF-κB.SUMO4 conjugates IκBα and stabilizes it from degradation.As a result NF-κB is kept inactive in the cytosol by forming a complex with IκBα and SUMO4.Moreover dissociated NF-κB binds to SUMO4 promoter and activates its transcription.S(o) SUMO4 can function as a negative feedback regulator for the NF-κB signaling pathway.Studies confirmed that SUMO4 M55V substitution results in significant higher levels of NF-κB activity and is associated with susceptibility to DM and CAD.This review focuses on the role of SUMO4 gene polymorphism in NF-κB signalling pathway and its research advances in genetic susceptibility to DM and CAD.%糖尿病( diabetes mellitus,DM)和冠心病(coronary artery disease,CAD)均为多基因介导、基因和环境相互作用的慢性复杂疾病.核因子κB(nuclear factor kappa B,NF- κB)是一类转录因子家族,在细胞内信号传导系统中起关键作用,参与包括DM和CAD在内的多种炎症性疾病的发生发展.研究证实小泛素样修饰蛋白4( small ubiquitin- like modi - fier 4,SUMO4)使NF- κB的抑制剂IκBα免于降解,负性调节NF- κB的转录活性,另外游离的NF- κB能够与SUMO4基因启动子结合并激活SUMO4基因的转录.而SUMO4基因多态性导致NF-κB活性显著增加,并与DM和CAD的易感性相关.此文主要就SUMO4基因多态性对NF-κB转录活性的影响及其在DM和CAD遗传易感性的关联研究进展作一概述.

  11. Expressions of SUMO4, NF-κB and IκB in kidney of rats with type 2diabetes mellitus and its significance%SUMO4、NF-κB、IκB在2型糖尿病大鼠肾脏中的表达及意义

    Institute of Scientific and Technical Information of China (English)

    陈思娇; 王大南; 徐锦春; 李红燕; 刘芙蓉; 宋今丹

    2012-01-01

    Objective:To study expressions of small ubiquitin-related modifier protein (SUMO) 4 (SUMO4),nuclear factor (NF) -κB and inhibitory factor of NF-κB (IκB) in kidneys of rats with type 2 diabetes mellitus (T2DM).Methods:A total of ten 40-week-old male Goto-Kakizaki (GK) rats (with spontaneous diabetes mellitus) of specificpathogen free (SPF) grade,and ten 40-week-old male Wistar rats of SPF grade were selected.The lesion of renal tissue was observed by hematoxylin eosin (HE) staining.Experssions of SUMO4,NF-κB and IκB in renal tissue were observed by immunohistochemistry methods.Results:In the GK rats,glomerular capillary ball hypertrophy,basilar membrane slightly thickening; glomerular mesangial cells hyperplasia,hypertrophy and renal tubular epithelial cells hypertrophy were observed.Compared with normal Wistar rats,expression levels of NF-κB [ (0.232 ±0.034) vs.(0.634± 0.058)],IκB [ (0.242 ± 0.027) vs.(0.712 ± 0.078)] and SUMO4 [ (0.160 ± 0.031) vs.(0.545 ± 0.045)] significantly increased in renal tissue of GK rats (P<0.01 all).Conclusion:Compared with Wistar rats,expressions of NF-κB,IκB and SUMO4 significantly increase in renal tissue of GK rats,suggesting that SUMO inhibiting transcriptional activity of NF-κB may exist in kidneys of T2DM rats.Therefore,sumoylation may be a new therapeutic target for inhibit renal microvascular lesion of diabetic disease.%目的:探讨2型糖尿病大鼠肾脏组织小泛素相关修饰蛋白4 (SUMO4)、核转录因子(NF) -κB、NF-κB的抑制因子(IκB)的表达及意义.方法:取10只40周龄的无特定病原体(SPF)级雄性自发性糖尿病(GK)大鼠,10只40周龄的SPF级雄性Wistar大鼠,通过HE染色法观察肾组织病变、免疫组化法观察肾组织的SUMO4与IκB、SUMO4与NF-κB表达情况.结果:GK大鼠的肾小球毛细血管球肥大,基底膜轻度增厚,肾小球系膜细胞增生、肥大,肾小管上皮细胞肥大.与正常Wistar大鼠比较,GK大鼠的肾脏NF-κB[ (0.232±0

  12. Catalytic Efficiency Is a Function of How Rhodium(I) (5 + 2) Catalysts Accommodate a Conserved Substrate Transition State Geometry: Induced Fit Model for Explaining Transition Metal Catalysis.

    Science.gov (United States)

    Mustard, Thomas J L; Wender, Paul A; Cheong, Paul Ha-Yeon

    2015-03-06

    The origins of differential catalytic reactivities of four Rh(I) catalysts and their derivatives in the (5 + 2) cycloaddition reaction were elucidated using density functional theory. Computed free energy spans are in excellent agreement with known experimental rates. For every catalyst, the substrate geometries in the transition state remained constant (Catalytic efficiency is shown to be a function of how well the catalyst accommodates the substrate transition state geometry and electronics. This shows that the induced fit model for explaining biological catalysis may be relevant to transition metal catalysis. This could serve as a general model for understanding the origins of efficiencies of catalytic reactions.

  13. Modeling the impact, flattening and solidification of a molten droplet on a solid substrate during plasma spraying

    Science.gov (United States)

    Zheng, Y. Z.; Li, Q.; Zheng, Z. H.; Zhu, J. F.; Cao, P. L.

    2014-10-01

    It is quite important to clearly understand the dynamic process of single splat formation for optimizing the plasma spraying process. In present study, a three-dimensional model including heat transfer and phase change was developed on Ansys Fluent 14 platform to simulate the impact, flattening and solidification of a molten droplet on a solid substrate during plasma spraying. The phase, contact pressure, temperature and velocity fields at different spreading times were presented to gain an insight into splat formation mechanism. The predicted splat morphology was in good agreement with the experimental photos. The effect of mushy zone constant, a parameter dominating the solidification behavior of fluid in Fluent, on the flattening of droplet was further investigated. Through comparing the calculated spread factor from present model with the experimental value, a mushy zone constant of 108 or 109 was found to be more appropriate for simulation on the solidification problem occurring in high-speed impact and flattening process, instead of the range of 104-107 recommended in Fluent.

  14. Mathematical modeling of poly[(R)-3-hydroxyalkanoate] synthesis by Cupriavidus necator DSM 545 on substrates stemming from biodiesel production.

    Science.gov (United States)

    Špoljarić, Ivna Vrana; Lopar, Markan; Koller, Martin; Muhr, Alexander; Salerno, Anna; Reiterer, Angelika; Malli, Karin; Angerer, Hannes; Strohmeier, Katharina; Schober, Sigurd; Mittelbach, Martin; Horvat, Predrag

    2013-04-01

    Two low structured mathematical models for fed-batch production of polyhydroxybutyrate and poly[hydroxybutyrate-co-hydroxyvalerate] by Cupriavidus necator DSM 545 on renewable substrates (glycerol and fatty acid methyl esters-FAME) combined with glucose and valeric acid, were established. The models were used for development/optimization of feeding strategies of carbon and nitrogen sources concerning PHA content and polymer/copolymer composition. Glycerol/glucose fermentation featured a max. specific growth rate of 0.171 h(-1), a max. specific production rate of 0.038 h(-1) and a PHB content of 64.5%, whereas the FAME/valeric acid fermentation resulted in a max. specific growth rate of 0.046 h(-1), a max. specific production rate of 0.07 h(-1) and 63.6% PHBV content with 4.3% of 3-hydroxyvalerate (3HV) in PHBV. A strong inhibition of glycerol consumption by glucose was confirmed (inhibition constant ki,G=4.28×10(-4) g L(-1)). Applied concentration of FAME (10-12 g L(-1)) positively influenced on PHBV synthesis. HV/PHBV ratio depends on applied VA concentration. Copyright © 2013 Elsevier Ltd. All rights reserved.

  15. KAP1 Is a Host Restriction Factor That Promotes Human Adenovirus E1B-55K SUMO Modification

    DEFF Research Database (Denmark)

    Bürck, Carolin; Mund, Andreas; Berscheminski, Julia

    2016-01-01

    characterized, but represent a decisive moment in establishing a productive infection. Here, we identify a novel host viral restriction factor, KAP1. This heterochromatin associated transcription factor regulates the dynamic organization of host chromatin structure via its ability to influence epigenetic marks...... epigenetic gene silencing and to promote SUMO modification of E1B-55K by a so far unknown mechanism. IMPORTANCE: Here we describe a novel cellular restriction factor for Human Adenovirus (HAdV) that sheds light on very early modulation processes in viral infection. We reported that chromatin formation...... and cellular SWI/SNF chromatin remodeling play a key role in HAdV transcriptional regulation (1-4). We observed that the cellular chromatin-associated factor, and epigenetic reader SPOC1 represses HAdV infection and gene expression. Here, we illustrate the role of the SPOC1 interacting factor KAP1 during...

  16. Incorporating an enzymatic model of effects of temperature, moisture, and substrate supply on soil respiration into an ecosystem model for two forests of northeastern USA

    Science.gov (United States)

    Sihi, Debjani; Davidson, Eric; Chen, Min; Savage, Kathleen; Richardson, Andrew; Keenan, Trevor; Hollinger, David

    2017-04-01

    Soils represent the largest terrestrial carbon (C) pool, and microbial decomposition of soil organic matter (SOM) to carbon dioxide, also called heterotrophic respiration (Rh), is an important component of the global C cycle. Temperature sensitivity of Rh is often represented with a simple Q10 function in ecosystem models and earth system models (ESMs), sometimes accompanied by an empirical soil moisture modifier. More explicit representation of the effects of soil moisture, substrate supply, and their interactions with temperature has been proposed to disentangle the confounding factors of apparent temperature sensitivity of SOM decomposition and improve performance of ecosystem models and ESMs. The objective of this work was to incorporate into an ecosystem model a more mechanistic, but still parsimonious, model of environmental factors controlling Rh. The Dual Arrhenius and Michaelis-Menten (DAMM) model simulates Rh using Michaelis-Menten, Arrhenius, and diffusion functions. Soil moisture affects Rh and its apparent temperature sensitivity in DAMM by regulating the diffusion of oxygen and soluble carbon substrates to the enzymatic reaction site. However, in its current configuration, DAMM depends on assumptions or inputs from other models regarding soil C inputs. Here we merged the DAMM soil flux model with a parsimonious ecosystem flux model, FöBAAR (Forest Biomass, Assimilation, Allocation and Respiration) by replacing FöBAAR's algorithms for Rh with those of DAMM. Classical root trenching experiments provided data to partition soil CO2 efflux into Rh (trenched plot) and root respiration (untrenched minus trenched plots). We used three years of high-frequency soil flux data from automated soil chambers (trenched and untrenched plots) and landscape-scale ecosystem fluxes from eddy covariance towers from two mid-latitude forests (Harvard Forest, MA and Howland Forest, ME) of northeastern USA to develop and validate the merged model and to quantify the

  17. The SUMO protease SENP1 is required for cohesion maintenance and mitotic arrest following spindle poison treatment

    Energy Technology Data Exchange (ETDEWEB)

    Era, Saho [Fondazione IFOM, Istituto FIRC di Oncologia Molecolare, IFOM-IEO campus, Via Adamello 16, 20139 Milan (Italy); Radiation Genetics, Graduate School of Medicine, Kyoto University, Yoshida-Konoe, Sakyo-ku, Kyoto 606-8501 (Japan); Abe, Takuya; Arakawa, Hiroshi [Fondazione IFOM, Istituto FIRC di Oncologia Molecolare, IFOM-IEO campus, Via Adamello 16, 20139 Milan (Italy); Kobayashi, Shunsuke [Radiation Genetics, Graduate School of Medicine, Kyoto University, Yoshida-Konoe, Sakyo-ku, Kyoto 606-8501 (Japan); Szakal, Barnabas [Fondazione IFOM, Istituto FIRC di Oncologia Molecolare, IFOM-IEO campus, Via Adamello 16, 20139 Milan (Italy); Yoshikawa, Yusuke; Motegi, Akira; Takeda, Shunichi [Radiation Genetics, Graduate School of Medicine, Kyoto University, Yoshida-Konoe, Sakyo-ku, Kyoto 606-8501 (Japan); Branzei, Dana, E-mail: dana.branzei@ifom.eu [Fondazione IFOM, Istituto FIRC di Oncologia Molecolare, IFOM-IEO campus, Via Adamello 16, 20139 Milan (Italy)

    2012-09-28

    Highlights: Black-Right-Pointing-Pointer SENP1 knockout chicken DT40 cells are hypersensitive to spindle poisons. Black-Right-Pointing-Pointer Spindle poison treatment of SENP1{sup -/-} cells leads to increased mitotic slippage. Black-Right-Pointing-Pointer Mitotic slippage in SENP1{sup -/-} cells associates with apoptosis and endoreplication. Black-Right-Pointing-Pointer SENP1 counteracts sister chromatid separation during mitotic arrest. Black-Right-Pointing-Pointer Plk1-mediated cohesion down-regulation is involved in colcemid cytotoxicity. -- Abstract: SUMO conjugation is a reversible posttranslational modification that regulates protein function. SENP1 is one of the six SUMO-specific proteases present in vertebrate cells and its altered expression is observed in several carcinomas. To characterize SENP1 role in genome integrity, we generated Senp1 knockout chicken DT40 cells. SENP1{sup -/-} cells show normal proliferation, but are sensitive to spindle poisons. This hypersensitivity correlates with increased sister chromatid separation, mitotic slippage, and apoptosis. To test whether the cohesion defect had a causal relationship with the observed mitotic events, we restored the cohesive status of sister chromatids by introducing the TOP2{alpha}{sup +/-} mutation, which leads to increased catenation, or by inhibiting Plk1 and Aurora B kinases that promote cohesin release from chromosomes during prolonged mitotic arrest. Although TOP2{alpha} is SUMOylated during mitosis, the TOP2{alpha}{sup +/-} mutation had no obvious effect. By contrast, inhibition of Plk1 or Aurora B rescued the hypersensitivity of SENP1{sup -/-} cells to colcemid. In conclusion, we identify SENP1 as a novel factor required for mitotic arrest and cohesion maintenance during prolonged mitotic arrest induced by spindle poisons.

  18. Construction of fusion expression vector pET22b-SUMO-FGFR4 and optimization of expression conditions in E.coli%融合表达载体 pET22b-SUMO-FGFR4的构建及其在大肠杆菌中表达条件的优化

    Institute of Scientific and Technical Information of China (English)

    刘微; 姚杨; 马萧萧; 邓裕宣; 梅迪; 刘磊; 王会岩

    2016-01-01

    目的:设计合成小泛素修饰物-成纤维细胞生长因子受体4(SUMO-FGFR4)基因,构建 pET22b-SUMO-FGFR4表达载体,并对其表达条件进行优化。方法:采用 Overlap PCR 方法制备 SUMO-FGFR4融合基因,并连接到原核表达载体 pET22b 中,获得 pET22b-SUMO-FGFR4重组表达载体。以乳糖为诱导剂,观察乳糖浓度、诱导时机、诱导温度、诱导时间和乳糖的添加方式等因素对 SUMO-FGFR4蛋白表达量的影响,确定最佳诱导条件,并进行重组蛋白的可溶性分析。结果:pET22b-SUMO-FGFR4表达的融合蛋白在相对分子质量40000处显示目标条带,并与 FGFR4抗体特异性结合。融合蛋白在乳糖终浓度为1.0 g·L-1、诱导时间为3 h、诱导时机 A (600)值为0.8、诱导温度为37℃时表达量最高,乳糖的添加方式对 SUMO-FGFR4融合蛋白的表达量无明显影响。乳糖作为诱导剂比传统诱导剂 IPTG 诱导 SUMO-FGFR4融合蛋白的表达量高7.5%,融合蛋白以包涵体形式为主。结论:以乳糖作为诱导剂,成功表达了 SUMO-FGFR4融合蛋白,确定了融合蛋白的最佳表达条件。%Objective:To design the small ubiquitin modification-fibroblast growth factor receptor 4 (SUMO-FGFR4) fusion gene and construct the expression vector pET22b-SUMO-FGFR4, to optimize the expression conditions. Methods:The SUMO-FGFR4 fusion gene was obtained by Overlap PCR and was connected to pET22b;the recombinant expression vector pET22b-SUMO-FGFR4 was obtained. The influence of lactose concentration, induction time,induction temperature,induction point and adding mode of lactose in the expression levels was observed,and the best induction condition was determined; then the solubility of recombinant protein was analyzed.Results:The SUMO-FGFR4 fusion protein was highly expressed,the molecular weight of the fusion protein was about 40 000 and it could bind with FGFR4 specific antibody.When the lactose concentration

  19. A model framework to describe growth-linked biodegradation of trace-level pollutants in the presence of coincidental carbon substrates and microbes.

    Science.gov (United States)

    Liu, Li; Helbling, Damian E; Kohler, Hans-Peter E; Smets, Barth F

    2014-11-18

    Pollutants such as pesticides and their degradation products occur ubiquitously in natural aquatic environments at trace concentrations (μg L(-1) and lower). Microbial biodegradation processes have long been known to contribute to the attenuation of pesticides in contaminated environments. However, challenges remain in developing engineered remediation strategies for pesticide-contaminated environments because the fundamental processes that regulate growth-linked biodegradation of pesticides in natural environments remain poorly understood. In this research, we developed a model framework to describe growth-linked biodegradation of pesticides at trace concentrations. We used experimental data reported in the literature or novel simulations to explore three fundamental kinetic processes in isolation. We then combine these kinetic processes into a unified model framework. The three kinetic processes described were: the growth-linked biodegradation of micropollutant at environmentally relevant concentrations; the effect of coincidental assimilable organic carbon substrates; and the effect of coincidental microbes that compete for assimilable organic carbon substrates. We used Monod kinetic models to describe substrate utilization and microbial growth rates for specific pesticide and degrader pairs. We then extended the model to include terms for utilization of assimilable organic carbon substrates by the specific degrader and coincidental microbes, growth on assimilable organic carbon substrates by the specific degrader and coincidental microbes, and endogenous metabolism. The proposed model framework enables interpretation and description of a range of experimental observations on micropollutant biodegradation. The model provides a useful tool to identify environmental conditions with respect to the occurrence of assimilable organic carbon and coincidental microbes that may result in enhanced or reduced micropollutant biodegradation.

  20. Structural characterization of mouse neutrophil serine proteases and identification of their substrate specificities: relevance to mouse models of human inflammatory diseases.

    Science.gov (United States)

    Kalupov, Timofey; Brillard-Bourdet, Michèle; Dadé, Sébastien; Serrano, Hélène; Wartelle, Julien; Guyot, Nicolas; Juliano, Luiz; Moreau, Thierry; Belaaouaj, Azzaq; Gauthier, Francis

    2009-12-01

    It is widely accepted that neutrophil serine proteases (NSPs) play a critical role in neutrophil-associated lung inflammatory and tissue-destructive diseases. To investigate NSP pathogenic role(s), various mouse experimental models have been developed that mimic acutely or chronically injured human lungs. We and others are using mouse exposure to cigarette smoke as a model for chronic obstructive pulmonary disease with or without exacerbation. However, the relative contribution of NSPs to lung disease processes as well as their underlying mechanisms remains still poorly understood. And the lack of purified mouse NSPs and their specific substrates have hampered advances in these studies. In this work, we compared mouse and human NSPs and generated three-dimensional models of murine NSPs based on three-dimensional structures of their human homologs. Analyses of these models provided compelling evidence that peptide substrate specificities of human and mouse NSPs are different despite their conserved cleft and close structural resemblance. These studies allowed us to synthesize for the first time novel sensitive fluorescence resonance energy transfer substrates for individual mouse NSPs. Our findings and the newly identified substrates should better our understanding about the role of NSPs in the pathogenesis of cigarette-associated chronic obstructive pulmonary disease as well as other neutrophils-associated inflammatory diseases.

  1. Small Ubiquitin-Like Modifier Conjugating Enzyme with Active Site Mutation Acts as Dominant Negative Inhibitor of SUMO Conjugation in Arabidopsis

    Institute of Scientific and Technical Information of China (English)

    Konstantin Tomanov; Christian Hardtke; Ruchika Budhiraja; Rebecca Hermkes; George Coupland; Andreas Bachmair

    2013-01-01

    Small ubiquitin-like modifier (SUMO) conjugation affects a broad range of processes in plants,including growth,flower initiation,pathogen defense,and responses to abiotic stress.Here,we investigate in vivo and in vitro a SUMO conjugating enzyme with a Cys to Ser change in the active site,and show that it has a dominant negative effect.In planta expression significantly perturbs normal development,leading to growth retardation,early flowering and gene expression changes.We suggest that the mutant protein can serve as a probe to investigate sumoylation,also in plants for which poor genetic infrastructure precludes analysis via loss-of-function mutants.

  2. High-level SUMO-mediated fusion expression of ABP-dHC-cecropin A from multiple joined genes in Escherichia coli.

    Science.gov (United States)

    Zhang, Jiaxin; Movahedi, Ali; Wei, Zhiheng; Sang, Ming; Wu, Xiaolong; Wang, Mengyang; Wei, Hui; Pan, Huixin; Yin, Tongming; Zhuge, Qiang

    2016-09-15

    The antimicrobial peptide ABP-dHC-cecropin A is a small cationic peptide with potent activity against a wide range of bacterial species. Evidence of antifungal activity has also been suggested; however, evaluation of this peptide has been limited due to the low expression of cecropin proteins in Escherichia coli. To improve the expression level of ABP-dHC-cecropin A in E. coli, tandem repeats of the ABP-dHC-cecropin A gene were constructed and expressed as fusion proteins (SUMO-nABP-dHC-cecropin, n = 1, 2, 3, 4) via pSUMO-nABP-dHC-cecropin A vectors (n = 1, 2, 3, 4). Comparison of the expression levels of soluble SUMO-nABP-dHC-cecropin A fusion proteins (n = 1, 2, 3, 4) suggested that BL21 (DE3)/pSUMO-3ABP-dHC-cecropin A is an ideal recombinant strain for ABP-dHC-cecropin A production. Under the selected conditions of cultivation and isopropylthiogalactoside (IPTG) induction, the expression level of ABP-dHC-cecropin A was as high as 65 mg/L, with ∼21.3% of the fusion protein in soluble form. By large-scale fermentation, protein production reached nearly 300 mg/L, which is the highest yield of ABP-dHC-cecropin A reported to date. In antibacterial experiments, the efficacy was approximately the same as that of synthetic ABP-dHC-cecropin A. This method provides a novel and effective means of producing large amounts of ABP-dHC-cecropin A.

  3. Recombinant Scorpine Produced Using SUMO Fusion Partner in Escherichia coli Has the Activities against Clinically Isolated Bacteria and Inhibits the Plasmodium falciparum Parasitemia In Vitro

    OpenAIRE

    Chao Zhang; Xinlong He; Yaping Gu; Huayun Zhou; Jun Cao; Qi Gao

    2014-01-01

    Scorpine, a small cationic peptide from the venom of Pandinus imperator, which has been shown to have anti-bacterial and anti-plasmodial activities, has potential important applications in the pharmaceutical industries. However, the isolation of scorpine from natural sources is inefficient and time-consuming. Here, we first report the expression and purification of recombinant scorpine in Escherichia coli, using small ubiquitin-related modifier (SUMO) fusion partner. The fusion protein was ex...

  4. The nucleolar SUMO-specific protease SMT3IP1/SENP3 attenuates Mdm2-mediated p53 ubiquitination and degradation

    Energy Technology Data Exchange (ETDEWEB)

    Nishida, Tamotsu, E-mail: nishida@gene.mie-u.ac.jp [Department of Human Functional Genomics, Life Science Research Center, Mie University, 1577 Kurima-machiya, Tsu 514-8507 (Japan); Yamada, Yoshiji [Department of Human Functional Genomics, Life Science Research Center, Mie University, 1577 Kurima-machiya, Tsu 514-8507 (Japan)

    2011-03-11

    Research highlights: {yields} SMT3IP1 interacts with p53 and Mdm2, and desumoylates both proteins. {yields} SMT3IP1 competes with p53 for binding to the central acidic domain of Mdm2. {yields} SMT3IP1 binding to Mdm2 inhibits Mdm2-mediated p53 ubiquitination and degradation. {yields} We postulate that SMT3IP1 acts as a new regulator of the p53-Mdm2 pathway. -- Abstract: SUMO (small ubiquitin-like modifier) modification plays multiple roles in several cellular processes. Sumoylation is reversibly regulated by SUMO-specific proteases. SUMO-specific proteases have recently been implicated in cell proliferation and early embryogenesis, but the underlying mechanisms remain unknown. Here, we show that a nucleolar SUMO-specific protease, SMT3IP1/SENP3, controls the p53-Mdm2 pathway. We found that SMT3IP1 interacts with p53 and Mdm2, and desumoylates both proteins. Overexpression of SMT3IP1 in cells resulted in the accumulation of Mdm2 in the nucleolus and increased stability of the p53 protein. In addition, SMT3IP1 bound to the acidic domain of Mdm2, which also mediates the p53 interaction, and competed with p53 for binding. Increasing expression of SMT3IP1 suppressed Mdm2-mediated p53 ubiquitination and subsequent proteasomal degradation. Interestingly, the desumoylation activity of SMT3IP1 was not necessary for p53 stabilization. These results suggest that SMT3IP1 is a new regulator of the p53-Mdm2 pathway.

  5. Role of the SUMO-interacting motif in HIPK2 targeting to the PML nuclear bodies and regulation of p53

    Energy Technology Data Exchange (ETDEWEB)

    Sung, Ki Sa; Lee, Yun-Ah [Department of Biological Sciences, Sungkyunkwan University, Suwon 440-746 (Korea, Republic of); Kim, Eui Tae [Department of Molecular Cell Biology, Samsung Biomedical Research Institute, Sungkyunkwan University School of Medicine, Suwon 440-746 (Korea, Republic of); Lee, Seung-Rock [Department of Biochemistry, Research Center for Aging and Geriatrics, Research Institute of Medical Sciences, Chonnam National University Medical School, Gwangju 501-190 (Korea, Republic of); Ahn, Jin-Hyun [Department of Molecular Cell Biology, Samsung Biomedical Research Institute, Sungkyunkwan University School of Medicine, Suwon 440-746 (Korea, Republic of); Choi, Cheol Yong, E-mail: choicy@skku.ac.kr [Department of Biological Sciences, Sungkyunkwan University, Suwon 440-746 (Korea, Republic of)

    2011-04-15

    Homeodomain-interacting protein kinase 2 (HIPK2) is a key regulator of various transcription factors including p53 and CtBP in the DNA damage signaling pathway. PML-nuclear body (NB) is required for HIPK2-mediated p53 phosphorylation at Ser46 and induction of apoptosis. Although PML-NB targeting of HIPK2 has been shown, much is not clear about the molecular mechanism of HIPK2 recruitment to PML-NBs. Here we show that HIPK2 colocalizes specifically with PML-I and PML-IV. Mutational analysis showed that HIPK2 recruitment to PML-IV-NBs is mediated by the SUMO-interaction motifs (SIMs) of both PML-IV and HIPK2. Wild-type HIPK2 associated with SUMO-conjugated PML-IV at a higher affinity than with un-conjugated PML-IV, while the association of a HIPK2 SIM mutant with SUMO-modified PML-IV was impaired. In colony formation assays, HIPK2 strongly suppressed cell proliferation, but HIPK2 SIM mutants did not. In addition, activation and phosphorylation of p53 at the Ser46 residue were impaired by HIPK2 SIM mutants. These results suggest that SIM-mediated HIPK2 targeting to PML-NBs is crucial for HIPK2-mediated p53 activation and induction of apoptosis.

  6. Recombinant scorpine produced using SUMO fusion partner in Escherichia coli has the activities against clinically isolated bacteria and inhibits the Plasmodium falciparum parasitemia in vitro.

    Directory of Open Access Journals (Sweden)

    Chao Zhang

    Full Text Available Scorpine, a small cationic peptide from the venom of Pandinus imperator, which has been shown to have anti-bacterial and anti-plasmodial activities, has potential important applications in the pharmaceutical industries. However, the isolation of scorpine from natural sources is inefficient and time-consuming. Here, we first report the expression and purification of recombinant scorpine in Escherichia coli, using small ubiquitin-related modifier (SUMO fusion partner. The fusion protein was expressed in soluble form in E. coli, and expression was verified by SDS-PAGE and western blotting analysis. The fusion protein was purified to 90% purity by nickel-nitrilotriacetic acid (Ni2+-NTA resin chromatography. After the SUMO-scorpine fusion protein was cleaved by the SUMO protease, the cleaved sample was reapplied to a Ni2+-NTA column. Tricine/SDS-PAGE gel results indicated that Scorpine had been purified successfully to more than 95% purity. The recombinantly expressed Scorpine showed anti-bacterial activity against two standard bacteria including Staphylococcus aureus ATCC 29213 and Acinetobacter baumannii ATCC 19606, and clinically isolated bacteria including S. aureus S, S. aureus R, A. baumannii S, and A. baumannii R. It also produced 100% reduction in Plasmodium falciparum parasitemia in vitro. Thus, the expression strategy presented in this study allowed convenient high yield and easy purification of recombinant Scorpine for pharmaceutical applications in the future.

  7. Recombinant scorpine produced using SUMO fusion partner in Escherichia coli has the activities against clinically isolated bacteria and inhibits the Plasmodium falciparum parasitemia in vitro.

    Science.gov (United States)

    Zhang, Chao; He, Xinlong; Gu, Yaping; Zhou, Huayun; Cao, Jun; Gao, Qi

    2014-01-01

    Scorpine, a small cationic peptide from the venom of Pandinus imperator, which has been shown to have anti-bacterial and anti-plasmodial activities, has potential important applications in the pharmaceutical industries. However, the isolation of scorpine from natural sources is inefficient and time-consuming. Here, we first report the expression and purification of recombinant scorpine in Escherichia coli, using small ubiquitin-related modifier (SUMO) fusion partner. The fusion protein was expressed in soluble form in E. coli, and expression was verified by SDS-PAGE and western blotting analysis. The fusion protein was purified to 90% purity by nickel-nitrilotriacetic acid (Ni2+-NTA) resin chromatography. After the SUMO-scorpine fusion protein was cleaved by the SUMO protease, the cleaved sample was reapplied to a Ni2+-NTA column. Tricine/SDS-PAGE gel results indicated that Scorpine had been purified successfully to more than 95% purity. The recombinantly expressed Scorpine showed anti-bacterial activity against two standard bacteria including Staphylococcus aureus ATCC 29213 and Acinetobacter baumannii ATCC 19606, and clinically isolated bacteria including S. aureus S, S. aureus R, A. baumannii S, and A. baumannii R. It also produced 100% reduction in Plasmodium falciparum parasitemia in vitro. Thus, the expression strategy presented in this study allowed convenient high yield and easy purification of recombinant Scorpine for pharmaceutical applications in the future.

  8. Studi Kinerja VANET Scenario Generators: SUMO dan VanetMobisim untuk Implementasi Routing Protocol AODV menggunakan Network Simulator 2 (NS-2

    Directory of Open Access Journals (Sweden)

    Firdaus Nutrihadi

    2016-04-01

    Full Text Available Vehicular Ad Hoc Network (VANET merupakan turunan dari MANET (Mobile Ad Hoc Network sebagai inovasi baru dalam dunia teknologi yang membantu kebutuhan manusia dalam berkomunikasi. VANET dapat mendukung komunikasi langsung antara kendaraan (Vehicle to Vehicle dan antara kendaraan-infrastruktur (Vehicle to Infrastructure dengan adanya infrastruktur jaringan nirkabel. Namun, implementasi VANET di dunia masih sulit dilakukan sehingga banyak penelitian dilakukan dengan membuat simulasi menggunakan mobility generator dan network simulator. Pada makalah ini yang diteliti yaitu performa skema VANET yang dihasilkan oleh mobility generator SUMO dan Vanetmobisim. Penelitian ini menggunakan NS-2 sebagai simulator VANET dengan protokol reaktif AODV sebagai routing protocol. Skenario VANET dengan peta berbentuk grid dan peta riil Sutomo,Surabaya digunakan pada kedua generator SUMO dan VanetMobisim dengan memvariasikan jumlah kendaraan simulasi. Matriks evaluasi kinerja yang digunakan dalam penelitian ini adalah packet delivery ratio, end-to-end delay, dan routing overhead. Dalam ketiga skenario, performa routing protokol SUMO-AODV lebih baik. VanetMobisim-AODV, dikarenakan lebih banyak lalu lintas dan rute yang putus, menghasilkan performa yang baik namun masih di bawah pesaingnya.

  9. Increased oxidative stress and apoptosis in the hypothalamus of diabetic male mice in the insulin receptor substrate-2 knockout model

    Science.gov (United States)

    Canelles, Sandra; Argente, Jesús; Barrios, Vicente

    2016-01-01

    ABSTRACT Insulin receptor substrate-2-deficient (IRS2−/−) mice are considered a good model to study the development of diabetes because IRS proteins mediate the pleiotropic effects of insulin-like growth factor-I (IGF-I) and insulin on metabolism, mitogenesis and cell survival. The hypothalamus might play a key role in the early onset of diabetes, owing to its involvement in the control of glucose homeostasis and energy balance. Because some inflammatory markers are elevated in the hypothalamus of diabetic IRS2−/− mice, our aim was to analyze whether the diabetes associated with the absence of IRS2 results in hypothalamic injury and to analyze the intracellular mechanisms involved. Only diabetic IRS2−/− mice showed increased cell death and activation of caspase-8 and -3 in the hypothalamus. Regulators of apoptosis such as FADD, Bcl-2, Bcl-xL and p53 were also increased, whereas p-IκB and c-FLIPL were decreased. This was accompanied by increased levels of Nox-4 and catalase, enzymes involved in oxidative stress. In summary, the hypothalamus of diabetic IRS2−/− mice showed an increase in oxidative stress and inflammatory markers that finally resulted in cell death via substantial activation of the extrinsic apoptotic pathway. Conversely, non-diabetic IRS2−/− mice did not show cell death in the hypothalamus, possibly owing to an increase in the levels of circulating IGF-I and in the enhanced hypothalamic IGF-IR phosphorylation that would lead to the stimulation of survival pathways. In conclusion, diabetes in IRS2-deficient male mice is associated with increased oxidative stress and apoptosis in the hypothalamus. PMID:27013528

  10. Modeling of the effects of different substrate materials on the residual thermal stresses in the aluminum nitride crystal grown by sublimation

    Science.gov (United States)

    Lee, R. G.; Idesman, A.; Nyakiti, L.; Chaudhuri, J.

    2009-02-01

    A three-dimensional numerical finite element modeling method is applied to compare interfacial residual thermal stress distribution in AlN single crystals grown by using different substrates such as silicon carbide, boron nitride, tungsten, tantalum carbide, and niobium carbide. A dimensionless coordinate system is used which reduces the numbers of computations and hence simplifies the stress analysis. All components of the stress distribution, both in the film and in the substrate, including the normal stress along the growth direction as well as in-plane normal stresses and shear stresses are fully investigated. This information about the stress distribution provides insight into understanding and controlling the AlN single crystal growth by the sublimation technique. The normal stress in the film at the interface along the growth direction and the shear stresses are zero except at the edges, whereas in-plane stresses are nonzero. The in-plane stresses are compressive when TaC and NbC substrates are used. A small compressive stress might be beneficial in prohibiting crack growth in the film. The compressive stress in the AlN is lower for the TaC substrate than that for the NbC. Tensile in-plane stresses are formed in the AlN for 6H-SiC, BN, and W substrates. This tensile stress in the film is detrimental as it will assist in the crack growth. The stress concentration at the edges of the AlN film at the interface is compressive in nature when TaC and NbC are used as a substrate. This causes the film to bend downward (i.e., convex shape) and assist it to adhere to the substrate. The AlN film curves upward or in a concave shape when SiC, BN, and W substrates are used since the stress concentration at the edges of the AlN film is tensile at the interface and this may cause detachment of the film from the substrate.

  11. Homology modeling of lanosterol 14alpha-demethylase of Candida albicans and Aspergillus fumigatus and insights into the enzyme-substrate Interactions

    Institute of Scientific and Technical Information of China (English)

    Sheng C; Zhou Y; Zhu J; Lu J; Zhang W; Zhang M; Song Y; Ji H; Zhu J; Yao J; Yu J; Yang S

    2005-01-01

    The crystal structure of 14alpha-sterol demethylase from Mycobacterium tuberculosis (MT_14DM) provides a good template for modeling the three dimensional structure of lanosterol 14alpha-demethylase, which is the target of azole antifungal agents. Homologous 3D models of lanosterol 14alpha-demethylase from Candida albicans (CA_14DM) and Aspergillus fumigatus (AF_14DM) were built on the basis of the crystal coordinates of MT_14DM in complex with 4-phenylimidazole and fluconazole. The reliability of the two models was assessed by Ramachandran plots, Profile-3D analysis, and by analyzing the consistency of the two models with the experimental data on the P450(14DM). The overall structures of the resulting CA_14DM model and AF_14DM model are similar to those of the template structures. The two models remain the core structure characteristic for cytochrome P450s and most of the insertions and deletions expose the molecular surface. The structurally and functionally important residues such as the heme binding residues, the residues lining the substrate access channel, and residues in active site were identified from the model. To explore the binding mode of the substrate with the two models, 24(28)-methylene-24,25-dihydrolanosterol was docked into the active site of the two models and hydrophobic interaction and hydrogen-bonding were found to play an important role in substrate recognition and orientation. These results provided a basis for experiments to probe structure-function relationships in the P450(14DM). Although CA_14DM and AF_14DM shared similar core structural character, the active site of the two models were quite different, thus allowing the rational design of specific inhibitors to the target enzyme and the discovery of novel antifungal agents with broad spectrum.

  12. SUMO 特异性蛋白酶1在心肌组织缺血/再灌注损伤中的保护作用%Protective Effect of SUMO-Specific Protease 1 on Ischemia/Reperfusion Injury of Myocardial Tissue

    Institute of Scientific and Technical Information of China (English)

    顾剑民; 薛松

    2014-01-01

    目的:探讨SUMO特异性蛋白酶1( SENP1)在心肌组织缺血/再灌注损伤中的保护作用。方法选取14例行体外循环心脏手术前、后的右心房组织,建立缺血再灌注小鼠模型,通过实时定量PCR检测患者和大鼠心肌组织SENP1的mRNA变化。对H9 C2细胞株进行缺氧复氧处理后检测 LDH释放率,观察细胞死亡情况。结果实时定量PCR结果显示,14例患者缺血再灌注后心肌组织内SENP1 mRNA含量为(4.845±1.248),较灌注前明显升高(P<0.01)。单纯缺血处理的小鼠心肌SENP1 mRNA含量较正常组小鼠略高,且随着灌注时间的延长,SENP1 mRNA含量呈逐渐递增的趋势。特异性siRNA敲除大鼠心肌细胞H9C2细胞内SENP148 h后SENP1 mRNA水平降至对照组的30%以下,敲除SENP1的H9C2细胞在缺氧及复氧处理后LDH释放增加。结论 SENP1在心肌缺血再灌注中有保护作用,缺乏SENP1可能会加重心肌损伤。%Objective To investigate the protective effect of SUMO -specific protease 1 ( SUMO-SENP1 ) on ischemia/reperfusion injury of myocardial tissue .Methods Fourteen cases of right atrial tissue before and after cardiopulmonary bypass heart operation were selected , the mouse model of ischemia reperfusion was established , and real-time quantitative polymerase chain reaction ( PCR) was used to detect the changes of SENP1 mRNA in myocardial tissue of patients and rats.The LDH release rate was detected after hypoxia reoxygenation on H 9C2 cell line,and the cell death was observed .Results The results of real-time PCR showed that the SENP1 mRNA content in myocardial tissue of14 cases after reperfusion was(4.845 ±1.248),which was significantly higher than that before reperfusion (P<0.01).The SENP1 mRNA content in myocardium of ischemia treatment rats was slightly higher than that of normal rats , and increased with the increasing reperfusion time .The mRNA level of SENP1 in H9C2 cells of specific si

  13. Three dimensional material removal model of laser-induced backside wet etching of sapphire substrate with CuSO4 solutions

    Science.gov (United States)

    Xie, Xiaozhu; Huang, Xiandong; Jiang, Wei; Wei, Xin; Hu, Wei; Ren, Qinglei

    2017-03-01

    The mechanism of laser-induced backside wet etching (LIBWE) of sapphire substrate with CuSO4 solution is considered as a two-step process. First, it deposits the layer from copper sulfate solution on the backside of sapphire substrate by 1064 nm laser irradiation. Then it is followed by the absorption of deposited layer to laser irradiation, resulting in the etching of the sapphire. Therefore, the material removal of LIBWE is based on laser interaction with multilayer materials (sapphire substrate-deposition layer-liquid solution). A three-dimensional thermal model is established to simulate the material removal during the LIBWE process by considering the material data variations of temperature, enthalpy change and latent heat fusion. The model can predict the groove shape influenced by the laser processing parameters (laser fluence, scanning velocity and scanning pass). The simulation results indicate that the groove depth increases with the decreasing of scanning velocity, the increasing of laser fluence and the scanning pass. The groove width is comparable with the focal beam diameter. Some peaks and valleys occur at the bottom of the groove. A comparison between the modeling and experiment indicates that the groove shape in simulation agrees well with the experiment data at laser pulse energy of 4.3 mJ/pulse, scanning velocity of 15 mm/s and the scanning pass of 4. i.e, the present physical model is effective and feasible.

  14. The Crystal Structure Analysis of Group B Streptococcus Sortase C1: A Model for the ;Lid; Movement upon Substrate Binding

    Energy Technology Data Exchange (ETDEWEB)

    Khare, Baldeep; Fu, Zheng-Qing; Huang, I-Hsiu; Ton-That, Hung; Narayana, Sthanam V.L. (UAB); (Georgia); (UTSMC)

    2012-02-07

    A unique feature of the class-C-type sortases, enzymes essential for Gram-positive pilus biogenesis, is the presence of a flexible 'lid' anchored in the active site. However, the mechanistic details of the 'lid' displacement, suggested to be a critical prelude for enzyme catalysis, are not yet known. This is partly due to the absence of enzyme-substrate and enzyme-inhibitor complex crystal structures. We have recently described the crystal structures of the Streptococcus agalactiae SAG2603 V/R sortase SrtC1 in two space groups (type II and type III) and that of its 'lid' mutant and proposed a role of the 'lid' as a protector of the active-site hydrophobic environment. Here, we report the crystal structures of SAG2603 V/R sortase C1 in a different space group (type I) and that of its complex with a small-molecule cysteine protease inhibitor. We observe that the catalytic Cys residue is covalently linked to the small-molecule inhibitor without lid displacement. However, the type I structure provides a view of the sortase SrtC1 lid displacement while having structural elements similar to a substrate sorting motif suitably positioned in the active site. We propose that these major conformational changes seen in the presence of a substrate mimic in the active site may represent universal features of class C sortase substrate recognition and enzyme activation.

  15. Discrimination among eight modified michaelis-menten kinetics models of cellulose hydrolysis with a large range of substrate/enzyme ratios: inhibition by cellobiose.

    Science.gov (United States)

    Bezerra, Rui M F; Dias, Albino A

    2004-03-01

    The kinetics of exoglucanase (Cel7A) from Trichoderma reesei was investigated in the presence of cellobiose and 24 different enzyme/Avicel ratios for 47 h, in order to establish which of the eight available kinetic models best explained the factors involved. The heterogeneous catalysis was studied and the kinetic parameters were estimated employing integrated forms of Michaelis-Menten equations through the use of nonlinear least squares. It was found that cellulose hydrolysis follows a model that takes into account competitive inhibition by cellobiose (final product) with the following parameters: Km = 3.8 mM, Kic = 0.041 mM, kcat = 2 h-1 (5.6 x 10-4 s-1). Other models, such as mixed type inhibition and those incorporating improvements concerning inhibition by substrate and parabolic inhibition, increased the modulation performance very slightly. The results support the hypothesis that nonproductive enzyme substrate complexes, parabolic inhibition, and enzyme inactivation (Selwyn test) are not the principal constraints in enzymatic cellulose hydrolysis. Under our conditions, the increment in hydrolysis was not significant for substrate/enzyme ratios <6.5.

  16. Modeling of Temperature and Residual Stress Fields Resulting from Impacting Process of a Molten Ni Particle onto a Flat Substrate

    Institute of Scientific and Technical Information of China (English)

    WANG Lu; FAN Qun-bo; WANG Fu-chi

    2003-01-01

    Seeveral effective numerical techniques, based on a finite element analysis, have been developed and computed independently. Restdts are presented describing the imppacting process, and the subsequent temperature and residual stress fields of a molten nickel particle impacting onto a flat substrate. Problems of this type , especially the prediction of the thermal residual stresses, are of major practical interest in thermal spray operations as a pioneering approach.

  17. Mycobacterium tuberculosis RecG binds and unwinds model DNA substrates with a preference for Holliday junctions.

    Science.gov (United States)

    Zegeye, Ephrem Debebe; Balasingham, Seetha V; Laerdahl, Jon K; Homberset, Håvard; Tønjum, Tone

    2012-08-01

    The RecG enzyme, a superfamily 2 helicase, is present in nearly all bacteria. Here we report for the first time that the recG gene is also present in the genomes of most vascular plants as well as in green algae, but is not found in other eukaryotes or archaea. The precise function of RecG is poorly understood, although ample evidence shows that it plays critical roles in DNA repair, recombination and replication. We further demonstrate that Mycobacterium tuberculosis RecG (RecG(Mtb)) DNA binding activity had a broad substrate specificity, whereas it only unwound branched-DNA substrates such as Holliday junctions (HJs), replication forks, D-loops and R-loops, with a strong preference for the HJ as a helicase substrate. In addition, RecG(Mtb) preferentially bound relatively long (≥40 nt) ssDNA, exhibiting a higher affinity for the homopolymeric nucleotides poly(dT), poly(dG) and poly(dC) than for poly(dA). RecG(Mtb) helicase activity was supported by hydrolysis of ATP or dATP in the presence of Mg(2+), Mn(2+), Cu(2+) or Fe(2+). Like its Escherichia coli orthologue, RecG(Mtb) is also a strictly DNA-dependent ATPase.

  18. Simulation of green roof runoff under different substrate depths and vegetation covers by coupling a simple conceptual and a physically based hydrological model.

    Science.gov (United States)

    Soulis, Konstantinos X; Valiantzas, John D; Ntoulas, Nikolaos; Kargas, George; Nektarios, Panayiotis A

    2017-09-15

    In spite of the well-known green roof benefits, their widespread adoption in the management practices of urban drainage systems requires the use of adequate analytical and modelling tools. In the current study, green roof runoff modeling was accomplished by developing, testing, and jointly using a simple conceptual model and a physically based numerical simulation model utilizing HYDRUS-1D software. The use of such an approach combines the advantages of the conceptual model, namely simplicity, low computational requirements, and ability to be easily integrated in decision support tools with the capacity of the physically based simulation model to be easily transferred in conditions and locations other than those used for calibrating and validating it. The proposed approach was evaluated with an experimental dataset that included various green roof covers (either succulent plants - Sedum sediforme, or xerophytic plants - Origanum onites, or bare substrate without any vegetation) and two substrate depths (either 8 cm or 16 cm). Both the physically based and the conceptual models matched very closely the observed hydrographs. In general, the conceptual model performed better than the physically based simulation model but the overall performance of both models was sufficient in most cases as it is revealed by the Nash-Sutcliffe Efficiency index which was generally greater than 0.70. Finally, it was showcased how a physically based and a simple conceptual model can be jointly used to allow the use of the simple conceptual model for a wider set of conditions than the available experimental data and in order to support green roof design. Copyright © 2017 Elsevier Ltd. All rights reserved.

  19. Determination of the photocatalytic activity of TiO2 coatings on clay roofing tile substrates methylene blue as model pollutant

    Directory of Open Access Journals (Sweden)

    Skapin Andrea S.

    2009-01-01

    Full Text Available The photocatalytically active mesoporous coatings, based on titanium dioxide sols (Degussa, of the fired clay roofing tiles substrate were prepared by using poly(ethylene glycol (PEG M-600 and M-4000, as the structure directing agents. The coatings were deposited using spray technique followed by thermal treatment. Photocatalytic activity of the TiO2 coatings was evaluated by aqueous solution of methylene blue as model dye, deposited on the top of the coatings, after irradiation with UV light. The results were compared with the photocatalytic efficiency of some commercial self-cleaning products (clay roofing tiles, glass. The newly design coatings showed an interesting decolourisation performance (over 30 % after 24 h. It appeared that the procedure of photocatalytic activity determination, in the case of porous substrates, should be renewed by a preadsorption process.

  20. Slip or not slip? A methodical examination of the interface formation model using two-dimensional droplet spreading on a horizontal planar substrate as a prototype system

    CERN Document Server

    Sibley, David N; Kalliadasis, Serafim

    2012-01-01

    We consider the spreading of a thin two-dimensional droplet on a planar substrate as a prototype system to compare the contemporary model for contact line motion based on interface formation of Shikhmurzaev [Int. J. Multiphas. Flow 19, 589 (1993)], to the more commonly used continuum fluid dynamical equations augmented with the Navier-slip condition. Considering quasistatic droplet evolution and using the method of matched asymptotics, we find that the evolution of the droplet radius using the interface formation model reduces to an equivalent expression for a slip model, where the prescribed microscopic dynamic contact angle has a velocity dependent correction to its static value. This result is found for both the original interface formation model formulation and for a more recent version, where mass transfer from bulk to surface layers is accounted for through the boundary conditions. Various features of the model, such as the pressure behaviour and rolling motion at the contact line, and their relevance, ...

  1. Membrane-bound human orphan cytochrome P450 2U1: Sequence singularities, construction of a full 3D model, and substrate docking.

    Science.gov (United States)

    Ducassou, Lionel; Dhers, Laura; Jonasson, Gabriella; Pietrancosta, Nicolas; Boucher, Jean-Luc; Mansuy, Daniel; André, François

    2017-09-01

    Human cytochrome P450 2U1 (CYP2U1) is an orphan CYP that exhibits several distinctive characteristics among the 57 human CYPs with a highly conserved sequence in almost all living organisms. We compared its protein sequence with those of the 57 human CYPs and constructed a 3D structure of a full-length CYP2U1 model bound to a POPC membrane. We also performed docking experiments of arachidonic acid (AA) and N-arachidonoylserotonin (AS) in this model. The protein sequence of CYP2U1 displayed two unique characteristics when compared to those of the human CYPs, the presence of a longer N-terminal region upstream of the putative trans-membrane helix (TMH) containing 8 proline residues, and of an insert of about 20 amino acids containing 5 arginine residues between helices A' and A. Its N-terminal part upstream of TMH involved an additional short terminal helix, in a manner similar to what was reported in the crystal structure of Saccharomyces cerevisiae CYP51. Our model also showed a specific interaction between the charged residues of insert AA' and phosphate groups of lipid polar heads, suggesting a possible role of this insert in substrate recruitment. Docking of AA and AS in this model showed these substrates in channel 2ac, with the terminal alkyl chain of AA or the indole ring of AS close to the heme, in agreement with the reported CYP2U1-catalyzed AA and AS hydroxylation regioselectivities. This model should be useful to find new endogenous or exogenous CYP2U1 substrates and to interpret the regioselectivity of their hydroxylation. Copyright © 2017 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

  2. Mathematical modeling of the ethanol fermentation of cashew apple juice by a flocculent yeast: the effect of initial substrate concentration and temperature.

    Science.gov (United States)

    Pinheiro, Álvaro Daniel Teles; da Silva Pereira, Andréa; Barros, Emanuel Meneses; Antonini, Sandra Regina Ceccato; Cartaxo, Samuel Jorge Marques; Rocha, Maria Valderez Ponte; Gonçalves, Luciana Rocha B

    2017-08-01

    In this work, the effect of initial sugar concentration and temperature on the production of ethanol by Saccharomyces cerevisiae CCA008, a flocculent yeast, using cashew apple juice in a 1L-bioreactor was studied. The experimental results were used to develop a kinetic model relating biomass, ethanol production and total reducing sugar consumption. Monod, Andrews, Levenspiel and Ghose and Tyagi models were investigated to represent the specific growth rate without inhibition, with inhibition by substrate and with inhibition by product, respectively. Model validation was performed using a new set of experimental data obtained at 34 °C and using 100 g L(-1) of initial substrate concentration. The model proposed by Ghose and Tyagi was able to accurately describe the dynamics of ethanol production by S. cerevisiae CCA008 growing on cashew apple juice, containing an initial reducing sugar concentration ranging from 70 to 170 g L(-1) and temperature, from 26 to 42 °C. The model optimization was also accomplished based on the following parameters: percentage volume of ethanol per volume of solution (%V ethanol/V solution), efficiency and reaction productivity. The optimal operational conditions were determined using response surface graphs constructed with simulated data, reaching an efficiency and a productivity of 93.5% and 5.45 g L(-1) h(-1), respectively.

  3. Production of aggregation prone human interferon gamma and its mutant in highly soluble and biologically active form by SUMO fusion technology.

    Science.gov (United States)

    Tileva, M; Krachmarova, E; Ivanov, I; Maskos, K; Nacheva, G

    2016-01-01

    The Escherichia coli expression system is a preferable choice for production of recombinant proteins. A disadvantage of this system is the target protein aggregation in "inclusion bodies" (IBs) that further requires solubilisation and refolding, which is crucial for the properties and the yield of the final product. In order to prevent aggregation, SUMO fusion tag technology has been successfully applied for expression of eukaryotic proteins, including human interferon gamma (hIFNγ) that was reported, however, with no satisfactory biological activity. We modified this methodology for expression and purification of both the wild type hIFNγ and an extremely prone to aggregation mutant hIFNγ-K88Q, whose recovery from IBs showed to be ineffective upon numerous conditions. By expression of the N-terminal His-SUMO fusion proteins in the E. coli strain BL21(DE3)pG-KJE8, co-expressing two chaperone systems, at 24 °C a significant increase in solubility of both target proteins (1.5-fold for hIFNγ and 8-fold for K88Q) was achieved. Two-step chromatography (affinity and ion-exchange) with on-dialysis His-SUMO-tag cleavage was applied for protein purification that yielded 6.0-7.0mg/g wet biomass for both proteins with >95% purity and native N-termini. The optimised protocol led to increased yields from 5.5 times for hIFNγ up to 100 times for K88Q in comparison to their isolation from IBs. Purified hIFNγ showed preserved thermal stability and antiproliferative activity corresponding to that of the native reference sample (3 × 10(7)IU/mg). The developed methodology represents an optimised procedure that can be successfully applied for large scale expression and purification of aggregation-prone proteins in soluble native form.

  4. Modeling, synthesis and study of highly efficient solar cells based on III-nitride nanowire arrays grown on Si substrates

    Science.gov (United States)

    Mozharov, A. M.; Bolshakov, A. D.; Kudryashov, D. A.; Kryzhanovskaya, N. V.; Cirlin, G. E.; Mukhin, I. S.; Harmand, J. C.; Tchernysheva, M.

    2015-11-01

    In this letter we investigate photovoltaic properties of GaN nanowires (NWs) - Si substrate heterostructure obtained by molecular beam epitaxy (MBE). Antireflection properties of the NW array were studied theoretically and experimentally to show an order of magnitude enhancement in antireflection comparing to the pure Si surface (2.5% vs. 33.8%). In order to determine optimal morphology and doping levels of the structure with maximum possible efficiency we simulated it's properties using a finite difference method. The carried out simulation showed that a maximum efficiency should be 20%.

  5. Semi-Analytical Modeling and Analysis in Three Dimensions of the Optical Carrier Injection and Diffusion in a Semiconductor Substrate

    Science.gov (United States)

    Gary, René; Arnould, Jean-Daniel; Vilcot, Anne

    2006-05-01

    In order to be faster and more precise than any numerical technique for the computation of the photo-induced plasma in semiconductor, an analytical solution has to be developed. In this paper, the Hankel transform is used to simplify the solution of the differential equation of second order with nonconstant coefficient, known as the diffusion equation. The resulting expression of the three-dimensional (3-D) carrier density includes all the physical parameters of the substrate and the laser beam as well. A parametric study was also feasible using the developed expressions.

  6. Steady state kinetic model for the binding of substrates and allosteric effectors to Escherichia coli phosphoribosyl-diphosphate synthase

    DEFF Research Database (Denmark)

    Willemoës, Martin; Hove-Jensen, Bjarne; Larsen, Sine

    2000-01-01

    saturation with ribose 5-phosphate leads to the binding of Mg2+ and substrates via a slow pathway where Pi binds to the enzyme last. The random mechanism for Pi binding was further supported by studies with competitive inhibitors of Mg2+, MgATP, and ribose 5-phosphate that all appeared noncompetitive when...... varying Pi at either saturating or unsaturating ribose 5-phosphate concentrations. Furthermore, none of the inhibitors induced inhibition at increasing Pi concentrations. Results from ADP inhibition of Pi activation suggest that these effectors compete for binding to a common regulatory site....

  7. Power electronics substrate for direct substrate cooling

    Science.gov (United States)

    Le, Khiet [Mission Viejo, CA; Ward, Terence G [Redondo Beach, CA; Mann, Brooks S [Redondo Beach, CA; Yankoski, Edward P [Corona, CA; Smith, Gregory S [Woodland Hills, CA

    2012-05-01

    Systems and apparatus are provided for power electronics substrates adapted for direct substrate cooling. A power electronics substrate comprises a first surface configured to have electrical circuitry disposed thereon, a second surface, and a plurality of physical features on the second surface. The physical features are configured to promote a turbulent boundary layer in a coolant impinged upon the second surface.

  8. La tradición oral de los indígenas Sumos: Características y temáticas

    OpenAIRE

    Arguedas, Gilda Rosa

    2015-01-01

    Este artículo se ocupa de la tradición oral de los "Sumo" indios, uno de los grupos que habitan la costa atlántica de Honduras y Nicaragua, también conocido como "Mosquitia".Las historias analizadas (36) son principalmente narraciones aunque hay algunas descripciones y explicaciones. De acuerdo con el momento en el que se establece la narración, estas historias se clasifican en tres grupos: los mitos, leyendas y cuentos. Los temas de las historias son variadas, aunque los elementos míticos pr...

  9. Atrial Heterogeneity Generates Re-entrant Substrate during Atrial Fibrillation and Anti-arrhythmic Drug Action: Mechanistic Insights from Canine Atrial Models.

    Science.gov (United States)

    Varela, Marta; Colman, Michael A; Hancox, Jules C; Aslanidi, Oleg V

    2016-12-01

    Anti-arrhythmic drug therapy is a frontline treatment for atrial fibrillation (AF), but its success rates are highly variable. This is due to incomplete understanding of the mechanisms of action of specific drugs on the atrial substrate at different stages of AF progression. We aimed to elucidate the role of cellular, tissue and organ level atrial heterogeneities in the generation of a re-entrant substrate during AF progression, and their modulation by the acute action of selected anti-arrhythmic drugs. To explore the complex cell-to-organ mechanisms, a detailed biophysical models of the entire 3D canine atria was developed. The model incorporated atrial geometry and fibre orientation from high-resolution micro-computed tomography, region-specific atrial cell electrophysiology and the effects of progressive AF-induced remodelling. The actions of multi-channel class III anti-arrhythmic agents vernakalant and amiodarone were introduced in the model by inhibiting appropriate ionic channel currents according to experimentally reported concentration-response relationships. AF was initiated by applied ectopic pacing in the pulmonary veins, which led to the generation of localized sustained re-entrant waves (rotors), followed by progressive wave breakdown and rotor multiplication in both atria. The simulated AF scenarios were in agreement with observations in canine models and patients. The 3D atrial simulations revealed that a re-entrant substrate was typically provided by tissue regions of high heterogeneity of action potential duration (APD). Amiodarone increased atrial APD and reduced APD heterogeneity and was more effective in terminating AF than vernakalant, which increased both APD and APD dispersion. In summary, the initiation and sustenance of rotors in AF is linked to atrial APD heterogeneity and APD reduction due to progressive remodelling. Our results suggest that anti-arrhythmic strategies that increase atrial APD without increasing its dispersion are

  10. Structure-Function Relationship of a Plant NCS1 Member – Homology Modeling and Mutagenesis Identified Residues Critical for Substrate Specificity of PLUTO, a Nucleobase Transporter from Arabidopsis

    Science.gov (United States)

    Witz, Sandra; Panwar, Pankaj; Schober, Markus; Deppe, Johannes; Pasha, Farhan Ahmad; Lemieux, M. Joanne; Möhlmann, Torsten

    2014-01-01

    Plastidic uracil salvage is essential for plant growth and development. So far, PLUTO, the plastidic nucleobase transporter from Arabidopsis thaliana is the only known uracil importer at the inner plastidic membrane which represents the permeability barrier of this organelle. We present the first homology model of PLUTO, the sole plant NCS1 member from Arabidopsis based on the crystal structure of the benzyl hydantoin transporter MHP1 from Microbacterium liquefaciens and validated by molecular dynamics simulations. Polar side chains of residues Glu-227 and backbones of Val-145, Gly-147 and Thr-425 are proposed to form the binding site for the three PLUTO substrates uracil, adenine and guanine. Mutational analysis and competition studies identified Glu-227 as an important residue for uracil and to a lesser extent for guanine transport. A differential response in substrate transport was apparent with PLUTO double mutants E227Q G147Q and E227Q T425A, both of which most strongly affected adenine transport, and in V145A G147Q, which markedly affected guanine transport. These differences could be explained by docking studies, showing that uracil and guanine exhibit a similar binding mode whereas adenine binds deep into the catalytic pocket of PLUTO. Furthermore, competition studies confirmed these results. The present study defines the molecular determinants for PLUTO substrate binding and demonstrates key differences in structure-function relations between PLUTO and other NCS1 family members. PMID:24621654

  11. Structure-function relationship of a plant NCS1 member--homology modeling and mutagenesis identified residues critical for substrate specificity of PLUTO, a nucleobase transporter from Arabidopsis.

    Science.gov (United States)

    Witz, Sandra; Panwar, Pankaj; Schober, Markus; Deppe, Johannes; Pasha, Farhan Ahmad; Lemieux, M Joanne; Möhlmann, Torsten

    2014-01-01

    Plastidic uracil salvage is essential for plant growth and development. So far, PLUTO, the plastidic nucleobase transporter from Arabidopsis thaliana is the only known uracil importer at the inner plastidic membrane which represents the permeability barrier of this organelle. We present the first homology model of PLUTO, the sole plant NCS1 member from Arabidopsis based on the crystal structure of the benzyl hydantoin transporter MHP1 from Microbacterium liquefaciens and validated by molecular dynamics simulations. Polar side chains of residues Glu-227 and backbones of Val-145, Gly-147 and Thr-425 are proposed to form the binding site for the three PLUTO substrates uracil, adenine and guanine. Mutational analysis and competition studies identified Glu-227 as an important residue for uracil and to a lesser extent for guanine transport. A differential response in substrate transport was apparent with PLUTO double mutants E227Q G147Q and E227Q T425A, both of which most strongly affected adenine transport, and in V145A G147Q, which markedly affected guanine transport. These differences could be explained by docking studies, showing that uracil and guanine exhibit a similar binding mode whereas adenine binds deep into the catalytic pocket of PLUTO. Furthermore, competition studies confirmed these results. The present study defines the molecular determinants for PLUTO substrate binding and demonstrates key differences in structure-function relations between PLUTO and other NCS1 family members.

  12. Structure-function relationship of a plant NCS1 member--homology modeling and mutagenesis identified residues critical for substrate specificity of PLUTO, a nucleobase transporter from Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Sandra Witz

    Full Text Available Plastidic uracil salvage is essential for plant growth and development. So far, PLUTO, the plastidic nucleobase transporter from Arabidopsis thaliana is the only known uracil importer at the inner plastidic membrane which represents the permeability barrier of this organelle. We present the first homology model of PLUTO, the sole plant NCS1 member from Arabidopsis based on the crystal structure of the benzyl hydantoin transporter MHP1 from Microbacterium liquefaciens and validated by molecular dynamics simulations. Polar side chains of residues Glu-227 and backbones of Val-145, Gly-147 and Thr-425 are proposed to form the binding site for the three PLUTO substrates uracil, adenine and guanine. Mutational analysis and competition studies identified Glu-227 as an important residue for uracil and to a lesser extent for guanine transport. A differential response in substrate transport was apparent with PLUTO double mutants E227Q G147Q and E227Q T425A, both of which most strongly affected adenine transport, and in V145A G147Q, which markedly affected guanine transport. These differences could be explained by docking studies, showing that uracil and guanine exhibit a similar binding mode whereas adenine binds deep into the catalytic pocket of PLUTO. Furthermore, competition studies confirmed these results. The present study defines the molecular determinants for PLUTO substrate binding and demonstrates key differences in structure-function relations between PLUTO and other NCS1 family members.

  13. Structure-function relationship of a plant NCS1 member - Homology modeling and mutagenesis identified residues critical for substrate specificity of PLUTO, a nucleobase transporter from arabidopsis

    KAUST Repository

    Witz, Sandra

    2014-03-12

    Plastidic uracil salvage is essential for plant growth and development. So far, PLUTO, the plastidic nucleobase transporter from Arabidopsis thaliana is the only known uracil importer at the inner plastidic membrane which represents the permeability barrier of this organelle. We present the first homology model of PLUTO, the sole plant NCS1 member from Arabidopsis based on the crystal structure of the benzyl hydantoin transporter MHP1 from Microbacterium liquefaciens and validated by molecular dynamics simulations. Polar side chains of residues Glu-227 and backbones of Val-145, Gly-147 and Thr-425 are proposed to form the binding site for the three PLUTO substrates uracil, adenine and guanine. Mutational analysis and competition studies identified Glu-227 as an important residue for uracil and to a lesser extent for guanine transport. A differential response in substrate transport was apparent with PLUTO double mutants E227Q G147Q and E227Q T425A, both of which most strongly affected adenine transport, and in V145A G147Q, which markedly affected guanine transport. These differences could be explained by docking studies, showing that uracil and guanine exhibit a similar binding mode whereas adenine binds deep into the catalytic pocket of PLUTO. Furthermore, competition studies confirmed these results. The present study defines the molecular determinants for PLUTO substrate binding and demonstrates key differences in structure-function relations between PLUTO and other NCS1 family members. 2014 Witz et al.

  14. Atomistic modeling of femtosecond laser-induced melting and atomic mixing in Au film - Cu substrate system

    Energy Technology Data Exchange (ETDEWEB)

    Thomas, Derek A.; Lin Zhibin [Department of Materials Science and Engineering, University of Virginia, 395 McCormick Road, Charlottesville, VA 22904-4745 (United States); Zhigilei, Leonid V., E-mail: lz2n@virginia.edu [Department of Materials Science and Engineering, University of Virginia, 395 McCormick Road, Charlottesville, VA 22904-4745 (United States); Gurevich, Evgeny L.; Kittel, Silke; Hergenroeder, Roland [ISAS - Institute for Analytical Sciences, Department of Material Analysis Bunsen-Kirchhoff-Str. 11, 44139 Dortmund (Germany)

    2009-09-30

    The mechanisms of femtosecond laser-induced transient melting and atomic mixing in a target composed of a 30 nm Au film deposited on a bulk Cu substrate are investigated in a series of atomistic simulations. The relative strength and the electron temperature dependence of the electron-phonon coupling of the metals composing the layered target are identified as major factors affecting the initial energy redistribution and the location of the region(s) undergoing transient melting and resolidification. The higher strength of the electron-phonon coupling in Cu, as compared to Au, results in a preferential sub-surface heating and melting of the Cu substrate, while the overlaying Au film largely retains its original crystalline structure. The large difference in the atomic mobility in the transiently melted and crystalline regions of the target makes it possible to connect the final distributions of the components in the resolidified targets to the history of the laser-induced melting process, thus allowing for experimental verification of the computational predictions.

  15. Atomistic modeling of femtosecond laser-induced melting and atomic mixing in Au film - Cu substrate system

    Science.gov (United States)

    Thomas, Derek A.; Lin, Zhibin; Zhigilei, Leonid V.; Gurevich, Evgeny L.; Kittel, Silke; Hergenröder, Roland

    2009-09-01

    The mechanisms of femtosecond laser-induced transient melting and atomic mixing in a target composed of a 30 nm Au film deposited on a bulk Cu substrate are investigated in a series of atomistic simulations. The relative strength and the electron temperature dependence of the electron-phonon coupling of the metals composing the layered target are identified as major factors affecting the initial energy redistribution and the location of the region(s) undergoing transient melting and resolidification. The higher strength of the electron-phonon coupling in Cu, as compared to Au, results in a preferential sub-surface heating and melting of the Cu substrate, while the overlaying Au film largely retains its original crystalline structure. The large difference in the atomic mobility in the transiently melted and crystalline regions of the target makes it possible to connect the final distributions of the components in the resolidified targets to the history of the laser-induced melting process, thus allowing for experimental verification of the computational predictions.

  16. Triptolide inhibits the proliferation of prostate cancer cells and down-regulates SUMO-specific protease 1 expression.

    Directory of Open Access Journals (Sweden)

    Weiwei Huang

    Full Text Available Recently, traditional Chinese medicine and medicinal herbs have attracted more attentions worldwide for its anti-tumor efficacy. Celastrol and Triptolide, two active components extracted from the Chinese herb Tripterygium wilfordii Hook F (known as Lei Gong Teng or Thunder of God Vine, have shown anti-tumor effects. Celastrol was identified as a natural 26 s proteasome inhibitor which promotes cell apoptosis and inhibits tumor growth. The effect and mechanism of Triptolide on prostate cancer (PCa is not well studied. Here we demonstrated that Triptolide, more potent than Celastrol, inhibited cell growth and induced cell death in LNCaP and PC-3 cell lines. Triptolide also significantly inhibited the xenografted PC-3 tumor growth in nude mice. Moreover, Triptolide induced PCa cell apoptosis through caspases activation and PARP cleavage. Unbalance between SUMOylation and deSUMOylation was reported to play an important role in PCa progression. SUMO-specific protease 1 (SENP1 was thought to be a potential marker and therapeutical target of PCa. Importantly, we observed that Triptolide down-regulated SENP1 expression in both mRNA and protein levels in dose-dependent and time-dependent manners, resulting in an enhanced cellular SUMOylation in PCa cells. Meanwhile, Triptolide decreased AR and c-Jun expression at similar manners, and suppressed AR and c-Jun transcription activity. Furthermore, knockdown or ectopic SENP1, c-Jun and AR expression in PCa cells inhibited the Triptolide anti-PCa effects. Taken together, our data suggest that Triptolide is a natural compound with potential therapeutic value for PCa. Its anti-tumor activity may be attributed to mechanisms involving down-regulation of SENP1 that restores SUMOylation and deSUMOyaltion balance and negative regulation of AR and c-Jun expression that inhibits the AR and c-Jun mediated transcription in PCa.

  17. Oxidation of dibenzothiophene as a model substrate for the removal of organic sulphur from fossil fuels by iron(III ions generated from pyrite by Acidithiobacillus ferrooxidans

    Directory of Open Access Journals (Sweden)

    VLADIMIR P. BESKOSKI

    2007-06-01

    Full Text Available Within this paper a new idea for the removal of organically bonded sulphur from fossil fuels is discussed. Dibenzothiophene (DBT was used as a model compound of organicmolecules containing sulphur. This form of (biodesulphurization was performed by an indirect mechanism in which iron(III ions generated from pyrite by Acidithiobacillus ferrooxidans performed the abiotic oxidation. The obtained reaction products, dibenzothiopene sulfoxide and dibenzothiophene sulfone, are more soluble in water than the basic substrate and the obtained results confirmed the basic hypothesis and give the posibility of continuing the experiments related to application of this (biodesulphurization process.

  18. Application of the finite difference method to model pH and substrate concentration in a double-chamber microbial fuel cell.

    Science.gov (United States)

    Zhang, Liwei; Deshusses, Marc

    2014-01-01

    The purpose of this study was to develop a mathematical model that can describe glucose degradation in a microbial fuel cell (MFC) with the use of finite difference approach. The dynamic model can describe both substrate and pH changes in the anode chamber of a double-chamber MFC. It was developed using finite differences and incorporates basic mass transfer concepts. Model simulation results could fit the experimental data for substrate consumption well, while there was a moderate discrepancy (maximum 0.11 pH unit) between the simulated pH and the experimental data. A parametric sensitivity analysis showed that increases in acetate and propionate consumption rates can cause great decrease in chemical oxygen demand (COD) in the anode chamber, while an increase in glucose consumption rate does not result in significant changes of COD reduction. Therefore, the rate limitation steps of glucose degradation are the oxidations of secondary degradation products of glucose (acetate and propionate). Due to the buffering effect of the nutrient solution, the increases in glucose, acetate and propionate consumption rates did not result in much change on pH of the anode chamber.

  19. Statistical evaluation and modeling of cheap substrate-based cultivation medium of Chlorella vulgaris to enhance microalgae lipid as new potential feedstock for biolubricant.

    Science.gov (United States)

    Mohammad Mirzaie, M A; Kalbasi, M; Mousavi, S M; Ghobadian, B

    2016-05-18

    Chlorella vulgaris (C. vulgaris) microalga was investigated as a new potential feedstock for the production of biodegradable lubricant. In order to enhance microalgae lipid for biolubricant production, mixotrophic growth of C. vulgaris was optimized using statistical analysis of Plackett-Burman (P-B) and response surface methodology (RSM). A cheap substrate-based medium of molasses and corn steep liquor (CSL) was used instead of expensive mineral salts to reduce the total cost of microalgae production. The effects of molasses and CSL concentration (cheap substrates) and light intensity on the growth of microalgae and their lipid content were analyzed and modeled. Designed models by RSM showed good compatibility with a 95% confidence level when compared to the cultivation system. According to the models, optimal cultivation conditions were obtained with biomass productivity of 0.123 g L(-1) day(-1) and lipid dry weight of 0.64 g L(-1) as 35% of dry weight of C. vulgaris. The extracted microalgae lipid presented useful fatty acid for biolubricant production with viscosities of 42.00 cSt at 40°C and 8.500 cSt at 100°C, viscosity index of 185, flash point of 185°C, and pour point of -6°C. These properties showed that microalgae lipid could be used as potential feedstock for biolubricant production.

  20. Laser fabricated discontinuous anisotropic microconical substrates as a new model scaffold to control the directionality of neuronal network outgrowth.

    Science.gov (United States)

    Simitzi, C; Efstathopoulos, P; Kourgiantaki, A; Ranella, A; Charalampopoulos, I; Fotakis, C; Athanassakis, I; Stratakis, E; Gravanis, A

    2015-10-01

    Patterning of neuronal outgrowth in vitro is important in tissue engineering as well as for the development of neuronal interfaces with desirable characteristics. To date, this has been achieved with the aid of micro- and nanofabrication techniques giving rise to various anisotropic topographies, either in the form of continuous or discontinuous structures. In this study we propose a currently unexplored geometry of a 3D culture substrate for neuronal cell growth comprising discontinuous subcellular microstructures with anisotropic geometrical cross-section. Specifically, using laser precision 3D micro/nano fabrication techniques, silicon substrates comprising arrays of parallel oriented elliptical microcones (MCs) were fabricated to investigate whether a discontinuous geometry comprising anisotropic features at the subcellular level could influence the alignment of peripheral nervous system cell populations. It was shown that both Schwann cells and axons of sympathetic neurons were parallel oriented onto the MCs of elliptical shape, while they exhibited a random orientation onto the MCs of arbitrary shape. Notably, this topography-induced guidance effect was also observed in more complex cell culture systems, such as the organotypic culture whole dorsal root ganglia (DRG) explants. Our results suggest that a discontinuous topographical pattern could promote Schwann cell and axonal alignment, provided that it hosts anisotropic geometrical features, even though the sizes of those range at the subcellular lengthscale. The laser-patterned arrays of MCs presented here could potentially be a useful platform for patterning neurons into artificial networks, allowing the study of neuronal cells interactions under 3D ex-vivo conditions.

  1. Droplet dynamics on patterned substrates

    Indian Academy of Sciences (India)

    A Dupuis; J M Yeomans

    2005-06-01

    We present a lattice Boltzmann algorithm which can be used to explore the spreading of droplets on chemically and topologically patterned substrates. As an example we use the method to show that the final configuration of a drop on a substrate comprising hydrophobic and hydrophilic stripes can depend sensitively on the dynamical pathway by which the state is reached. We also consider a substrate covered with micron-scale posts and investigate how this can lead to superhydrophobic behaviour. Finally we model how a Namibian desert beetle collects water from the wind.

  2. Substrate noise coupling in RFICs

    CERN Document Server

    Helmy, Ahmed

    2008-01-01

    Substrate Noise Coupling in RFICs addresses substrate noise coupling in RF and mixed signal ICs when used in a system on chip (SoC) containing digital ICs as well. This trend of integrating RF, mixed signal ICs with large digital ICs is found in many of today's commercial ICs such as single chip Wi-Fi or Bluetooth solutions and is expected to grow rapidly in the future. The book reports modeling and simulation techniques for substrate noise coupling effects in RFICs and introduces isolation structures and design guides to mitigate such effects with the ultimate goal of enhancing the yield of R

  3. Bridged bis(beta-cyclodextrin)s possessing coordinated metal center(s) and their inclusion complexation behavior with model substrates: enhanced molecular binding ability by multiple recognition.

    Science.gov (United States)

    Liu, Y; Chen, Y; Li, L; Zhang, H Y; Liu, S X; Guan, X D

    2001-12-14

    To investigate quantitatively the cooperative binding ability of several beta-cyclodextrin oligomers bearing single or multiligated metal center(s), the inclusion complexation behavior of four bis(beta-cyclodextrin)s (2-5) linked by 2,2'-bipyridine-4,4'-dicarboxy tethers and their copper(II) complexes (6-9) with representative dye guests, i.e., methyl orange (MO), acridine red (AR), rhodamine B (RhB), ammonium 8-anilino-1-naphthalenesulfonic acid (ANS), and sodium 6-(p-toludino)-2-naphthalenesulfonate (TNS), have been examined in aqueous solution at 25 degrees C by means of UV-vis, circular dichroism, fluorescence, and 2D NMR spectroscopy. The results obtained indicate that bis(beta-cyclodextrin)s 2-5 can associate with one or three copper(II) ion(s) producing 2:1 or 2:3 bis(beta-cyclodextrin)-copper(II) complexes. These metal-ligated oligo(beta-cyclodextrin)s can bind two model substrates to form intramolecular 2:2 host-guest inclusion complexes and thus significantly enhance the original binding abilities of parent beta-cyclodextrin and bis(beta-cyclodextrin) toward model substrates through the cooperative binding of two guest molecules by four tethered cyclodextrin moieties, as well as the additional binding effect supplied by ligated metal center(s). Host 6 showed the highest enhancement of the stability constant, up to 38.3 times for ANS as compared with parent beta-cyclodextrin. The molecular binding mode and stability constant of substrates by bridged bis- and oligo(beta-cyclodextrin)s 2-9 are discussed from the viewpoint of the size/shape-fit interaction and molecular multiple recognition between host and guest.

  4. Development of novel rifampicin-derived P-glycoprotein activators/inducers. synthesis, in silico analysis and application in the RBE4 cell model, using paraquat as substrate.

    Directory of Open Access Journals (Sweden)

    Vânia Vilas-Boas

    Full Text Available P-glycoprotein (P-gp is a 170 kDa transmembrane protein involved in the outward transport of many structurally unrelated substrates. P-gp activation/induction may function as an antidotal pathway to prevent the cytotoxicity of these substrates. In the present study we aimed at testing rifampicin (Rif and three newly synthesized Rif derivatives (a mono-methoxylated derivative, MeORif, a peracetylated derivative, PerAcRif, and a reduced derivative, RedRif to establish their ability to modulate P-gp expression and activity in a cellular model of the rat's blood-brain barrier, the RBE4 cell line P-gp expression was assessed by western blot using C219 anti-P-gp antibody. P-gp function was evaluated by flow cytometry measuring the accumulation of rhodamine123. Whenever P-gp activation/induction ability was detected in a tested compound, its antidotal effect was further tested using paraquat as cytotoxicity model. Interactions between Rif or its derivatives and P-gp were also investigated by computational analysis. Rif led to a significant increase in P-gp expression at 72 h and RedRif significantly increased both P-gp expression and activity. No significant differences were observed for the other derivatives. Pre- or simultaneous treatment with RedRif protected cells against paraquat-induced cytotoxicity, an effect reverted by GF120918, a P-gp inhibitor, corroborating the observed P-gp activation ability. Interaction of RedRif with P-gp drug-binding pocket was consistent with an activation mechanism of action, which was confirmed with docking studies. Therefore, RedRif protection against paraquat-induced cytotoxicity in RBE4 cells, through P-gp activation/induction, suggests that it may be useful as an antidote for cytotoxic substrates of P-gp.

  5. Detection of O-propargyl-puromycin with SUMO and ubiquitin by click chemistry at PML-nuclear bodies during abortive proteasome activities

    Energy Technology Data Exchange (ETDEWEB)

    Uozumi, Naoki; Matsumoto, Hotaru [Course for Biological Sciences, Faculty of Science, Kumamoto University, Kumamoto (Japan); Saitoh, Hisato, E-mail: hisa@kumamoto-u.ac.jp [Course for Biological Sciences, Faculty of Science, Kumamoto University, Kumamoto (Japan); Department of Biological Sciences, Graduate School of Science and Technology, Kumamoto University, Kumamoto (Japan)

    2016-05-27

    The amino-nucleoside antibiotic, puromycin, acts by covalently linking to elongating polypeptide chains on ribosomes to generate prematurely terminated immature polypeptides. The trafficking of puromycin-conjugated (puromycylated) immature polypeptides within cell has, however, remained elusive. In this study, using O-propargyl-puromycin (OP-Puro), the distribution of puromycylated polypeptides was assessed in HeLa cells by click chemistry. Under standard culture conditions, OP-Puro signals were detected in the cytoplasm and nucleus with the highest concentrations in the nucleolus. Intriguingly, when proteasome activities were aborted using MG132, OP-Puro signals began to accumulate at promyelocytic leukemia nuclear bodies (PML-NBs) in addition to the nucleolus. We also found promiscuous association of OP-Puro signals with SUMO-2/3 and ubiquitin at PML-NBs, but not at the nucleolus, during abortive proteasome activities. This study reveals a previously unknown distribution of OP-Puro that argues for a nuclear function in regulating immature protein homeostasis. -- Highlights: •Click chemistry detects O-propargyl-puromycin (OP-Puro) signals in the nucleus. •OP-Puro accumulates at PML-NBs during abortive proteasome activities. •SUMO and ubiquitin are promiscuously associated with OP-Puro at PML-NBs. •The nucleus may function in immature protein homeostasis.

  6. Effects of Nephrolithiasis on Serum DNase (Deoxyribonuclease I and II) Activity and E3 SUMO-Protein Ligase NSE2 (NSMCE2) in Malaysian Individuals

    Institute of Scientific and Technical Information of China (English)

    Faridah Yusof; Atheer Awad Mehde; Wesen Adel Mehdi; Raha Ahmed Raus; Hamid Ghazali; Azlina Abd Rahman

    2015-01-01

    Objective Nephrolithiasis is one of the most common disorders of the urinary tract. The aim of this study was to examine a possible relationship between DNase I/II activity and E3 SUMO-protein ligase NSE2 in the sera of nephrolithiasis patients to evaluate the possibility of a new biomarker for evaluating kidney damage. Methods Sixty nephrolithiasis patients and 50 control patients were enrolled in a case-control study. Their blood urea, creatinine, protein levels and DNase I/II activity levels were measured by spectrometry. Serum NSMCE2 levels were measured by ELISA. Blood was collected from patients of the government health clinics in Kuantan-Pahang and fulfilled the inclusion criteria. Results The result indicated that mean levels of sera NSMCE2 have a significantly increase (P Conclusion This study suggests that an increase in serum concentrations of DNase I/II and E3 SUMO-protein ligase NSE2 level can be used as indicators for the diagnosis of kidney injury in patients with nephrolithiasis.

  7. Sgs1's roles in DNA end resection, HJ dissolution, and crossover suppression require a two-step SUMO regulation dependent on Smc5/6.

    Science.gov (United States)

    Bermúdez-López, Marcelino; Villoria, María Teresa; Esteras, Miguel; Jarmuz, Adam; Torres-Rosell, Jordi; Clemente-Blanco, Andres; Aragon, Luis

    2016-06-01

    The RecQ helicase Sgs1 plays critical roles during DNA repair by homologous recombination, from end resection to Holliday junction (HJ) dissolution. Sgs1 has both pro- and anti-recombinogenic roles, and therefore its activity must be tightly regulated. However, the controls involved in recruitment and activation of Sgs1 at damaged sites are unknown. Here we show a two-step role for Smc5/6 in recruiting and activating Sgs1 through SUMOylation. First, auto-SUMOylation of Smc5/6 subunits leads to recruitment of Sgs1 as part of the STR (Sgs1-Top3-Rmi1) complex, mediated by two SUMO-interacting motifs (SIMs) on Sgs1 that specifically recognize SUMOylated Smc5/6. Second, Smc5/6-dependent SUMOylation of Sgs1 and Top3 is required for the efficient function of STR. Sgs1 mutants impaired in recognition of SUMOylated Smc5/6 (sgs1-SIMΔ) or SUMO-dead alleles (sgs1-KR) exhibit unprocessed HJs at damaged replication forks, increased crossover frequencies during double-strand break repair, and severe impairment in DNA end resection. Smc5/6 is a key regulator of Sgs1's recombination functions.

  8. Effects of Nephrolithiasis on Serum DNase (Deoxyribonuclease I and II) Activity and E3 SUMO-Protein Ligase NSE2 (NSMCE2) in Malaysian Individuals.

    Science.gov (United States)

    Yusof, Faridah; Mehde, Atheer Awad; Mehdi, Wesen Adel; Raus, Raha Ahmed; Ghazali, Hamid; Rahman, Azlina Abd

    2015-09-01

    Nephrolithiasis is one of the most common disorders of the urinary tract. The aim of this study was to examine a possible relationship between DNase I/II activity and E3 SUMO-protein ligase NSE2 in the sera of nephrolithiasis patients to evaluate the possibility of a new biomarker for evaluating kidney damage. Sixty nephrolithiasis patients and 50 control patients were enrolled in a case-control study. Their blood urea, creatinine, protein levels and DNase I/II activity levels were measured by spectrometry. Serum NSMCE2 levels were measured by ELISA. Blood was collected from patients of the government health clinics in Kuantan-Pahang and fulfilled the inclusion criteria. The result indicated that mean levels of sera NSMCE2 have a significantly increase (P<0.01) in patients compared to control group. Compared with control subjects, activities and specific activities of serum DNase I and II were significantly elevated in nephrolithiasis patients (P$lt;0.01). This study suggests that an increase in serum concentrations of DNase I/II and E3 SUMO-protein ligase NSE2 level can be used as indicators for the diagnosis of kidney injury in patients with nephrolithiasis. Copyright © 2015 The Editorial Board of Biomedical and Environmental Sciences. Published by China CDC. All rights reserved.

  9. A Compact P⁺ Contact Resistance Model for Characterization of Substrate Coupling in Modern Lightly Doped CMOS Processes

    DEFF Research Database (Denmark)

    Shen, Ming; Mikkelsen, Jan H.; Jensen, Ole Kiel;

    2012-01-01

    lightly-doped CMOS processes with P-well layers are still unavailable. This paper presents a new compact resistance model aiming at solving this problem. A Conformal Mapping(CM) method was used to derive the closed-form expressions for the resistances in the model. The model requires no fitting factors...

  10. Analytical modeling of the lattice and thermo-elastic coefficient mismatch-induced stress into silicon nanowires horizontally embedded on insulator-on-silicon substrates

    Science.gov (United States)

    Chatterjee, Sulagna; Chattopadhyay, Sanatan

    2017-01-01

    In the current work, an analytical model has been developed to estimate the amount of induced stress in nanowires which are horizontally embedded with different fractions within an Insulator-on-Silicon substrate. For estimating such stress, different crystallographic orientations of substrates and embedded nanowires have been considered. The induced stress for both the difference in thermo-elastic constants and lattice-mismatch is included and accuracy of the analytical model has been verified with the similar results obtained from ANSYS Multiphysics. Induced stress is observed to be insensitive of the nanowire size, however, depends significantly on the fractional insertion of the nanowires. A tensile stress of 1.95 GPa and a compressive stress of -1.0719 GPa have been obtained for the oriented Si-nanowires. Hole mobility of 850 cm2/Vs can be achieved for the 3/4th insertion of the nanowires which is comparable to electron mobility and therefore can be utilized for the design of symmetric nano-electronic devices.

  11. Signal sensitivity of alternating current potential drop measurement for crack detection of conductive substrate with tunable coating materials through finite element modeling

    Science.gov (United States)

    Sandeep Rao, Simha; Liu, Ming; Peng, Fei; Zhang, Bo; Zhao, Huijuan

    2016-12-01

    We adopt a finite element numerical modeling approach to investigate the electromagnetic coupling effect of two parallel electric conductors with tunable electric conductivity σ and magnetic permeability μ. For two parallel conductors C and S (μ C   ṡ  σ C   ≤  μ S   ṡ  σ S), we find that the shape of current density profile of conductor S is dependent on the product of μ C   ṡ  σ C, while the magnitude is determined by the AC current frequency f. On the other hand, the frequency f affects not only the shape but also the magnitude of the current density profile of conductor C. We further adopt a coplanar model to investigate the signal sensitivity of alternating current potential drop (ACPD) measurement for both surface crack and inner crack detection. We find that with modified coating materials (lower electric conductivity and higher magnetic permeability, compared with the substrate material properties), the crack detection signal sensitivity can be greatly enhanced for both the cracks within the coating and at the coating/substrate interface, where cracks are most commonly encountered in real situations.

  12. Substrate specificity of pyrimidine nucleoside phosphorylases of NP-II family probed by X-ray crystallography and molecular modeling

    Science.gov (United States)

    Balaev, V. V.; Lashkov, A. A.; Prokofev, I. I.; Gabdulkhakov, A. G.; Seregina, T. A.; Mironov, A. S.; Betzel, C.; Mikhailov, A. M.

    2016-09-01

    Pyrimidine nucleoside phosphorylases, which are widely used in the biotechnological production of nucleosides, have different substrate specificity for pyrimidine nucleosides. An interesting feature of these enzymes is that the three-dimensional structure of thymidine-specific nucleoside phosphorylase is similar to the structure of nonspecific pyrimidine nucleoside phosphorylase. The three-dimensional structures of thymidine phosphorylase from Salmonella typhimurium and nonspecific pyrimidine nucleoside phosphorylase from Bacillus subtilis in complexes with a sulfate anion were determined for the first time by X-ray crystallography. An analysis of the structural differences between these enzymes demonstrated that Lys108, which is involved in the phosphate binding in pyrimidine nucleoside phosphorylase, corresponds to Met111 in thymidine phosphorylases. This difference results in a decrease in the charge on one of the hydroxyl oxygens of the phosphate anion in thymidine phosphorylase and facilitates the catalysis through SN2 nucleophilic substitution. Based on the results of X-ray crystallography, the virtual screening was performed for identifying a potent inhibitor (anticancer agent) of nonspecific pyrimidine nucleoside phosphorylase, which does not bind to thymidine phosphorylase. The molecular dynamics simulation revealed the stable binding of the discovered compound—2-pyrimidin-2-yl-1H-imidazole-4-carboxylic acid—to the active site of pyrimidine nucleoside phosphorylase.

  13. Hutchinson-Gilford progeria syndrome alters nuclear shape and reduces cell motility in three dimensional model substrates.

    Science.gov (United States)

    Booth-Gauthier, Elizabeth A; Du, Vicard; Ghibaudo, Marion; Rape, Andrew D; Dahl, Kris Noel; Ladoux, Benoit

    2013-03-01

    Cell migration through tight interstitial spaces in three dimensional (3D) environments impacts development, wound healing and cancer metastasis and is altered by the aging process. The stiffness of the extracellular matrix (ECM) increases with aging and affects the cells and cytoskeletal processes involved in cell migration. However, the nucleus, which is the largest and densest organelle, has not been widely studied during cell migration through the ECM. Additionally, the nucleus is stiffened during the aging process through the accumulation of a mutant nucleoskeleton protein lamin A, progerin. By using microfabricated substrates to mimic the confined environment of surrounding tissues, we characterized nuclear movements and deformation during cell migration into micropillars where interspacing can be tuned to vary nuclear confinement. Cell motility decreased with decreased micropillar (μP) spacing and correlated with increased dysmorphic shapes of nuclei. We examined the effects of increased nuclear stiffness which correlates with cellular aging by studying Hutchinson-Gilford progeria syndrome cells which are known to accumulate progerin. With the expression of progerin, cells showed a threshold response to decreased μP spacing. Cells became trapped in the close spacing, possibly from visible micro-defects in the nucleoskeleton induced by cell crawling through the μP and from reduced force generation, measured independently. We suggest that ECM changes during aging could be compounded by the increasing stiffness of the nucleus and thus changes in cell migration through 3D tissues.

  14. Substrate compositional variation with tissue/region and Gba1 mutations in mouse models--implications for Gaucher disease.

    Directory of Open Access Journals (Sweden)

    Ying Sun

    Full Text Available Gaucher disease results from GBA1 mutations that lead to defective acid β-glucosidase (GCase mediated cleavage of glucosylceramide (GC and glucosylsphingosine as well as heterogeneous manifestations in the viscera and CNS. The mutation, tissue, and age-dependent accumulations of different GC species were characterized in mice with Gba1 missense mutations alone or in combination with isolated saposin C deficiency (C*. Gba1 heteroallelism for D409V and null alleles (9V/null led to GC excesses primarily in the visceral tissues with preferential accumulations of lung GC24∶0, but not in liver, spleen, or brain. Age-dependent increases of different GC species were observed. The combined saposin C deficiency (C* with V394L homozygosity (4L;C* showed major GC18:0 degradation defects in the brain, whereas the analogous mice with D409H homozygosity and C* (9H;C* led to all GC species accumulating in visceral tissues. Glucosylsphingosine was poorly degraded in brain by V394L and D409H GCases and in visceral tissues by D409V GCase. The neonatal lethal N370S/N370S genotype had insignificant substrate accumulations in any tissue. These results demonstrate age, organ, and mutation-specific quantitative differences in GC species and glucosylsphingosine accumulations that can have influence in the tissue/regional expression of Gaucher disease phenotypes.

  15. [Determination of the parameters for producing a biobinder from wood: a mathematical modeling of the transformation of lignocellulose substrate by the fungus Panus tigrinus].

    Science.gov (United States)

    Kondrashchenko, V I; Manukovskiĭ, N S; Kovalev, V S

    2006-01-01

    A biochemical scheme for the transformation of wood lignocellulose during enzymatic hydrolysis of polysaccharides and lignin destruction in reactions involving free radicals was developed, and a corresponding mathematical model was constructed. Processing (fermentation) of wood particles by the fungus Panus tigrinus in a submerged culture for producing a biobinder of wood composites--woodchip boards and fiber-boards--is considered. The mathematical model was used to study the technological parameters that influence the production of enzymes and fungal biomass and the level of free radical accumulation in the substrate, i.e., the factors determining the production of the biobinder. The optimal values of these parameters were determined, namely: the specific surface of wood particles, amounting to 2000 cm2/g; processing time of 56 h; and an initial concentration of 3.0 g/l of fungal biomass in the submerged culture.

  16. Modeling Geometric Arrangements of TiO2-Based Catalyst Substrates and Isotropic Light Sources to Enhance the Efficiency of a Photocatalystic Oxidation (PCO) Reactor

    Science.gov (United States)

    Richards, Jeffrey T.; Levine, Lanfang H.; Husk, Geoffrey K.

    2011-01-01

    The closed confined environments of the ISS, as well as in future spacecraft for exploration beyond LEO, provide many challenges to crew health. One such challenge is the availability of a robust, energy efficient, and re-generable air revitalization system that controls trace volatile organic contaminants (VOCs) to levels below a specified spacecraft maximum allowable concentration (SMAC). Photocatalytic oxidation (PCO), which is capable of mineralizing VOCs at room temperature and of accommodating a high volumetric flow, is being evaluated as an alternative trace contaminant control technology. In an architecture of a combined air and water management system, placing a PCO unit before a condensing heat exchanger for humidity control will greatly reduce the organic load into the humidity condensate loop ofthe water processing assembly (WPA) thereby enhancing the life cycle economics ofthe WPA. This targeted application dictates a single pass efficiency of greater than 90% for polar VOCs. Although this target was met in laboratory bench-scaled reactors, no commercial or SBIR-developed prototype PCO units examined to date have achieved this goal. Furthermore, the formation of partial oxidation products (e.g., acetaldehyde) was not eliminated. It is known that single pass efficiency and partial oxidation are strongly dependent upon the contact time and catalyst illumination, hence the requirement for an efficient reactor design. The objective of this study is to maximize the apparent contact time and illuminated catalyst surface area at a given reactor volume and volumetric flow. In this study, a Ti02-based photocatalyst is assumed to be immobilized on porous substrate panels and illumination derived from linear isotropic light sources. Mathematical modeling using computational fluid dynamics (CFD) analyses were performed to investigate the effect of: 1) the geometry and configuration of catalyst-coated substrate panels, 2) porosity of the supporting substrate, and 3

  17. Neuroanatomical and cellular substrates of hypergrooming induced by microinjection of oxytocin in central nucleus of amygdala, an experimental model of compulsive behavior.

    Science.gov (United States)

    Marroni, S S; Nakano, F N; Gati, C D C; Oliveira, J A C; Antunes-Rodrigues, J; Garcia-Cairasco, N

    2007-12-01

    Oxytocin (OT) is a neurosecretory nonapeptide synthesized in hypothalamic cells that project to the neurohypophysis as well as to widely distributed sites in the central nervous system. Central OT microinjections induce a variety of cognitive, sexual, reproductive, grooming and affiliative behaviors in animals. Obsessive-compulsive disorder (OCD) includes a range of cognitive and behavioral symptoms that bear some relationship with OT. Here, we study the neuroanatomical and cellular substrates of the hypergrooming induced by administration of OT in the central nucleus of amygdala (CeA). In this context, this hypergrooming is considered as a model of compulsive behavior. Our data suggest a link between the CeA and the hypothalamic grooming area (HGA). The HGA includes parts of the paraventricular nucleus and the dorsal hypothalamic area. Our data on colocalization of OT (immunohistochemistry for peptide), OT receptor (binding assay) and its retrogradely labeled cells after Fluoro-Gold injection in the CeA suggest that CeA and connections are important substrates of the circuit underlying this OT-dependent compulsive behavioral pattern.

  18. Adding mucins to an in vitro batch fermentation model of the large intestine induces changes in microbial population isolated from porcine feces depending on the substrate.

    Science.gov (United States)

    Tran, T H T; Boudry, C; Everaert, N; Théwis, A; Portetelle, D; Daube, G; Nezer, C; Taminiau, B; Bindelle, J

    2016-02-01

    Adding mucus to in vitro fermentation models of the large intestine shows that some genera, namely lactobacilli, are dependent on host-microbiota interactions and that they rely on mucosal layers to increase their activity. This study investigated whether this dependence on mucus is substrate dependent and to what extent other genera are impacted by the presence of mucus. Inulin and cellulose were fermented in vitro by a fecal inoculum from pig in the presence or not of mucin beads in order to compare fermentation patterns and bacterial communities. Mucins increased final gas production with inulin and shifted short-chain fatty acid molar ratios (P inulin was fermented. A more in-depth community analysis indicated that the mucins increased Proteobacteria (0.55 vs 0.25%, P = 0.013), Verrucomicrobia (5.25 vs 0.03%, P = 0.032), Ruminococcaceae, Bacteroidaceae and Akkermansia spp. Proteobacteria (5.67 vs 0.55%, P < 0.001) and Lachnospiraceae (33 vs 10.4%) were promoted in the mucus compared with the broth, while Ruminococcaceae decreased. The introduction of mucins affected many microbial genera and fermentation patterns, but from PCA results, the impact of mucus was independent of the fermentation substrate.

  19. Iron deposition following chronic myocardial infarction as a substrate for cardiac electrical anomalies: initial findings in a canine model.

    Directory of Open Access Journals (Sweden)

    Ivan Cokic

    evolve as an arrhythmogenic substrate in the post MI period.

  20. Real-time quantitative monitoring of hiPSC-based model of macular degeneration on Electric Cell-substrate Impedance Sensing microelectrodes.

    Science.gov (United States)

    Gamal, W; Borooah, S; Smith, S; Underwood, I; Srsen, V; Chandran, S; Bagnaninchi, P O; Dhillon, B

    2015-09-15

    Age-related macular degeneration (AMD) is the leading cause of blindness in the developed world. Humanized disease models are required to develop new therapies for currently incurable forms of AMD. In this work, a tissue-on-a-chip approach was developed through combining human induced pluripotent stem cells, Electric Cell-substrate Impedance Sensing (ECIS) and reproducible electrical wounding assays to model and quantitatively study AMD. Retinal Pigment Epithelium (RPE) cells generated from a patient with an inherited macular degeneration and from an unaffected sibling were used to test the model platform on which a reproducible electrical wounding assay was conducted to model RPE damage. First, a robust and reproducible real-time quantitative monitoring over a 25-day period demonstrated the establishment and maturation of RPE layers on the microelectrode arrays. A spatially controlled RPE layer damage that mimicked cell loss in AMD disease was then initiated. Post recovery, significant differences (P model-on-a-chip is a powerful platform for translational studies with considerable potential to investigate novel therapies by enabling real-time, quantitative and reproducible patient-specific RPE cell repair studies. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.

  1. Evaluation of nitrogenous substrates such as peptones from fish:a new method based on Gompertz modeling of microbial growth.

    Science.gov (United States)

    Dufossé, L; De La Broise, D; Guerard, F

    2001-01-01

    Fish peptones from tuna, cod, salmon, and unspecified fish were compared with a casein one by using a new method based on Gompertz modeling of microbial growth. Cumulative results obtained from six species of bacteria, yeasts, and fungi showed that, in most cases, these fish peptones are very effective. Nevertheless, this study raised some questions about the standardization of fish raw material, the enzymatic hydrolysis of fish proteins, and the composition of the culture medium used for testing the peptones.

  2. Numerical modelling of sliding wear caused by pin-on-disk method over copper coated ABS plastic substrate

    Science.gov (United States)

    Nigam, S.; Mahapatra, S. S.; Patel, S. K.

    2016-09-01

    The coating of metal increases the face value of the plastic and inhibits other properties like conductivity, hardness and lustre. Thus the combination of plastic and metal coating results in a material that is light in weight because of the presence of plastic as the base material and; electrical and thermal conductive because of the presence of metal on the surface. The requirement of such materials is growing day by day. Copper coated plastic has various applications such as in fabrication of printed circuit boards (PCB's) and various automobile parts and in electromagnetic interference shielding. It is important to analyse the tribological aspect of the same in order to broaden its range of application. The present work contains 3D modelling of thermally sprayed copper on ABS plastic and simulation of sliding wear test by pin-on-disc method. The Johnson cook flow stress model is selected for the coating material. Archard's wear model has provided the best results for calculating the wear rate. The results obtained are in good agreement with the experimental values.

  3. Dewetting on microstructured substrates

    Science.gov (United States)

    Kim, Taehong; Kim, Wonjung

    2016-11-01

    A thin liquid film has an equilibrium thickness in such a way as to minimize the free energy. When a liquid film thickness is out of its equilibrium, the film seeks its equilibrium state, resulting in dynamics of liquid film, which are referred to as wetting and dewetting, depending on the flow direction. We here present a combined experimental and theoretical investigation of dewetting on a substrate with parallel microstructures. Our experiments show that residue may remain on the substrate after dewetting, and residue morphologies can be classified into three modes. Based on our experimental observations, we elucidate how the modes depend on the pattern morphology and contact angle, and develop a model for the contact line motion. Our results provide a basis for controlling the thickness film, which is important for many practical applications such as oil recovery, detergency, lithography, and cleaning. This work was supported by the National Research Foundation of Korea (NRF) Grant funded by the Korea government (MSIP) (No.2015R1A2A2A04006181).

  4. Discovery of 1-[4-(N-benzylamino)phenyl]-3-phenylurea derivatives as non-peptidic selective SUMO-sentrin specific protease (SENP)1 inhibitors.

    Science.gov (United States)

    Uno, Masaharu; Koma, Yosuke; Ban, Hyun Seung; Nakamura, Hiroyuki

    2012-08-15

    We developed 1-[4-(N-benzylamino)phenyl]-3-phenylurea derivative 4 (GN6958) as a non-peptidic selective SUMO-sentrin specific protease (SENP)1 protease inhibitor based on the hypoxia inducible factor (HIF)-1α inhibitor 1 (GN6767). The direct interaction of compound 1 with SENP1 protein in cells was observed by the pull-down experiments using the biotin-tagged compound 2 coated on the streptavidin affinity column. Among the various 1-[4-(N-benzylamino)phenyl]-3-phenylurea derivatives tested, compounds 3 and 4 suppressed HIF-1α accumulation in a concentration-dependent manner without affecting the expression level of tubulin protein in HeLa cells. Both compounds inhibited SENP1 protease activity in a concentration-dependent manner, and compound 4 exhibited more potent inhibition than compound 3. Compound 4 exhibited selective inhibition against SENP1 protease activity without inhibiting other protease enzyme activities in vitro.

  5. Papel de SUMO en la regulación de PKR y la vía molecular PI3K/AKT

    OpenAIRE

    2015-01-01

    Tesis Doctoral inédita leída en la Universidad Autónoma de Madrid, Facultad de Ciencias, Departamento de Biología Molecular. Fecha de lectura: 17-07-2015 La SUMOilación es una modificación post-traduccional que consiste en que las proteínas SUMO se conjugan a sus proteínas diana. Esta modificación regula proteínas celulares involucradas en múltiples procesos como transcripción, replicación, reparación del ADN, apoptosis, senescencia, transporte nuclear y transducción de señales. De esta fo...

  6. Homo-timeric structural model of human microsomal prostaglandin E synthase-1 and characterization of its substrate/inhibitor binding interactions

    Science.gov (United States)

    Xing, Li; Kurumbail, Ravi G.; Frazier, Ronald B.; Davies, Michael S.; Fujiwara, Hideji; Weinberg, Robin A.; Gierse, James K.; Caspers, Nicole; Carter, Jeffrey S.; McDonald, Joseph J.; Moore, William M.; Vazquez, Michael L.

    2009-01-01

    Inducible, microsomal prostaglandin E synthase 1 (mPGES-1), the terminal enzyme in the prostaglandin (PG) biosynthetic pathway, constitutes a promising therapeutic target for the development of new anti-inflammatory drugs. To elucidate structure-function relationships and to enable structure-based design, an mPGES-1 homology model was developed using the three-dimensional structure of the closest homologue of the MAPEG family (Membrane Associated Proteins in Eicosanoid and Glutathione metabolism), mGST-1. The ensuing model of mPGES-1 is a homo-trimer, with each monomer consisting of four membrane-spanning segments. Extensive structure refinement revealed an inter-monomer salt bridge (K26-E77) as well as inter-helical interactions within each monomer, including polar hydrogen bonds (e.g. T78-R110-T129) and hydrophobic π-stacking (F82-F103-F106), all contributing to the overall stability of the homo-trimer of mPGES-1. Catalytic co-factor glutathione (GSH) was docked into the mPGES-1 model by flexible optimization of both the ligand and the protein conformations, starting from the initial location ascertained from the mGST-1 structure. Possible binding site for the substrate, prostaglandin H2 (PGH2), was identified by systematically probing the refined molecular structure of mPGES-1. A binding model was generated by induced fit docking of PGH2 in the presence of GSH. The homology model prescribes three potential inhibitor binding sites per mPGES-1 trimer. This was further confirmed experimentally by equilibrium dialysis study which generated a binding stoichiometric ratio of approximately three inhibitor molecules to three mPGES-1 monomers. The structural model that we have derived could serve as a useful tool for structure-guided design of inhibitors for this emergently important therapeutic target.

  7. Effect of substrate preparation on the structure and chemisorption properties of Pd/MgO model catalyst

    Science.gov (United States)

    Henry, C. R.; Poppa, H.

    1988-01-01

    Pd/MgO model catalysts are prepared by vapor deposition of Pd particles on MgO thin films which are deposited, at different temperatures, on UHV cleaved mica. MgO films and Pd particles are characterized in situ by Auger electron spectroscopy and energy-loss spectroscopy and ex situ by transmission electron microscopy. Succesive temperature programmed desorption (TPD) of CO shows a different evolution of the morphology of the Pd particles deposited on MgO films prepared at low and high temperature. In addition, on MgO prepared at low temperature, the smallest particles show a continuous decay of CO adsorption during TPD due to contamination by support species.

  8. AMPK activation through mitochondrial regulation results in increased substrate oxidation and improved metabolic parameters in models of diabetes.

    Directory of Open Access Journals (Sweden)

    Yonchu Jenkins

    Full Text Available Modulation of mitochondrial function through inhibiting respiratory complex I activates a key sensor of cellular energy status, the 5'-AMP-activated protein kinase (AMPK. Activation of AMPK results in the mobilization of nutrient uptake and catabolism for mitochondrial ATP generation to restore energy homeostasis. How these nutrient pathways are affected in the presence of a potent modulator of mitochondrial function and the role of AMPK activation in these effects remain unclear. We have identified a molecule, named R419, that activates AMPK in vitro via complex I inhibition at much lower concentrations than metformin (IC50 100 nM vs 27 mM, respectively. R419 potently increased myocyte glucose uptake that was dependent on AMPK activation, while its ability to suppress hepatic glucose production in vitro was not. In addition, R419 treatment of mouse primary hepatocytes increased fatty acid oxidation and inhibited lipogenesis in an AMPK-dependent fashion. We have performed an extensive metabolic characterization of its effects in the db/db mouse diabetes model. In vivo metabolite profiling of R419-treated db/db mice showed a clear upregulation of fatty acid oxidation and catabolism of branched chain amino acids. Additionally, analyses performed using both (13C-palmitate and (13C-glucose tracers revealed that R419 induces complete oxidation of both glucose and palmitate to CO2 in skeletal muscle, liver, and adipose tissue, confirming that the compound increases mitochondrial function in vivo. Taken together, our results show that R419 is a potent inhibitor of complex I and modulates mitochondrial function in vitro and in diabetic animals in vivo. R419 may serve as a valuable molecular tool for investigating the impact of modulating mitochondrial function on nutrient metabolism in multiple tissues and on glucose and lipid homeostasis in diabetic animal models.

  9. Substrate recognition and motion mode analyses of PFV integrase in complex with viral DNA via coarse-grained models.

    Directory of Open Access Journals (Sweden)

    Jianping Hu

    Full Text Available HIV-1 integrase (IN is an important target in the development of drugs against the AIDS virus. Drug design based on the structure of IN was markedly hampered due to the lack of three-dimensional structure information of HIV-1 IN-viral DNA complex. The prototype foamy virus (PFV IN has a highly functional and structural homology with HIV-1 IN. Recently, the X-ray crystal complex structure of PFV IN with its cognate viral DNA has been obtained. In this study, both Gaussian network model (GNM and anisotropy network model (ANM have been applied to comparatively investigate the motion modes of PFV DNA-free and DNA-bound IN. The results show that the motion mode of PFV IN has only a slight change after binding with DNA. The motion of this enzyme is in favor of association with DNA, and the binding ability is determined by its intrinsic structural topology. Molecular docking experiments were performed to gain the binding modes of a series of diketo acid (DKA inhibitors with PFV IN obtained from ANM, from which the dependability of PFV IN-DNA used in the drug screen for strand transfer (ST inhibitors was confirmed. It is also found that the functional groups of keto-enol, bis-diketo, tetrazole and azido play a key role in aiding the recognition of viral DNA, and thus finally increase the inhibition capability for the corresponding DKA inhibitor. Our study provides some theoretical information and helps to design anti-AIDS drug based on the structure of IN.

  10. Computational modeling analysis of mitochondrial superoxide production under varying substrate conditions and upon inhibition of different segments of the electron transport chain.

    Science.gov (United States)

    Markevich, Nikolai I; Hoek, Jan B

    2015-01-01

    A computational mechanistic model of superoxide (O2•-) formation in the mitochondrial electron transport chain (ETC) was developed to facilitate the quantitative analysis of factors controlling mitochondrial O2•- production and assist in the interpretation of experimental studies. The model takes into account all individual electron transfer reactions in Complexes I and III. The model accounts for multiple, often seemingly contradictory observations on the effects of ΔΨ and ΔpH, and for the effects of multiple substrate and inhibitor conditions, including differential effects of Complex III inhibitors antimycin A, myxothiazol and stigmatellin. Simulation results confirm that, in addition to O2•- formation in Complex III and at the flavin site of Complex I, the quinone binding site of Complex I is an additional superoxide generating site that accounts for experimental observations on O2•- production during reverse electron transfer. However, our simulation results predict that, when cytochrome c oxidase is inhibited during oxidation of succinate, ROS production at this site is eliminated and almost all superoxide in Complex I is generated by reduced FMN, even when the redox pressure for reverse electron transfer from succinate is strong. In addition, the model indicates that conflicting literature data on the kinetics of electron transfer in Complex III involving the iron-sulfur protein-cytochrome bL complex can be resolved in favor of a dissociation of the protein only after electron transfer to cytochrome bH. The model predictions can be helpful in understanding factors driving mitochondrial superoxide formation in intact cells and tissues.

  11. P-glycoprotein Mediated Efflux Limits Substrate and Drug Uptake in a Preclinical Brain Metastases of Breast Cancer Model

    Directory of Open Access Journals (Sweden)

    Chris E Adkins

    2013-11-01

    Full Text Available The blood-brain barrier (BBB is a specialized vascular interface that restricts the entry of many compounds into brain. This is accomplished through the sealing of vascular endothelial cells together with tight junction proteins to prevent paracellular diffusion. In addition, the BBB has a high degree of expression of numerous efflux transporters which actively extrude compounds back into blood. However, when a metastatic lesion develops in brain the vasculature is typically compromised with increases in passive permeability (blood-tumor barrier; BTB. What is not well documented is to what degree active efflux retains function at the BTB despite the changes observed in passive permeability. In addition, there have been previous reports documenting both increased and decreased expression of P-gp in lesion vasculature. Herein, we simultaneously administer a passive diffusion marker (14C-AIB and a tracer subject to P-gp efflux (rhodamine 123 into a murine preclinical model of brain metastases of breast cancer. We observed that the metastatic lesions had similar expression (p>0.05; n=756-1214 vessels evaluated at the BBB and the BTB. Moreover, tissue distribution of R123 was not significantly (p>0.05 different between normal brain and the metastatic lesion. It is possible that the similar expression of P-gp on the BBB and the BTB contribute to this phenomenon. Additionally we observed P-gp expression at the metastatic cancer cells adjacent to the vasculature which may also contribute to reduced R123 uptake into the lesion. The data suggest that despite the disrupted integrity of the BTB, efflux mechanisms appear to be intact, and may be functionally comparable to the normal BBB. The BTB is a significant hurdle to delivering drugs to brain metastasis.

  12. Aqueous-Phase Preparation of Model HDS Catalysts on Planar Alumina Substrates: Support Effect on Mo Adsorption and Sulfidation.

    Science.gov (United States)

    Bara, Cédric; Plais, Lucie; Larmier, Kim; Devers, Elodie; Digne, Mathieu; Lamic-Humblot, Anne-Félicie; Pirngruber, Gerhard D; Carrier, Xavier

    2015-12-23

    The role of the oxide support on the structure of the MoS2 active phase (size, morphology, orientation, sulfidation ratio, etc.) remains an open question in hydrotreating catalysis and biomass processing with important industrial implications for the design of improved catalytic formulations. The present work builds on an aqueous-phase surface-science approach using four well-defined α-alumina single crystal surfaces (C (0001), A (112̅0), M (101̅0), and R (11̅02) planes) as surrogates for γ-alumina (the industrial support) in order to discriminate the specific role of individual support facets. The reactivity of the various surface orientations toward molybdenum adsorption is controlled by the speciation of surface hydroxyls that determines the surface charge at the oxide/water interface. The C (0001) plane is inert, and the R (11̅02) plane has a limited Mo adsorption capacity while the A (112̅0) and M (101̅0) surfaces are highly reactive. Sulfidation of model catalysts reveals the highest sulfidation degree for the A (112̅0) and M (101̅0) planes suggesting weak metal/support interactions. Conversely, a low sulfidation rate and shorter MoS2 slabs are found for the R (11̅02) plane implying stronger Mo-O-Al bonds. These limiting cases are reminiscent of type I/type II MoS2 nanostructures. Structural analogies between α- and γ- alumina surfaces allow us to bridge the material gap with real Al2O3-supported catalysts. Hence, it can be proposed that Mo distribution and sulfidation rate are heterogeneous and surface-dependent on industrial γ-Al2O3-supported high-surface-area catalysts. These results demonstrate that a proper control of the γ-alumina morphology is a strategic lever for a molecular-scale design of hydrotreating catalysts.

  13. In vitro generation of three-dimensional substrate-adherent embryonic stem cell-derived neural aggregates for application in animal models of neurological disorders.

    Science.gov (United States)

    Hargus, Gunnar; Cui, Yi-Fang; Dihné, Marcel; Bernreuther, Christian; Schachner, Melitta

    2012-05-01

    In vitro-differentiated embryonic stem (ES) cells comprise a useful source for cell replacement therapy, but the efficiency and safety of a translational approach are highly dependent on optimized protocols for directed differentiation of ES cells into the desired cell types in vitro. Furthermore, the transplantation of three-dimensional ES cell-derived structures instead of a single-cell suspension may improve graft survival and function by providing a beneficial microenvironment for implanted cells. To this end, we have developed a new method to efficiently differentiate mouse ES cells into neural aggregates that consist predominantly (>90%) of postmitotic neurons, neural progenitor cells, and radial glia-like cells. When transplanted into the excitotoxically lesioned striatum of adult mice, these substrate-adherent embryonic stem cell-derived neural aggregates (SENAs) showed significant advantages over transplanted single-cell suspensions of ES cell-derived neural cells, including improved survival of GABAergic neurons, increased cell migration, and significantly decreased risk of teratoma formation. Furthermore, SENAs mediated functional improvement after transplantation into animal models of Parkinson's disease and spinal cord injury. This unit describes in detail how SENAs are efficiently derived from mouse ES cells in vitro and how SENAs are isolated for transplantation. Furthermore, methods are presented for successful implantation of SENAs into animal models of Huntington's disease, Parkinson's disease, and spinal cord injury to study the effects of stem cell-derived neural aggregates in a disease context in vivo.

  14. Impact of bacterial activity on turnover of insoluble hydrophobic substrates (phenanthrene and pyrene)-Model simulations for prediction of bioremediation success.

    Science.gov (United States)

    Rein, Arno; Adam, Iris K U; Miltner, Anja; Brumme, Katja; Kästner, Matthias; Trapp, Stefan

    2016-04-05

    Many attempts for bioremediation of polycyclic aromatic hydrocarbon (PAH) contaminated sites failed in the past, but the reasons for this failure are not well understood. Here we apply and improve a model for integrated assessment of mass transfer, biodegradation and residual concentrations for predicting the success of remediation actions. First, we provide growth parameters for Mycobacterium rutilum and Mycobacterium pallens growing on phenanthrene (PHE) or pyrene (PYR) degraded the PAH completely at all investigated concentrations. Maximum metabolic rates vmax and growth rates μ were similar for the substrates PHE and PYR and for both strains. The investigated Mycobacterium species were not superior in PHE degradation to strains investigated earlier with this method. Real-world degradation scenario simulations including diffusive flux to the microbial cells indicate: that (i) bioaugmentation only has a small, short-lived effect; (ii) Increasing sorption shifts the remaining PAH to the adsorbed/sequestered PAH pool; (iii) mobilizing by solvents or surfactants resulted in a significant decrease of the sequestered PAH, and (iv) co-metabolization e.g. by compost addition can contribute significantly to the reduction of PAH, because active biomass is maintained at a high level by the compost. The model therefore is a valuable contribution to the assessment of potential remediation action at PAH-polluted sites.

  15. Modeling of photodegradation in solar cell modules of substrate and superstrate design made with ethylene-vinyl acetate as pottant material

    Science.gov (United States)

    Somersall, A. C.; Guillet, J. E.

    1981-01-01

    The rates of hydrogen abstraction by peroxy radicals were proven to be too slow for significant oxidation of the alkane substrate to be important. The numerical procedure, independent of our particular data base was verified by reproducing concentration time profiles for a model reaction set describing the cesium flare system in the upper atmosphere. Simulation was identical to that given in the literature. Experimental verification of the data base is to be attempted by weatherometry studies in the coming year. Work on the new diagnostic techniques was completed. The adapted automated viscometer was demonstrated to be an efficient and reliable tool for routine measurements of viscosity (molecular weight) changes in solid samples after batch solutions have been made up. The laser photolysis GC method for monitoring extremely low levels of oxidation in polymers proved to be impractical because the yields of carbon monoxide were too low for quantification. Much progress was made with the computer model. The reaction matrix was completely revised, resulting in a new scheme of 31 reactions and time, lifetimes in excess of ten years. The results to date lead us to some tentative observations.

  16. Substrate curvature regulates cell migration

    Science.gov (United States)

    He, Xiuxiu; Jiang, Yi

    2017-06-01

    Cell migration is essential in many aspects of biology. Many basic migration processes, including adhesion, membrane protrusion and tension, cytoskeletal polymerization, and contraction, have to act in concert to regulate cell migration. At the same time, substrate topography modulates these processes. In this work, we study how substrate curvature at micrometer scale regulates cell motility. We have developed a 3D mechanical model of single cell migration and simulated migration on curved substrates with different curvatures. The simulation results show that cell migration is more persistent on concave surfaces than on convex surfaces. We have further calculated analytically the cell shape and protrusion force for cells on curved substrates. We have shown that while cells spread out more on convex surfaces than on concave ones, the protrusion force magnitude in the direction of migration is larger on concave surfaces than on convex ones. These results offer a novel biomechanical explanation to substrate curvature regulation of cell migration: geometric constrains bias the direction of the protrusion force and facilitates persistent migration on concave surfaces.

  17. Nanowires and nanoneedles nucleation on vicinal substrate

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Xu, E-mail: zhangxubetter@gmail.com [Henan Key Laboratory of Laser and Opto-electric Information Technology, School of Information Engineering, Zhengzhou University, Zhengzhou 450052 (China); Xie, Dan; Huang, Genling [Zhengzhou Railway Vocational and Technical College, Zhengzhou 450052 (China); Sun, Xiao-Hong [Henan Key Laboratory of Laser and Opto-electric Information Technology, School of Information Engineering, Zhengzhou University, Zhengzhou 450052 (China)

    2015-01-01

    An analytic stress-driven nucleation model of nanowires (NWs) and nanoneedles (NNs) growing on a mismatched vicinal substrate is proposed. It is demonstrated that the formation enthalpy of NWs and NNs is a function of three independent variables, the base radius, aspect ratio and miscut angle of the vicinal surface. Theoretical analysis shows that the minimum nucleation barrier of an island decreases with increment of substrate misorientation, which means the nucleation of islands on a vicinal substrate is more favorable than that on a flat substrate.

  18. Produtividade do tomateiro em diferentes substratos e modelos de casas de vegetação Tomato crop production under different substrates and greenhouse models

    Directory of Open Access Journals (Sweden)

    Osmar A. Carrijo

    2004-03-01

    Full Text Available Um experimento com a cultura do tomate, foi instalado na Embrapa Hortaliças em Brasília, durante os anos de 2000 e 2001, para avaliar a produção do tomateiro em diferentes substratos e casas de vegetação. Os substratos utilizados foram casca de arroz, casca de arroz parcialmente carbonizada, fibra de coco verde, lã de rocha, maravalha, serragem e substrato para produção de mudas utilizado na Embrapa Hortaliças (150 L de terra de subsolo, 50 L de casca de arroz parcialmente carbonizada e 17 L de esterco de galinha. Os modelos de casas de vegetação utilizados foram teto em arco, arco com teto convectivo e capela. Não foi verificada diferença estatística significativa quanto a produção de frutos comerciais entre os substratos fibra de coco (10,4 kg m-2, serragem (9,3 kg m-2, casca de arroz carbonizada (9,3 kg m-2 e maravalha (9,0 kg m-2. A menor produção foi obtida com o substrato lã de rocha (6,4 kg m-2. Houve redução da produção entre os anos de cultivo, em torno de 33%, em decorrência de um intenso ataque de traça do tomateiro (Tuta absoluta em todas as casas de vegetação, prejudicando a produtividade. O maior peso médio dos frutos foi obtido sobre a fibra de coco (128 g m-2 e casca de arroz carbonizada (123 g m-2, independente do modelo de casa de vegetação utilizado.The trial was carried out at Embrapa Hortaliças, in Brasilia, Brazil, to evaluate the performance of tomato crop production during two years (2000 and 2001, under three greenhouse models and different types of substrates. The greenhouse models were arch roof; even span and an arch roof with upper convective aperture. The substrates were rice husk, carbonized rice husk, coconut fiber, sawdust, coarsed sawdust, rockwool and a substrate for seedling production used at Embrapa Hortaliças. No significant statistical difference was verified for tomatoes cultivated in coconut fiber (10,4 kg m-2, sawdust (9,9 kg m-2, carbonized rice husk (9,3 kg m-2 and

  19. Designing specificity of protein-substrate interactions

    NARCIS (Netherlands)

    Coluzza, I.; Frenkel, D.

    2004-01-01

    One of the key properties of biological molecules is that they can bind strongly to certain substrates yet interact only weakly with the very large number of other molecules that they encounter. Using a simple lattice model, we test several methods to design molecule-substrate binding specificity. W

  20. Designing specificity of protein-substrate interactions

    NARCIS (Netherlands)

    Coluzza, I.; Frenkel, D.

    2004-01-01

    One of the key properties of biological molecules is that they can bind strongly to certain substrates yet interact only weakly with the very large number of other molecules that they encounter. Using a simple lattice model, we test several methods to design molecule-substrate binding specificity.

  1. Foot-and-mouth disease virus-like particles produced by a SUMO fusion protein system in Escherichia coli induce potent protective immune responses in guinea pigs, swine and cattle.

    Science.gov (United States)

    Guo, Hui-Chen; Sun, Shi-Qi; Jin, Ye; Yang, Shun-Li; Wei, Yan-Quan; Sun, De-Hui; Yin, Shuang-Hui; Ma, Jun-Wu; Liu, Zai-Xin; Guo, Jian-Hong; Luo, Jian-Xun; Yin, Hong; Liu, Xiang-Tao; Liu, Ding Xiang

    2013-07-04

    Foot-and-mouth disease virus (FMDV) causes a highly contagious infection in cloven-hoofed animals. The format of FMD virus-like particles (VLP) as a non-replicating particulate vaccine candidate is a promising alternative to conventional inactivated FMDV vaccines. In this study, we explored a prokaryotic system to express and assemble the FMD VLP and validated the potential of VLP as an FMDV vaccine candidate. VLP composed entirely of FMDV (Asia1/Jiangsu/China/2005) capsid proteins (VP0, VP1 and VP3) were simultaneously produced as SUMO fusion proteins by an improved SUMO fusion protein system in E. coli. Proteolytic removal of the SUMO moiety from the fusion proteins resulted in the assembly of VLP with size and shape resembling the authentic FMDV. Immunization of guinea pigs, swine and cattle with FMD VLP by intramuscular inoculation stimulated the FMDV-specific antibody response, neutralizing antibody response, T-cell proliferation response and secretion of cytokine IFN-γ. In addition, immunization with one dose of the VLP resulted in complete protection of these animals from homologous FMDV challenge. The 50% protection dose (PD50) of FMD VLP in cattle is up to 6.34. These results suggest that FMD VLP expressed in E. coli are an effective vaccine in guinea pigs, swine and cattle and support further development of these VLP as a vaccine candidate for protection against FMDV.

  2. M atom (M = Cu, Ag and Au) interaction with Ag and Au substrates: a first-principles study using cluster and slab models.

    Science.gov (United States)

    Nigam, Sandeep; Majumder, Chiranjib

    2010-11-03

    Using state-of-the-art first-principles calculations we report the interaction of M atoms (M = Cu, Ag and Au) with small Ag(n), Au(n) clusters (n = 3 and 6) and periodic Ag(111) and Au(111) surfaces. All calculations were performed using the plane wave pseudo-potential approach under the spin polarized version of the generalized gradient approximation scheme. The result shows that the equilibrium geometry of all MAg(3) and MAu(3) clusters favor a planar rhombus structure. From the charge distribution analysis of MAg(n)/MAu(n) clusters it is found that, while Cu and Ag donates electronic charge towards the host clusters, the Au atom acts as an acceptor, thus creating charge polarization in the system. The difference in orbital decomposed charges before and after the M interaction reveals that enhanced s-d hybridization is responsible for keeping the MAu(6) cluster planar, and increased p-orbital participation induces three-dimensional configurations in MAg(6) clusters. The optimization of M atom deposition on the Ag(111) and Au(111) surfaces shows that M atoms prefer to adsorb on the threefold fcc site over other well-defined sites. From the orbital decomposed charge analysis it is inferred that, although there is significant difference in the absolute magnitude of the interaction energy between M atoms and the Ag or Au substrates, the nature of chemical bonding is similar for the finite size clusters as well as in slab models.

  3. Evaluation of silicon nitride as a substrate for culture of PC12 cells: an interfacial model for functional studies in neurons.

    Directory of Open Access Journals (Sweden)

    Johan Jaime Medina Benavente

    Full Text Available Silicon nitride is a biocompatible material that is currently used as an interfacial surface between cells and large-scale integration devices incorporating ion-sensitive field-effect transistor technology. Here, we investigated whether a poly-L-lysine coated silicon nitride surface is suitable for the culture of PC12 cells, which are widely used as a model for neural differentiation, and we characterized their interaction based on cell behavior when seeded on the tested material. The coated surface was first examined in terms of wettability and topography using contact angle measurements and atomic force microscopy and then, conditioned silicon nitride surface was used as the substrate for the study of PC12 cell culture properties. We found that coating silicon nitride with poly-L-lysine increased surface hydrophilicity and that exposing this coated surface to an extracellular aqueous environment gradually decreased its roughness. When PC12 cells were cultured on a coated silicon nitride surface, adhesion and spreading were facilitated, and the cells showed enhanced morphological differentiation compared to those cultured on a plastic culture dish. A bromodeoxyuridine assay demonstrated that, on the coated silicon nitride surface, higher proportions of cells left the cell cycle, remained in a quiescent state and had longer survival times. Therefore, our study of the interaction of the silicon nitride surface with PC12 cells provides important information for the production of devices that need to have optimal cell culture-supporting properties in order to be used in the study of neuronal functions.

  4. Climate Change Projection with Reduced Model Systematic Error over Tropic Pacific

    Science.gov (United States)

    Keenlyside, Noel; Shen, Mao-Lin; Selten, Frank; Wiegerinck, Wim; Duane, Gregory

    2014-05-01

    The tropical Pacific is considered as a major driver of the global climate system. However, realistic representation of the equatorial Pacific remains a challenge for state-of-the-art global circulation models (GCMs). For example, the multi-model ensemble mean of the CMIP5 historical simulation exhibits large biases of sea surface temperature. Here we construct an interactive model ensemble (SUMO) by coupling two atmospheric GCMs (AGCM) with one ocean GCM (OGCM). Through optimal coupling weights, synchronization of the atmospheric models over tropical Pacific is enhanced and the dynamic and thermodynamic feedback over Pacific of the GCM become realistic. A set of climate change projections is performed with SUMO and results will be contrasted with conventional multi-model scenario simulations and a standard flux corrected model version to identify main differences.

  5. P-gp substrate-induced neurotoxicity in an Abcb1a knock-in/Abcb1b knock-out mouse model with a mutated canine ABCB1 targeted insertion.

    Science.gov (United States)

    Swain, M D; Orzechowski, K L; Swaim, H L; Jones, Y L; Robl, M G; Tinaza, C A; Myers, M J; Jhingory, M V; Buckely, L E; Lancaster, V A; Yancy, H F

    2013-06-01

    Certain dog breeds, especially Collies, are observed to exhibit neurotoxicity to avermectin drugs, which are P-glycoprotein (P-gp) substrates. This neurotoxicity is due to an ABCB1 gene mutation (ABCB1-1Δ) that results in non-functional P-gp expression. A developed Abcb1a knock-in/Abcb1b knock-out mouse model expressing the ABCB1-1Δ canine gene was previously reported and mice exhibited sensitivity upon ivermectin administration. Here, model and wild-type mice were administered P-gp substrates doramectin, moxidectin, and digoxin. While knock-in/knock-out mice exhibited ataxia, lethargy and tremor, wild-type mice remained unaffected. In addition, no neurotoxic clinical signs were observed in either mouse type administered domperidone, a P-gp substrate with no reported neurotoxicity in ABCB1-1Δ Collies. Overall, neurotoxic signs displayed by model mice closely paralleled those observed in ivermectin-sensitive Collies. This model can be used to identify toxic P-gp substrates with altered safety in dog populations and may reduce dog use in safety studies that are part of the drug approval process.

  6. Moving college students to a better understanding of substrate specificity of enzymes through utilizing multimedia pre-training and an interactive enzyme model

    Science.gov (United States)

    Saleh, Mounir R.

    Scientists' progress in understanding enzyme specificity uncovered a complex natural phenomenon. However, not all of the currently available biology textbooks seem to be up to date on this progress. Students' understanding of how enzymes work is a core requirement in biochemistry and biology tertiary education. Nevertheless, current pre-college science education does not provide students with enough biochemical background to enable them to understand complex material such as this. To bridge this gap, a multimedia pre-training presentation was prepared to fuel the learner's prior knowledge with discrete facts necessary to understand the presented concept. This treatment is also known to manage intrinsic cognitive load during the learning process. An interactive instructional enzyme model was also built to motivate students to learn about substrate specificity of enzymes. Upon testing the effect of this combined treatment on 111 college students, desirable learning outcomes were found in terms of cognitive load, motivation, and achievement. The multimedia pre-training group reported significantly less intrinsic cognitive load, higher motivation, and demonstrated higher transfer performance than the control and post-training groups. In this study, a statistical mediation model is also proposed to explain how cognitive load and motivation work in concert to foster learning from multimedia pre-training. This type of research goes beyond simple forms of "what works" to a deeper understanding of "how it works", thus enabling informed decisions for multimedia instructional design. Multimedia learning plays multiple roles in science education. Therefore, science learners would be some of the first to benefit from improving multimedia instructional design. Accordingly, complex scientific phenomena can be introduced to college students in a motivating, informative, and cognitively efficient learning environment.

  7. Measurement and modelling of process impacts of substrates and filter media to the operation of wastewater treatment plants with fixed biomass

    OpenAIRE

    Drev, Darko; Panjan, Jože

    2006-01-01

    Substrates and filter media are built in wastewater treatment plants in various ways, and can have a high impact on their operation effect. As such, they can either only intensify the treatment processes, or they can act as a main treatment carrier. The specific treatment effects, which can be achieved by substrates or filter media application, were until now rarely exploited in the wastewater treatment plants design. The usual reason for this was insufficient knowledge about these materials ...

  8. 堆肥体系中有机物动态模拟模型研究进展%Composting models describing the dynamics of organic substrates

    Institute of Scientific and Technical Information of China (English)

    张园; 耿春女

    2012-01-01

    堆肥作为生活垃圾处理的一项重要手段,以其无害化程度高、减量化效果明显,以及可以最大限度地实现生活垃圾处理资源化的特点,被越来越多的国家和地区广泛利用。该文的研究目的在于归纳描述堆肥动态变化的模型,分析模型中的关键环境参数,从而借助模型对堆肥过程的有机质动态变化获得更清晰的理解,并预测堆肥产品的质量。为此,文中概述了堆肥模型中环境参数的影响,并列举了目前几种实用的有机质模型。研究结果表明,大多数的堆肥模型一般都用于描述和预测干物质、温度、湿度、氧气和二氧化碳的动态变化过程。随着堆肥模型领域的发展,关于有机质的生化降解以及微生物的作用等方面的研究逐渐展开。%The urban waste composting as an important organic wastes treatment is widely used as for its special effects of high harmlessness, maximum recycling, and obvious quantity reduction. This paper reviews the models describing the composting dynamics, and analyses the key environmental parameters in the models to obtain a clearer understanding of the dynamics of organic fractions during composting, and to predict the final quality of compost products. The findings indicate that most of the composting models have described and predicted the dynamic changes of dry mass, temperature, moisture, oxygen and carbon dioxide. With the development of com- posting modeling, the studies on the biodegradation of organic substrates and microbial activities have been focused on.

  9. PIC-1/SUMO-1-Modified PML-Retinoic Acid Receptor α Mediates Arsenic Trioxide-Induced Apoptosis in Acute Promyelocytic Leukemia

    Science.gov (United States)

    Sternsdorf, Thomas; Puccetti, Elena; Jensen, Kirsten; Hoelzer, Dieter; Will, Hans; Ottmann, Oliver Gerhard; Ruthardt, Martin

    1999-01-01

    Fusion proteins involving the retinoic acid receptor α (RARα) and PML or PLZF nuclear protein are the genetic markers of acute promyelocytic leukemia (APL). APLs with PML-RARα or PLZF-RARα fusion protein differ only in their response to retinoic acid (RA) treatment: the t(15;17) (PML-RARα-positive) APL blasts are sensitive to RA in vitro, and patients enter disease remission after RA treatment, while those with t(11;17) (PLZF-RARα-positive) APLs do not. Recently it has been shown that complete remission can be achieved upon treatment with arsenic trioxide (As2O3) in PML-RARα-positive APL, even when the patient has relapsed and the disease is RA resistant. This appears to be due to apoptosis induced by As2O3 in the APL blasts by poorly defined mechanisms. Here we report that (i) As2O3 induces apoptosis only in cells expressing the PML-RARα, not the PLZF-RARα, fusion protein; (ii) PML-RARα is partially modified by covalent linkage with a PIC-1/SUMO-1-like protein prior to As2O3 treatment, whereas PLZF-RARα is not; (iii) As2O3 treatment induces a change in the modification pattern of PML-RARα toward highly modified forms; (iv) redistribution of PML nuclear bodies (PML-NBs) upon As2O3 treatment is accompanied by recruitment of PIC-1/SUMO-1 into PML-NBs, probably due to hypermodification of both PML and PML-RARα; (v) As2O3-induced apoptosis is independent of the DNA binding activity located in the RARα portion of the PML-RARα fusion protein; and (vi) the apoptotic process is bcl-2 and caspase 3 independent and is blocked only partially by a global caspase inhibitor. Taken together, these data provide novel insights into the mechanisms involved in As2O3-induced apoptosis in APL and predict that treatment of t(11;17) (PLZF-RARα-positive) APLs with As2O3 will not be successful. PMID:10373566

  10. C2株蓝氏贾第鞭毛虫SUMO基因的克隆、原核表达和生物信息学分析%Cloning, prokaryotic expression and bioinformatics analysis of Giardia lamblia C2 strain SUMO gene

    Institute of Scientific and Technical Information of China (English)

    王洋; 王沂; LI Wei-wei; 李冀; 李思瑾; 楚晗; 田喜凤

    2013-01-01

    Small ubiquitin-like modifier (or SUMO) proteins are a family of small proteins that are covalently attached to and detached from other proteins in cells to modify their function.In order to express Giardia lamblia (C2 strain) SUMO prorein in E.coli,the full-length open reading frame of SUMO was amplified by PCR from Giardia lamblia genome DNA.The PCR product about 320 bp in length was cloned into prokaryotic expression vector pET 28a(+) with restriction enzymes Nco I and Xho I.Sequencing result showed the sequence of SUMO in C2 strain was identical with that in WB strain.Bioinformatics analysis showed that SUMO protein displayed an ubiquitin-fold structure in three dimensional space.Phylogenetic analysis showed a distant relationship of SUMO protein between Giardia lamblia and other eukaryotes.The recombinant vector pET-28a(+)-SUMO was transformed into E.coli Rosetta(DE3),then the recombinant SUMO protein was expressed by IPTG induction.SDS-PAGE and western blot using anti-His Tag antibody showed that the expression product of SUMO was a fusion protein about 12.7 kD.The successful prokaryotic expression and bioinformatics analysis of Giardia lamblia SUMO protein provide basis for antibody preparation and functional study of SUMO related protein.%目的 克隆、原核表达C2株蓝氏贾第鞭毛虫(Giardia lamblia,简称贾第虫)的小泛素相关修饰物(small ubiquit in-related modifier,SUMO)蛋白,并对其序列进行生物信息学分析.方法 提取C2株贾第虫基因组DNA,以基因组DNA为模板PCR获得SUMO基因片段,双酶切连入原核表达载体pET28a(+),经测序验证并进行生物信息学分析,将重组质粒pET 28a(+)-SUMO转化大肠杆菌Rosetta(DE3).IPTG诱导后收集菌体,裂解后进行SDS-PAGE及Western blot检测.结果 成功构建了C2株贾第虫SUMO基因原核表达载体pET 28a(+)-SUMO,经IPTG诱导后,在大肠杆菌中高效表达,SDSPAGE及Western blot分析显示,在相对分子量约12.7 kD的位置出现目的蛋

  11. Robust plasmonic substrates

    DEFF Research Database (Denmark)

    Kostiučenko, Oksana; Fiutowski, Jacek; Tamulevicius, Tomas

    2014-01-01

    substrates is presented, which relies on the coverage of gold nanostructures with diamond-like carbon (DLC) thin films of thicknesses 25, 55 and 105 nm. DLC thin films were grown by direct hydrocarbon ion beam deposition. In order to find the optimum balance between optical and mechanical properties...... and breaking. DLC coating with thicknesses between 25 and 105 nm is found to considerably increase the mechanical strength of the substrates while at the same time ensuring conservation of sufficient field enhancements of the gold plasmonic substrates....

  12. Impact of nickel and cobalt on biogas production and process stability during semi-continuous anaerobic fermentation of a model substrate for maize silage.

    Science.gov (United States)

    Pobeheim, Herbert; Munk, Bernhard; Lindorfer, Harald; Guebitz, Georg M

    2011-01-01

    The importance of nickel and cobalt on anaerobic degradation of a defined model substrate for maize was demonstrated. Five semi-continuous reactors were operated for 250 days at 35 °C and a well-defined trace metal solution was added to all reactors. Two reactors each were limited regarding the concentration of Ni(2+) and Co(2+), respectively, for certain time intervals. The required nickel concentration was depending on the organic loading rates (OLR) while, for example, above 2.6 g ODM L(-1) d(-1) nickel concentrations below 0.06 mg kg(-1) FM in the process significantly decreased biogas production by up to 25% compared to a control reactor containing 0.8 mg Ni(2+) kg(-1) FM. Similarly, limitation of cobalt to 0.02 mg kg(-1) FM decreased biogas production by about 10%. Limitations of nickel as well as cobalt lead to process instability. However, after gradual addition of nickel till 0.6 mg and cobalt till 0.05 mg kg(-1) FM the OLR was again increased to 4.3 g ODM L(-1) d(-1) while process stability was recovered and a fast metabolisation of acetic and propionic acid was detected. An increase of nickel to 0.88 mg kg(-1) FM did not enhance biogas performance. Furthermore, the increase of cobalt from 0.05 mg kg(-1) FM up to 0.07 mg kg(-1) FM did not exhibit a change in anaerobic fermentation and biogas production.

  13. Interleukin-32α downregulates the activity of the B-cell CLL/lymphoma 6 protein by inhibiting protein kinase Cε-dependent SUMO-2 modification.

    Science.gov (United States)

    Park, Yun Sun; Kang, Jeong-Woo; Lee, Dong Hun; Kim, Man Sub; Bak, Yesol; Yang, Young; Lee, Hee Gu; Hong, JinTae; Yoon, Do-Young

    2014-09-30

    A proinflammatory cytokine IL-32 acts as an intracellular mediator. IL-32α interacts with many intracellular molecules, but there are no reports of interaction with a transcriptional repressor BCL6. In this study, we showed that PMA induces an interaction between IL-32α, PKCε, and BCL6, forming a trimer. To identify the mechanism of the interaction, we treated cells with various inhibitors. In HEK293 and THP-1 cell lines, treatment with a pan-PKC inhibitor, PKCε inhibitor, and PKCδ inhibitor decreased BCL6 and IL-32α protein expression. MAPK inhibitors and classical PKC inhibitor did not decrease PMA-induced BCL6 and IL-32α protein expression. Further, the pan-PKC inhibitor and PKCε inhibitor disrupted PMA-induced interaction between IL-32α and BCL6. These data demonstrate that the intracellular interaction between IL-32α and BCL6 is induced by PMA-activated PKCε. PMA induces post-translational modification of BCL6 by conjugation to SUMO-2, while IL-32α inhibits. PKCε inhibition eliminated PMA-induced SUMOylation of BCL6. Inhibition of BCL6 SUMOylation by IL-32α affected the cellular function and activity of the transcriptional repressor BCL6 in THP-1 cells. Thus, we showed that IL-32α is a negative regulator of the transcriptional repressor BCL6. IL-32α inhibits BCL6 SUMOylation by activating PKCε, resulting in the modulation of BCL6 target genes and cellular functions of BCL6.

  14. A role for the Cajal-body-associated SUMO isopeptidase USPL1 in snRNA transcription mediated by RNA polymerase II.

    Science.gov (United States)

    Hutten, Saskia; Chachami, Georgia; Winter, Ulrike; Melchior, Frauke; Lamond, Angus I

    2014-03-01

    Cajal bodies are nuclear structures that are involved in biogenesis of snRNPs and snoRNPs, maintenance of telomeres and processing of histone mRNA. Recently, the SUMO isopeptidase USPL1 was identified as a component of Cajal bodies that is essential for cellular growth and Cajal body integrity. However, a cellular function for USPL1 is so far unknown. Here, we use RNAi-mediated knockdown in human cells in combination with biochemical and fluorescence microscopy approaches to investigate the function of USPL1 and its link to Cajal bodies. We demonstrate that levels of snRNAs transcribed by RNA polymerase (RNAP) II are reduced upon knockdown of USPL1 and that downstream processes such as snRNP assembly and pre-mRNA splicing are compromised. Importantly, we find that USPL1 associates directly with U snRNA loci and that it interacts and colocalises with components of the Little Elongation Complex, which is involved in RNAPII-mediated snRNA transcription. Thus, our data indicate that USPL1 plays a key role in RNAPII-mediated snRNA transcription.

  15. Detection of O-propargyl-puromycin with SUMO and ubiquitin by click chemistry at PML-nuclear bodies during abortive proteasome activities.

    Science.gov (United States)

    Uozumi, Naoki; Matsumoto, Hotaru; Saitoh, Hisato

    2016-05-27

    The amino-nucleoside antibiotic, puromycin, acts by covalently linking to elongating polypeptide chains on ribosomes to generate prematurely terminated immature polypeptides. The trafficking of puromycin-conjugated (puromycylated) immature polypeptides within cell has, however, remained elusive. In this study, using O-propargyl-puromycin (OP-Puro), the distribution of puromycylated polypeptides was assessed in HeLa cells by click chemistry. Under standard culture conditions, OP-Puro signals were detected in the cytoplasm and nucleus with the highest concentrations in the nucleolus. Intriguingly, when proteasome activities were aborted using MG132, OP-Puro signals began to accumulate at promyelocytic leukemia nuclear bodies (PML-NBs) in addition to the nucleolus. We also found promiscuous association of OP-Puro signals with SUMO-2/3 and ubiquitin at PML-NBs, but not at the nucleolus, during abortive proteasome activities. This study reveals a previously unknown distribution of OP-Puro that argues for a nuclear function in regulating immature protein homeostasis.

  16. Graphene folding on flat substrates

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Xiaoming; Zhao, Yadong; Ke, Changhong, E-mail: cke@binghamton.edu [Department of Mechanical Engineering, State University of New York at Binghamton, Binghamton, New York 13902 (United States); Zhang, Liuyang; Wang, Xianqiao [College of Engineering, University of Georgia, Athens, Georgia 30602 (United States)

    2014-10-28

    We present a combined experimental-theoretical study of graphene folding on flat substrates. The structure and deformation of the folded graphene sheet are experimentally characterized by atomic force microscopy. The local graphene folding behaviors are interpreted based on nonlinear continuum mechanics modeling and molecular dynamics simulations. Our study on self-folding of a trilayer graphene sheet reports a bending stiffness of about 6.57 eV, which is about four times the reported values for monolayer graphene. Our results reveal that an intriguing free sliding phenomenon occurs at the interlayer van der Waals interfaces during the graphene folding process. This work demonstrates that it is a plausible venue to quantify the bending stiffness of graphene based on its self-folding conformation on flat substrates. The findings reported in this work are useful to a better understanding of the mechanical properties of graphene and in the pursuit of its applications.

  17. PREFACE: Cell-substrate interactions Cell-substrate interactions

    Science.gov (United States)

    Gardel, Margaret; Schwarz, Ulrich

    2010-05-01

    not on the amount of ligand for adhesion receptors, but on its spatial distribution [1]. New protocols for the preparation of soft elastic substrates were essential to show that adhesion structures and cytoskeleton of adherent cells strongly adapt to substrate stiffness [2], with dramatic effects for cellular decision making. For example, it has been shown recently that differentiation of mesenchymal stem cells is strongly influenced by substrate stiffness [3]. Thus, physical factors appear to be equally important as biochemical ones in determining the cellular response to its substrate [4]. The introduction of novel physical techniques not only opened up completely new perspectives regarding biological function, it also introduced a new quantitative element into this field. For example, the availability of soft elastic substrates with controlled stiffness allows us to reconstruct cellular traction forces and to correlate them with other cellular features. This development enables modeling approaches to work in close contact with experimental data, thus opening up the perspective that the field of cell-substrate interactions will become a quantitative and predictive science in the future. Because physical research into cell-substrate interactions has become one of the fastest growing research areas in cellular biophysics and materials science, we believe that it is very timely that this special issue gathers some of the on-going research effort in this field. In contrast to the non-living world, cellular systems usually interact with their environment through specific adhesion, mainly based on adhesion receptors from the integrin family. During recent years, force spectroscopy has emerged as one of the main methods to study the physics of specific adhesion. In this special issue, single cell force spectroscopy is used by Boettiger and Wehrle-Haller to characterize the strength of cell-matrix adhesion and how it is modulated by the glycocalyx [5], while Chirasatitsin

  18. Substrate-induced bulk alignment of liquid crystals

    DEFF Research Database (Denmark)

    Zhang, Zhengping; Chakrabarti, A.; Mouritsen, Ole G.;

    1996-01-01

    The Gay-Berne model for liquid crystals in the presence of a substrate surface is studied using the hybrid Monte Carlo method. A simple non-mean-field substrate-molecule potential is proposed to describe the effects of rubbed polymer-coated substrates on the liquid crystals. Effects...

  19. Atrial Heterogeneity Generates Re-entrant Substrate during Atrial Fibrillation and Anti-arrhythmic Drug Action: Mechanistic Insights from Canine Atrial Models

    National Research Council Canada - National Science Library

    Varela, Marta; Colman, Michael A; Hancox, Jules C; Aslanidi, Oleg V

    2016-01-01

    ...], which contributes the suboptimal success rates of clinical therapies [3]. Available anti-arrhythmic drugs have major limitations, including poor long-term effectiveness and, for some, high pro-arrhythmic risk [1,3]. Multiple studies have suggested that AF can be sustained by re-entrant waves propagating in an abnormal atrial substrate [2...

  20. Substrate stress relaxation regulates cell spreading

    Science.gov (United States)

    Chaudhuri, Ovijit; Gu, Luo; Darnell, Max; Klumpers, Darinka; Bencherif, Sidi A.; Weaver, James C.; Huebsch, Nathaniel; Mooney, David J.

    2015-02-01

    Studies of cellular mechanotransduction have converged upon the idea that cells sense extracellular matrix (ECM) elasticity by gauging resistance to the traction forces they exert on the ECM. However, these studies typically utilize purely elastic materials as substrates, whereas physiological ECMs are viscoelastic, and exhibit stress relaxation, so that cellular traction forces exerted by cells remodel the ECM. Here we investigate the influence of ECM stress relaxation on cell behaviour through computational modelling and cellular experiments. Surprisingly, both our computational model and experiments find that spreading for cells cultured on soft substrates that exhibit stress relaxation is greater than cells spreading on elastic substrates of the same modulus, but similar to that of cells spreading on stiffer elastic substrates. These findings challenge the current view of how cells sense and respond to the ECM.

  1. A Hybrid Multi-gate Model of a Gallium Nitride (GaN) High Electron Mobility Transistor (HEMT) Device Incorporating GaN-substrate Thermal Boundary Resistance

    Science.gov (United States)

    2012-10-01

    coordinates and elliptical cylinder coordinates (8, 9). Douglas et al. studied the effects of several HEMT design parameters, including substrate thermal...Determination of Thermal Resistance of FETs. IEEE Transaction on Microwave Theory and Techniques 2005, 53 (1), 306‒313. 10. Douglas , E. A.; Ren, F...Physics Letters 2010, 97, 232107‒1-3. 29. Taking, S.; Banerjee, A.; Zhou, H.; Li, X.; Khokhar, A. Z.; Oxland, R.; McGregor , I.; Bentley, S.; Rahman, F

  2. Multiple alternative substrate kinetics.

    Science.gov (United States)

    Anderson, Vernon E

    2015-11-01

    The specificity of enzymes for their respective substrates has been a focal point of enzyme kinetics since the initial characterization of metabolic chemistry. Various processes to quantify an enzyme's specificity using kinetics have been utilized over the decades. Fersht's definition of the ratio kcat/Km for two different substrates as the "specificity constant" (ref [7]), based on the premise that the important specificity existed when the substrates were competing in the same reaction, has become a consensus standard for enzymes obeying Michaelis-Menten kinetics. The expansion of the theory for the determination of the relative specificity constants for a very large number of competing substrates, e.g. those present in a combinatorial library, in a single reaction mixture has been developed in this contribution. The ratio of kcat/Km for isotopologs has also become a standard in mechanistic enzymology where kinetic isotope effects have been measured by the development of internal competition experiments with extreme precision. This contribution extends the theory of kinetic isotope effects to internal competition between three isotopologs present at non-tracer concentrations in the same reaction mix. This article is part of a special issue titled: Enzyme Transition States from Theory and Experiment.

  3. Known Good Substrates Year 1

    Science.gov (United States)

    2007-12-05

    Monotonic Decrease of VC-VSi, SN1, and SN2 with ↑ ρ (↓ [N])” Known Good Substrates Contractor: Dow Corning Corporation Contract No. N00014-05-C-0324...Vapor Transport (CVT). The key essence of the CVT approach is to control the growth process by strategic controlled chemical reactions of the source...efficacy for its use in 4H n+ SiC crystal growth. The approach consists of the following steps: • Reaction cell design/modeling and chemical

  4. Evaluation of a SUMO E2 conjugating enzyme involved in resistance to Clavibacter michiganensis subsp. michiganensis in Solanum peruvianum, through a tomato mottle virus VIGS assay

    Directory of Open Access Journals (Sweden)

    Mayra Janeth Esparza-Araiza

    2015-12-01

    Full Text Available Clavibacter michiganensis subsp. michiganensis (Cmm causes bacterial wilt and canker of tomato. Currently, no Solanum lycopersicum resistant varieties are commercially available, but some degree of Cmm resistance has been identified in Solanum peruvianum. Previous research showed up-regulation of a SUMO E2 conjugating enzyme (SCEI transcript in resistant S. peruvianum compared to susceptible S. lycopersicum following infection by Cmm. In order to test the role of SCEI in resistance to Cmm, a fragment of the gene from S. peruvianum was cloned into a novel virus-induced gene-silencing (VIGS vector based on the geminivirus Tomato Mottle Virus (ToMoV. Using biolistic inoculation, the ToMoV-based VIGS vector was shown to be effective in S. peruvianum by silencing the magnesium chelatase gene, which resulted in leaf bleaching. The ToMoV_SCEI construct resulted in approx. 61% silencing of SCEI in leaves of S. peruvianum as determined by quantitative RT-PCR. VIGS of SCEI in S. peruvianum resulted in unilateral wilting (15 dpi and subsequent death (20 dpi of the entire plant after Cmm inoculation, whereas empty vector-treated plants only showed wilting in the Cmm-inoculated leaf. SCEI-silenced plants also showed higher Cmm colonization with an average of 4.5 times more damaged tissue compared to the empty vector control plants. SCEI appears to play an important role in the innate immunity of S. peruvianum against Cmm, perhaps through the regulation of WRKY transcription factors, which may lead to expression of proteins involved in salicylic acid-dependent defense responses.

  5. Evaluation of a SUMO E2 Conjugating Enzyme Involved in Resistance to Clavibacter michiganensis Subsp. michiganensis in Solanum peruvianum, Through a Tomato Mottle Virus VIGS Assay.

    Science.gov (United States)

    Esparza-Araiza, Mayra J; Bañuelos-Hernández, Bernardo; Argüello-Astorga, Gerardo R; Lara-Ávila, José P; Goodwin, Paul H; Isordia-Jasso, María I; Castillo-Collazo, Rosalba; Rougon-Cardoso, Alejandra; Alpuche-Solís, Ángel G

    2015-01-01

    Clavibacter michiganensis subsp. michiganensis (Cmm) causes bacterial wilt and canker of tomato. Currently, no Solanum lycopersicum resistant varieties are commercially available, but some degree of Cmm resistance has been identified in Solanum peruvianum. Previous research showed up-regulation of a SUMO E2 conjugating enzyme (SCEI) transcript in S. peruvianum compared to S. lycopersicum following infection with Cmm. In order to test the role of SCEI in resistance to Cmm, a fragment of SCEI from S. peruvianum was cloned into a novel virus-induced gene-silencing (VIGS) vector based on the geminivirus, Tomato Mottle Virus (ToMoV). Using biolistic inoculation, the ToMoV-based VIGS vector was shown to be effective in S. peruvianum by silencing the magnesium chelatase gene, resulting in leaf bleaching. VIGS with the ToMoV_SCEI construct resulted in ~61% silencing of SCEI in leaves of S. peruvianum as determined by quantitative RT-PCR. The SCEI-silenced plants showed unilateral wilting (15 dpi) and subsequent death (20 dpi) of the entire plant after Cmm inoculation, whereas the empty vector-treated plants only showed wilting in the Cmm-inoculated leaf. The SCEI-silenced plants showed higher Cmm colonization and an average of 4.5 times more damaged tissue compared to the empty vector control plants. SCEI appears to play an important role in the innate immunity of S. peruvianum against Cmm, perhaps through the regulation of transcription factors, leading to expression of proteins involved in salicylic acid-dependent defense responses.

  6. First aminoacetone chelate: [Co(tren){NH2CH2C(O)CH3}]3+-a substrate binding and activation model for zinc(II)-dependent 5-aminolaevulinic acid dehydratase.

    Science.gov (United States)

    Gumm, Andreas; Hammershøi, Anders; Kofod-Hansen, Mikael; Mønsted, Ole; Osholm Sørensen, Henning

    2007-08-14

    The complex p-[Co(tren){NH(2)CH(2)C(O)CH(3)}](ClO(4))(3).H(2)O was produced stereoselectively from [Co(tren)(O(3)SCF(3))(2)]O(3)SCF(3) () and 2-(aminomethyl)-2-methyl-1,3-dioxolane. The structure of was determined by X-ray crystallography. The complex is the first aminoacetone chelate to be reported and the first structurally characterized example of a non-conjugated ketone moiety coordinated to cobalt(iii). The robust complex was stable to aquation in strong acid and behaved as an acid with pK(a) = 4.99(1) indicative of a strong activation of the aminoacetone ligand towards deprotonation. The complex constitutes a structural model for a proposed substrate binding mode relevant for substrate activation of the zinc(ii)-dependent enzyme 5-aminolaevulinic acid dehydratase.

  7. Traffic modelling framework for electric vehicles

    Science.gov (United States)

    Schlote, Arieh; Crisostomi, Emanuele; Kirkland, Stephen; Shorten, Robert

    2012-07-01

    This article reviews and improves a recently proposed model of road network dynamics. The model is also adapted and generalised to represent the patterns of battery consumption of electric vehicles travelling in the road network. Simulations from the mobility simulator SUMO are given to support and to illustrate the efficacy of the proposed approach. Applications relevant in the field of electric vehicles, such as optimal routing and traffic load control, are provided to illustrate how the proposed model can be used to address typical problems arising in contemporary road network planning and electric vehicle mobility.

  8. Experimental analysis of green roof substrate detention characteristics.

    Science.gov (United States)

    Yio, Marcus H N; Stovin, Virginia; Werdin, Jörg; Vesuviano, Gianni

    2013-01-01

    Green roofs may make an important contribution to urban stormwater management. Rainfall-runoff models are required to evaluate green roof responses to specific rainfall inputs. The roof's hydrological response is a function of its configuration, with the substrate - or growing media - providing both retention and detention of rainfall. The objective of the research described here is to quantify the detention effects due to green roof substrates, and to propose a suitable hydrological modelling approach. Laboratory results from experimental detention tests on green roof substrates are presented. It is shown that detention increases with substrate depth and as a result of increasing substrate organic content. Model structures based on reservoir routing are evaluated, and it is found that a one-parameter reservoir routing model coupled with a parameter that describes the delay to start of runoff best fits the observed data. Preliminary findings support the hypothesis that the reservoir routing parameter values can be defined from the substrate's physical characteristics.

  9. Small droplets on superhydrophobic substrates.

    Science.gov (United States)

    Gross, Markus; Varnik, Fathollah; Raabe, Dierk; Steinbach, Ingo

    2010-05-01

    We investigate the wetting behavior of liquid droplets on rough hydrophobic substrates for the case of droplets that are of comparable size to the surface asperities. Using a simple three-dimensional analytical free-energy model, we have shown in a recent letter [M. Gross, F. Varnik, and D. Raabe, EPL 88, 26002 (2009)] that, in addition to the well-known Cassie-Baxter and Wenzel states, there exists a further metastable wetting state where the droplet is immersed into the texture to a finite depth, yet not touching the bottom of the substrate. Due to this new state, a quasistatically evaporating droplet can be saved from going over to the Wenzel state and instead remains close to the top of the surface. In the present paper, we give an in-depth account of the droplet behavior based on the results of extensive computer simulations and an improved theoretical model. In particular, we show that releasing the assumption that the droplet is pinned at the outer edges of the pillars improves the analytical results for larger droplets. Interestingly, all qualitative aspects, such as the existence of an intermediate minimum and the "reentrant transition," remain unchanged. We also give a detailed description of the evaporation process for droplets of varying sizes. Our results point out the role of droplet size for superhydrophobicity and give hints for achieving the desired wetting properties of technically produced materials.

  10. Toward reduction in animal sacrifice for drugs: molecular modeling of Macaca fascicularis P450 2C20 for virtual screening of Homo sapiens P450 2C8 substrates.

    Science.gov (United States)

    Rua, Francesco; Di Nardo, Giovanna; Sadeghi, Sheila J; Gilardi, Gianfranco

    2012-01-01

    Macaca fascicularis P450 2C20 shares 92% identity with human cytochrome P450 2C8, which is involved in the metabolism of more than 8% of all prescribed drugs. To date, only paclitaxel and amodiaquine, two substrate markers of the human P450 2C8, have been experimentally confirmed as M. fascicularis P450 2C20 drugs. To bridge the lack of information on the ligands recognized by M. fascicularis P450 2C20, in this study, a three-dimensional homology model of this enzyme was generated on the basis of the available crystal structure of the human homologue P450 2C8 using YASARA. The results indicated that 90.0%, 9.0%, 0.5%, and 0.5% of the residues of the P450 2C20 model were located in the most favorable, allowed, generously allowed, and disallowed regions, respectively. The root-mean-square deviation of the C-alpha superposition of the M. fascicularis P450 2C20 model with the Homo sapiens P450 2C8 was 0.074 Å, indicating a very high similarity of the two structures. Subsequently, the 2C20 model was used for in silico screening of 58 known P450 2C8 substrates and 62 inhibitors. These were also docked in the active site of the crystal structure of the human P450 2C8. The affinity of each compound for the active site of both cytochromes proved to be very similar, meaning that the few key residues that are mutated in the active site of the M. fascicularis P450 do not prevent the P450 2C20 from recognizing the same substrates as the human P450 2C8.

  11. Substrate quality alters the microbial mineralization of added substrate and soil organic carbon

    Science.gov (United States)

    Jagadamma, S.; Mayes, M. A.; Steinweg, J. M.; Schaeffer, S. M.

    2014-09-01

    The rate and extent of decomposition of soil organic carbon (SOC) is dependent, among other factors, on substrate chemistry and microbial dynamics. Our objectives were to understand the influence of substrate chemistry on microbial decomposition of carbon (C), and to use model fitting to quantify differences in pool sizes and mineralization rates. We conducted an incubation experiment for 270 days using four uniformly labeled 14C substrates (glucose, starch, cinnamic acid and stearic acid) on four different soils (a temperate Mollisol, a tropical Ultisol, a sub-arctic Andisol, and an arctic Gelisol). The 14C labeling enabled us to separate CO2 respired from added substrates and from native SOC. Microbial gene copy numbers were quantified at days 4, 30 and 270 using quantitative polymerase chain reaction (qPCR). Substrate C respiration was always higher for glucose than other substrates. Soils with cinnamic and stearic acid lost more native SOC than glucose- and starch-amended soils. Cinnamic and stearic acid amendments also exhibited higher fungal gene copy numbers at the end of incubation compared to unamended soils. We found that 270 days were sufficient to model the decomposition of simple substrates (glucose and starch) with three pools, but were insufficient for more complex substrates (cinnamic and stearic acid) and native SOC. This study reveals that substrate quality exerts considerable control on the microbial decomposition of newly added and native SOC, and demonstrates the need for multi-year incubation experiments to constrain decomposition parameters for the most recalcitrant fractions of SOC and complex substrates.

  12. Overexpression of the Rice SUMO E3 Ligase Gene OsSIZ1 in Cotton Enhances Drought and Heat Tolerance, and Substantially Improves Fiber Yields in the Field under Reduced Irrigation and Rainfed Conditions

    Science.gov (United States)

    Mishra, Neelam; Sun, Li; Zhu, Xunlu; Smith, Jennifer; Prakash Srivastava, Anurag; Yang, Xiaojie; Pehlivan, Necla; Esmaeili, Nardana; Luo, Hong; Shen, Guoxin; Jones, Don; Auld, Dick; Burke, John

    2017-01-01

    The Arabidopsis SUMO E3 ligase gene AtSIZ1 plays important roles in plant response to abiotic stresses as loss of function in AtSIZ1 leads to increased sensitivity to drought, heat and salt stresses. Overexpression of the AtSIZ1 rice homolog, OsSIZ1, leads to increased heat and drought tolerance in bentgrass, suggesting that the function of the E3 ligase SIZ1 is highly conserved in plants and it plays a critical role in abiotic stress responses. To test the possibility that the SUMO E3 ligase could be used to engineer drought- and heat-tolerant crops, the rice gene OsSIZ1 was overexpressed in cotton. We report here that overexpression of OsSIZ1 in cotton results in higher net photosynthesis and better growth than wild-type cotton under drought and thermal stresses in growth chamber and greenhouse conditions. Additionally, this tolerance to abiotic stresses was correlated with higher fiber yield in both controlled-environment and field trials carried out under reduced irrigation and rainfed conditions. These results suggest that OsSIZ1 is a viable candidate gene to improve crop yields under water-limited and rainfed agricultural production systems. PMID:28340002

  13. Nanomechanics of hard films on compliant substrates.

    Energy Technology Data Exchange (ETDEWEB)

    Reedy, Earl David, Jr. (Sandia National Laboratories, Albuquerque, NM); Emerson, John Allen (Sandia National Laboratories, Albuquerque, NM); Bahr, David F. (Washington State University, Pullman, WA); Moody, Neville Reid; Zhou, Xiao Wang; Hales, Lucas (University of Minnesota, Minneapolis, MN); Adams, David Price (Sandia National Laboratories, Albuquerque, NM); Yeager,John (Washington State University, Pullman, WA); Nyugen, Thao D. (Johns Hopkins University, Baltimore, MD); Corona, Edmundo (Sandia National Laboratories, Albuquerque, NM); Kennedy, Marian S. (Clemson University, Clemson, SC); Cordill, Megan J. (Erich Schmid Institute, Leoben, Austria)

    2009-09-01

    a result, our understanding of the critical relationship between adhesion, properties, and fracture for hard films on compliant substrates is limited. To address this issue, we integrated nanomechanical testing and mechanics-based modeling in a program to define the critical relationship between deformation and fracture of nanoscale films on compliant substrates. The approach involved designing model film systems and employing nano-scale experimental characterization techniques to isolate effects of compliance, viscoelasticity, and plasticity on deformation and fracture of thin hard films on substrates that spanned more than two orders of compliance magnitude exhibit different interface structures, have different adhesion strengths, and function differently under stress. The results of this work are described in six chapters. Chapter 1 provides the motivation for this work. Chapter 2 presents experimental results covering film system design, sample preparation, indentation response, and fracture including discussion on the effects of substrate compliance on fracture energies and buckle formation from existing models. Chapter 3 describes the use of analytical and finite element simulations to define the role of substrate compliance and film geometry on the indentation response of thin hard films on compliant substrates. Chapter 4 describes the development and application of cohesive zone model based finite element simulations to determine how substrate compliance affects debond growth. Chapter 5 describes the use of molecular dynamics simulations to define the effects of substrate compliance on interfacial fracture of thin hard tungsten films on silicon substrates. Chapter 6 describes the Workshops sponsored through this program to advance understanding of material and system behavior.

  14. PVA降解中菌体生长与基质消耗的动力学模型%Dynamic Model of Poly(vinyl alcohol) Degradation in Growth of Microbes and Substrate Consumption

    Institute of Scientific and Technical Information of China (English)

    刘晓娟

    2011-01-01

    研究了PVA降解菌的分批发酵动力学,并首次建立了发酵过程菌体生长、基质消耗随培养时间变化的数学模型。Logistic方程及与Luedeking-Piret方程相似的基质消耗方程能够很好的分别描述聚乙烯醇降解菌的生长及基质的消耗。研究中还论证了在聚乙烯醇降解过程中,其菌体生长与基质降解是偶联的。%The fermentation kinetics of poly(vinyl alcohol)-degradated-strain were studied in a batch sustem.Mathematical model of the growth of microbes in fermentation process and substrate consumption with the culture time was proposed for the first time.Using the Logistic equation for describing PVA degradation microbes growth and the Luedeking-Piret equation for substrate consumption were reviewed.Data gained from various experiments showed that the growth of microbes was coupled with the substrate degradation in the poly(vinyl alcohol) degradation process.

  15. Potential of enzymatic kinetic resolution using solid substrates suspension: improved yield, productivity, substrate concentration, and recovery

    Science.gov (United States)

    Wolff; van Asperen V; Straathof; Heijnen

    1999-03-01

    In the literature the enzymatic kinetic resolution of a suspension of a solid substrate has largely been treated as a conventional kinetic resolution of a fully dissolved substrate. In this paper it is shown that this type of kinetic resolution is different in several important aspects. Quantitative models are developed for two types of such suspension processes. These models are used to compare the merits of these processes with the conventional kinetic resolution process where fully dissolved substrate is used. In the suspension processes the liquid phase concentration of substrate enantiomer that should be converted can be kept close to the maximum value, i.e., the solubility, when process conditions are properly chosen, whereas in a conventional process this concentration gradually decreases. Calculations show that this leads to a productivity that is about 6-fold higher in the suspension processes. Also, for enzymes with a low enantioselectivity, a severalfold increase in yield of remaining enantiopure substrate is predicted compared to the conventional kinetic resolution of dissolved enantiomers. Other potential advantages of using suspension reactions are that the initial substrate concentration may be higher (up to 25% (w/w)) and that the desired remaining substrate may be recovered by simply filtering off the solid crystals. Experimental evidence that these merits can be exploited is only partly given, using the few available examples from the literature.

  16. Biomechanics of substrate boring by fig wasps.

    Science.gov (United States)

    Kundanati, Lakshminath; Gundiah, Namrata

    2014-06-01

    Female insects of diverse orders bore into substrates to deposit their eggs. Such insects must overcome several biomechanical challenges to successfully oviposit, which include the selection of suitable substrates through which the ovipositor can penetrate without itself fracturing. In many cases, the insect may also need to steer and manipulate the ovipositor within the substrate to deliver eggs at desired locations before rapidly retracting her ovipositor to avoid predation. In the case of female parasitoid ichneumonid wasps, this process is repeated multiple times during her lifetime, thus testing the ability of the ovipositioning apparatus to endure fracture and fatigue. What specific adaptations does the ovipositioning apparatus of a female ichneumonoid wasp possess to withstand these challenges? We addressed this question using a model system composed of parasitoid and pollinator fig wasps. First, we show that parasitoid ovipositor tips have teeth-like structures, preferentially enriched with zinc, unlike the smooth morphology of pollinator ovipositors. We describe sensillae present on the parasitoid ovipositor tip that are likely to aid in the detection of chemical species and mechanical deformations and sample microenvironments within the substrate. Second, using atomic force microscopy, we show that parasitoid tip regions have a higher modulus compared with regions proximal to the abdomen in parasitoid and pollinator ovipositors. Finally, we use videography to film wasps during substrate boring and analyse buckling of the ovipositor to estimate the forces required for substrate boring. Together, these results allow us to describe the biomechanical principles underlying substrate boring in parasitoid ichneumonid wasps. Such studies may be useful for the biomimetic design of surgical tools and in the use of novel mechanisms to bore through hard substrates.

  17. Modeling of Lamb wave propagation in plate with two-dimensional phononic crystal layer coated on uniform substrate using plane-wave-expansion method

    Energy Technology Data Exchange (ETDEWEB)

    Hou Zhilin [Laboratoire de Physique des Milieux Ionises et Applications (LPMIA), Nancy University, CNRS Boulevard des Aiguillettes, BP 239 F-54506, Vandoeuvre-les-Nancy (France)], E-mail: zhilin.hou@lpmi.uhp-nancy.fr; Assouar, Badreddine M. [Laboratoire de Physique des Milieux Ionises et Applications (LPMIA), Nancy University, CNRS Boulevard des Aiguillettes, BP 239 F-54506, Vandoeuvre-les-Nancy (France)

    2008-03-17

    We show that the conversional three-dimensional plane wave expansion method can be revised to investigate the lamb wave propagation in the plate with two-dimensional phononic crystal layer coated on uniform substrate. We find that an imaginary three-dimensional periodic system can be constructed by stacking the studied plates and vacuum layers alternately, and then the Fourier series expansion can be performed. The difference between our imaginary periodic system and the true three-dimensional one is that, in our system, the Bloch feature of the wave along the thickness direction is broken. Three different systems are investigated by the proposed method as examples. The principle and reliability of the method are also discussed.

  18. SUMO1基因单核苷酸多态性分析及rs7599810多态性与非综合征型唇/腭裂的关联研究%Analysis of the HapMap data on SNPs in SUMO1 and association study of rs7599810 in trios with non-syndromic cleft lip with or without cleft palate

    Institute of Scientific and Technical Information of China (English)

    岳青; 王红; 张博; 赵凯平

    2014-01-01

    目的:对HapMap数据库四个不同人群SUMO1基因的单核苷酸多态性(single nucleotide polymorphisms,SNPs)进行分析和比较,并探讨山东汉族人群rs7599810位点多态性与非综合征型唇/腭裂(non-syndromic cleft lip with or without cleft palate,NSCL/P)的关联.方法:首先用Haploview软件确定HapMap四个人群,分别为:中国北京汉族(Han Chinese in Beijing,China,CHB)、日本东京(Japanese in Tokyo,Japan,JPT)、西北欧后裔(Utah residents with Northern and Western European ancestry from the CEPH collection,CEU)和尼日利亚约鲁巴(Yoruban in Ibadan,Nigeria,YRI),四个人群中SUMO1基因SNPs中最小等位基因频率(minor allele frequency,MAF)>0.01且符合Hardy-Weinberg平衡的SNPs为合格SNPs,用MAF全距相对比指标衡量人群间最小等位基因相同的共有合格SNPs的MAFs差异;然后对四个人群共有合格SNPs进行单体域和单体型的分析及比较;最后对本研究所测该基因中的rs7599810在山东地区183个NSCL/P核心家系中进行传递不平衡检验(transmission/disequilibrium test,TDT).结果:HapMap提供的24个SNPs中,纯合SNPs在CHB和JPT均为9个,在CEU为8个,在YRI为6个,四个人群共有合格SNPs为12个.这些SNPs在四个人群均形成了一个单体域,其中CHB、JPT、CEU人群的单体域包括全部12个SNPs,YRI人群的单体域包括第3~11个SNPs.CHB、JPT、CEU和YRI人群的单体域分别确定了3、3、5、6种单体型,且均以前两种单体型为主,频率为0.634~ 0.922.TDT结果显示SUMO1基因rs7599810多态性与NSCL/P未见统计学关联(x2 =0.485,P=0.486,OR=0.898,95% CI:0.663 ~1.021).结论:CHB、JPT和CEU人群SUMO1基因SNPs在等位基因组成、MAF、单体域及单体型构成等方面以共性特征为主,与之相比,YRI人群SNPs表现出较大的独特性.山东汉族人群中SUMO1基因rs7599810多态性与NSCL/P未见统计学关联.

  19. Model experiments on growth modes and interface electronics of CuInS{sub 2}: Ultrathin epitaxial films on GaAs(100) substrates

    Energy Technology Data Exchange (ETDEWEB)

    Calvet, Wolfram [Institute for Heterogeneous Materials Systems, Helmholtz-Zentrum Berlin, Hahn-Meitner-Platz 1, 14109, Berlin (Germany); Lewerenz, Hans-Joachim [Joint Center for Artificial Photosynthesis, California Institute of Technology, 1200 E. California Blvd., Pasadena, CA, 91101 (United States); Pettenkofer, Christian [Institute Silicon Photovoltaics, Helmholtz-Zentrum Berlin, Kekulestrasse 5, 12489, Berlin (Germany)

    2014-09-15

    The heterojunction formation between GaAs(100) and CuInS{sub 2} is investigated using ultraviolet photoelectron spectroscopy (UPS), X-ray photoelectron spectroscopy (XPS), and low energy electron diffraction (LEED). Thin layers of CuInS{sub 2} films were deposited in a step-by-step process on wet chemically pre-treated GaAs(100) surfaces by molecular beam epitaxy (MBE) with a total upper thickness limit of the films of 60 nm. The film growth starts from a sulfur-rich GaAs(100) surface. XPS core level analysis of the substrate and film reveals initially a transitory growth regime with the formation of a Ga containing chalcopyrite phase. With increasing film thickness, a change in stoichiometry from Cu-poor to Cu-rich composition is observed. The evaluation of the LEED data shows the occurrence of a recrystallization process where the film orientation follows that of the substrate with the epitaxial relation GaAs{100} parallel CuInS{sub 2}{001}. On the completed junction with a CuInS{sub 2} film thickness of 60 nm, the band discontinuities of the GaAs(100)/CuInS{sub 2} structure measured with XPS and UPS were determined as ΔE{sub V} = 0.1 ± 0.1 eV and ΔE{sub C} = 0.0 ± 0.1 eV, thus showing a type II band alignment. (copyright 2014 WILEY-VCH Verlag GmbH and Co. KGaA, Weinheim)

  20. LAMP-2 deficiency leads to hippocampal dysfunction but normal clearance of neuronal substrates of chaperone-mediated autophagy in a mouse model for Danon disease.

    Science.gov (United States)

    Rothaug, Michelle; Stroobants, Stijn; Schweizer, Michaela; Peters, Judith; Zunke, Friederike; Allerding, Mirka; D'Hooge, Rudi; Saftig, Paul; Blanz, Judith

    2015-01-31

    The Lysosomal Associated Membrane Protein type-2 (LAMP-2) is an abundant lysosomal membrane protein with an important role in immunity, macroautophagy (MA) and chaperone-mediated autophagy (CMA). Mutations within the Lamp2 gene cause Danon disease, an X-linked lysosomal storage disorder characterized by (cardio)myopathy and intellectual dysfunction. The pathological hallmark of this disease is an accumulation of glycogen and autophagic vacuoles in cardiac and skeletal muscle that, along with the myopathy, is also present in LAMP-2-deficient mice. Intellectual dysfunction observed in the human disease suggests a pivotal role of LAMP-2 within brain. LAMP-2A, one specific LAMP-2 isoform, was proposed to be important for the lysosomal degradation of selective proteins involved in neurodegenerative diseases such as Huntington's and Parkinson's disease. To elucidate the neuronal function of LAMP-2 we analyzed knockout mice for neuropathological changes, MA and steady-state levels of CMA substrates. The absence of LAMP-2 in murine brain led to inflammation and abnormal behavior, including motor deficits and impaired learning. The latter abnormality points to hippocampal dysfunction caused by altered lysosomal activity, distinct accumulation of p62-positive aggregates, autophagic vacuoles and lipid storage within hippocampal neurons and their presynaptic terminals. The absence of LAMP-2 did not apparently affect MA or steady-state levels of selected CMA substrates in brain or neuroblastoma cells under physiological and prolonged starvation conditions. Our data contribute to the understanding of intellectual dysfunction observed in Danon disease patients and highlight the role of LAMP-2 within the central nervous system, particularly the hippocampus.

  1. Kinetic characterization, optimum conditions for catalysis and substrate preference of secretory phospholipase A2 from Glycine max in model membrane systems.

    Science.gov (United States)

    Mariani, María Elisa; Madoery, Ricardo Román; Fidelio, Gerardo Daniel

    2015-01-01

    Two secretory phospholipase A2 (sPLA2s) from Glycine max, GmsPLA2-IXA-1 and GmsPLA2-XIB-2, have been purified as recombinant proteins and the activity was evaluated in order to obtain the optimum conditions for catalysis using mixed micelles and lipid monolayers as substrate. Both sPLA2s showed a maximum enzyme activity at pH 7 and a requirement of Ca(2+) in the micromolar range. These parameters were similar to those found for animal sPLA2s but a surprising optimum temperature for catalysis at 60 °C was observed. The effect of negative interfacial charges on the hydrolysis of organized substrates was evaluated through initial rate measurements using short chain phospholipids with different head groups. The enzymes showed subtle differences in the specificity for phospholipids with different head groups (DLPC, DLPG, DLPE, DLPA) in presence or absence of NaCl. Both recombinant enzymes showed lower activity toward anionic phospholipids and a preference for the zwitterionic ones. The values of the apparent kinetic parameters (Vmax and KM) demonstrated that these enzymes have more affinity for phosphatidylcholine compared with phosphatidylglycerol, in contrast with the results observed for pancreatic sPLA2. A hopping mode of catalysis was proposed for the action of these sPLA2 on mixed phospholipid/triton micelles. On the other hand, Langmuir-monolayers assays indicated an optimum lateral surface pressure for activity in between 13 and 16 mN/m for both recombinant enzymes. Copyright © 2014 Elsevier B.V. and Société française de biochimie et biologie Moléculaire (SFBBM). All rights reserved.

  2. Spectroscopic investigation of new water soluble Mn(II)(2) and Mg(II)(2) complexes for the substrate binding models of xylose/glucose isomerases.

    Science.gov (United States)

    Patra, Ayan; Bera, Manindranath

    2014-01-30

    In methanol, the reaction of stoichiometri