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Sample records for ajuba lim proteins

  1. Ajuba LIM proteins are Snail/Slug corepressors required for neural crest development in Xenopus

    PubMed Central

    Langer, Ellen M.; Feng, Yunfeng; Zhaoyuan, Hou; Rauscher, Frank J.; Kroll, Kristen L.; Longmore, Gregory D.

    2008-01-01

    Snail family transcriptional repressors regulate epithelial mesenchymal transitions during physiological and pathological processes. A conserved SNAG repression domain present in all vertebrate Snail proteins is necessary for repressor complex assembly. Here, we identify the Ajuba family of LIM proteins as functional corepressors of the Snail family via an interaction with the SNAG domain. Ajuba LIM proteins interact with Snail in the nucleus on endogenous E-cadherin promoters and contribute to Snail-dependent repression of E-cadherin. Using Xenopus neural crest as a model of in vivo Snail- or Slug-induced EMT, we demonstrate that Ajuba LIM proteins contribute to neural crest development as Snail/Slug corepressors and are required for in vivo Snail/Slug function. Because Ajuba LIM proteins are also components of adherens junction and contribute to their assembly or stability, their functional interaction with Snail proteins in the nucleus suggests that Ajuba LIM proteins are important regulators of epithelia dynamics communicating surface events with nuclear responses. PMID:18331720

  2. The Dictyostelium LIM Domain-containing Protein LIM2 Is Essential for Proper Chemotaxis and Morphogenesis

    PubMed Central

    Chien, Sharon; Chung, Chang Y.; Sukumaran, Sujatha; Osborne, Nicholas; Lee, Susan; Ellsworth, Charlene; McNally, James G.; Firtel, Richard A.

    2000-01-01

    We have identified limB, a gene encoding a novel LIM domain-containing protein, LIM2, in a screen for genes required for morphogenesis. limB null cells aggregate, although poorly, but they are unable to undergo morphogenesis, and the aggregates arrest at the mound stage. limB null cells exhibit an aberrant actin cytoskeleton and have numerous F-actin–enriched microspikes. The cells exhibit poor adhesion to a substratum and do not form tight cell–cell agglomerates in suspension. Furthermore, limB null cells are unable to properly polarize in chemoattractant gradients and move very poorly. Expression of limB from a prestalk-specific but not a prespore-specific promoter complements the morphogenetic defects of the limB null strain, suggesting that the limB null cell developmental defect results from an inability to properly sort prestalk cells. LIM2 protein is enriched in the cortex of wild-type cells, although it does not colocalize with the actin cytoskeleton. Our analysis indicates that LIM2 is a new regulatory protein that functions to control rearrangements of the actin cytoskeleton and is required for cell motility and chemotaxis. Our findings may be generally applicable to understanding pathways that control cell movement and morphogenesis in all multicellular organisms. Structure function studies on the LIM domains are presented. PMID:10749929

  3. Cytoskeletal Tension inhibits Hippo signaling through an Ajuba-Warts complex

    PubMed Central

    Sun, Gongping; Pan, Yuanwang; Irvine, Kenneth D.

    2014-01-01

    Mechanical forces have been proposed to modulate organ growth, but a molecular mechanism that links them to growth regulation in vivo has been lacking. We report that increasing tension within the cytoskeleton increases Drosophila wing growth, whereas decreasing cytoskeletal tension decreases wing growth. These changes in growth can be accounted for by changes in the activity of Yorkie, a transcription factor regulated by the Hippo pathway. The influence of myosin activity on Yorkie depends genetically on the Ajuba LIM protein Jub, a negative regulator of Warts within the Hippo pathway. We further show that Jub associates with α-catenin, and that its localization to adherens junctions and association with α-catenin are promoted by cytoskeletal tension. Jub recruits Warts to junctions in a tension-dependent manner. Our observations delineate a mechanism that links cytoskeletal tension to regulation of Hippo pathway activity, providing a molecular understanding of how mechanical forces can modulate organ growth. PMID:24995985

  4. Nucleo-cytoplasmic functions of the PDZ-LIM protein family: new insights in organ development

    PubMed Central

    Krcmery, Jennifer; Camarata, Troy; Kulisz, Andre; Simon, Hans-Georg

    2010-01-01

    Summary Recent work on the PDZ-LIM protein family has revealed important activities at the cellular level, mediating signals between the nucleus and the cytoskeleton, with significant impact on organ development. We review and integrate current knowledge about the PDZ-LIM protein family and propose a new functional role, sequestering nuclear factors in the cytoplasm. Characterized by their PDZ and LIM domains, the PDZ-LIM family is comprised of evolutionarily conserved proteins found throughout the animal kingdom, from worms to humans. Combining two functional domains in one protein, PDZ-LIM proteins have wide-ranging and multi-compartmental cell functions during development and homeostasis while, in contrast, misregulation can lead to cancer formation and progression. New emerging roles include interactions with integrins, T-box transcription factors, and receptor tyrosine kinases. Facilitating the assembly of protein complexes, PDZ-LIM proteins can act as signal modulators, influence actin dynamics, regulate cell architecture and control gene transcription. PMID:20091751

  5. [Cloning and expression analysis of a LIM-domain protein gene from cotton (Gossypium hirsuturm L.)].

    PubMed

    Luo, Ming; Xiao, Yue-Hua; Hou, Lei; Luo, Xiao-Ying; Li, De-Mou; Pei, Yan

    2003-02-01

    LIM-domain protein plays an important role in various cellular processes, including construction of cytoskeleton, transcription control and signal transduction. Based on cotton fiber EST database and contig analysis, the coding region of a cotton LIM-domain protein gene (GhLIM1) was obtained by RT-PCR from 4DPA (day post anthesis) ovule with fiber. The cloned fragment of 848 bp contains an open reading frame of 570 bp, coding for a polypeptide of 189 amino acids. It was demonstrated that the deduced GhLIM1 protein was highly homologous to the LIM-domain protein of sunflower (Helianthus annuus), tobacco (Nicotiana tabacum) and Arabidopsis thaliana. Two intact LIM-domains, with the conserved sequence of a double zinc-finger structure (C-X2-C-X17-19-H-X2-C-X2-C-X2-C-X16-24-C-X2-H), were found in the GhLIM1 protein. RT-PCR and Northern blot analysis showed that GhLIM1 gene expressed in root, shoot tip, hypocotyls, bud, leaf, anther, ovule and fiber (4DPA, 12DPA, 18DPA). However it was preferentially expressed in the shoot tip, fiber and ovule. It was proposed that the express of GhLIM1 gene is related to cotton fiber development. PMID:12776607

  6. The Evolutionarily Conserved LIM Homeodomain Protein LIM-4/LHX6 Specifies the Terminal Identity of a Cholinergic and Peptidergic C. elegans Sensory/Inter/Motor Neuron-Type

    PubMed Central

    Choi, Seong-Kyoon; Huh, Yang Hoon; Fang, Zi; Park, Seo Jin; Kim, Myoung Ok; Ryoo, Zae Young; Kang, Kyeongjin; Kweon, Hee-Seok; Jeon, Won Bae; Li, Chris; Kim, Kyuhyung

    2015-01-01

    The expression of specific transcription factors determines the differentiated features of postmitotic neurons. However, the mechanism by which specific molecules determine neuronal cell fate and the extent to which the functions of transcription factors are conserved in evolution are not fully understood. In C. elegans, the cholinergic and peptidergic SMB sensory/inter/motor neurons innervate muscle quadrants in the head and control the amplitude of sinusoidal movement. Here we show that the LIM homeobox protein LIM-4 determines neuronal characteristics of the SMB neurons. In lim-4 mutant animals, expression of terminal differentiation genes, such as the cholinergic gene battery and the flp-12 neuropeptide gene, is completely abolished and thus the function of the SMB neurons is compromised. LIM-4 activity promotes SMB identity by directly regulating the expression of the SMB marker genes via a distinct cis-regulatory motif. Two human LIM-4 orthologs, LHX6 and LHX8, functionally substitute for LIM-4 in C. elegans. Furthermore, C. elegans LIM-4 or human LHX6 can induce cholinergic and peptidergic characteristics in the human neuronal cell lines. Our results indicate that the evolutionarily conserved LIM-4/LHX6 homeodomain proteins function in generation of precise neuronal subtypes. PMID:26305787

  7. Molecular and expression analysis of a LIM protein gene family from flowering plants.

    PubMed

    Eliasson, A; Gass, N; Mundel, C; Baltz, R; Kräuter, R; Evrard, J L; Steinmetz, A

    2000-10-01

    LIM-domain proteins participate in important cellular processes in eukaryotes, including gene transcription and actin cytoskeleton organization. They are predominantly found in animals, but have also been identified in yeast and plants. Following the characterization ofa LIM-domain protein in sunflower pollen, we carried out an extensive search for these proteins in flowering plants. We have isolated and studied cDNAs and/or genomic sequences for two novel LIM-domain proteins from sunflower, three from tobacco, and one from Arabidopsis. The plant proteins are structurally related to the cytoskeleton-associated CRP class of LIM proteins in animals, but show several distinctive features, including a second, atypical, LIM domain. We have performed comparative expression studies of these genes, as well as of one other gene from tobacco and two additional Arabidopsis genes whose sequences are available from databases. These studies, carried out by RT-PCR in the presence of gene-specific primers, showed that, in sunflower and tobacco, pollen grains and sporophytic tissues express different sets of LIM proteins. With the exception of one Arabidopsis gene--which has two introns--all the genes analyzed contain four introns at conserved positions, indicating that the ancestral gene from which the various copies evolved in higher plants allready had this split structure. PMID:11085265

  8. PDZ motifs in PTP-BL and RIL bind to internal protein segments in the LIM domain protein RIL.

    PubMed

    Cuppen, E; Gerrits, H; Pepers, B; Wieringa, B; Hendriks, W

    1998-03-01

    The specificity of protein-protein interactions in cellular signaling cascades is dependent on the sequence and intramolecular location of distinct amino acid motifs. We used the two-hybrid interaction trap to identify proteins that can associate with the PDZ motif-rich segment in the protein tyrosine phosphatase PTP-BL. A specific interaction was found with the Lin-11, Isl-1, Mec-3 (LIM) domain containing protein RIL. More detailed analysis demonstrated that the binding specificity resides in the second and fourth PDZ motif of PTP-BL and the LIM domain in RIL. Immunohistochemistry on various mouse tissues revealed a submembranous colocalization of PTP-BL and RIL in epithelial cells. Remarkably, there is also an N-terminal PDZ motif in RIL itself that can bind to the RIL-LIM domain. We demonstrate here that the RIL-LIM domain can be phosphorylated on tyrosine in vitro and in vivo and can be dephosphorylated in vitro by the PTPase domain of PTP-BL. Our data point to the presence of a double PDZ-binding interface on the RIL-LIM domain and suggest tyrosine phosphorylation as a regulatory mechanism for LIM-PDZ associations in the assembly of multiprotein complexes. These findings are in line with an important role of PDZ-mediated interactions in the shaping and organization of submembranous microenvironments of polarized cells. PMID:9487134

  9. PDZ Motifs in PTP-BL and RIL Bind to Internal Protein Segments in the LIM Domain Protein RIL

    PubMed Central

    Cuppen, Edwin; Gerrits, Herlinde; Pepers, Barry; Wieringa, Bé; Hendriks, Wiljan

    1998-01-01

    The specificity of protein–protein interactions in cellular signaling cascades is dependent on the sequence and intramolecular location of distinct amino acid motifs. We used the two-hybrid interaction trap to identify proteins that can associate with the PDZ motif-rich segment in the protein tyrosine phosphatase PTP-BL. A specific interaction was found with the Lin-11, Isl-1, Mec-3 (LIM) domain containing protein RIL. More detailed analysis demonstrated that the binding specificity resides in the second and fourth PDZ motif of PTP-BL and the LIM domain in RIL. Immunohistochemistry on various mouse tissues revealed a submembranous colocalization of PTP-BL and RIL in epithelial cells. Remarkably, there is also an N-terminal PDZ motif in RIL itself that can bind to the RIL-LIM domain. We demonstrate here that the RIL-LIM domain can be phosphorylated on tyrosine in vitro and in vivo and can be dephosphorylated in vitro by the PTPase domain of PTP-BL. Our data point to the presence of a double PDZ-binding interface on the RIL-LIM domain and suggest tyrosine phosphorylation as a regulatory mechanism for LIM-PDZ associations in the assembly of multiprotein complexes. These findings are in line with an important role of PDZ-mediated interactions in the shaping and organization of submembranous microenvironments of polarized cells. PMID:9487134

  10. A LIM domain protein from tobacco involved in actin-bundling and histone gene transcription.

    PubMed

    Moes, Danièle; Gatti, Sabrina; Hoffmann, Céline; Dieterle, Monika; Moreau, Flora; Neumann, Katrin; Schumacher, Marc; Diederich, Marc; Grill, Erwin; Shen, Wen-Hui; Steinmetz, André; Thomas, Clément

    2013-03-01

    The two LIM domain-containing proteins from plants (LIMs) typically exhibit a dual cytoplasmic-nuclear distribution, suggesting that, in addition to their previously described roles in actin cytoskeleton organization, they participate in nuclear processes. Using a south-western blot-based screen aimed at identifying factors that bind to plant histone gene promoters, we isolated a positive clone containing the tobacco LIM protein WLIM2 (NtWLIM2) cDNA. Using both green fluorescent protein (GFP) fusion- and immunology-based strategies, we provide clear evidence that NtWLIM2 localizes to the actin cytoskeleton, the nucleus, and the nucleolus. Interestingly, the disruption of the actin cytoskeleton by latrunculin B significantly increases NtWLIM2 nuclear fraction, pinpointing a possible novel cytoskeletal-nuclear crosstalk. Biochemical and electron microscopy experiments reveal the ability of NtWLIM2 to directly bind to actin filaments and to crosslink the latter into thick actin bundles. Electrophoretic mobility shift assays show that NtWLIM2 specifically binds to the conserved octameric cis-elements (Oct) of the Arabidopsis histone H4A748 gene promoter and that this binding largely relies on both LIM domains. Importantly, reporter-based experiments conducted in Arabidopsis and tobacco protoplasts confirm the ability of NtWLIM2 to bind to and activate the H4A748 gene promoter in live cells. Expression studies indicate the constitutive presence of NtWLIM2 mRNA and NtWLIM2 protein during tobacco BY-2 cell proliferation and cell cycle progression, suggesting a role of NtWLIM2 in the activation of basal histone gene expression. Interestingly, both live cell and in vitro data support NtWLIM2 di/oligomerization. We propose that NtWLIM2 functions as an actin-stabilizing protein, which, upon cytoskeleton remodeling, shuttles to the nucleus in order to modify gene expression. PMID:22930731

  11. Sulindac modulates secreted protein expression from LIM1215 colon carcinoma cells prior to apoptosis.

    PubMed

    Greening, David W; Ji, Hong; Kapp, Eugene A; Simpson, Richard J

    2013-11-01

    Colorectal cancer (CRC) is a major cause of mortality in Western populations. Growing evidence from human and rodent studies indicate that nonsteroidal anti-inflammatory drugs (NSAIDs) cause regression of existing colon tumors and act as effective chemopreventive agents in sporadic colon tumor formation. Although much is known about the action of the NSAID sulindac, especially its role in inducing apoptosis, mechanisms underlying these effects is poorly understood. In previous secretome-based proteomic studies using 2D-DIGE/MS and cytokine arrays we identified over 150 proteins released from the CRC cell line LIM1215 whose expression levels were dysregulated by treatment with 1mM sulindac over 16h; many of these proteins are implicated in molecular and cellular functions such as cell proliferation, differentiation, adhesion, angiogenesis and apoptosis (Ji et al., Proteomics Clin. Appl. 2009, 3, 433-451). We have extended these studies and describe here an improved protein/peptide separation strategy that facilitated the identification of 987 proteins and peptides released from LIM1215 cells following 1mM sulindac treatment for 8h preceding the onset of apoptosis. This peptidome separation strategy involved fractional centrifugal ultrafiltration of concentrated cell culture media (CM) using nominal molecular weight membrane filters (NMWL 30K, 3K and 1K). Proteins isolated in the >30K and 3-30K fractions were electrophoretically separated by SDS-PAGE and endogenous peptides in the 1-3K membrane filter were fractioned by RP-HPLC; isolated proteins and peptides were identified by nanoLC-MS-MS. Collectively, our data show that LIM1215 cells treated with 1mM sulindac for 8h secrete decreased levels of proteins associated with extracellular matrix remodeling (e.g., collagens, perlecan, syndecans, filamins, dyneins, metalloproteinases and endopeptidases), cell adhesion (e.g., cadherins, integrins, laminins) and mucosal maintenance (e.g., glycoprotein 340 and mucins 5AC, 6

  12. Silk Gland Factor-2, Involved in Fibroin Gene Transcription, Consists of LIM Homeodomain, LIM-interacting, and Single-stranded DNA-binding Proteins*

    PubMed Central

    Ohno, Kaoru; Sawada, Jun-ichi; Takiya, Shigeharu; Kimoto, Mai; Matsumoto, Akiko; Tsubota, Takuya; Uchino, Keiro; Hui, Chi-chung; Sezutsu, Hideki; Handa, Hiroshi; Suzuki, Yoshiaki

    2013-01-01

    SGF-2 binds to promoter elements governing posterior silk gland-specific expression of the fibroin gene in Bombyx mori. We purified SGF-2 and showed that SGF-2 contains at least four gene products: the silkworm orthologues of LIM homeodomain protein Awh, LIM domain-binding protein (Ldb), a sequence-specific single-stranded DNA-binding protein (Lcaf), and the silk protein P25/fibrohexamerin (fhx). Using co-expression of these factors in Sf9 cells, Awh, Ldb, and Lcaf proteins were co-purified as a ternary complex that bound to the enhancer sequence in vitro. Lcaf interacts with Ldb as well as Awh through the conserved regions to mediate transcriptional activation in yeast. Misexpression of Awh in transgenic silkworms induces ectopic expression of the fibroin gene in the middle silk glands, where Ldb and Lcaf are expressed. Taken together, this study demonstrates that SGF-2 is a multisubunit activator complex containing Awh. Moreover, our results suggest that the Ldb·Lcaf protein complex serves as a scaffold to facilitate communication between transcriptional control elements. PMID:24022586

  13. The LIM/homeodomain protein islet-1 modulates estrogen receptor functions.

    PubMed

    Gay, F; Anglade, I; Gong, Z; Salbert, G

    2000-10-01

    LIM/Homeodomain (HD) proteins are classically considered as major transcriptional regulators which, in cooperation with other transcription factors, play critical roles in the developing nervous system. Among LIM/HD proteins, Islet-1 (ISL1) is the earliest known marker of motoneuron differentiation and has been extensively studied in this context. However, ISL1 expression is not restricted to developing motoneurons. In both embryonic and adult central nervous system of rodent and fish, ISL1 is found in discrete brain areas known to express the estrogen receptor (ER). These observations led us to postulate the possible involvement of ISL1 in the control of brain functions by steroid hormones. Dual immunohistochemistry for ISL1 and ER provided evidence for ISL1-ER coexpression by the same neuronal subpopulation within the rat hypothalamic arcuate nucleus. The relationship between ER and ISL1 was further analyzed at the molecular level and we could show that 1) ISL1 directly interacts in vivo and in vitro with the rat ER, as well as with various other nuclear receptors; 2) ISL1-ER interaction is mediated, at least in part, by the ligand binding domain of ER and is significantly strengthened by estradiol; 3) as a consequence, ISL1 prevents ER dimerization in solution, thus leading to a strong and specific inhibition of ER DNA binding activity; 4) ISL1, via its N-terminal LIM domains, specifically inhibits the ER-driven transcriptional activation in some promoter contexts, while ER can serve as a coactivator for ISL1 in other promoter contexts. Taken together, these data suggest that ISL1-ER cross-talk could differentially regulate the expression of ER and ISL1 target genes. PMID:11043578

  14. Degradation of LIM domain-binding protein three during processing of Spanish dry-cured ham.

    PubMed

    Gallego, Marta; Mora, Leticia; Fraser, Paul D; Aristoy, María-Concepción; Toldrá, Fidel

    2014-04-15

    Extensive proteolysis takes place during the processing of dry-cured ham due to the action of muscle peptidases. The aim of this work was to study the degradation of LIM domain binding protein 3 (LDB3), which is located at the Z-lines of the sarcomere, at different times during the Spanish dry-cured ham processing (2, 3.5, 5, 6.5, and 9 months). A total of 107 peptides have been identified by mass spectrometry, most of them generated from the first region of the protein sequence (position 1-90) providing evidence for the complexity and variability of proteolytic reactions throughout the whole process of dry-curing. Methionine oxidation has been observed in several peptides by the end of the process. The potential of some of the identified peptides to be used as biomarkers of dry-cured ham processing has also been considered. PMID:24295685

  15. Arabidopsis LIM Proteins: A Family of Actin Bundlers with Distinct Expression Patterns and Modes of Regulation[W][OA

    PubMed Central

    Papuga, Jessica; Hoffmann, Céline; Dieterle, Monika; Moes, Danièle; Moreau, Flora; Tholl, Stéphane; Steinmetz, André; Thomas, Clément

    2010-01-01

    Recently, a number of two LIM-domain containing proteins (LIMs) have been reported to trigger the formation of actin bundles, a major higher-order cytoskeletal assembly. Here, we analyzed the six Arabidopsis thaliana LIM proteins. Promoter-β-glucuronidase reporter studies revealed that WLIM1, WLIM2a, and WLIM2b are widely expressed, whereas PLIM2a, PLIM2b, and PLIM2c are predominantly expressed in pollen. LIM-green fluorescent protein (GFP) fusions all decorated the actin cytoskeleton and increased actin bundle thickness in transgenic plants and in vitro, although with different affinities and efficiencies. Remarkably, the activities of WLIMs were calcium and pH independent, whereas those of PLIMs were inhibited by high pH and, in the case of PLIM2c, by high [Ca2+]. Domain analysis showed that the C-terminal domain is key for the responsiveness of PLIM2c to pH and calcium. Regulation of LIM by pH was further analyzed in vivo by tracking GFP-WLIM1 and GFP-PLIM2c during intracellular pH modifications. Cytoplasmic alkalinization specifically promoted release of GFP-PLIM2c but not GFP-WLIM1, from filamentous actin. Consistent with these data, GFP-PLIM2c decorated long actin bundles in the pollen tube shank, a region of relatively low pH. Together, our data support a prominent role of Arabidopsis LIM proteins in the regulation of actin cytoskeleton organization and dynamics in sporophytic tissues and pollen. PMID:20817848

  16. The LIM-Only Protein FHL2 Attenuates Lung Inflammation during Bleomycin-Induced Fibrosis

    PubMed Central

    Schied, Tanja; Chiquet-Ehrismann, Ruth; Loser, Karin; Vogl, Thomas; Ludwig, Stephan; Wixler, Viktor

    2013-01-01

    Fibrogenesis is usually initiated when regenerative processes have failed and/or chronic inflammation occurs. It is characterised by the activation of tissue fibroblasts and dysregulated synthesis of extracellular matrix proteins. FHL2 (four-and-a-half LIM domain protein 2) is a scaffolding protein that interacts with numerous cellular proteins, regulating signalling cascades and gene transcription. It is involved in tissue remodelling and tumour progression. Recent data suggest that FHL2 might support fibrogenesis by maintaining the transcriptional expression of alpha smooth muscle actin and the excessive synthesis and assembly of matrix proteins in activated fibroblasts. Here, we present evidence that FHL2 does not promote bleomycin-induced lung fibrosis, but rather suppresses this process by attenuating lung inflammation. Loss of FHL2 results in increased expression of the pro-inflammatory matrix protein tenascin C and downregulation of the macrophage activating C-type lectin receptor DC-SIGN. Consequently, FHL2 knockout mice developed a severe and long-lasting lung pathology following bleomycin administration due to enhanced expression of tenascin C and impaired activation of inflammation-resolving macrophages. PMID:24260575

  17. Neuronal Expression of Muscle LIM Protein in Postnatal Retinae of Rodents

    PubMed Central

    Levin, Evgeny; Leibinger, Marco; Andreadaki, Anastasia; Fischer, Dietmar

    2014-01-01

    Muscle LIM protein (MLP) is a member of the cysteine rich protein family and has so far been regarded as a muscle-specific protein that is mainly involved in myogenesis and the organization of cytoskeletal structure in myocytes, respectively. The current study demonstrates for the first time that MLP expression is not restricted to muscle tissue, but is also found in the rat naive central nervous system. Using quantitative PCR, Western blot and immunohistochemical analyses we detected MLP in the postnatal rat retina, specifically in the somas and dendritic arbors of cholinergic amacrine cells (AC) of the inner nuclear layer and the ganglion cell layer (displaced AC). Induction of MLP expression started at embryonic day 20 and peaked between postnatal days 7 and 14. It subsequently decreased again to non-detectable protein levels after postnatal day 28. MLP was identified in the cytoplasm and dendrites but not in the nucleus of AC. Thus, retinal MLP expression correlates with the morphologic and functional development of cholinergic AC, suggesting a potential role of this protein in postnatal maturation and making MLP a suitable marker for these neurons. PMID:24945278

  18. Is the LIM-domain protein HaWLIM1 associated with cortical microtubules in sunflower protoplasts?

    PubMed

    Brière, Christian; Bordel, Anne-Claire; Barthou, Henri; Jauneau, Alain; Steinmetz, André; Alibert, Gilbert; Petitprez, Michel

    2003-10-01

    Flowering plants express several LIM-domain proteins related to the animal cystein-rich proteins. The expression of sunflower LIM genes was followed by RT-PCR in cultured sunflower protoplasts. A transcript was detected only for HaWLIM1, but not for the other two genes HaPLIM1 and HaPLIM2. Polyclonal antibodies raised against either full length recombinant HaWLIM1 protein or peptides recognized a 27 kDa polypeptide on Western blots. Immunocytolocalization studies showed that HaWLIM1 is located in the cytoplasm and in the nucleus. In the cytoplasm, HaWLIM1 is localized in punctate structures, distributed along microtubule bundles. Depolymerizing microtubules with oryzalin resulted in a strong modification of the HaWLIM1 cortical pattern. In contrast, treatment of protoplasts with latrunculin B, which disrupts actin filaments, had no effect on HaWLIM1 localization. HaWLIM1 was also located within the nucleus of interphase protoplasts. During mitosis, nuclear labelling was observed in prophase, which decreased in metaphase, disappeared in anaphase, and recovered in telophase. These results suggest a dual role for HaWLIM1: in the cytoplasm, as a component of molecular complexes which may interact with microtubules, and in the nucleus, as a partner of transcription factors during interphase. PMID:14581630

  19. The LIM domain-containing Dbm1 GTPase-activating protein is required for normal cellular morphogenesis in Saccharomyces cerevisiae.

    PubMed Central

    Chen, G C; Zheng, L; Chan, C S

    1996-01-01

    Normal cell growth in the yeast Saccharomyces cerevisiae involves the selection of genetically determined bud sites where most growth is localized. Previous studies have shown that BEM2, which encodes a GTPase-activating protein (GAP) that is specific for the Rho-type GTPase Rho1p in vitro, is required for proper bud site selection and bud emergence. We show here that DBM1, which encodes another putative Rho-type GAP with two tandemly arranged cysteine-rich LIM domains, also is needed for proper bud site selection, as haploid cells lacking Dbm1p bud predominantly in a bipolar, rather than the normal axial, manner. Furthermore, yeast cells lacking both Bem2p and Dbm1p are inviable. The nonaxial budding defect of dbm1 mutants can be rescued partially by overproduction of Bem3p and is exacerbated by its absence. Since Bem3p has previously been shown to function as a GAP for Cdc42p, and also less efficiently for Rho1p, our results suggest that Dbm1p, like Bem2p and Bem3p, may function in vivo as a GAP for Cdc42p and/or Rho1p. Both LIM domains of Dbm1p are essential for its normal function. Point mutations that alter single conserved cysteine residues within either LIM domain result in mutant forms of Dbm1p that can no longer function in bud site selection but instead are capable of rescuing the inviability of bem2 mutants at 35 degrees C. PMID:8657111

  20. Characterization of a unique motif in LIM mineralization protein-1 that interacts with jun activation-domain-binding protein 1.

    PubMed

    Sangadala, Sreedhara; Yoshioka, Katsuhito; Enyo, Yoshio; Liu, Yunshan; Titus, Louisa; Boden, Scott D

    2014-01-01

    Development and repair of the skeletal system and other organs are highly dependent on precise regulation of the bone morphogenetic protein (BMP) pathway. The use of BMPs clinically to induce bone formation has been limited in part by the requirement of much higher doses of recombinant proteins in primates than were needed in cell culture or rodents. Therefore, increasing cellular responsiveness to BMPs has become our focus. We determined that an osteogenic LIM mineralization protein, LMP-1 interacts with Smurf1 (Smad ubiquitin regulatory factor 1) and prevents ubiquitination of Smads resulting in potentiation of BMP activity. In the region of LMP-1 responsible for bone formation, there is a motif that directly interacts with the Smurf1 WW2 domain and thus effectively competes for binding with Smad1 and Smad5, key signaling proteins in the BMP pathway. Here we show that the same region also contains a motif that interacts with Jun activation-domain-binding protein 1 (Jab1) which targets a common Smad, Smad4, shared by both the BMP and transforming growth factor-β (TGF-β) pathways, for proteasomal degradation. Jab1 was first identified as a coactivator of the transcription factor c-Jun. Jab1 binds to Smad4, Smad5, and Smad7, key intracellular signaling molecules of the TGF-β superfamily, and causes ubiquitination and/or degradation of these Smads. We confirmed a direct interaction of Jab1 with LMP-1 using recombinantly expressed wild-type and mutant proteins in slot-blot-binding assays. We hypothesized that LMP-1 binding to Jab1 prevents the binding and subsequent degradation of these Smads causing increased accumulation of osteogenic Smads in cells. We identified a sequence motif in LMP-1 that was predicted to interact with Jab1 based on the MAME/MAST sequence analysis of several cellular signaling molecules that are known to interact with Jab-1. We further mutated the potential key interacting residues in LMP-1 and showed loss of binding to Jab1 in binding

  1. Characterization of a unique motif in LIM mineralization protein-1 that interacts with jun activation-domain-binding protein 1

    PubMed Central

    Sangadala, Sreedhara; Yoshioka, Katsuhito; Enyo, Yoshio; Liu, Yunshan; Titus, Louisa; Boden, Scott D.

    2014-01-01

    Development and repair of the skeletal system and other organs are highly dependent on precise regulation of the bone morphogenetic protein (BMP) pathway. The use of BMPs clinically to induce bone formation has been limited in part by the requirement of much higher doses of recombinant proteins in primates than were needed in cell culture or rodents. Therefore, increasing cellular responsiveness to BMPs has become our focus. We determined that an osteogenic LIM mineralization protein, LMP-1 interacts with Smurf1 (Smad ubiquitin regulatory factor 1) and prevents ubiquitination of Smads resulting in potentiation of BMP activity. In the region of LMP-1 responsible for bone formation, there is a motif that directly interacts with the Smurf1 WW2 domain and thus effectively competes for binding with Smad1 and Smad5, key signaling proteins in the BMP pathway. Here we show that the same region also contains a motif that interacts with Jun activation-domain-binding protein 1 (Jab1) which targets a common Smad, Smad4, shared by both the BMP and transforming growth factor-β (TGF-β) pathways, for proteasomal degradation. Jab1 was first identified as a coactivator of the transcription factor c-Jun. Jab1 binds to Smad4, Smad5, and Smad7, key intracellular signaling molecules of the TGF-β superfamily, and causes ubiquiti-nation and/or degradation of these Smads. We confirmed a direct interaction of Jab1 with LMP-1 using recombinantly expressed wild-type and mutant proteins in slot-blot-binding assays. We hypothesized that LMP-1 binding to Jab1 prevents the binding and subsequent degradation of these Smads causing increased accumulation of osteogenic Smads in cells. We identified a sequence motif in LMP-1 that was predicted to interact with Jab1 based on the MAME/MAST sequence analysis of several cellular signaling molecules that are known to interact with Jab-1. We further mutated the potential key interacting residues in LMP-1 and showed loss of binding to Jab1 in binding

  2. LIM homeobox protein 5 (Lhx5) is essential for mamillary body development

    PubMed Central

    Miquelajáuregui, Amaya; Sandoval-Schaefer, Teresa; Martínez-Armenta, Miriam; Pérez-Martínez, Leonor; Cárabez, Alfonso; Zhao, Yangu; Heide, Michael; Alvarez-Bolado, Gonzalo; Varela-Echavarría, Alfredo

    2015-01-01

    The mamillary body (MM) is a group of hypothalamic nuclei related to memory and spatial navigation that interconnects hippocampal, thalamic, and tegmental regions. Here we demonstrate that Lhx5, a LIM-HD domain transcription factor expressed early in the developing posterior hypothalamus, is required for the generation of the MM and its derived tracts. The MM markers Foxb1, Sim2, and Lhx1 are absent in Lhx5 knock-out mice from early embryonic stages, suggesting abnormal specification of this region. This was supported by the absence of Nkx2.1 and expansion of Shh in the prospective mamillary area. Interestingly, we also found an ectopic domain expressing Lhx2 and Lhx9 along the anterio-posterior hypothalamic axis. Our results suggest that Lhx5 controls early aspects of hypothalamic development by regulating gene expression and cellular specification in the prospective MM. PMID:26578897

  3. LIM homeobox protein 5 (Lhx5) is essential for mamillary body development.

    PubMed

    Miquelajáuregui, Amaya; Sandoval-Schaefer, Teresa; Martínez-Armenta, Miriam; Pérez-Martínez, Leonor; Cárabez, Alfonso; Zhao, Yangu; Heide, Michael; Alvarez-Bolado, Gonzalo; Varela-Echavarría, Alfredo

    2015-01-01

    The mamillary body (MM) is a group of hypothalamic nuclei related to memory and spatial navigation that interconnects hippocampal, thalamic, and tegmental regions. Here we demonstrate that Lhx5, a LIM-HD domain transcription factor expressed early in the developing posterior hypothalamus, is required for the generation of the MM and its derived tracts. The MM markers Foxb1, Sim2, and Lhx1 are absent in Lhx5 knock-out mice from early embryonic stages, suggesting abnormal specification of this region. This was supported by the absence of Nkx2.1 and expansion of Shh in the prospective mamillary area. Interestingly, we also found an ectopic domain expressing Lhx2 and Lhx9 along the anterio-posterior hypothalamic axis. Our results suggest that Lhx5 controls early aspects of hypothalamic development by regulating gene expression and cellular specification in the prospective MM. PMID:26578897

  4. The molecular interaction of heart LIM protein (HLP) with RyR2 and caveolin-3 is essential for Ca(2+)-induced Ca(2+) release in the heart.

    PubMed

    Song, Dong Woo; Lee, Kyung-Eun; Ryu, Jae Yong; Jeon, Hyesung; Kim, Do Han

    2015-08-01

    The heart LIM protein (HLP) is a LIM-only protein family member that mediates protein-protein interactions. To date, no studies have yet been conducted regarding its function in the heart. In the present study, we have identified that HLP binds the cytosolic region of RyR2 in the heart using a bacterial two-hybrid system, LC-MS/MS, co-immunoprecipitation, and GST-pull down assays. Microscopy revealed that HLP forms a triple complex with RyR2 and caveolin-3. siRNA and adenovirus-mediated KD of HLP decreased the electrically evoked Ca(2+) release from the sarcoplasmic reticulum without directly affecting SERCA2 and RyR2 activities. Collectively, the HLP-RyR2 interaction in the cell surface caveolae region may be essential for efficient excitation-contraction coupling in the heart. PMID:26071359

  5. Dral Is a P53-Responsive Gene Whose Four and a Half Lim Domain Protein Product Induces Apoptosis

    PubMed Central

    Scholl, Florence A.; McLoughlin, Patricia; Ehler, Elisabeth; de Giovanni, Carla; Schäfer, Beat W.

    2000-01-01

    DRAL is a four and a half LIM domain protein identified because of its differential expression between normal human myoblasts and the malignant counterparts, rhabdomyosarcoma cells. In the current study, we demonstrate that transcription of the DRAL gene can be stimulated by p53, since transient expression of functional p53 in rhabdomyosarcoma cells as well as stimulation of endogenous p53 by ionizing radiation in wild-type cells enhances DRAL mRNA levels. In support of these observations, five potential p53 target sites could be identified in the promoter region of the human DRAL gene. To obtain insight into the possible functions of DRAL, ectopic expression experiments were performed. Interestingly, DRAL expression efficiently triggered apoptosis in three cell lines of different origin to the extent that no cells could be generated that stably overexpressed this protein. However, transient transfection experiments as well as immunofluorescence staining of the endogenous protein allowed for the localization of DRAL in different cellular compartments, namely cytoplasm, nucleus, focal contacts, as well as Z-discs and to a lesser extent the M-bands in cardiac myofibrils. These data suggest that downregulation of DRAL might be involved in tumor development. Furthermore, DRAL expression might be important for heart function. PMID:11062252

  6. Modeling and analysis of molecularinteraction between Smurf1-WW2 domain and various isoforms of LIM mineralization protein.

    PubMed

    Sangadala, Sreedhara; Boden, Scott D; Metpally, Raghu Prasad Rao; Reddy, Boojala Vijay B

    2007-08-15

    LIM Mineralization Protein-1 (LMP-1) has been cloned and shown to be osteoinductive. Our efforts to understand the mode of action of LMP-1 led to the determination that LMP-1 interacts with Smad Ubiquitin Regulatory Factor-1 (Smurf1). Smurf1 targets osteogenic Smads, Smad1/5, for ubiquitin-mediated proteasomal degradation. Smurf1 interaction with LMP-1 or Smads is based on the presence of unique WW-domain interacting motif in these target molecules. By performing site-directed mutagenesis and binding studies in vitro on purified recombinant proteins, we identified a specific motif within the osteogenic region of several LMP isoforms that is necessary for Smurf1 interaction. Similarly, we have identified that the WW2 domain of Smurf1 is necessary for target protein interaction. Here, we present a homology-based modeling of the Smurf1 WW2 domain and its interacting motif of LMP-1. We performed computational docking of the interacting domains in Smurf1 and LMPs to identify the key amino acid residues involved in their binding regions. In support of the computational predictions, we also present biochemical evidence supporting the hypothesis that the physical interaction of Smurf1 and osteoinductive forms of LMP may prevent Smurf1 from targeting osteogenic Smads by ubiquitin-mediated proteasomal degradation. PMID:17510966

  7. The transcriptional co-factor Chip acts with LIM-homeodomain proteins to set the boundary of the eye field in Drosophila

    PubMed Central

    Roignant, Jean-Yves; Legent, Kevin; Janody, Florence; Treisman, Jessica E.

    2010-01-01

    Development involves the establishment of boundaries between fields specified to differentiate into distinct tissues. The Drosophila larval eye-antennal imaginal disc must be subdivided into regions that differentiate into the adult eye, antenna and head cuticle. We have found that the transcriptional co-factor Chip is required for cells at the ventral eye-antennal disc border to take on a head cuticle fate; clones of Chip mutant cells in this region instead form outgrowths that differentiate into ectopic eye tissue. Chip acts independently of the transcription factor Homothorax, which was previously shown to promote head cuticle development in the same region. Chip and its vertebrate CLIM homologues have been shown to form complexes with LIM-homeodomain transcription factors, and the domain of Chip that mediates these interactions is required for its ability to suppress the eye fate. We show that two LIM-homeodomain proteins, Arrowhead and Lim1, are expressed in the region of the eye-antennal disc affected in Chip mutants, and that both require Chip for their ability to suppress photoreceptor differentiation when misexpressed in the eye field. Loss-of-function studies support the model that Arrowhead and Lim1 act redundantly, using Chip as a co-factor, to prevent retinal differentiation in regions of the eye disc destined to become ventral head tissue. PMID:20040493

  8. Cytoskeleton-interacting LIM-domain protein CRP1 suppresses cell proliferation and protects from stress-induced cell death

    SciTech Connect

    Latonen, Leena; Jaervinen, Paeivi M.; Laiho, Marikki

    2008-02-15

    Members of the cysteine-rich protein (CRP) family are actin cytoskeleton-interacting LIM-domain proteins known to act in muscle cell differentiation. We have earlier found that CRP1, a founding member of this family, is transcriptionally induced by UV radiation in human diploid fibroblasts [M. Gentile, L. Latonen, M. Laiho, Cell cycle arrest and apoptosis provoked by UV radiation-induced DNA damage are transcriptionally highly divergent responses, Nucleic Acids Res. 31 (2003) 4779-4790]. Here we show that CRP1 is induced by growth-inhibitory signals, such as increased cellular density, and cytotoxic stress induced by UV radiation or staurosporine. We found that high levels of CRP1 correlate with differentiation-associated morphology towards the myofibroblast lineage and that expression of ectopic CRP1 suppresses cell proliferation. Following UV- and staurosporine-induced stresses, expression of CRP1 provides a survival advantage evidenced by decreased cellular death and increased cellular metabolic activity and attachment. Our studies identify that CRP1 is a novel stress response factor, and provide evidence for its growth-inhibitory and cytoprotective functions.

  9. Ex-vivo transduced autologous skin fibroblasts expressing human Lim Mineralization Protein-3 efficiently form new bone in animal models

    PubMed Central

    Lattanzi, Wanda; Parrilla, Claudio; Fetoni, Annarita; Logroscino, Giandomenico; Straface, Giuseppe; Pecorini, Giovanni; Stigliano, Egidio; Tampieri, Anna; Bedini, Rossella; Pecci, Raffaella; Michetti, Fabrizio; Gambotto, Andrea; Robbins, Paul D.; Pola, Enrico

    2012-01-01

    Local gene transfer of the human LIM Mineralization Protein (LMP), a novel intracellular positive regulator of the osteoblast differentiation program, can induce efficient bone formation in rodents. In order to develop a clinically relevant gene therapy approach to facilitate bone healing, we have used primary dermal fibroblasts transduced ex vivo with Ad.LMP3 and seeded on an hydroxyapatite/collagen matrix prior to autologous implantation. Here we demonstrate that genetically modified autologous dermal fibroblasts expressing Ad.LMP-3 are able to induce ectopic bone formation following implantation of the matrix into the mouse triceps and paravertebral muscles. Moreover, implantation of the Ad.LMP-3-modified dermal fibroblasts into a rat mandibular bone critical size defect model results in efficient healing as determined by X-ray, histology and three dimensional micro computed tomography (3DμCT). These results demonstrate the effectiveness of the non-secreted intracellular osteogenic factor LMP-3, in inducing bone formation in vivo. Moreover, the utilization of autologous dermal fibroblasts implanted on a biomaterial represents a promising approach for possible future clinical applications aimed at inducing new bone formation. PMID:18633445

  10. Drosophila, a genetic model system to study cocaine-related behaviors: A review with focus on LIM-only proteins

    PubMed Central

    Heberlein, Ulrike; Tsai, Linus T.–Y.; Kapfhamer, David; Lasek, Amy W.

    2010-01-01

    Summary In the last decade, the fruit fly Drosophila melanogaster, highly accessible to genetic, behavioral and molecular analyses, has been introduced as a novel model organism to help decipher the complex genetic, neurochemical, and neuroanatomical underpinnings of behaviors induced by drugs of abuse. Here we review these data, focusing specifically on cocaine-related behaviors. Several of cocaine's most characteristic properties have been recapitulated in Drosophila. First, cocaine induces motor behaviors in flies that are remarkably similar to those observed in mammals. Second, repeated cocaine administration induces behavioral sensitization a form of behavioral plasticity believed to underlie certain aspects of addiction. Third, a key role for dopaminergic systems in mediating cocaine's effects has been demonstrated through both pharmacological and genetic methods. Finally, and most importantly, unbiased genetic screens, feasible because of the simplicity and scale with which flies can be manipulated in the laboratory, have identified several novel genes and pathways whose role in cocaine behaviors had not been anticipated. Many of these genes and pathways have been validated in mammalian models of drug addiction. We focus in this review on the role of LIM-only proteins in cocaine-induced behaviors. PMID:18694769

  11. Deficiency in four and one half LIM domain protein 2 (FHL2) aggravates liver fibrosis in mice

    PubMed Central

    2013-01-01

    Background Four and one half LIM domain protein 2 (FHL2) has been reported to be a key regulator in many cellular processes being associated with fibrogenesis such as cell migration and contraction. Moreover, hepatic FHL2 is involved in regulation pathways mediating proliferation and cell death machineries. We here investigated the role of FHL2 in the setting of experimental and clinical liver fibrosis. Methods FHL2−/− and wild type (wt) mice were challenged with CCl4. Fibrotic response was assessed by quantitative real time PCR (qRT-PCR) of fibrotic marker genes, measurement of hydroxyproline content and histological methods. Murine FHL2−/− and hepatic stellate cells (HSC) were isolated and investigated via immunofluorescence. Human fibrotic and normal liver samples were analysed immunohistochemically using antibodies directed against FHL2. Results FHL2−/− mice displayed aggravated liver fibrosis compared to wt mice. However, immunofluorescence revealed no significant morphological changes in cultured FHL2−/− and wt myofibroblasts (MFB). In human liver samples, FHL2 was strongly expressed both in the nucleus and cytoplasm in MFB of fibrotic livers. In contrast, FHL2 expression was absent in normal liver tissue. Conclusions Deficiency of FHL2 results in aggravation of murine liver fibrosis. In human liver samples, FHL2 is expressed in activated HSCs and portal fibroblasts in human fibrotic livers, pointing to a central role of FHL2 for human hepatic fibrogenesis as well. PMID:23311569

  12. The Ras suppressor Rsu-1 binds to the LIM 5 domain of the adaptor protein PINCH1 and participates in adhesion-related functions

    SciTech Connect

    Dougherty, Gerard W.; Chopp, Treasa; Qi Shengmei; Cutler, Mary Lou . E-mail: mcutler@usuhs.mil

    2005-05-15

    Rsu-1 is a highly conserved leucine rich repeat (LRR) protein that is expressed ubiquitously in mammalian cells. Rsu-1 was identified based on its ability to inhibit transformation by Ras, and previous studies demonstrated that ectopic expression of Rsu-1 inhibited anchorage-independent growth of Ras-transformed cells and human tumor cell lines. Using GAL4-based yeast two-hybrid screening, the LIM domain protein, PINCH1, was identified as the binding partner of Rsu-1. PINCH1 is an adaptor protein that localizes to focal adhesions and it has been implicated in the regulation of adhesion functions. Subdomain mapping in yeast revealed that Rsu-1 binds to the LIM 5 domain of PINCH1, a region not previously identified as a specific binding domain for any other protein. Additional testing demonstrated that PINCH2, which is highly homologous to PINCH1, except in the LIM 5 domain, does not interact with Rsu-1. Glutathione transferase fusion protein binding studies determined that the LRR region of Rsu-1 interacts with PINCH1. Transient expression studies using epitope-tagged Rsu-1 and PINCH1 revealed that Rsu-1 co-immunoprecipitated with PINCH1 and colocalized with vinculin at sites of focal adhesions in mammalian cells. In addition, endogenous P33 Rsu-1 from 293T cells co-immunoprecipitated with transiently expressed myc-tagged PINCH1. Furthermore, RNAi-induced reduction in Rsu-1 RNA and protein inhibited cell attachment, and while previous studies demonstrated that ectopic expression of Rsu-1 inhibited Jun kinase activation, the depletion of Rsu-1 resulted in activation of Jun and p38 stress kinases. These studies demonstrate that Rsu-1 interacts with PINCH1 in mammalian cells and functions, in part, by altering cell adhesion.

  13. The Structure of an LIM-Only Protein 4 (LMO4) and Deformed Epidermal Autoregulatory Factor-1 (DEAF1) Complex Reveals a Common Mode of Binding to LMO4

    PubMed Central

    Joseph, Soumya; Kwan, Ann H.; Stokes, Philippa H.; Mackay, Joel P.; Cubeddu, Liza; Matthews, Jacqueline M.

    2014-01-01

    LIM-domain only protein 4 (LMO4) is a widely expressed protein with important roles in embryonic development and breast cancer. It has been reported to bind many partners, including the transcription factor Deformed epidermal autoregulatory factor-1 (DEAF1), with which LMO4 shares many biological parallels. We used yeast two-hybrid assays to show that DEAF1 binds both LIM domains of LMO4 and that DEAF1 binds the same face on LMO4 as two other LMO4-binding partners, namely LIM domain binding protein 1 (LDB1) and C-terminal binding protein interacting protein (CtIP/RBBP8). Mutagenic screening analysed by the same method, indicates that the key residues in the interaction lie in LMO4LIM2 and the N-terminal half of the LMO4-binding domain in DEAF1. We generated a stable LMO4LIM2-DEAF1 complex and determined the solution structure of that complex. Although the LMO4-binding domain from DEAF1 is intrinsically disordered, it becomes structured on binding. The structure confirms that LDB1, CtIP and DEAF1 all bind to the same face on LMO4. LMO4 appears to form a hub in protein-protein interaction networks, linking numerous pathways within cells. Competitive binding for LMO4 therefore most likely provides a level of regulation between those different pathways. PMID:25310299

  14. LIM mineralization protein-1 potentiates bone morphogenetic protein responsiveness via a novel interaction with Smurf1 resulting in decreased ubiquitination of Smads.

    PubMed

    Sangadala, Sreedhara; Boden, Scott D; Viggeswarapu, Manjula; Liu, Yunshan; Titus, Louisa

    2006-06-23

    Development and repair of the skeletal system and other organs is highly dependent on precise regulation of bone morphogenetic proteins (BMPs), their receptors, and their intracellular signaling proteins known as Smads. The use of BMPs clinically to induce bone formation has been limited in part by the requirement of much higher doses of recombinant proteins in primates than were needed in cell culture or rodents. Therefore, control of cellular responsiveness to BMPs is now a critical area that is poorly understood. We determined that LMP-1, a LIM domain protein capable of inducing de novo bone formation, interacts with Smurf1 (Smad ubiquitin regulatory factor 1) and prevents ubiquitination of Smads. In the region of LMP responsible for bone formation, there is a motif that directly interacts with the Smurf1 WW2 domain and can effectively compete with Smad1 and Smad5 for binding. We have shown that small peptides containing this motif can mimic the ability to block Smurf1 from binding Smads. This novel interaction of LMP-1 with the WW2 domain of Smurf1 to block Smad binding results in increased cellular responsiveness to exogenous BMP and demonstrates a novel regulatory mechanism for the BMP signaling pathway. PMID:16611643

  15. Human ESP1/CRP2, a member of the LIM domain protein family: Characterization of the cDNA and assignment of the gene locus to chromosome 14q32.3

    SciTech Connect

    Karim, Mohammad Azharul; Ohta, Kohji; Matsuda, Ichiro

    1996-01-15

    The LIM domain is present in a wide variety of proteins with diverse functions and exhibits characteristic arrangements of Cys and His residues with a novel zinc-binding motif. LIM domain proteins have been implicated in development, cell regulation, and cell structure. A LIM domain protein was identified by screening a human cDNA library with rat cysteine-rich intestinal protein (CRIP) as a probe, under conditions of low stringency. Comparison of the predicted amino acid sequence with several LIM domain proteins revealed 93% of the residues to be identical to rat LIM domain protein, termed ESP1 or CRP2. Thus, the protein is hereafter referred to as human ESP1/CRP2. The cDNA encompasses a 1171-base region, including 26, 624, and 521 bases in the 5{prime}-noncoding region, coding region, and 3{prime}-noncoding regions, respectively, and encodes the entire ESP1/CRP2 protein has two LIM domains, and each shares 35.1% and 77 or 79% identical residues with human cysteine-rich protein (CRP) and rat CRIP, respectively. Northern blot analysis of ESP1/CRP2 in various human tissues showed distinct tissue distributions compared with CRP and CRIP, suggesting that each might serve related but specific roles in tissue organization or function. Using a panel of human-rodent somatic cell hybrids, the ESP1/CRP2 locus was assigned to chromosome 14. Fluorescence in situ hybridization, using cDNA and a genome DNA fragment of the ESP1/CRP2 as probes, confirms this assignment and relegates regional localization to band 14q32.3 47 refs., 7 figs.

  16. MamK, a bacterial actin, forms dynamic filaments in vivo that are regulated by the acidic proteins MamJ and LimJ

    PubMed Central

    Draper, Olga; Byrne, Meghan E.; Li, Zhuo; Keyhani, Sepehr; Cueto Barrozo, Joyce; Jensen, Grant; Komeili, Arash

    2011-01-01

    SUMMARY Bacterial actins, in contrast to their eukaryotic counterparts, are highly divergent proteins whose wide-ranging functions are thought to correlate with their evolutionary diversity. One clade, represented by the MamK protein of magnetotactic bacteria, is required for the subcellular organization of magnetosomes, membrane-bound organelles that aid in navigation along the earth’s magnetic field. Using a fluorescence recovery after photobleaching assay in Magnetospirillum magneticum AMB-1, we find that, like traditional actins, MamK forms dynamic filaments that require an intact NTPase motif for their turnover in vivo. We also uncover two proteins, MamJ and LimJ, which perform a redundant function to promote the dynamic behavior of MamK filaments in wildtype cells. The absence of both MamJ and LimJ leads to static filaments, a disrupted magnetosome chain, and an anomalous build-up of cytoskeletal filaments between magnetosomes. Our results suggest that MamK filaments, like eukaryotic actins, are intrinsically stable and rely on regulators for their dynamic behavior, a feature that stands in contrast to some classes of bacterial actins characterized to date. PMID:21883528

  17. RNA-LIM: a novel procedure for analyzing protein/single-stranded RNA propensity data with concomitant estimation of interface structure.

    PubMed

    Hall, Damien; Li, Songling; Yamashita, Kazuo; Azuma, Ryuzo; Carver, John A; Standley, Daron M

    2015-03-01

    RNA-LIM is a procedure that can analyze various pseudo-potentials describing the affinity between single-stranded RNA (ssRNA) ribonucleotides and surface amino acids to produce a coarse-grained estimate of the structure of the ssRNA at the protein interface. The search algorithm works by evolving an ssRNA chain, of known sequence, as a series of walks between fixed sites on a protein surface. Optimal routes are found by application of a set of minimal "limiting" restraints derived jointly from (i) selective sampling of the ribonucleotide amino acid affinity pseudo-potential data, (ii) limited surface path exploration by prior determination of surface arc lengths, and (iii) RNA structural specification obtained from a statistical potential gathered from a library of experimentally determined ssRNA structures. We describe the general approach using a NAST (Nucleic Acid Simulation Tool)-like approximation of the ssRNA chain and a generalized pseudo-potential reflecting the location of nucleic acid binding residues. Minimum and maximum performance indicators of the methodology are established using both synthetic data, for which the pseudo-potential defining nucleic acid binding affinity is systematically degraded, and a representative real case, where the RNA binding sites are predicted by the amplified antisense RNA (aaRNA) method. Some potential uses and extensions of the routine are discussed. RNA-LIM analysis programs along with detailed instructions for their use are available on request from the authors. PMID:25479604

  18. PARPs database: A LIMS systems for protein-protein interaction data mining or laboratory information management system

    PubMed Central

    Droit, Arnaud; Hunter, Joanna M; Rouleau, Michèle; Ethier, Chantal; Picard-Cloutier, Aude; Bourgais, David; Poirier, Guy G

    2007-01-01

    Background In the "post-genome" era, mass spectrometry (MS) has become an important method for the analysis of proteins and the rapid advancement of this technique, in combination with other proteomics methods, results in an increasing amount of proteome data. This data must be archived and analysed using specialized bioinformatics tools. Description We herein describe "PARPs database," a data analysis and management pipeline for liquid chromatography tandem mass spectrometry (LC-MS/MS) proteomics. PARPs database is a web-based tool whose features include experiment annotation, protein database searching, protein sequence management, as well as data-mining of the peptides and proteins identified. Conclusion Using this pipeline, we have successfully identified several interactions of biological significance between PARP-1 and other proteins, namely RFC-1, 2, 3, 4 and 5. PMID:18093328

  19. The LIM domain only 4 protein is a metabolic responsive inhibitor of protein tyrosine phosphatase 1B that controls hypothalamic leptin signaling.

    PubMed

    Pandey, Nihar R; Zhou, Xun; Qin, Zhaohong; Zaman, Tariq; Gomez-Smith, Mariana; Keyhanian, Kianoosh; Anisman, Hymie; Brunel, Jean Michel; Stewart, Alexandre F R; Chen, Hsiao-Huei

    2013-07-31

    Protein tyrosine phosphatase 1B (PTP1B) counteracts leptin signaling and is a therapeutic target for obesity and diabetes. Here we found that LIM domain only 4 (LMO4) inhibits PTP1B activity by increasing the oxidized inactive form of PTP1B. Mice with neuronal ablation of LMO4 have elevated PTP1B activity and impaired hypothalamic leptin signaling, and a PTP1B inhibitor normalized PTP1B activity and restored leptin control of circulating insulin levels. LMO4 is palmitoylated at its C-terminal cysteine, and deletion of this residue prevented palmitoylation and retention of LMO4 at the endoplasmic reticulum and abolished its inhibitory effect on PTP1B. Importantly, LMO4 palmitoylation is sensitive to metabolic stress; mice challenged with a brief high-fat diet or acute intracerebroventricular infusion of saturated fatty acid had less palmitoylated LMO4, less oxidized PTP1B, and increased PTP1B activity in the hypothalamus. Thus, unleashed PTP1B activity attributable to loss of LMO4 palmitoylation may account for rapid loss of central leptin signaling after acute exposure to saturated fat. PMID:23904601

  20. LIM-domain protein cysteine- and glycine-rich protein 2 (CRP2) is a novel marker of hepatic stellate cells and binding partner of the protein inhibitor of activated STAT1.

    PubMed Central

    Weiskirchen, R; Moser, M; Weiskirchen, S; Erdel, M; Dahmen, S; Buettner, R; Gressner, A M

    2001-01-01

    Activation of hepatic stellate cells is considered to be the main step in the development of liver fibrosis, which is characterized by the transition of quiescent vitamin-A-rich cells to proliferative, fibrogenic and contractile myofibroblasts. The identification of regulatory genes during early cell activation and transdifferentiation is essential to extend our knowledge of hepatic fibrogenesis. In liver, the gene CSRP2 is exclusively expressed by stellate cells, whereas no transcripts are detectable in hepatocytes, sinusoidal endothelial cells or Kupffer cells. The early activation of stellate cells induced by platelet-derived growth factor is accompanied by an enhanced expression of CSRP2. During later stages of transdifferentiation, the expression of CSRP2 in these cells is suppressed in vitro and in vivo. The CSRP2-encoded cysteine- and glycine-rich double-LIM-domain protein (CRP)2 is proposed to function as a molecular adapter, arranging two or more as yet unidentified protein constituents into a macromolecular complex. To identify these proteins and assign a cellular function to CRP2, a human cDNA library was screened with full-length CRP2 as bait in a yeast two-hybrid screen. The protein inhibitor of activated STAT1 ('PIAS1') was shown to associate selectively with the C-terminal LIM domain of CRP2. Physical interaction of both proteins in the cellular environment was confirmed by co-localization experiments with confocal laser scanning microscopy and co-immunoprecipitation analysis. These results establish CRP2 as a potential new factor in the JAK/STAT-signalling pathway and suggest that the suppression of CSRP2 might be a prerequisite for the myofibroblastic transition of hepatic stellate cells. PMID:11672422

  1. Genome Wide Identification of LIM Genes in Cicer arietinum and Response of Ca-2LIMs in Development, Hormone and Pathogenic Stress

    PubMed Central

    Srivastava, Vikas; Verma, Praveen Kumar

    2015-01-01

    The eukaryotic lineage-specific LIM protein (LIN11, ISL1, and MEC3) family play pivotal role in modulation of actin dynamics and transcriptional regulation. The systematic investigation of this family has not been carried in detail and rare in legumes. Current study involves the mining of Cicer arietinum genome for the genes coding for LIM domain proteins and displayed significant homology with LIM genes of other species. The analysis led to the identification of 15 members, which were positioned on chickpea chromosomes. The phylogenetic and motif analysis suggested their categorization into two sub-families i.e., Ca-2LIMs and Ca-DA1/DAR, which comprised of nine and six candidates, respectively. Further sub-categories of Ca-2LIMs were recognised as αLIM, βLIM, δLIM and γLIM. The LIM genes within their sub-families displayed conserved genomic and motif organization. The expression pattern of Ca-2LIMs across developmental and reproductive tissues demonstrated strong correlation with established consensus. The Ca-2LIM belongs to PLIM and GLIM (XLIM) was found highly expressed in floral tissue. Others showed ubiquitous expression pattern with their dominance in stem. Under hormonal and pathogenic conditions these LIMs were found to up-regulate during salicylic acid, abscisic acid and Ascochyta rabiei treatment or infection; and down-regulated in response to jasmonic acid treatment. The findings of this work, particularly in terms of modulation of LIM genes under biotic stress will open up the way to further explore and establish the role of chickpea LIMs in plant defense response. PMID:26418014

  2. Genome Wide Identification of LIM Genes in Cicer arietinum and Response of Ca-2LIMs in Development, Hormone and Pathogenic Stress.

    PubMed

    Srivastava, Vikas; Verma, Praveen Kumar

    2015-01-01

    The eukaryotic lineage-specific LIM protein (LIN11, ISL1, and MEC3) family play pivotal role in modulation of actin dynamics and transcriptional regulation. The systematic investigation of this family has not been carried in detail and rare in legumes. Current study involves the mining of Cicer arietinum genome for the genes coding for LIM domain proteins and displayed significant homology with LIM genes of other species. The analysis led to the identification of 15 members, which were positioned on chickpea chromosomes. The phylogenetic and motif analysis suggested their categorization into two sub-families i.e., Ca-2LIMs and Ca-DA1/DAR, which comprised of nine and six candidates, respectively. Further sub-categories of Ca-2LIMs were recognised as αLIM, βLIM, δLIM and γLIM. The LIM genes within their sub-families displayed conserved genomic and motif organization. The expression pattern of Ca-2LIMs across developmental and reproductive tissues demonstrated strong correlation with established consensus. The Ca-2LIM belongs to PLIM and GLIM (XLIM) was found highly expressed in floral tissue. Others showed ubiquitous expression pattern with their dominance in stem. Under hormonal and pathogenic conditions these LIMs were found to up-regulate during salicylic acid, abscisic acid and Ascochyta rabiei treatment or infection; and down-regulated in response to jasmonic acid treatment. The findings of this work, particularly in terms of modulation of LIM genes under biotic stress will open up the way to further explore and establish the role of chickpea LIMs in plant defense response. PMID:26418014

  3. Induction of senescence-like phenotypes by forced expression of hic-5, which encodes a novel LIM motif protein, in immortalized human fibroblasts.

    PubMed Central

    Shibanuma, M; Mochizuki, E; Maniwa, R; Mashimo, J; Nishiya, N; Imai, S; Takano, T; Oshimura, M; Nose, K

    1997-01-01

    The hic-5 gene encodes a novel protein with Zn finger-like (LIM) motifs, the expression of which increases during cellular senescence. The ectopic expression of hic-5 in nontumorigenic immortalized human fibroblasts, whose expression levels of hic-5 were significantly reduced in comparison with those of mortal cells, decreased colony-forming efficiency. Stable clones expressing high levels of hic-5 mRNA showed higher levels of mRNAs for several extracellular matrix-related proteins, along with the alteration of an alternative splicing as seen in senescent cells and decreased c-fos inducibility. Furthermore, these clones acquired a senescence-like phenotype, such as growth retardation; senescence-like morphology; and increased expression of Cip1/WAF1/sdi1 after 20 to 40 population doublings. On the other hand, antisense RNA expression of hic-5 in human normal diploid fibroblasts delayed the senescence process. HIC-5 was localized in nuclei and had affinity for DNA. Based on these observations, we speculated that HIC-5 affected the expression of senescence-related genes through interacting with DNA and thereby induced the senescence-like phenotypes. To our knowledge, hic-5 is the first single gene that could induce senescence-like phenotypes in a certain type of immortalized human cell and mediate the normal process of senescence. PMID:9032249

  4. The LIM protein leupaxin is enriched in smooth muscle and functions as an serum response factor cofactor to induce smooth muscle cell gene transcription.

    PubMed

    Sundberg-Smith, Liisa J; DiMichele, Laura A; Sayers, Rebecca L; Mack, Christopher P; Taylor, Joan M

    2008-06-20

    Leupaxin is a LIM domain-containing adapter protein belonging to the paxillin family that has been previously reported to be preferentially expressed in hematopoietic cells. Herein, we identified leupaxin in a screen for focal adhesion kinase binding partners in aortic smooth muscle, and we show that leupaxin is enriched in human and mouse vascular smooth muscle and that leupaxin expression is dynamically regulated during development. In addition, our studies reveal that leupaxin can undergo cytoplasmic/nuclear shuttling and functions as an serum response factor cofactor in the nucleus. We found that leupaxin forms a complex with serum response factor and associates with CArG-containing regions of smooth muscle promoters and that ectopic expression of leupaxin induces smooth muscle marker gene expression in both 10T1/2 cells and rat aortic smooth muscle cells. Subsequent studies indicated that enhanced focal adhesion kinase activity (induced by fibronectin or expression of constitutively active focal adhesion kinase) attenuates the nuclear accumulation of leupaxin and limits the ability of leupaxin to enhance serum response factor-dependent gene transcription. Thus, these studies indicate that modulation of the subcellular localization of serum response factor cofactors is 1 mechanism by which extracellular matrix-dependent signals may regulate phenotypic switching of smooth muscle cells. PMID:18497331

  5. Exome sequencing identifies variants in two genes encoding the LIM-proteins NRAP and FHL1 in an Italian patient with BAG3 myofibrillar myopathy.

    PubMed

    D'Avila, Francesca; Meregalli, Mirella; Lupoli, Sara; Barcella, Matteo; Orro, Alessandro; De Santis, Francesca; Sitzia, Clementina; Farini, Andrea; D'Ursi, Pasqualina; Erratico, Silvia; Cristofani, Riccardo; Milanesi, Luciano; Braga, Daniele; Cusi, Daniele; Poletti, Angelo; Barlassina, Cristina; Torrente, Yvan

    2016-06-01

    Myofibrillar myopathies (MFMs) are genetically heterogeneous dystrophies characterized by the disintegration of Z-disks and myofibrils and are associated with mutations in genes encoding Z-disk or Z-disk-related proteins. The c.626 C > T (p.P209L) mutation in the BAG3 gene has been described as causative of a subtype of MFM. We report a sporadic case of a 26-year-old Italian woman, affected by MFM with axonal neuropathy, cardiomyopathy, rigid spine, who carries the c.626 C > T mutation in the BAG3 gene. The patient and her non-consanguineous healthy parents and brother were studied with whole exome sequencing (WES) to further investigate the genetic basis of this complex phenotype. In the patient, we found that the BAG3 mutation is associated with variants in the NRAP and FHL1 genes that encode muscle-specific, LIM domain containing proteins. Quantitative real time PCR, immunohistochemistry and Western blot analysis of the patient's muscular biopsy showed the absence of NRAP expression and FHL1 accumulation in aggregates in the affected skeletal muscle tissue. Molecular dynamic analysis of the mutated FHL1 domain showed a modification in its surface charge, which could affect its capability to bind its target proteins. To our knowledge this is the first study reporting, in a BAG3 MFM, the simultaneous presence of genetic variants in the BAG3 and FHL1 genes (previously described as independently associated with MFMs) and linking the NRAP gene to MFM for the first time. PMID:27443559

  6. Nuclear localization of lymphocyte-specific protein tyrosine kinase (Lck) and its role in regulating LIM domain only 2 (Lmo2) gene

    SciTech Connect

    Venkitachalam, Srividya; Chueh, Fu-Yu; Yu, Chao-Lan

    2012-01-20

    Highlights: Black-Right-Pointing-Pointer Lmo2 expression is elevated in Lck-transformed cells. Black-Right-Pointing-Pointer Both endogenous and exogenous Lck localize in the nucleus. Black-Right-Pointing-Pointer Nuclear Lck is active in Lck-transformed cells. Black-Right-Pointing-Pointer Lck binds to the promoter region of Lmo2 gene in vivo. Black-Right-Pointing-Pointer In contrast to JAK2, Lck does not increase histone H3 phosphorylation on Tyr 41. -- Abstract: LIM domain only protein 2 (Lmo2) is a transcription factor that plays a critical role in the development of T-acute lymphoblastic leukemia (T-ALL). A previous report established a link between Lmo2 expression and the nuclear presence of oncogenic Janus kinase 2 (JAK2), a non-receptor protein tyrosine kinase. The oncogenic JAK2 kinase phosphorylates histone H3 on Tyr 41 that leads to the relief of Lmo2 promoter repression and subsequent gene expression. Similar to JAK2, constitutive activation of lymphocyte-specific protein tyrosine kinase (Lck) has been implicated in lymphoid malignancies. However, it is not known whether oncogenic Lck regulates Lmo2 expression through a similar mechanism. We show here that Lmo2 expression is significantly elevated in T cell leukemia LSTRA overexpressing active Lck kinase and in HEK 293 cells expressing oncogenic Y505FLck kinase. Nuclear localization of active Lck kinase was confirmed in both Lck-transformed cells by subcellular fractionation and immunofluorescence microscopy. More importantly, in contrast to oncogenic JAK2, oncogenic Lck kinase does not result in significant increase in histone H3 phosphorylation on Tyr 41. Instead, chromatin immunoprecipitation experiment shows that oncogenic Y505FLck kinase binds to the Lmo2 promoter in vivo. This result raises the possibility that oncogenic Lck may activate Lmo2 promoter through direct interaction.

  7. Androgen receptor coactivators lysine-specific histone demethylase 1 and four and a half LIM domain protein 2 predict risk of prostate cancer recurrence.

    PubMed

    Kahl, Philip; Gullotti, Lucia; Heukamp, Lukas Carl; Wolf, Susanne; Friedrichs, Nicolaus; Vorreuther, Roland; Solleder, Gerold; Bastian, Patrick J; Ellinger, Jörg; Metzger, Eric; Schüle, Roland; Buettner, Reinhard

    2006-12-01

    Prostate cancer biology varies from locally confined tumors with low risk for relapse to tumors with high risk for progression even after radical prostatectomy. Currently, there are no reliable biomarkers to predict tumor relapse and poor clinical outcome. In this study, we correlated expression patterns of the androgen receptor (AR) coactivators lysine-specific histone demethylase 1 (LSD1) and four and a half LIM-domain protein 2 (FHL2), AR, Gleason score, Gleason grade, and p53 expression in clinically organ confined prostate cancers with relapse after radical prostatectomy. Our data reveal that high levels of LSD1, nuclear expression of the FHL2 coactivator, high Gleason score and grade, and very strong staining of nuclear p53 correlate significantly with relapse during follow-up. No correlation exists with relapse and the expression of AR and cytoplasmic expression of FHL2. To confirm these data, we did quantitative reverse transcription-PCR and Western blot analyses in a subset of tumor specimens. Consistently, both LSD1 mRNA and protein levels were significantly up-regulated in high-risk tumors. We previously identified LSD1 and FHL2 as nuclear cofactors interacting specifically with the AR in prostate cells and showed that both stimulate androgen-dependent gene transcription. Our present study suggests that LSD1 and nuclear FHL2 may serve as novel biomarkers predictive for prostate cancer with aggressive biology and point to a role of LSD1 and FHL2 in constitutive activation of AR-mediated growth signals. PMID:17145880

  8. The LIM domain protein nTRIP6 acts as a co-repressor for the transcription factor MEF2C in myoblasts

    PubMed Central

    Kemler, Denise; Dahley, Oliver; Roßwag, Sven; Litfin, Margarethe; Kassel, Olivier

    2016-01-01

    The transcription factor Myocyte enhancer factor 2C (MEF2C) plays a key role in the late differentiation of skeletal muscle progenitor cells, the so-called myoblasts. During myoblast differentiation, both MEF2C expression and transcriptional activity are regulated. We have reported that nTRIP6, the nuclear isoform of the focal adhesion LIM domain protein TRIP6, acts as an adaptor transcriptional co-activator for several transcription factors. It interacts with the promoter-bound transcription factors and consequently mediates the recruitment of other co-activators. Based on a described interaction between MEF2C and TRIP6 in a yeast-two-hybrid screen, we hypothesised a co-regulatory function of nTRIP6 for MEF2C. In proliferating myoblasts, nTRIP6 interacted with MEF2C and was recruited together with MEF2C to the MEF2-binding regions of the MEF2C target genes Myom2, Mb, Tnni2 and Des. Silencing nTRIP6 or preventing its interaction with MEF2C increased MEF2C transcriptional activity and increased the expression of these MEF2C target genes. Thus, nTRIP6 acts as a co-repressor for MEF2C. Mechanistically, nTRIP6 mediated the recruitment of the class IIa histone deacetylase HDAC5 to the MEF2C-bound promoters. In conclusion, our results unravel a transcriptional co-repressor function for nTRIP6. This adaptor co-regulator can thus exert either co-activator or co-repressor functions, depending on the transcription factor it interacts with. PMID:27292777

  9. The LIM domain protein FHL2 interacts with the NR5A family of nuclear receptors and CREB to activate the inhibin-α subunit gene in ovarian granulosa cells.

    PubMed

    Matulis, Christina K; Mayo, Kelly E

    2012-08-01

    Nuclear receptor transcriptional activity is enhanced by interaction with coactivators. The highly related nuclear receptor 5A (NR5A) subfamily members liver receptor homolog 1 and steroidogenic factor 1 bind to and activate several of the same genes, many of which are important for reproductive function. To better understand transcriptional activation by these nuclear receptors, we sought to identify interacting proteins that might function as coactivators. The LIM domain protein four and a half LIM domain 2 (FHL2) was identified as interacting with the NR5A receptors in a yeast two-hybrid screen of a human ovary cDNA library. FHL2, and the closely related FHL1, are both expressed in the rodent ovary and in granulosa cells. Small interfering RNA-mediated knockdown of FHL1 and FHL2 in primary mouse granulosa cells reduced expression of the NR5A target genes encoding inhibin-α and P450scc. In vitro assays confirmed the interaction between the FHL and NR5A proteins and revealed that a single LIM domain of FHL2 is sufficient for this interaction, whereas determinants in both the ligand binding domain and DNA binding domain of NR5A proteins are important. FHL2 enhances the ability of both liver receptor homolog 1 and steroidogenic factor 1 to activate the inhibin-α subunit gene promoter in granulosa cells and thus functions as a transcriptional coactivator. FHL2 also interacts with cAMP response element-binding protein and substantially augments activation of inhibin gene expression by the combination of NR5A receptors and forskolin, suggesting that FHL2 may facilitate integration of these two signals. Collectively these results identify FHL2 as a novel coactivator of NR5A nuclear receptors in ovarian granulosa cells and suggest its involvement in regulating target genes important for mammalian reproduction. PMID:22734036

  10. LIMS strategy review

    SciTech Connect

    Ibsen, T.G.

    1995-03-30

    The evaluation team found that the LABCORE program has a viable Laboratory Information Management System (LIMS) that will handle the sample tracking and reporting necessary to accomplish the requirements established by the system specifications. The team also found that the LABCORE program has made real accomplishments and that it has been moving in the right direction, for example the phased implementation at the 222-S laboratory. The LABCORE program has many strengths but it also has short comings that need to be addressed. The strengths are that the hardware, software, staff, and the computer and training facilities are all first rate. The shortcomings are that it is missing a strategy outlining the goals which would clearly define when the project implementation has been successfully completed. It also had excessive cost during its acquisition. Specific recommendations are made. As a whole, the evaluation team believes that LABCORE is the system that was specified and will meet the needs of both laboratories, 222-S and the Waste Sampling and Characterization Facility. LABCORE has experienced all the challenges of a new computer system but with continued effort on the part of the LABCORE team and laboratory personnel the system will operate as needed.

  11. A Perl toolkit for LIMS development

    PubMed Central

    Morris, James A; Gayther, Simon A; Jacobs, Ian J; Jones, Christopher

    2008-01-01

    Background High throughput laboratory techniques generate huge quantities of scientific data. Laboratory Information Management Systems (LIMS) are a necessary requirement, dealing with sample tracking, data storage and data reporting. Commercial LIMS solutions are available, but these can be both costly and overly complex for the task. The development of bespoke LIMS solutions offers a number of advantages, including the flexibility to fulfil all a laboratory's requirements at a fraction of the price of a commercial system. The programming language Perl is a perfect development solution for LIMS applications because of Perl's powerful but simple to use database and web interaction, it is also well known for enabling rapid application development and deployment, and boasts a very active and helpful developer community. The development of an in house LIMS from scratch however can take considerable time and resources, so programming tools that enable the rapid development of LIMS applications are essential but there are currently no LIMS development tools for Perl. Results We have developed ArrayPipeline, a Perl toolkit providing object oriented methods that facilitate the rapid development of bespoke LIMS applications. The toolkit includes Perl objects that encapsulate key components of a LIMS, providing methods for creating interactive web pages, interacting with databases, error tracking and reporting, and user and session management. The MT_Plate object provides methods for manipulation and management of microtitre plates, while a given LIMS can be encapsulated by extension of the core modules, providing system specific methods for database interaction and web page management. Conclusion This important addition to the Perl developer's library will make the development of in house LIMS applications quicker and easier encouraging laboratories to create bespoke LIMS applications to meet their specific data management requirements. PMID:18353174

  12. Proliferating cell nuclear antigen (PCNA) interacts with a meiosis-specific RecA homologues, Lim15/Dmc1, but does not stimulate its strand transfer activity

    SciTech Connect

    Hamada, Fumika N.; Koshiyama, Akiyo; Namekawa, Satoshi H.; Ishii, Satomi; Iwabata, Kazuki; Sugawara, Hiroko; Nara, Takayuki Y.; Sakaguchi, Kengo . E-mail: kengo@rs.noda.tus.ac.jp; Sawado, Tomoyuki

    2007-01-26

    PCNA is a multi-functional protein that is involved in various nuclear events. Here we show that PCNA participates in events occurring during early meiotic prophase. Analysis of protein-protein interactions using surface plasmon resonance indicates that Coprinus cinereus PCNA (CoPCNA) specifically interacts with a meiotic specific RecA-like factor, C. cinereus Lim15/Dmc1 (CoLim15) in vitro. The binding efficiency increases with addition of Mg{sup 2+} ions, while ATP inhibits the interaction. Co-immunoprecipitation experiments indicate that the CoLim15 protein interacts with the CoPCNA protein in vitro and in the cell extracts. Despite the interaction between these two factors, no enhancement of CoLim15-dependent strand transfer activity by CoPCNA was found in vitro. We propose that the interaction between Lim15/Dmc1 and PCNA mediates the recombination-associated DNA synthesis during meiosis.

  13. Structural Basis of Focal Adhesion Localization of LIM-only Adaptor PINCH by Integrin-linked Kinase*S⃞

    PubMed Central

    Yang, Yanwu; Wang, Xiaoxia; Hawkins, Cheryl A.; Chen, Kan; Vaynberg, Julia; Mao, Xian; Tu, Yizeng; Zuo, Xiaobing; Wang, Jinbu; Wang, Yun-xing; Wu, Chuanyue; Tjandra, Nico; Qin, Jun

    2009-01-01

    The LIM-only adaptor PINCH (the particularly interesting cysteine- and histidine-rich protein) plays a pivotal role in the assembly of focal adhesions (FAs), supramolecular complexes that transmit mechanical and biochemical information between extracellular matrix and actin cytoskeleton, regulating diverse cell adhesive processes such as cell migration, cell spreading, and survival. A key step for the PINCH function is its localization to FAs, which depends critically on the tight binding of PINCH to integrin-linked kinase (ILK). Here we report the solution NMR structure of the core ILK·PINCH complex (28 kDa, KD ∼ 68 nm) involving the N-terminal ankyrin repeat domain (ARD) of ILK and the first LIM domain (LIM1) of PINCH. We show that the ILK ARD exhibits five sequentially stacked ankyrin repeat units, which provide a large concave surface to grip the two contiguous zinc fingers of the PINCH LIM1. The highly electrostatic interface is evolutionally conserved but differs drastically from those of known ARD and LIM bound to other types of protein domains. Consistently mutation of a hot spot in LIM1, which is not conserved in other LIM domains, disrupted the PINCH binding to ILK and abolished the PINCH targeting to FAs. These data provide atomic insight into a novel modular recognition and demonstrate how PINCH is specifically recruited by ILK to mediate the FA assembly and cell-extracellular matrix communication. PMID:19117955

  14. Structural Basis of Focal Adhesion Localization of LIM-only Adaptor PINCH by Integrin-linked Kinase

    SciTech Connect

    Yang, Yanwu; Wang, Xiaoxia; Hawkins, Cheryl A.; Chen, Kan; Vaynberg, Julia; Mao, Xian; Tu, Yizeng; Zuo, Xiaobing; Wang, Jinbu; Wang, Yun-xing; Wu, Chuanyue; Tjandra, Nico; Qin, Jun

    2010-11-22

    The LIM-only adaptor PINCH (the particularly interesting cysteine- and histidine-rich protein) plays a pivotal role in the assembly of focal adhesions (FAs), supramolecular complexes that transmit mechanical and biochemical information between extracellular matrix and actin cytoskeleton, regulating diverse cell adhesive processes such as cell migration, cell spreading, and survival. A key step for the PINCH function is its localization to FAs, which depends critically on the tight binding of PINCH to integrin-linked kinase (ILK). Here we report the solution NMR structure of the core ILK {center_dot} PINCH complex (28 kDa, K{sub D} {approx} 68 nm) involving the N-terminal ankyrin repeat domain (ARD) of ILK and the first LIM domain (LIM1) of PINCH. We show that the ILK ARD exhibits five sequentially stacked ankyrin repeat units, which provide a large concave surface to grip the two contiguous zinc fingers of the PINCH LIM1. The highly electrostatic interface is evolutionally conserved but differs drastically from those of known ARD and LIM bound to other types of protein domains. Consistently mutation of a hot spot in LIM1, which is not conserved in other LIM domains, disrupted the PINCH binding to ILK and abolished the PINCH targeting to FAs. These data provide atomic insight into a novel modular recognition and demonstrate how PINCH is specifically recruited by ILK to mediate the FA assembly and cell-extracellular matrix communication.

  15. LIMS Version 6 Level 3 Dataset

    NASA Technical Reports Server (NTRS)

    Remsberg, Ellis E.; Lingenfelser, Gretchen

    2010-01-01

    This report describes the Limb Infrared Monitor of the Stratosphere (LIMS) Version 6 (V6) Level 3 data products and the assumptions used for their generation. A sequential estimation algorithm was used to obtain daily, zonal Fourier coefficients of the several parameters of the LIMS dataset for 216 days of 1978-79. The coefficients are available at up to 28 pressure levels and at every two degrees of latitude from 64 S to 84 N and at the synoptic time of 12 UT. Example plots were prepared and archived from the data at 10 hPa of January 1, 1979, to illustrate the overall coherence of the features obtained with the LIMS-retrieved parameters.

  16. Spectroscopy and transmittances for the LIMS experiment

    NASA Technical Reports Server (NTRS)

    Drayson, S. R.; Bailey, P. L.; Gille, J. C.; Fischer, H.; Girard, A.; Gordley, L. L.; Harries, J. E.; Planet, W. G.; Remsberg, E. E.; Russell, J. M., III

    1984-01-01

    The Limb Infrared Monitor of the Stratosphere (LIMS) instrument on the Nimbus 7 spacecraft sensed limb emission profiles for 7 months, starting in October 1979. Vertical temperature profiles are inferred from measurements in two CO2 channels, and constituent profiles are obtained form the O3, HNO3, NO2, and water vapor channels. The line parameters used to calculate transmittances in the retrieval algorithm are presented. Their adequacy for LIMS and for other applications are assessed, and recommendations for future investigations are outlined.

  17. Coordinate roles for LIM homeobox genes in directing the dorsoventral trajectory of motor axons in the vertebrate limb.

    PubMed

    Kania, A; Johnson, R L; Jessell, T M

    2000-07-21

    Motor neurons extend axons along specific trajectories, but the molecules that control their pathfinding remain poorly defined. We show that two LIM homeodomain transcription factors, Lim1 and Lmx1b, control the initial trajectory of motor axons in the developing mammalian limb. The expression of Lim1 by a lateral set of lateral motor column (LMC) neurons ensures that their axons select a dorsal trajectory in the limb. In a complementary manner, the expression of Lmx1b by dorsal limb mesenchymal cells controls the dorsal and ventral axonal trajectories of medial and lateral LMC neurons. In the absence of these two proteins, motor axons appear to select dorsal and ventral trajectories at random. Thus, LIM homeodomain proteins act within motor neurons and cells that guide motor axons to establish the fidelity of a binary choice in axonal trajectory. PMID:10943837

  18. Gene expression and association analysis of LIM (PDLIM5) in major depression.

    PubMed

    Iga, Jun-ichi; Ueno, Shu-ichi; Yamauchi, Ken; Numata, Shusuke; Motoki, Ikuyo; Tayoshi, Sumiko; Kinouchi, Sawako; Ohta, Koshi; Song, Hongwei; Morita, Kyoko; Rokutan, Kazuhito; Tanabe, Hirotaka; Sano, Akira; Ohmori, Tetsuro

    2006-06-12

    LIM (PDLIM5) is a small protein that interacts with protein kinase C-epsilon and the N-type calcium channel alpha-1B subunit and modulates neuronal calcium signaling. Recently, the LIM mRNA expression in postmortem brains and immortalized lymphoblastoid cells from mood disorder patients was reported to be changed and seems to be involved in its pathophysiology. We hypothesized that the expression of the LIM mRNA in the native peripheral leukocytes may be a good candidate for the biological marker for mood disorders. Twenty patients with major depression and age- and sex-matched control subjects were included in this expression study. The LIM mRNA levels in the peripheral leukocytes from drug-naive depressive patients were significantly lower than those from control subjects and increased significantly after 4-week paroxetine treatments, to almost the same level as controls'. Hamilton depressive scores (HAM-D) were improved about 50% after 4-week treatment but neither paroxetine concentrations nor the changes of HAM-D scores showed significant correlation with the change of the mRNA levels. Then, we genotyped three single nucleotide polymorphic markers of LIM gene, which were reported to be associated with bipolar disorder in patients with major depression and control subjects (n=130, each), but there were no associations between these SNPs and major depression. Our investigation indicates that the lower expression levels of LIM mRNA in the peripheral leukocytes are associated with the depressive state and that its recovery after treatment may be an adaptive change induced by the antidepressant. PMID:16595163

  19. Principles and application of LIMS in mouse clinics.

    PubMed

    Maier, Holger; Schütt, Christine; Steinkamp, Ralph; Hurt, Anja; Schneltzer, Elida; Gormanns, Philipp; Lengger, Christoph; Griffiths, Mark; Melvin, David; Agrawal, Neha; Alcantara, Rafael; Evans, Arthur; Gannon, David; Holroyd, Simon; Kipp, Christian; Raj, Navis Pretheeba; Richardson, David; LeBlanc, Sophie; Vasseur, Laurent; Masuya, Hiroshi; Kobayashi, Kimio; Suzuki, Tomohiro; Tanaka, Nobuhiko; Wakana, Shigeharu; Walling, Alison; Clary, David; Gallegos, Juan; Fuchs, Helmut; de Angelis, Martin Hrabě; Gailus-Durner, Valerie

    2015-10-01

    Large-scale systemic mouse phenotyping, as performed by mouse clinics for more than a decade, requires thousands of mice from a multitude of different mutant lines to be bred, individually tracked and subjected to phenotyping procedures according to a standardised schedule. All these efforts are typically organised in overlapping projects, running in parallel. In terms of logistics, data capture, data analysis, result visualisation and reporting, new challenges have emerged from such projects. These challenges could hardly be met with traditional methods such as pen & paper colony management, spreadsheet-based data management and manual data analysis. Hence, different Laboratory Information Management Systems (LIMS) have been developed in mouse clinics to facilitate or even enable mouse and data management in the described order of magnitude. This review shows that general principles of LIMS can be empirically deduced from LIMS used by different mouse clinics, although these have evolved differently. Supported by LIMS descriptions and lessons learned from seven mouse clinics, this review also shows that the unique LIMS environment in a particular facility strongly influences strategic LIMS decisions and LIMS development. As a major conclusion, this review states that there is no universal LIMS for the mouse research domain that fits all requirements. Still, empirically deduced general LIMS principles can serve as a master decision support template, which is provided as a hands-on tool for mouse research facilities looking for a LIMS. PMID:26208973

  20. Mapping of four mouse genes encoding eye lens-specific structural, gap junction, and integral membrane proteins: Cryba1 (crystallin{beta}A3/A1), Crybb2 (crystallin{beta}B2), Gja8 (MP70), and Lim2 (MP19)

    SciTech Connect

    Kerscher, S.; Boyd, Y.; Lyon, M.F.

    1995-09-20

    Four genes encoding eye lens-specific proteins, potential candidate genes for congenital cataract (CC) mutations, were mapped in the mouse genome using a panel of somatic cell hybrids and DNAs from the EUCIB (European Collaborative Interspecific Backcross). Two of them are lens fiber cell structural proteins: the Cryba1 locus encoding crystallin{beta}A3/A1 maps to chromosome 11, 2.5 {+-} 2.5 cM distal to D11Mit31, and the Crybb2 locus encoding crystallin{beta}B2 maps to chromosome 5, 9.1 {+-} 4.3 cM distal to D5Mit88. The other two genes encode lens-specific gap junction and integral membrane proteins, respectively: the Gja8 locus encoding gap junction membrane channel protein {alpha}8, also called connexin50 or MP70, maps to chromosome 3, 11.9 {+-} 5.0 cM distal to D3Mit22, and the Lim2 locus encoding lens intrinsic membrane protein 2, also call MP19, maps to chromosome 7, 2.5 {+-} 2.5 cM proximal to Ngfg. All four map positions, when compared with the corresponding positions in human, lie within known regions of conserved synteny between mouse and human chromosomes. 43 refs., 2 figs., 1 tab.

  1. Principles of Contour Information: Reply to Lim and Leek (2012)

    ERIC Educational Resources Information Center

    Singh, Manish; Feldman, Jacob

    2012-01-01

    Lim and Leek (2012) presented a formalization of information along object contours, which they argued was an alternative to the approach taken in our article (Feldman & Singh, 2005). Here, we summarize the 2 approaches, showing that--notwithstanding Lim and Leek's (2012) critical rhetoric--their approach is substantially identical to ours, except…

  2. A dedicated LIMS for routine gamma-ray spectrometry.

    PubMed

    Bruggeman, M; Verheyen, L; Vidmar, T

    2014-05-01

    We developed a Microsoft(®) Access-based LIMS (Laboratory Information and Management Systems), γ-LIMS, for the management of our gamma-spectrometry laboratory, in which thousands of routine, but high-quality analyses are performed each year. This paper explains the main features of the γ-LIMS and puts special attention on the interfacing methods and solutions for using the Genie™2000 spectrometry software in conjunction with the EFFTRAN package, which serves for efficiency transfer calculations, coincidence summing corrections and a procedure for uncertainty estimation. PMID:24332338

  3. Laboratory Information Management System (LIMS): A case study

    NASA Technical Reports Server (NTRS)

    Crandall, Karen S.; Auping, Judith V.; Megargle, Robert G.

    1987-01-01

    In the late 70's, a refurbishment of the analytical laboratories serving the Materials Division at NASA Lewis Research Center was undertaken. As part of the modernization efforts, a Laboratory Information Management System (LIMS) was to be included. Preliminary studies indicated a custom-designed system as the best choice in order to satisfy all of the requirements. A scaled down version of the original design has been in operation since 1984. The LIMS, a combination of computer hardware, provides the chemical characterization laboratory with an information data base, a report generator, a user interface, and networking capabilities. This paper is an account of the processes involved in designing and implementing that LIMS.

  4. The accuracy of Nimbus 7 LIMS temperatures in the mesosphere

    NASA Technical Reports Server (NTRS)

    Remsberg, E. E.

    1986-01-01

    Zonal-mean temperatures from the Nimbus 7 Limb Infrared Monitor of the Stratosphere (LIMS) experiment are compared with a temperature 'climatology' derived from 4 years of Rayleigh backscatter lidar measurements at 44 N and for the months of March, April, and May, when wave activity and thus atmospheric variability is very weak. The mean difference between the two data sets is less than 3.5 K between 37 and 64 km and is consistent with the theoretical estimates of accuracy reported for the LIMS data. The effects of longitudinal and year-to-year variaiations in this comparison were considered, but they do not change the conclusions significantly. These validation results extend the altitude range of accurate LIMS temperatures in the mesosphere some 10 km higher than previously reported. It also means the LIMS ozone and NO2 retrievals in the lower mesosphere should be free of any major temperature bias error effects.

  5. Genetic variations in GJA3, GJA8, LIM2, and age-related cataract in the Chinese population: a mutation screening study

    PubMed Central

    Zhou, Zhou; Wang, Binbin; Hu, Shanshan; Zhang, Chunmei; Ma, Xu

    2011-01-01

    Purpose To investigate the role of genetic variations in three known cataract-associated genes, gap junction protein α3 (GJA3), gap junction protein α8 (GJA8), lens intrinsic membrane protein 2 (LIM2), encoding lens fiber cell membrane proteins in the development of age-related cataracts. Methods One hundred and forty-five sporadic age-related cataract patients and one hundred and fifty-six unrelated random healthy controls participated in this study. Genomic DNA was extracted from peripheral blood leukocytes. All exons of GJA3, GJA8, and LIM2 were sequenced after being amplified by polymerase chain reaction (PCR). The functional consequences of the mutations were analyzed using PolyPhen. Results We found five novel variations in 145 patients and none of them presented in the 156 controls. There are two variations in GJA3 (c.-39C>G, c. 415G>A); one in GJA8 (c. 823G>A), and two in LIM2 (c.57G>A, c.67A>C). PolyPhen predicted that the LIM2 c.67A>C mutation may have potential pathogenicity. Conclusions The genetic mutation in GJA3, GJA8, and LIM2 may slightly contribute to the development of age-related cataracts. This study showed a potential relationship between lens fiber cell membrane protein genes and the development of age-related cataracts in the Chinese population. PMID:21386927

  6. Limb Infrared Monitor of the Stratosphere /LIMS/ experiment

    NASA Technical Reports Server (NTRS)

    Drozewski, R. W.; Hatch, M. R.

    1978-01-01

    The Limb Infrared Monitor of the Stratosphere (LIMS) radiometer is a satellite-borne six-channel multispectral scanning radiometer using a two-stage methane and ammonia solid-cryogen cooler to cool the (Hg, Cd)Te focal plane to 65 K. The LIMS experiment employs thermal IR limb sounding to provide vertical-profile measurement of temperature and of concentrations of O3, NO3, and H2O in the stratosphere on a global scale. The goals of the experiment and the expected accuracies of the measurements are outlined, and the radiance measured by the radiometer is analyzed. A program of correlative measurements designed for verification and augmentation of the LIMS data is discussed. Detailed descriptions are given of the LIMS components and optics, the detector capsule assembly, the solid-cryogen cooler, the four operational modes of the LIMS system, and the data-processor electronics. It is noted that prelaunch acceptance testing thus far completed indicates that the LIMS specification design requirements have been met, and the experiment's performance goals can be achieved.

  7. Energetic Particle Precipitation Effects Observed in LIMS Data

    NASA Astrophysics Data System (ADS)

    Holt, L. A.; Randall, C. E.; Harvey, V. L.; Stiller, G. P.; Funke, B.; López-Puertas, M.; Remsberg, E. E.

    2008-12-01

    The Limb Infrared Monitor of the Stratosphere (LIMS) observed stratospheric enhancements in NO2 inside the Arctic polar vortex during the winter of 1978-1979. These enhancements were attributed to the descent of NOx originally produced by precipitating energetic particles in the upper atmosphere. Although few observations of such stratospheric NOx enhancements were made during the decade succeeding the LIMS measurements, investigations in the last decade have shown abundant evidence for these enhancements. Interannual variability in the enhancements appears to be controlled both by the amount of particle precipitation and the prevailing meteorological conditions, which dictate the efficiency with which NOx is transported from the upper atmosphere into the stratosphere. In this presentation, recent satellite measurements of the temporal evolution of NOx in the polar vortex are compared to the LIMS measurements. Our goal is to investigate whether the enhancements were observed by LIMS because of enhanced geomagnetic activity and/or anomalous dynamical conditions, or whether the nighttime observing capability of LIMS simply enabled it to detect the NOx enhancements under nominal conditions.

  8. Case study: Accelerated schedule for MULTI LIMS installation

    SciTech Connect

    Ibsen, T.G.

    1994-05-01

    This presentation focuses on the steps taken by the Westinghouse Hanford Company to meet an accelerated schedule for configuration and implementation of the MULTI LIMS in a multiple laboratory environment. The Westinghouse Hanford Company purchased the MULTI LIMS Laboratory Information Management System in August, 1993. Hardware delivery began in October, 1993. Less than four months later, the initial configuration was released for use in two Westinghouse Hanford Company laboratories. Several major obstacles were overcome during implementation. These include information gathering for base table loading, user training, acceptance of the new system by users of a legacy system, and hardware configuration issues. In summary, steps needed to be taken to meet the accelerated implementation schedule of the MULTI LIMS at the Hanford Site. The obstacles faced were overcome through the in-depth knowledge and help of the vendor and the dedication and drive of the technical staff.

  9. LimsPortal and BonsaiLIMS: development of a lab information management system for translational medicine

    PubMed Central

    2011-01-01

    Background Laboratory Information Management Systems (LIMS) are an increasingly important part of modern laboratory infrastructure. As typically very sophisticated software products, LIMS often require considerable resources to select, deploy and maintain. Larger organisations may have access to specialist IT support to assist with requirements elicitation and software customisation, however smaller groups will often have limited IT support to perform the kind of iterative development that can resolve the difficulties that biologists often have when specifying requirements. Translational medicine aims to accelerate the process of treatment discovery by bringing together multiple disciplines to discover new approaches to treating disease, or novel applications of existing treatments. The diverse set of disciplines and complexity of processing procedures involved, especially with the use of high throughput technologies, bring difficulties in customizing a generic LIMS to provide a single system for managing sample related data within a translational medicine research setting, especially where limited IT support is available. Results We have designed and developed a LIMS, BonsaiLIMS, around a very simple data model that can be easily implemented using a variety of technologies, and can be easily extended as specific requirements dictate. A reference implementation using Oracle 11 g database and the Python framework, Django is presented. Conclusions By focusing on a minimal feature set and a modular design we have been able to deploy the BonsaiLIMS system very quickly. The benefits to our institute have been the avoidance of the prolonged implementation timescales, budget overruns, scope creep, off-specifications and user fatigue issues that typify many enterprise software implementations. The transition away from using local, uncontrolled records in spreadsheet and paper formats to a centrally held, secured and backed-up database brings the immediate benefits of

  10. Inclusion through Access to Outdoor Education: Learning in Motion (LIM)

    ERIC Educational Resources Information Center

    Brodin, Jane

    2009-01-01

    Learning in Motion (LIM) was a European project involving seven partners in five countries: Sweden, Finland, Latvia, Germany and Greece. The project focused on inclusion and access to outdoor education and was financed by the European Commission within the framework of the Socrates-Grundtvig Programme. The aim of the project was to explore if and…

  11. Nucleolar localization signals of LIM kinase 2 function as a cell-penetrating peptide.

    PubMed

    Kobayashi, Nahoko; Niwa, Mikio; Hao, Yang; Yoshida, Tetsuhiko

    2010-12-01

    LIM Kinase 2 (LIMK2) is a LIM domain-containing protein kinase which regulates actin polymerization thorough phosphorylation of the actin depolymerizing factor cofilin. It is also known to function as a shuttle between the cytoplasm and nucleus in endothelial cells. A basic amino acid-rich motif in LIMK2 was previously identified to be responsible for this shuttling function, as a nucleolar localization signal (NoLS). Here it is shown that this nucleolar localization signal sequence also has the characteristic function of a cell-penetrating peptide (CPP). We synthesized LIMK2 NoLS-conjugated peptides and a protein and analyzed their cell-penetrating abilities in various types of cells. The BC-box motif of the Von Hippel-Lindau (VHL) protein was used for the peptide. This motif previously has been reported to be involved in the neural differentiation of bone marrow stromal cells and skin-derived precursor cells. Green fluorescence protein (GFP) was used as a large biologically active biomolecule for the protein. The LIMK2 NoLS-conjugated peptides and protein translocated across the cell membranes of fibroblast cells, neural stem cells, and even iPS cells. These results suggest that LIMK2 NoLS acts as a cell-penetrating peptide and its cell-penetrating ability is not restricted by cell type. Moreover, from an in vivo assay using a mouse brain, it was confirmed that NoLS has potential for transporting biomolecules across the blood-brain barrier. PMID:20937035

  12. SMITH: a LIMS for handling next-generation sequencing workflows

    PubMed Central

    2014-01-01

    Background Life-science laboratories make increasing use of Next Generation Sequencing (NGS) for studying bio-macromolecules and their interactions. Array-based methods for measuring gene expression or protein-DNA interactions are being replaced by RNA-Seq and ChIP-Seq. Sequencing is generally performed by specialized facilities that have to keep track of sequencing requests, trace samples, ensure quality and make data available according to predefined privileges. An integrated tool helps to troubleshoot problems, to maintain a high quality standard, to reduce time and costs. Commercial and non-commercial tools called LIMS (Laboratory Information Management Systems) are available for this purpose. However, they often come at prohibitive cost and/or lack the flexibility and scalability needed to adjust seamlessly to the frequently changing protocols employed. In order to manage the flow of sequencing data produced at the Genomic Unit of the Italian Institute of Technology (IIT), we developed SMITH (Sequencing Machine Information Tracking and Handling). Methods SMITH is a web application with a MySQL server at the backend. Wet-lab scientists of the Centre for Genomic Science and database experts from the Politecnico of Milan in the context of a Genomic Data Model Project developed SMITH. The data base schema stores all the information of an NGS experiment, including the descriptions of all protocols and algorithms used in the process. Notably, an attribute-value table allows associating an unconstrained textual description to each sample and all the data produced afterwards. This method permits the creation of metadata that can be used to search the database for specific files as well as for statistical analyses. Results SMITH runs automatically and limits direct human interaction mainly to administrative tasks. SMITH data-delivery procedures were standardized making it easier for biologists and analysts to navigate the data. Automation also helps saving time. The

  13. Analysis of LIMS data by potential vorticity inversion

    NASA Technical Reports Server (NTRS)

    Robinson, Walter A.

    1988-01-01

    This paper analyzes LIMS data for a minor warming of January 1979 by potential vorticity inversion, to investigate how flows in the middle atmosphere were determined. The inversion problem of calculating the flow from the potential vorticity is solved repeatedly, including and excluding different portions of the potential vorticity, thus revealing which bits of potential vorticity were important in determining the flow at a time and location of interest. The results of the analysis of LIMS data indicate that the middle stratospheric flow is dominated by potential vorticity that is local in height and in latitude, while planetary-scale mesospheric flows are primarily induced by potential vorticity in the stratosphere. A possible explanation for this difference is proposed.

  14. Principles of contour information: Reply to Lim and Leek (2012).

    PubMed

    Singh, Manish; Feldman, Jacob

    2012-07-01

    Lim and Leek (2012) presented a formalization of information along object contours, which they argued was an alternative to the approach taken in our article (Feldman & Singh, 2005). Here, we summarize the 2 approaches, showing that--notwithstanding Lim and Leek's (2012) critical rhetoric--their approach is substantially identical to ours, except for the technical details of the formalism. Following the logic of our article point by point, Lim and Leek (a) defined probabilistic expectations about the geometry of smooth contours (which they based on differential contour geometry, while we used a discrete approximation--the only essential difference in their approach), (b) assumed that information along the contour was proportional to the negative logarithm of probability, following standard information theory, and then (c) extended this formulation to closed contours. We analyze what they described as errors in our approach, all of which rest on mathematical misunderstandings or bizarre misreadings of our article. We also show that their extension to 3-dimensional surfaces and their "modified minima rule" contain fatal deficiencies. PMID:22775501

  15. The validation of Nimbus 7 LIMS measurements of ozone

    NASA Technical Reports Server (NTRS)

    Remsberg, E. E.; Russell, J. M., III; Gille, J. C.; Bailey, P. L.; Gordley, L. L.; Planet, W. G.; Harries, J. E.

    1984-01-01

    Approximately seven and one half months of stratospheric ozone profiles have been processed from the LIMS (Limb Infrared Monitor of the Stratosphere) experiment on the Nimbus 7 satellite. Data profiles cover the stratosphere and mesosphere from 100 to 0.1 mbar and from 84 degrees N to 64 degrees S latitude. Topics covered include the ozone channel characteristics, precision, systematic uncertainties, and comparisons with data from balloon and rocket underflights, Umkehr soundings, and Dobson measurements. Comparisons with Dobson total ozone are made by integrating combined LIMS plus balloon profiles. The estimated on-orbit precision is 0.02-0.16 ppmv. Simulations of the experiment indicate potential systematic uncertainties ranging from 15 percent in the 1-mbar region to an upper limit of 40 percent at 100 mbar and 0.1 mbar. Results are well within the uncertainties for the correlative sensors themselves. LIMS detects significant vertical structure in the ozone profile even below the ozone mixing ratio peak. Several other comparisons showing good agreement are noted.

  16. Hydroxyl radical regeneration in isoprene oxidation: upgraded mechanism LIM1

    NASA Astrophysics Data System (ADS)

    Peeters, Jozef; Son Nguyen, Vinh; Nguyen, Thanh Lam; Stravrakou, Trissevgeni; Muller, Jean-Francois

    2013-04-01

    The OH regeneration known to occur in isoprene oxidation at low/moderate NO is attributed in the Leuven Isoprene Mechanism to novel, theoretically characterized chemical pathways (LIM0: Peeters et al. 2009; Peeters and Muller 2010). Its key new features are (i) quasi-equilibration of the thermally labile beta-OH- and delta-OH-isoprenylperoxy isomers; (ii) 1,6-H shift isomerisation of the Z-delta-OH-peroxy isomers to yield hydroperoxy-methyl-butenals (HPALDs); (iii) fast photolysis of the HPALDs resulting overall in several OH radicals per HPALD. The OH-regeneration through photolabile HPALDs has recently found experimental support, but the peroxy isomerisation rate, HPALD yield and extent of OH recycling are still uncertain (Crounse et al. 2011; Wolfe et al. 2012). In this work, the upgraded LIM1 mechanism is presented. Based on much higher levels of theory that fully account for dispersion effects, the crucial equilibrium ratio of the isomerising Z-delta-OH-peroxys over the majority beta-OH-isoprenylperoxys is reduced by a factor ≈5 and the isomerisation rate of the Z-delta-OH-peroxys by a factor ≈1.5 compared to LIM0. The chemistry following the 1,6-H shift of the Z-delta-OH-peroxys is also much expanded and extended. Firstly, LIM1 introduces other pathways beside HPALD formation following the Z-delta-OH-peroxy isomerisation, but resulting likewise in OH recycling. This, together with the revised Z-delta-OH- equilibrium and isomerisation data above, affords a fair model-reproduction of the HPALD and other product yields observed by Crounse et al. (2011). Secondly, LIM1 proposes new fast reactions of HO2 with the alpha-oxoketene products from the peroxy isomerisation routes; these reactions are shown to efficiently convert HO2 into OH and are prime candidates for the unknown X + HO2 → OH + ... hydroxyl-recycling routes invoked in recent studies (Hofzumahaus et al.2009; Whalley et al. 2011). Modeling results using the IMAGES global CTM will be presented on

  17. The validation of LIMS ozone profiles using correlative rocket, ECC, and Dobson data sets

    NASA Technical Reports Server (NTRS)

    Remsberg, E. E.; Russell, J. M., III; Gordley, L. L.; Gille, J. C.; Bailey, P. L.

    1981-01-01

    The Limb Infrared Monitor of the Stratosphere (LIMS) experiment is a limb scanning infrared sounder designed to measure vertical temperature profiles and the concentrations of key chemical compounds which are important in the stratospheric ozone-nitrogen photochemistry. Preliminary ozone profiles are being intercompared with correlative rocket and balloonsonde ozone data sets as part of the Nimbus 7 LIMS data validation activity. The LIMS ozone retrievals extend from the lower mesosphere to the tropopause (or cloud-top altitudes). In this paper, preliminary statistics of comparisons of ECC (electrochemical concentration cell) ozonesonde and rocket ozone profiles with LIMS ozone profiles are presented. ECC ozonesonde data taken at times of LIMS overpasses are also used to extend the LIMS retrieval down to ground level. Each combined profile is then integrated to obtain total column ozone. These integrated results are compared to Dobson data obtained at approximately the same time and location.

  18. LIMS2 mutations are associated with a novel muscular dystrophy, severe cardiomyopathy and triangular tongues.

    PubMed

    Chardon, Jodi Warman; Smith, A C; Woulfe, J; Pena, E; Rakhra, K; Dennie, C; Beaulieu, C; Huang, Lijia; Schwartzentruber, J; Hawkins, C; Harms, M B; Dojeiji, S; Zhang, M; Majewski, J; Bulman, D E; Boycott, K M; Dyment, D A

    2015-12-01

    Limb girdle muscular dystrophy (LGMD) is a heterogeneous group of genetic disorders leading to progressive muscle degeneration and often associated with cardiac complications. We present two adult siblings with childhood-onset of weakness progressing to a severe quadriparesis with the additional features of triangular tongues and biventricular cardiac dysfunction. Whole exome sequencing identified compound heterozygous missense mutations that are predicted to be pathogenic in LIMS2. Biopsy of skeletal muscle demonstrated disrupted immunostaining of LIMS2. This is the first report of mutations in LIMS2 and resulting disruption of the integrin linked kinase (ILK)-LIMS-parvin complex associated with LGMD. PMID:25589244

  19. Topics in Chemical Instrumentation: Information Management Systems in the Undergraduate Instrumental Analysis Laboratory: Part I. Introduction to LIMS.

    ERIC Educational Resources Information Center

    Merrer, Robert J.

    1985-01-01

    Provides an overview of Laboratory Information Management Systems (LIMS) and their implementation in undergraduate analytical laboratories. Basic components of a well-constructed LIMS system, hardware considerations, and software considerations are addressed. (JN)

  20. LIMS HNO3 data above 5 mbar - Corrections based on simultaneous observations of other species

    NASA Technical Reports Server (NTRS)

    Jackman, C. H.; Kaye, J. A.; Guthrie, P. D.

    1985-01-01

    The Limb Infrared Monitor of the Stratosphere (LIMS) instrument monitored four trace gases (O3, H2O, HNO3, and NO2) and the temperature in the stratosphere and mesosphere. Recently, the seven months LIMS data, including the time from Oct. 25, 1978 to May 29, 1979, have become available to the scientific community. It is pointed out that the measurements are of great use in studying the dynamics and chemistry of the stratosphere. The LIMS HNO3 data appear to show a realistic behavior below 5 mbar, but above 5 mbar, the data appear to be too high. In the present paper, the LIMS HNO3 behavior is discussed, taking into account the reason for the unrealistic appearance of the data. Other LIMS data are used to derive HNO3 above 5 mbar through the intermediate species OH.

  1. openBIS ELN-LIMS: an open-source database for academic laboratories

    PubMed Central

    Barillari, Caterina; Ottoz, Diana S. M.; Fuentes-Serna, Juan Mariano; Ramakrishnan, Chandrasekhar; Rinn, Bernd; Rudolf, Fabian

    2016-01-01

    Summary: The open-source platform openBIS (open Biology Information System) offers an Electronic Laboratory Notebook and a Laboratory Information Management System (ELN-LIMS) solution suitable for the academic life science laboratories. openBIS ELN-LIMS allows researchers to efficiently document their work, to describe materials and methods and to collect raw and analyzed data. The system comes with a user-friendly web interface where data can be added, edited, browsed and searched. Availability and implementation: The openBIS software, a user guide and a demo instance are available at https://openbis-eln-lims.ethz.ch. The demo instance contains some data from our laboratory as an example to demonstrate the possibilities of the ELN-LIMS (Ottoz et al., 2014). For rapid local testing, a VirtualBox image of the ELN-LIMS is also available. Contact: brinn@ethz.ch or fabian.rudolf@bsse.ethz.ch PMID:26508761

  2. Early evolution of the LIM homeobox gene family

    SciTech Connect

    Srivastava, Mansi; Larroux, Claire; Lu, Daniel R; Mohanty, Kareshma; Chapman, Jarrod; Degnan, Bernard M; Rokhsar, Daniel S

    2010-01-01

    LIM homeobox (Lhx) transcription factors are unique to the animal lineage and have patterning roles during embryonic development in flies, nematodes and vertebrates, with a conserved role in specifying neuronal identity. Though genes of this family have been reported in a sponge and a cnidarian, the expression patterns and functions of the Lhx family during development in non-bilaterian phyla are not known. We identified Lhx genes in two cnidarians and a placozoan and report the expression of Lhx genes during embryonic development in Nematostella and the demosponge Amphimedon. Members of the six major LIM homeobox subfamilies are represented in the genomes of the starlet sea anemone, Nematostella vectensis, and the placozoan Trichoplax adhaerens. The hydrozoan cnidarian, Hydra magnipapillata, has retained four of the six Lhx subfamilies, but apparently lost two others. Only three subfamilies are represented in the haplosclerid demosponge Amphimedon queenslandica. A tandem cluster of three Lhx genes of different subfamilies and a gene containing two LIM domains in the genome of T. adhaerens (an animal without any neurons) indicates that Lhx subfamilies were generated by tandem duplication. This tandem cluster in Trichoplax is likely a remnant of the original chromosomal context in which Lhx subfamilies first appeared. Three of the six Trichoplax Lhx genes are expressed in animals in laboratory culture, as are all Lhx genes in Hydra. Expression patterns of Nematostella Lhx genes correlate with neural territories in larval and juvenile polyp stages. In the aneural demosponge, A. queenslandica, the three Lhx genes are expressed widely during development, including in cells that are associated with the larval photosensory ring. The Lhx family expanded and diversified early in animal evolution, with all six subfamilies already diverged prior to the cnidarian-placozoan-bilaterian last common ancestor. In Nematostella, Lhx gene expression is correlated with neural

  3. Global lower mesospheric water vapor revealed by LIMS observations

    NASA Technical Reports Server (NTRS)

    Gordley, L. L.; Russell, J. M., III; Remsberg, E. E.

    1985-01-01

    The Limb Infrared Monitor of the Stratospheric water vapor channel data analysis has been extended from the 1. mb level (about 48 km) to the .3 mb level (about 60 km) through a radiance averaging procedure and better understanding of systematic errors. The data show H2O mixing ratio peaks near the .5 mb level varying from 4 to 7 ppmv with latitude and season. Above this level the mixing ratio drops off quickly with altitude, but, due to experimental uncertainties, at an uncertain rate. The stratospheric results are virtually the same as determined from the archived LIMS results with a tropical hygropause and enhanced H2O concentration in the lower levels at high winter latitudes.

  4. Tissue specific characterisation of Lim-kinase 1 expression during mouse embryogenesis.

    PubMed

    Lindström, Nils O; Neves, Carlos; McIntosh, Rebecca; Miedzybrodzka, Zosia; Vargesson, Neil; Collinson, J Martin

    2011-01-01

    The Lim-kinase (LIMK) proteins are important for the regulation of the actin cytoskeleton, in particular the control of actin nucleation and depolymerisation via regulation of cofilin, and hence may control a large number of processes during development, including cell tensegrity, migration, cell cycling, and axon guidance. LIMK1/LIMK2 knockouts disrupt spinal cord morphogenesis and synapse formation but other tissues and developmental processes that require LIMK are yet to be fully determined. To identify tissues and cell-types that may require LIMK, we characterised the pattern of LIMK1 protein during mouse embryogenesis. We showed that LIMK1 displays an expression pattern that is temporally dynamic and tissue-specific. In several tissues LIMK1 is detected in cell-types that also express Wilms' tumour protein 1 and that undergo transitions between epithelial and mesenchymal states, including the pleura, epicardium, kidney nephrons, and gonads. LIMK1 was also found in a subset of cells in the dorsal retina, and in mesenchymal cells surrounding the peripheral nerves. This detailed study of the spatial and temporal expression of LIMK1 shows that LIMK1 expression is more dynamic than previously reported, in particular at sites of tissue-tissue interactions guiding multiple developmental processes. PMID:21167960

  5. The Nimbus 7 LIMS (Limb Infrared Monitor of the Stratosphere) water vapor measurements

    NASA Technical Reports Server (NTRS)

    Remsberg, Ellis E.; Russell, James M., III

    1988-01-01

    Earth orbital instruments, designed to measure the vertical and spatial distribution of atmospheric water vapor is discussed. Specifically, the operation of the Limb Infrared Monitor of the Stratosphere (LIMS) experiment is examined. The LIMS is a six channel limb scanning radiometer that was launched aboard Nimbus 7 in 1978. Profiles of stratospheric and mesospheric temperature, water vapor, and various other constituents were obtained by inverting the LIMS radiance measurements. This same technique was used in 1981 to analyze the data returned from another limb scanning radiometer aboard the Solar Mesosphere Explorer.

  6. Comparison of LIMS temperatures and geostrophic winds with Berlin radiosonde temperature and wind measurements

    NASA Technical Reports Server (NTRS)

    Grose, W. L.; Miles, T.; Labitzke, K.; Pantzke, E.

    1988-01-01

    The temperature and the derived winds obtained from the LIMS Map Archival Tape data for the period of October 25, 1978, to May 28, 1979, were compared with corresponding data from the Berin (Tempelhof) radiosonde station at several representative levels in the stratosphere, to assess the quality of the LIMS satellite data for use in dynamics and transport studies. It was found, on the basis of this single-station time series comparison, that the synoptically mapped LIMS temperature and wind analyses are of a sufficiently high quality for investigating large-scale dynamics in the stratosphere in conjunction with high-resolution radiosonde measurements.

  7. Intercomparison of nitrogen-containing species in Nimbus 7 LIMS and SAMS Data

    SciTech Connect

    Jackman, C.H.; Guthrie, P.D.; Kaye, J.A.

    1987-01-20

    Nitrogen dioxide (NO/sub 2/) and nitric acid (HNO/sub 3/) distributions were derived with a two-dimensional time-dependent model using N/sub 2/O, CH/sub 4/, and temperature measurements from the stratospheric and mesospheric sounder (SAMS) along with H/sub 2/O measurements from the limb infrared monitor of the stratosphere (LIMS) and O/sub 3/ measurements from the solar backscatter ultraviolet (SBUV) instrument. All three instruments (SAMS, LIMS, and SBUV) were aboard the Nimbus 7 satellite. The computed NO/sub 2/ and HNO/sub 3/ were compared with LIMS NO/sub 2/ and HNO/sub 3/ measurements. Calculated NO/sub 2/ is lower than the LIMS NO/sub 2/ in much of the lower stratosphere by about a factor of 4 or more. Differences in the upper stratosphere between derived NO/sub 2/ and LIMS NO/sub 2/ are within the photochemical uncertainties of the computation. Derived NO/sub 2/ is much larger than LIMS NO/sub 2/ at high latitudes in the upper stratosphere in December, indicating that dynamics and/or photochemistry in the atmosphere are different from those used in the model. Derived HNO/sub 3/ is in fairly good agreement with LIMS HNO/sub 3/ in the upper stratosphere. Derived and LIMS HNO/sub 3/ gradually become more different through the middle stratosphere and reach a fairly substantial disagreement in the lower stratosphere. The high-latitude maxima observed in LIMS HNO/sub 3/ are present at lower altitudes and at higher concentrations than those derived in the model computations. More significantly, the derived HNO/sub 3/ at polar latitudes is highest in the summer while LIMS HNO/sub 3/ is highest in the winter. Disagreements in the lower stratosphere between the computed and LIMS NO/sub 2/ and HNO/sub 3/ can only be reduced if two changes are made in the model computations: (1) additional lower stratospheric sources of odd nitrogen (other than N/sub 2/O+O(/sup 1/D)) are included and (2) a modified chemistry to allow the formation of HNO/sub 3/ at the expense of N/sub 2

  8. Comparison of Southern Hemisphere radiosonde and LIMS temperatures at 100 mb. [limb infrared monitor of stratosphere

    NASA Technical Reports Server (NTRS)

    Miles, T.; Grose, W. L.; Russell, J. M., III; Remsberg, E. E.

    1987-01-01

    Radiosonde (RS)and satellite-derived (Nimbus-7 LIMS) 100-mb temperatures over New Zealand at 12 GMT are compared for the 1978-79 summer. The colocated LIMS temperature information consists of synoptically mapped values (for 12 GMT), as well as the primary nighttime orbital retrievals valid at about 1030 GMT. The RS time series of temperature is dominated by temporal fluctuations associated mainly with the eastward passage of waves which have characteristic periods of 4-5 and 11-12 days and peak-to-peak amplitudes of 10-15 K. The LIMS temperatures and the corresponding temperature time series are also found to exhibit quite close agreement (in terms of temporal phase for the latter) with the RS data. However, the LIMS-mapped temperature fluctuations suffer from a noticeable attenuation in amplitude (approaching 50 percent for higher-frequency fluctuations), which will affect the accuracy of LIMS-derived estimates of dynamical quantities such as wind velocity and relative vorticity in the lower stratosphere.

  9. The geometry structures and electronic properties of LimBn (m + n = 12) clusters

    NASA Astrophysics Data System (ADS)

    Ruan, Wen; Xie, An-Dong; Wu, Dong-Lan; Luo, Wen-Lang; Yu, Xiao-Guang

    2014-03-01

    The geometric structures, electronic properties, total and binding energies, harmonic frequencies, the highest occupied molecular orbital to the lowest unoccupied molecular orbital energy gaps, and the vertical ionization potential energies of small LimBn (m+n = 12) clusters were investigated by the density functional theory B3LYP with a 6-311+G (2d, 2p) basis set. All the calculations were performed using the Gaussian09 program. For the study of the LimBn clusters, the global minimum of the B12 cluster was chosen as the starting point and the boron atoms were gradually replaced by Li atoms. The results showed that as the number of Li atoms increased, the stability of the LimBn cluster decreased and the physical and chemical properties became more active. In addition, on average there was a large charge transfer from the Li atoms to the B atoms.

  10. An intercomparison of ozone profile measurements from LIMS, SAGE, and SBUV

    NASA Technical Reports Server (NTRS)

    Fleig, A. J.; Bhartia, P. K.; Wong, C. K.; Klenk, K. F.

    1983-01-01

    Ozone profile measurements by the Limb IR Monitor of Stratosphere (LIMS) and the Solar Backscatter UV (SBUV) aboard the Nimbus satellite, and the Stratospheric Aerosol and Gas Experiment (SAGE) aboard the AEM-2 satellite are intercompared in an effort to assess the quality of satellite ozone retrieval techniques. Good correlation between LIMS and SBUV observations is noted in the ozone layer bounded by pressures of 31.2-15.6 mb; absolute differences are generally less than 10 percent, with similar zonal variations. Above 35 km SAGE values are systematically larger than SBUV or LIMS. The differences generally increase with height and are largest in the tropics. Finally, excellent agreement is noted to exist among the three data sets between 10 and 35 km of altitude with respect to the latitude dependence of the ozone profiles.

  11. On the Quality of the Nimbus 7 LIMS Version 6 Water Vapor Profiles and Distributions

    NASA Technical Reports Server (NTRS)

    Remsberg, E. E.; Natarajan, M.; Lingenfelser, G. S.; Thompson, R. E.; Marshall, B. T.; Gordley, L. L.

    2009-01-01

    This report describes the quality of the Nimbus 7 Limb Infrared Monitor of the Stratosphere (LIMS) water vapor (H2O) profiles of 1978/79 that were processed with a Version 6 (V6) algorithm and archived in 2002. The V6 profiles incorporate a better knowledge of the instrument attitude for the LIMS measurements along its orbits, leading to improvements for its temperature profiles and for the registration of its water vapor radiances with pressure. As a result, the LIMS V6 zonal-mean distributions of H2O exhibit better hemispheric symmetry than was the case from the original Version 5 (V5) dataset that was archived in 1982. Estimates of the precision and accuracy of the V6 H2O profiles are developed and provided. Individual profiles have a precision of order 5% and an estimated accuracy of about 19% at 3 hPa, 14% at 10 hPa, and 26% at 50 hPa. Profile segments within about 2 km of the tropopause are often affected by emissions from clouds that appear in the finite field-of-view of the detector for the LIMS H2O channel. Zonally-averaged distributions of the LIMS V6 H2O are compared with those from the more recent Microwave Limb Sounder (MLS) satellite experiment for November, February, and May of 2004/2005. The patterns and values of their respective distributions are similar in many respects. Effects of a strengthened Brewer-Dobson circulation are indicated in the MLS distributions of the recent decade versus those of LIMS from 1978/79. A tropical tape recorder signal is present in the 7-month time series of LIMS V6 H2O with lowest values in February 1979, and the estimated, annually-averaged "entry-level" H2O is 3.5 to 3.8 ppmv. It is judged that this historic LIMS water vapor dataset is of good quality for studies of the near global-scale chemistry and transport for pressure levels from 3 hPa to about 70 to 100 hPa.

  12. Evidence for equatorial Kelvin modes in Nimbus-7 LIMS. [Limb Infrared Monitor of Stratosphere

    NASA Technical Reports Server (NTRS)

    Salby, M. L.; Hartmann, D. L.; Bailey, P. L.; Gille, J. C.

    1984-01-01

    Equatorial Kelvin waves have emerged as an important source of westerly momentum for the tropical stratosphere. The present investigation is concerned with an analysis of remotely derived temperatures from Nimbus-7 LIMS (Limb Infrared Monitor of the Stratosphere). It is found that eastward propagating disturbances are present in two independent LIMS data sets. The disturbances manifest themselves consistently at several levels throughout the stratosphere and account for much of the tropical temperature variance out to latitudes where extratropical signals become large. Compelling evidence is found for several distinct phase velocities. Spectral analysis indicates isolated eastward variance at principally two frequencies for both wavenumbers.

  13. On the quality of the Nimbus 7 LIMS Version 6 water vapor profiles and distributions

    NASA Astrophysics Data System (ADS)

    Remsberg, E. E.; Natarajan, M.; Lingenfelser, G. S.; Thompson, R. E.; Marshall, B. T.; Gordley, L. L.

    2009-09-01

    This report describes the quality of the Nimbus 7 Limb Infrared Monitor of the Stratosphere (LIMS) water vapor (H2O) profiles of 1978/1979 that were processed with a Version 6 (V6) algorithm and archived in 2002. The V6 profiles incorporate a better knowledge of the instrument attitude for the LIMS measurements along its orbits, leading to improvements for its temperature profiles and for the registration of its water vapor radiances with pressure. As a result, the LIMS V6 zonal-mean distributions of H2O exhibit better hemispheric symmetry than was the case from the original Version 5 (V5) dataset that was archived in 1982. Estimates of the precision and accuracy of the V6 H2O profiles are developed and provided. Individual profiles have a precision of order 5% and an estimated accuracy of about 19% at 3 hPa, 14% at 10 hPa, and 26% at 50 hPa. Profile segments within about 2 km of the tropopause are often affected by emissions from clouds that appear in the finite field-of-view of the detector for the LIMS H2O channel. Zonally-averaged distributions of the LIMS V6 H2O are compared with those from the more recent Microwave Limb Sounder (MLS) satellite experiment for November, February, and May of 2004/2005. The patterns and values of their respective distributions are similar in many respects. Effects of a strengthened Brewer-Dobson circulation are indicated in the MLS distributions of the recent decade versus those of LIMS from 1978/1979. A tropical tape recorder signal is present in the 7-month time series of LIMS V6 H2O with lowest values in February 1979, and the estimated, annually-averaged "entry-level" H2O is 3.5 to 3.8 ppmv. It is judged that this historic LIMS water vapor dataset is of good quality for studies of the near global-scale chemistry and transport for pressure levels from 3 hPa to about 70 to 100 hPa.

  14. On the quality of the Nimbus 7 LIMS Version 6 water vapor profiles and distributions

    NASA Astrophysics Data System (ADS)

    Remsberg, E. E.; Natarajan, M.; Lingenfelser, G. S.; Thompson, R. E.; Marshall, B. T.; Gordley, L. L.

    2009-12-01

    This report describes the quality of the Nimbus 7 Limb Infrared Monitor of the Stratosphere (LIMS) water vapor (H2O) profiles of 1978/79 that were processed with a Version 6 (V6) algorithm and archived in 2002. The V6 profiles incorporate a better knowledge of the instrument attitude for the LIMS measurements along its orbits, leading to improvements for its temperature profiles and for the registration of its water vapor radiances with pressure. As a result, the LIMS V6 zonal-mean distributions of H2O exhibit better hemispheric symmetry than was the case from the original Version 5 (V5) dataset that was archived in 1982. Estimates of the precision and accuracy of the V6 H2O profiles are developed and provided. Individual profiles have a precision of order 5% and an estimated accuracy of about 19% at 3 hPa, 14% at 10 hPa, and 26% at 50 hPa. Profile segments within about 2 km of the tropopause are often affected by emissions from clouds that appear in the finite field-of-view of the detector for the LIMS H2O channel. Zonally-averaged distributions of the LIMS V6 H2O are compared with those from the more recent Microwave Limb Sounder (MLS) satellite experiment for November, February, and May of 2004/05. The patterns and values of their respective distributions are similar in many respects. Effects of a strengthened Brewer-Dobson circulation are indicated in the MLS distributions of the recent decade versus those of LIMS from 1978/79. A tropical tape recorder signal is present in the 7-month time series of LIMS V6 H2O with lowest values in February 1979, and the estimated, annually-averaged "entry-level" H2O is 3.5 to 3.8 ppmv. It is judged that this historic LIMS water vapor dataset is of good quality for studies of the near global-scale chemistry and transport for pressure levels from 3 hPa to about 70 to 100 hPa.

  15. Cofactors of LIM Domains Associate with Estrogen Receptor α to Regulate the Expression of Noncoding RNA H19 and Corneal Epithelial Progenitor Cell Function.

    PubMed

    Klein, Rachel Herndon; Stephens, Denise N; Ho, Hsiang; Chen, Jefferson K; Salmans, Michael L; Wang, Winnie; Yu, Zhengquan; Andersen, Bogi

    2016-06-17

    Cofactors of LIM domain proteins, CLIM1 and CLIM2, are widely expressed transcriptional cofactors that are recruited to gene regulatory regions by DNA-binding proteins, including LIM domain transcription factors. In the cornea, epithelium-specific expression of a dominant negative (DN) CLIM under the keratin 14 (K14) promoter causes blistering, wounding, inflammation, epithelial hyperplasia, and neovascularization followed by epithelial thinning and subsequent epidermal-like differentiation of the corneal epithelium. The defects in corneal epithelial differentiation and cell fate determination suggest that CLIM may regulate corneal progenitor cells and the transition to differentiation. Consistent with this notion, the K14-DN-Clim corneal epithelium first exhibits increased proliferation followed by fewer progenitor cells with decreased proliferative potential. In vivo ChIP-sequencing experiments with corneal epithelium show that CLIM binds to and regulates numerous genes involved in cell adhesion and proliferation, including limbally enriched genes. Intriguingly, CLIM associates primarily with non-LIM homeodomain motifs in corneal epithelial cells, including that of estrogen receptor α. Among CLIM targets is the noncoding RNA H19 whose deregulation is associated with Silver-Russell and Beckwith-Wiedemann syndromes. We demonstrate here that H19 negatively regulates corneal epithelial proliferation. In addition to cell cycle regulators, H19 affects the expression of multiple cell adhesion genes. CLIM interacts with estrogen receptor α at the H19 locus, potentially explaining the higher expression of H19 in female than male corneas. Together, our results demonstrate an important role for CLIM in regulating the proliferative potential of corneal epithelial progenitors and identify CLIM downstream target H19 as a regulator of corneal epithelial proliferation and adhesion. PMID:27129775

  16. Properties and catalytic activities of MICAL1, the flavoenzyme involved in cytoskeleton dynamics, and modulation by its CH, LIM and C-terminal domains.

    PubMed

    Vitali, Teresa; Maffioli, Elisa; Tedeschi, Gabriella; Vanoni, Maria A

    2016-03-01

    MICAL1 is a cytoplasmic 119 kDa protein participating in cytoskeleton dynamics through the NADPH-dependent oxidase and F-actin depolymerizing activities of its N-terminal flavoprotein domain, which is followed by calponin homology (CH), LIM domains and a C-terminal region with Pro-, Glu-rich and coiled-coil motifs. MICAL1 and truncated forms lacking the C-terminal, LIM and/or CH regions have been produced and characterized. The CH, LIM and C-terminal regions cause an increase of Km,NADPH exhibited by the NADPH oxidase activity of the flavoprotein domain, paralleling changes in the overall protein charge. The C-terminus also determines a ∼ 10-fold decrease of kcat, revealing its role in establishing an inactive/active conformational equilibrium, which is at the heart of the regulation of MICAL1 in cells. F-actin lowers Km,NADPH (10-50 μM) and increases kcat (10-25 s(-1)) to similar values for all MICAL forms. The apparent Km,actin of MICAL1 is ∼ 10-fold higher than that of the other forms (3-5 μM), reflecting the fact that F-actin binds to the flavoprotein domain in the MICAL's active conformation and stabilizes it. Analyses of the reaction in the presence of F-actin indicate that actin depolymerization is mediated by H2O2 produced by the NADPH oxidase reaction, rather than due to direct hydroxylation of actin methionine residues. PMID:26845023

  17. The Louvain-La-Neuve sea ice model LIM3.6: global and regional capabilities

    NASA Astrophysics Data System (ADS)

    Rousset, C.; Vancoppenolle, M.; Madec, G.; Fichefet, T.; Flavoni, S.; Barthélemy, A.; Benshila, R.; Chanut, J.; Levy, C.; Masson, S.; Vivier, F.

    2015-10-01

    The new 3.6 version of the Louvain-la-Neuve sea ice model (LIM) is presented, as integrated in the most recent stable release of Nucleus for European Modelling of the Ocean (NEMO) (3.6). The release will be used for the next Climate Model Inter-comparison Project (CMIP6). Developments focussed around three axes: improvements of robustness, versatility and sophistication of the code, which involved numerous changes. Robustness was improved by enforcing exact conservation through the inspection of the different processes driving the air-ice-ocean exchanges of heat, mass and salt. Versatility was enhanced by implementing lateral boundary conditions for sea ice and more flexible ice thickness categories. The latter includes a more practical computation of category boundaries, parameterizations to use LIM3.6 with a single ice category and a flux redistributor for coupling with atmospheric models that cannot handle multiple sub-grid fluxes. Sophistication was upgraded by including the effect of ice and snow weight on the sea surface. We illustrate some of the new capabilities of the code in two standard simulations. One is an ORCA2-LIM3 global simulation at a nominal 2° resolution, forced by reference atmospheric climatologies. The other one is a regional simulation at 2 km resolution around the Svalbard Archipelago in the Arctic Ocean, with open boundaries and tides. We show that the LIM3.6 forms a solid and flexible base for future scientific studies and model developments.

  18. A Couple of "Lim (h[right arrow]0)-Is-Missing" Problems

    ERIC Educational Resources Information Center

    Lau, Ko Hin

    2007-01-01

    Since most students "hate" the concept of limit, in order to make them "happier," this article suggests a couple of naive "lim (h[right arrow]0)-is-missing" problems for them to try for fun. Indeed, differential functional equations that are related to difference quotients in calculus are studied in this paper. In particular, two interesting…

  19. Autism, Accommodation and Treatment: A Rejoinder to Chong-Ming Lim's Critique.

    PubMed

    Jaarsma, Pier; Welin, Stellan

    2015-11-01

    We are very grateful to Chong-Ming Lim for his thoughtful reply published in this journal on one of our articles, which motivated us to think more carefully about accommodating autistic individuals and treating autism. However we believe there are some confusions in Lim's argument. Lim uses the accommodation thesis, according to which we should accommodate autistic individuals rather than treat autism, as the starting point for his reasoning. He claims that if the accommodation thesis is right, then we should not treat autistic individuals for their autism, not even low-functioning (i.e. intellectually disabled) ones, because this would be disrespectful to all autistic individuals. We should instead limit ourselves to accommodate all autistic individuals. However, the opposition between accommodation and treatment is not valid in the case of autism, because of ambiguity in the concepts of accommodation and treatment. Moreover there is confusion in Lim's reasoning caused by omitting important facts about the practice of treating autism. PMID:26307242

  20. Life-Involvement Model (LIM) Project for the 1973-74 Project Year. Final Report.

    ERIC Educational Resources Information Center

    Kapfer, Philip G.; Kapfer, Miriam B.

    The Life-Involvement Model (LIM) Project consisted of four major thrusts: a school-based operational program; a university-based, but field-centered, preservice teacher education program; a school-based in-service teacher education program; and curricular and instructional developmental work designed to support the above three operational programs…

  1. Energetic particle precipitation effects on the Northern Hemisphere stratosphere observed by LIMS

    NASA Astrophysics Data System (ADS)

    Holt, L. A.; Randall, C. E.; Harvey, L.; Funke, B.; Stiller, G. P.

    2009-12-01

    Energetic particle precipitation (EPP) in the upper atmosphere contributes to polar stratospheric enhancements of NOx. The first experimental evidence of this emerged when the Limb Infrared Monitor of the Stratosphere (LIMS) observed stratospheric NOx enhancements during the Arctic winter of 1978/79. Such enhancements have since been observed on numerous occasions, but until recently were much less obvious in the Northern Hemisphere than in the Southern Hemisphere. It is now understood that the magnitude of these stratospheric NOx enhancements depends on both the level of EPP and dynamical conditions. Three out of the last six Arctic winters have seen much larger than average polar stratospheric NOx enhancements due to EPP that have been attributed to extraordinary meteorological events. These are unique events on record and affirm that even with low levels of EPP, the stratosphere can still be influenced to a large degree by EPP. In this study, data from the LIMS instrument is revisited in order to understand the NOx enhancement it observed with respect to meteorological conditions and EPP activity. The temporal evolution of NOx in the polar vortex as measured by LIMS is compared to more recent satellite data, including the Michelson Interferometer for Passive Atmospheric Sounding (MIPAS) instrument. Preliminary results indicate that the enhancements are caused neither by elevated EPP nor unusual dynamical conditions, but rather are seen because of the ability of LIMS to observe in the polar night.

  2. Examination of wintertime latitudinal gradients in stratospheric NO2 using theory and LIMS observations

    NASA Technical Reports Server (NTRS)

    Callis, L. B.; Russell, J. M., III; Haggard, K. V.; Natarajan, M.

    1983-01-01

    Nimbus 7 LIMS data and a photochemical model are used to show that the observed sharp latitudinal gradients in stratospheric wintertime NO2 are consistent with the conversion of NO2 to N2O5 at high latitudes. This conversion, and the sharp gradients, are brought about by the interaction between transport and photochemistry. Calculated variations show good agreement with observations.

  3. The Transcription Factors Islet and Lim3 Combinatorially Regulate Ion Channel Gene Expression

    PubMed Central

    Wolfram, Verena; Southall, Tony D.; Günay, Cengiz; Prinz, Astrid A.; Brand, Andrea H.

    2014-01-01

    Expression of appropriate ion channels is essential to allow developing neurons to form functional networks. Our previous studies have identified LIM-homeodomain (HD) transcription factors (TFs), expressed by developing neurons, that are specifically able to regulate ion channel gene expression. In this study, we use the technique of DNA adenine methyltransferase identification (DamID) to identify putative gene targets of four such TFs that are differentially expressed in Drosophila motoneurons. Analysis of targets for Islet (Isl), Lim3, Hb9, and Even-skipped (Eve) identifies both ion channel genes and genes predicted to regulate aspects of dendritic and axonal morphology. Significantly, some ion channel genes are bound by more than one TF, consistent with the possibility of combinatorial regulation. One such gene is Shaker (Sh), which encodes a voltage-dependent fast K+ channel (Kv1.1). DamID reveals that Sh is bound by both Isl and Lim3. We used body wall muscle as a test tissue because in conditions of low Ca2+, the fast K+ current is carried solely by Sh channels (unlike neurons in which a second fast K+ current, Shal, also contributes). Ectopic expression of isl, but not Lim3, is sufficient to reduce both Sh transcript and Sh current level. By contrast, coexpression of both TFs is additive, resulting in a significantly greater reduction in both Sh transcript and current compared with isl expression alone. These observations provide evidence for combinatorial activity of Isl and Lim3 in regulating ion channel gene expression. PMID:24523544

  4. [Spectra characteristics of LiM (M = Ca, Sr, Ba) BO3 : Tb3+ phosphor].

    PubMed

    Wang, Zhi-Jun; Li, Pan-Lai; Yang, Zhi-Ping; Guo, Qing-Lin

    2009-11-01

    LiM (M = Ca, Sr, Ba) BO3 : Tb3+ phosphors were synthesized by solid state reaction. The starting materials CaCO3, SrCO3, BaCO3, H3 BO3, Li2 CO3, Na2 CO3, K2 CO3 and Tb4 O7 (99.99% in mass) in appropriate stoichiometric ratio were mixed in the alumina crucible, then the mixed powders were calcined at 700 degrees C for 2 h, and LiCaBO3 : Tb3+, LiSrBO3 : Tb3+ and LiB-aBO3 : Tb3+ phosphors were obtained. The emission and excitation spectra were measured by a Shimadzu RF-540 ultraviolet spectrophotometer. All the photoluminescence properties of these phosphors were measured at room temperature. The emission spectra of LiM (M = Ca, Sr, Ba) BO3 : Tb3+ phosphors show several bands, and the main emission peaks correspond to the 5D4 --> 7F6(486, 486, 488 nm), 5D4 --> F5 (544, 544, 544 nm), 5D4 --> 7F4 (590, 595, 593 nm) and 5D4 --> 7F3 (620, 620, 616 nm) typical transitions of Tb3+, and the typical transitions of Tb3+ happens to split because of the effects of LiM (M = Ca, Sr, Ba) BO3 crystals field. The excitation spectra for the 544 nm green emission of LiM (M = Ca, Sr, Ba)BO3 : Tb3+ phosphors illuminate that these kinds of phosphors can be effectively excited by ultraviolet (350-410 nm) light, and emit green light, therefore, they are promising phosphors for white light emitting diodes. Effects of activation and charge compensation on the luminescence intensities of LiM (M = Ca, Sr, Ba) BO3 : Tb3+ phosphors were studied, and the results show that the intensities were obviously effected. PMID:20101952

  5. On the Validation of the Nimbus 7 LIMS Version 6 Stratospheric Ozone

    NASA Astrophysics Data System (ADS)

    Remsberg, E.; Natarajan, M.; Thomason, L.; Lingenfelser, G.

    2005-12-01

    The Nimbus 7 LIMS experiment obtained daily distributions of ozone from late October 1978 through late May 1979 and with good vertical resolution and spatial sampling along its orbital tangent tracks. The historic LIMS Version 5 (V5) dataset was archived in 1982. Studies of those data revealed detailed information about the roles of chemistry and transport on the distribution of ozone in the stratosphere for the latitudes of 64S to 84N. Although its profiles were of good quality throughout most of the stratosphere, both the simulation and validation studies indicated that the accuracy of the V5 ozone became less good below about the 30-hPa level at low latitudes and below the 50-hPa level at middle and high latitudes, especially during polar winter. A LIMS Version 6 (V6) algorithm was developed some years later, in part, to improve on the quality of the ozone profiles in the lower stratosphere. That V6 profile dataset was created in 2001 and archived in 2002. In this paper we present comparisons of the LIMS V6 ozone with the SBUV Version 8 (V8) data, with the preliminary SAGE I Version 6 data, and with balloon ECC ozonesonde soundings from several stations. The findings indicate significant improvements in accuracy for LIMS V6 ozone in the lower stratosphere and better vertical and horizontal sampling for the changes in ozone at polar latitudes and throughout the Arctic winter period. This revised ozone dataset should provide for a better assessment of the effects of chemistry and transport on polar and middle latitude ozone for the Northern Hemisphere winter of 1978-79, when the loss of ozone due to reactive chlorine from heterogeneous processes was not as significant as it is today.

  6. Hydroxyl Radical Regeneration in Isoprene Oxidation: the Upgraded Mechanism LIM1

    NASA Astrophysics Data System (ADS)

    Peeters, J.; Nguyen, S.; Nguyen, T.; Stavrakou, T.; Muller, J. J.

    2012-12-01

    Measured hydroxyl radical concentrations in isoprene-rich areas are much higher than predicted by existing chemical models, to the extent that the global oxidizing capacity of our atmosphere should be significantly revised upwards. The OH regeneration that clearly occurs in isoprene oxidation at low/moderate NO is attributed in the Leuven Isoprene Mechanism to novel, theoretically characterized chemical pathways (LIM0: Peeters et al. 2009; Peeters and Muller 2010). The key new features of LIM0 are (i) thermal equilibration of the labile beta-OH- and delta-OH-isoprenylperoxy isomers; (ii) 1,6-H shift isomerisation of the Z-delta-OH-peroxy isomers to yield hydroperoxy-methyl-butenals (HPALDs); (iii) fast photolysis of the HPALDs resulting overall in several OH radicals per HPALD. The OH-regeneration through photolabile HPALDs has recently found experimental support, but the peroxy isomerisation rate, HPALD yield and the extent of OH recycling are still uncertain (Crounse et al. 2011; Wolfe et al. 2012). In this work, the upgraded LIM1 mechanism is presented. Based on better levels of theory, the crucial equilibrium ratio of the isomerising Z-delta-OH-peroxys over the majority beta-OH-isoprenylperoxys had to be reduced by a factor of about 5 compared to LIM0, while the isomerisation rate of the Z-delta-OH-peroxys adopted from Taraborrelli et al. (2012) is about 3 times lower than in LIM0. The chemistry following the 1,6-H shift of the Z-delta-OH-peroxys is much expanded and extended. Firstly, LIM1 introduces other pathways beside HPALD formation following the Z-delta-OH-peroxy isomerisation, resulting likewise in OH recycling. This, together with the revised Z-delta-OH- equilibrium and isomerisation data above, affords a close model-reproduction of the HPALD and other product yields observed by Crounse et al. (2011). Secondly, LIM1 proposes new fast reactions of HO2 with the alpha-oxoketene products from the peroxy isomerisation routes; these reactions are shown to

  7. The rhombotin family of cysteine-rich LIM-domain oncogenes: Distinct members are involved in T-cell translocations to human chromosomes 11p15 and 11p13

    SciTech Connect

    Boehm, T.; Foroni, L.; Perutz, M.F.; Rabbitts, T.H. ); Kaneko, Y. )

    1991-05-15

    A chromosomal translocation in a T-cell leukemia involving the short arm of human chromosome 11 at band 11p15 disrupts the rhombotin gene. This gene encodes a protein with duplicated cysteine-rich regions called LIM domains, which show homology to zinc-binding proteins and to iron-sulfur centers of ferredoxins. Two homologues of the rhombotin gene have now been isolated. One of these, designated Rhom-2, is located on human chromosome 11 at band 11p13, where a cluster of T-cell leukemia-specific translocations occur; all translocation breakpoints at 11p13 are upstream of the Rhom-2 gene. Human and mouse Rhom-2 are highly conserved and, like rhombotin, encode two tandem cysteine-rich LIM domains. Rhom-2 mRNA is expressed in early mouse development in central nervous system, lung, kidney, liver, and spleen but only very low levels occur in thymus. The other gene, designated Rhom-3, is not on chromosome 11 but also retains homology to the LIM domain of rhombotin. Since the Rhom-2 gene is such a common site of chromosomal damage in T-cell tumors, the consistency of translocations near the rhombotin gene was further examined. A second translocation adjacent to rhombotin was found and at the same position as in the previous example. Therefore, chromosome bands 11p15 (rhombotin) and 11p13 (Rhom-2) are consistent sites of chromosome translocation in T-cell leukemia, with the 11p15 target more rarely involved. The results define the rhombotin gene family as a class of T-cell oncogenes with duplicated cysteine-rich LIM domains.

  8. A comparison of U.S. and USSR rocketsondes using LIMS satellite temperature sounding as a transfer standard

    NASA Technical Reports Server (NTRS)

    Gille, J. C.; Bailey, P. L.; Beck, S. A.

    1984-01-01

    In connection with the reduction of the number of sounding rockets launched per year, there is a general need to make the most effective use of each sounding. In particular, it is often necessary to combine the results from different types of instruments to obtain a meaningful and complete picture of the atmosphere. For this reason, data from different rockets must be compatible. The present investigation is concerned with the utilization of the temperature soundings of the Limb Infrared Monitor of the Stratosphere (LIMS) as the uniform comparison standard. Gille et al. (1984) have compared the LIMS temperatures to a set of 122 U.S. datasonde rockets. In the current study, a comparison between soundings by rocketborne instruments of the USSR and LIMS is combined with the results of the previous LIMS-U.S. rocketsonde comparison in the same latitude bands to provide an updated estimate of the differences between U.S. and USSR rocketborne temperature measurements.

  9. The variability of stratospheric nitrogen compounds observed by LIMS in the winter of 1978-1979

    NASA Technical Reports Server (NTRS)

    Russell, J. M., III; Remsberg, E. E.; Gordley, L. L.; Gille, J. C.; Bailey, P. L.

    1982-01-01

    The LIMS experiment was launched on the Nimbus 7 satellite for the purpose of sounding the vertical structure of temperature and key upper atmosphere trace gases on a global scale. The technique of thermal infrared limb sounding was used to obtain measurements of O3, H2O, NO2, and HNO3. LIMS collected data almost continuously from late October to late May over the latitude range from 64 deg S to 84 deg N. Two of the gases, NO2 and HNO3, are important elements in the NO(x) chain of chemical reactions leading to ozone destruction. Results for these gases are described in terms of zonal mean profiles and latitudinal distributions. The period selected for study is January-May 1979, when a major stratospheric warming occurred.

  10. Variability of stratospheric nitrogen compounds observed by LIMS in the winter of 1978-1979

    SciTech Connect

    Russell, J.M. III; Remsberg, E.E.; Gordley, L.L.

    1982-01-01

    The LIMS experiment was launched on the Nimbus 7 satellite for the purpose of sounding the vertical structure of temperature and key upper atmosphere trace gases on a global scale. The technique of thermal infrared limb sounding was used to obtain measurements of O3, H2O, NO2, and HNO3. LIMS collected data almost continuously from late October to late May over the latitude range from 64 deg S to 84 deg N. Two of the gases, NO2 and HNO3, are important elements in the NO(x) chain of chemical reactions leading to ozone destruction. Results for these gases are described in terms of zonal mean profiles and latitudinal distributions. The period selected for study is January-May 1979, when a major stratospheric warming occurred.

  11. Optimized conditions for selective gold flotation by ToF-SIMS and ToF-LIMS

    NASA Astrophysics Data System (ADS)

    Chryssoulis, S. L.; Dimov, S. S.

    2004-06-01

    This work describes a comprehensive characterization of the factors controlling the floatability of free gold from flotation test using reagents (collectors) at plant concentration levels. A relationship between the collectors loadings on gold particles and their surface composition has been established. The findings of this study show that silver activates gold flotation and there is a strong correlation between the surface concentration of silver and the loading of certain collectors. The organic surface analysis was done by ToF-SIMS while the inorganic surface analysis was carried out by time-of-flight laser ionization mass spectrometry (ToF-LIMS). The developed testing protocol based on ToF-LIMS and ToF-SIMS complementary surface analysis allows for optimization of the flotation scheme and hence improved gold recovery.

  12. Diurnal tide in the equatorial middle atmosphere as seen in LIMS temperatures

    NASA Technical Reports Server (NTRS)

    Hitchman, M. H.; Leovy, C. B.

    1985-01-01

    The distribution of day-night temperature differences in the middle atmosphere determined by the Nimbus 7 LIMS experiment is described. Day-night differences maximize at and are approximately symmetric about the equator. Successive centers of opposite sign increase in amplitude with altitude, the pattern having a vertical wavelength of approximately 25 km. Profiles of rocket meridional wind at Kwajalein (8.7 deg N) and Ascension Island (8.0 deg S) taken near local noon and averaged over the LIMS data period, exhibit maxima which support the tidal interpretation of the equatorial temperature pattern. These characteristics are in general agreement with previous observational and theoretical results for the solar driven diurnal tide. Substantial time variations in amplitude and in location of the temperature maxima are observed. The diurnal tide near the equatorial stratopause appears to be influenced by the phase of the semiannual oscillation.

  13. Ozone budget in the upper stratosphere: Model studies using the reprocessed LIMS and the HALOE datasets

    NASA Astrophysics Data System (ADS)

    Natarajan, Murali; Remsberg, Ellis E.; Gordley, Larry L.

    2002-04-01

    Recently reprocessed LIMS dataset has been used with a contemporary photochemical model to study the balance between photochemical production and destruction of ozone in the upper stratosphere. Model results corresponding to January 1979 indicate that the ozone deficit is less than 15% in the pressure range of 5 to 0.5 mb between 50°S and 50°N latitude. The imbalance at 40 km is much smaller than reported by the earliest studies with the archived LIMS data. The same model, when initialized with HALOE (version 19) data for January, 1996, shows similar results with peak ozone deficits being less than 10%. For both cases, the model shows a near balance in the ozone budget above 1 mb, contrary to recent studies based on balloon-borne measurements. The magnitude of the ozone imbalance seen in this study is within the uncertainties of the data and model.

  14. Comparison of horizontal winds from the LIMS satellite instrument with rocket measurements

    NASA Technical Reports Server (NTRS)

    Smith, A. K.; Bailey, P. L.

    1985-01-01

    Statistical results are given for a comparison between horizontal geostrophic winds computed from satellite height data and all available in situ rocket wind soundings during a 7-month period. The satellite data are the daily mapped fields from the Nimbus 7 Limb Infrared Monitor of the Stratosphere (LIMS) instrument, which extend from 100 to 0.1 mbar. Results indicate that in both the tropics and the extratropical Northern Hemisphere, the average zonal and meridional wind speeds agree to within 2-4 m/s throughout the stratosphere. The rms differences are much larger, with values of 5-10 m/s in the lower stratosphere, increasing to 20-40 m/s in the lower mesosphere. Time series show that LIMS and rocketsonde zonal wind speeds show coherent variations with temporal periods of 1-2 weeks and more, and both exhibit irregular variations on time scales of less than one week.

  15. Estimation of synoptic fields of middle atmosphere parameters from Nimbus-7 LIMS profile data

    NASA Technical Reports Server (NTRS)

    Remsberg, Ellis E.; Haggard, Kenneth V.; Russell, James M., III

    1990-01-01

    The most widely used version of the Nimbus-7 middle atmosphere dataset is the set of high quality, daily, and zonal Fourier coefficients that resolve information out to six wavenumbers at 12 UTC. A Kalman filter algorithm was applied to the original profile data in order to generate those fields for the data archive or LAMAT product. The characteristics and implementation of the algorithm are described in some detail, along with examples of the output for each of the LIMS parameters.

  16. An intercomparison of nitrogen-containing species in Nimbus 7 LIMS and SAMS data

    NASA Technical Reports Server (NTRS)

    Jackman, Charles H.; Guthrie, Paul D.; Kaye, Jack A.

    1987-01-01

    Odd-nitrogen chemistry and transport are analyzed using a two-dimensional model and Nimbus 7 data. The Nimbus 7 data include: measurements of O3, NO2, HNO3, H2O, and temperature by the LIMS instrument; N2O, CH4, and temperature measurements by the SAMS instrument; and O3 data from the SBUV instrument. The characteristics of the two-dimensional model used in the study, a modified Guthrie et al. (1984) model, are described. NO2 and HNO3 are calculated using the two-dimensional model, and the computed data are compared with the LIMS NO2 and HNO3 measurements. The model uncertainties are computed based on the photochemical equilibrium assumption, and the role of stratospheric dynamics in determining NO2 and HNO3 is discussed. It is observed that there is good correlation between the LIMS and model data in the upper stratosphere; however, the data do not correspond in the lower stratosphere. The effect of nitrogen sources, such as lightning, on the stratospheric odd nitrogen distribution is examined.

  17. Gravity Wave Variance in LIMS Temperatures. Part II: Comparison with the Zonal-Mean Momentum Balance.

    NASA Astrophysics Data System (ADS)

    Fetzer, Eric J.; Gille, John C.

    1996-02-01

    Zonal-mean gravity wave variance in the Limb Infrared Monitor of the Stratosphere (LIMS) temperature data is seen to correlate strongly with the residual term in the LIMS zonal-mean momentum budget throughout much of the observed mesosphere. This momentum residual is attributed to gravity wave momentum transport at scales that cannot be directly sampled by the LIMS instrument Correlation is highest in the vicinity of the fall and winter mesospheric jets, where both gravity wave variance and momentum residual reach their largest values. Correlation is also high in the Southern Hemisphere subtropical mesophere, where gravity wave variance and the momentum residual have broad temporal maxima during the easterly acceleration of the stratopause semi-annual oscillation (SAO). This subtropical correlation has important implications for the SAO eastward acceleration, which several studies suggest is forced by gravity wave momentum flux divergence. Correlation between gravity wave variance and inferred gravity wave momentum flux divergence is unexpected because variance is dominated by large scales and long periods (inertio-gravity waves), while both theoretical arguments and ground-based observations indicate that momentum transport is dominated by periods under 1 h. The results of this study suggest a broadband gravity wave field experiencing forcing and loss processes, which are largely independent of frequency.

  18. A Reassessment of the Middle Atmosphere Heat Budget Using Nimbus 7 LIMS Data

    NASA Astrophysics Data System (ADS)

    Mertens, C. J.; Remsberg, E. E.; Mlynczak, M. G.; Natarajan, M.; Gordley, L. L.

    2002-05-01

    A reprocessing of the Nimbus 7 LIMS Level 2 dataset (its Version 6 or V6 profiles) is now complete. The revised algorithm employs a number of changes that affect its temperature T(p) and species profiles. Comparisons with the archived (Version 5 or V5) dataset show that the V6 temperatures are warmer in the mesosphere and in better agreement with correlative rocket profiles. V6 T(p) is cooler by several degrees at high latitudes of the winter upper stratosphere and by about 1 K at all latitudes in the middle stratosphere. V6 ozone is larger than V5 by about 10 percent at 2 to 3 mb, but smaller by about 5 to 10 percent in the middle mesosphere. These changes in temperature and ozone reduce the radiative imbalances that were reported for the stratosphere by Mlynczak et al. (JGR, p. 6039, 1999). Imbalances decrease but are still present in the middle mesosphere because of the non-LTE effects in CO2 and O3 that have not been accounted for in the retrieval of daytime LIMS ozone, in particular. The radiative heating and cooling and the net radiative balance for the V6 dataset will be presented along with revised estimates of the induced diabatic circulation. These new findings are an excellent indication of the quality of LIMS V6 temperature and ozone profiles.

  19. MAPKAPK-2-mediated LIM-kinase activation is critical for VEGF-induced actin remodeling and cell migration

    PubMed Central

    Kobayashi, Miho; Nishita, Michiru; Mishima, Toshiaki; Ohashi, Kazumasa; Mizuno, Kensaku

    2006-01-01

    Vascular endothelial growth factor-A (VEGF-A) induces actin reorganization and migration of endothelial cells through a p38 mitogen-activated protein kinase (MAPK) pathway. LIM-kinase 1 (LIMK1) induces actin remodeling by phosphorylating and inactivating cofilin, an actin-depolymerizing factor. In this study, we demonstrate that activation of LIMK1 by MAPKAPK-2 (MK2; a downstream kinase of p38 MAPK) represents a novel signaling pathway in VEGF-A-induced cell migration. VEGF-A induced LIMK1 activation and cofilin phosphorylation, and this was inhibited by the p38 MAPK inhibitor SB203580. Although p38 phosphorylated LIMK1 at Ser-310, it failed to activate LIMK1 directly; however, MK2 activated LIMK1 by phosphorylation at Ser-323. Expression of a Ser-323-non-phosphorylatable mutant of LIMK1 suppressed VEGF-A-induced stress fiber formation and cell migration; however, expression of a Ser-323-phosphorylation-mimic mutant enhanced these processes. Knockdown of MK2 by siRNA suppressed VEGF-A-induced LIMK1 activation, stress fiber formation, and cell migration. Expression of kinase-dead LIMK1 suppressed VEGF-A-induced tubule formation. These findings suggest that MK2-mediated LIMK1 phosphorylation/activation plays an essential role in VEGF-A-induced actin reorganization, migration, and tubule formation of endothelial cells. PMID:16456544

  20. PP2A binds to the LIM domains of lipoma-preferred partner through its PR130/B″ subunit to regulate cell adhesion and migration.

    PubMed

    Janssens, Veerle; Zwaenepoel, Karen; Rossé, Carine; Petit, Marleen M R; Goris, Jozef; Parker, Peter J

    2016-04-15

    Here, we identify the LIM protein lipoma-preferred partner (LPP) as a binding partner of a specific protein phosphatase 2A (PP2A) heterotrimer that is characterised by the regulatory PR130/B″α1 subunit (encoded byPPP2R3A). The PR130 subunit interacts with the LIM domains of LPP through a conserved Zn(2+)-finger-like motif in the differentially spliced N-terminus of PR130. Isolated LPP-associated PP2A complexes are catalytically active. PR130 colocalises with LPP at multiple locations within cells, including focal contacts, but is specifically excluded from mature focal adhesions, where LPP is still present. An LPP-PR130 fusion protein only localises to focal adhesions upon deletion of the domain of PR130 that binds to the PP2A catalytic subunit (PP2A/C), suggesting that PR130-LPP complex formation is dynamic and that permanent recruitment of PP2A activity might be unfavourable for focal adhesion maturation. Accordingly, siRNA-mediated knockdown of PR130 increases adhesion of HT1080 fibrosarcoma cells onto collagen I and decreases their migration in scratch wound and Transwell assays. Complex formation with LPP is mandatory for these PR130-PP2A functions, as neither phenotype can be rescued by re-expression of a PR130 mutant that no longer binds to LPP. Our data highlight the importance of specific, locally recruited PP2A complexes in cell adhesion and migration dynamics. PMID:26945059

  1. Role of the New South Wales Department of Primary Industries' Laboratory Information Management System (LIMS) in the 2007 equine influenza emergency animal disease response.

    PubMed

    Croft, M G; Fraser, G C; Gaul, W N

    2011-07-01

    A Laboratory Information Management System (LIMS) was used to manage the laboratory data and support planning and field activities as part of the response to the equine influenza outbreak in Australia in 2007. The database structure of the LIMS and the system configurations that were made to best handle the laboratory implications of the disease response are discussed. The operational aspects of the LIMS and the related procedures used at the laboratory to process the increased sample throughput are reviewed, as is the interaction of the LIMS with other corporate systems used in the management of the response. Outcomes from this tailored configuration and operation of the LIMS resulted in effective provision and control of the laboratory and laboratory information aspects of the response. The extent and immediate availability of the information provided from the LIMS was critical to some of the activities of key operatives involved in controlling the response. PMID:21711289

  2. Highly-purified exosomes and shed microvesicles isolated from the human colon cancer cell line LIM1863 by sequential centrifugal ultrafiltration are biochemically and functionally distinct.

    PubMed

    Xu, Rong; Greening, David W; Rai, Alin; Ji, Hong; Simpson, Richard J

    2015-10-01

    Secretion and exchange of extracellular vesicles (EVs) by most cell types is emerging as a fundamental biological process. Although much is known about EVs, there is still a lack of definition as to how many naturally occurring EV subtypes there are and how their properties and functionalities might differ. This vexing issue is critical if EVs are to be fully harnessed for therapeutic applications. To address this question we have developed and describe here a sequential centrifugal ultrafiltration (SCUF) method to examine, in an unbiased manner, what EV subtypes are released in vitro into cell culture medium using the human colon carcinoma cell line LIM1863 as a model system. Using the culture medium from ∼7.2×10(9) LIM1863 cells, SCUF was performed using hydrophilic PVDF membranes with low protein binding properties (Millipore Durapore™ Ultrafree-CL filters with 0.1, 0.22, 0.45 and 0.65 μm pore size). EV particle sizing was measured using both dynamic light scattering and cryo-electron microscopy. Comparative proteome profiling was performed by GeLC-MS/MS and qualitative protein differences between EV subtypes determined by label-free spectral counting. The results showed essentially two EV subtypes; one subtype (fraction Fn1) comprised heterogeneous EVs with particle diameters of 30-1300 nm, the other (fraction Fn5) being homogeneous EVs of 30-100 nm diameter; based on cryo-EM both EV subtypes were round shaped. Western blot analysis showed Fn5 (SCUF-Exos) contained traditional exosome marker proteins (Alix(+), TSG101(+), CD81(+), CD63(+)), while Fn1 (SCUF-sMVs) lacked these protein markers. These findings were consistent with sMVs isolated by differential centrifugation (10,000 g, DC-sMVs) and exosomes (100,000 g EVs depleted of 10,000 g material). The buoyant density of sMVs determined by OptiPrep™ density gradient centrifugation was 1.18-1.19 g/mL and exosomes 1.10-1.11 g/mL. Comparative protein profiling of SCUF-Exos/-sMVs revealed 354 and 606

  3. On the Quality of the Nimbus 7 LIMS Version 6 Ozone for Studies of the Middle Atmosphere

    NASA Technical Reports Server (NTRS)

    Remsberg, Ellis; Lingenfelser, Gretchen; Natarajan, Murali; Gordley, Larry; Thompson, Earl

    2006-01-01

    The Nimbus 7 Limb Infrared Monitor of the Stratosphere (LIMS) radiance profile dataset of 1978/79 was reconditioned and reprocessed to Version 6 (V6) profiles of temperature and species that are improved significantly over those from Version 5 (V5). The LIMS V6 dataset was archived for public use in 2002. Improvements for its ozone include: (1) a more accurate accounting for instrument and spacecraft motion effects in the radiances, (2) the use of better spectroscopic line parameters for its ozone forward model, (3) retrievals of all its scans, (4) more accurate and compatible temperature versus pressure profiles (or T(p)), which are needed for the registration of the ozone radiances and for the removal of temperature effects from them, and (5) a better accounting for interfering species in the lower stratosphere. The retrieved V6 ozone profiles extend from near cloud top altitudes to about 80 km and from 64S to 84N latitude with better sampling along the orbit than for the V5 dataset. Calculated estimates of the single-profile precision and accuracy are provided for the V6 ozone from this study. Precision estimates based on the data themselves are of order 3% or better from 1 to 30 hPa. Estimates of total systematic error for a single profile are hard to generalize because the separate sources of error may not all be of the same sign and they depend somewhat on the atmospheric state. It is estimated that the V6 zonal mean ozone distributions are accurate to within 9% to 7% from 50 hPa to 3 hPa, respectively. Effects of a temperature bias can be significant and may be present at 1 to 2 hPa though. There may be ozone biases of order 10% at those levels due to possible biases of up to +2 K, but there is no indication of a similar problem elsewhere in the stratosphere. Simulation studies show that the LIMS retrievals are also underestimating slightly the small amplitudes of the atmospheric temperature tides, which affect its retrieved day/night ozone differences

  4. adLIMS: a customized open source software that allows bridging clinical and basic molecular research studies

    PubMed Central

    2015-01-01

    Background Many biological laboratories that deal with genomic samples are facing the problem of sample tracking, both for pure laboratory management and for efficiency. Our laboratory exploits PCR techniques and Next Generation Sequencing (NGS) methods to perform high-throughput integration site monitoring in different clinical trials and scientific projects. Because of the huge amount of samples that we process every year, which result in hundreds of millions of sequencing reads, we need to standardize data management and tracking systems, building up a scalable and flexible structure with web-based interfaces, which are usually called Laboratory Information Management System (LIMS). Methods We started collecting end-users' requirements, composed of desired functionalities of the system and Graphical User Interfaces (GUI), and then we evaluated available tools that could address our requirements, spanning from pure LIMS to Content Management Systems (CMS) up to enterprise information systems. Our analysis identified ADempiere ERP, an open source Enterprise Resource Planning written in Java J2EE, as the best software that also natively implements some highly desirable technological advances, such as the high usability and modularity that grants high use-case flexibility and software scalability for custom solutions. Results We extended and customized ADempiere ERP to fulfil LIMS requirements and we developed adLIMS. It has been validated by our end-users verifying functionalities and GUIs through test cases for PCRs samples and pre-sequencing data and it is currently in use in our laboratories. adLIMS implements authorization and authentication policies, allowing multiple users management and roles definition that enables specific permissions, operations and data views to each user. For example, adLIMS allows creating sample sheets from stored data using available exporting operations. This simplicity and process standardization may avoid manual errors and

  5. Interaction of subway LIM vehicle with ballasted track in polygonal wheel wear development

    NASA Astrophysics Data System (ADS)

    Li, Ling; Xiao, Xin-Biao; Jin, Xue-Song

    2011-04-01

    This paper develops a coupled dynamics model for a linear induction motor (LIM) vehicle and a subway track to investigate the influence of polygonal wheels of the vehicle on the dynamic behavior of the system. In the model, the vehicle is modeled as a multi-body system with 35 degrees of freedom. A Timoshenko beam is used to model the rails which are discretely supported by sleepers. The sleepers are modeled as rigid bodies with their vertical, lateral, and rolling motions being considered. In order to simulate the vehicle running along the track, a moving sleeper support model is introduced to simulate the excitation by the discrete sleeper supporters, in which the sleepers are assumed to move backward at a constant speed that is the same as the train speed. The Hertzian contact theory and the Shen-Hedrick-Elkins' model are utilized to deal with the normal dynamic forces and the tangential forces between wheels and rails, respectively. In order to better characterize the linear metro system (LMS), Euler beam theory based on modal superposition method is used to model LIM and RP. The vertical electric magnetic force and the lateral restoring force between the LIM and RP are also taken into consideration. The former has gap-varying nonlinear characteristics, whilst the latter is considered as a constant restoring force of 1 kN. The numerical analysis considers the effect of the excitation due to polygonal wheels on the dynamic behavior of the system at different wear stages, in which the used data regarding the polygonal wear on the wheel tread are directly measured at the subway site.

  6. The Louvain-la-Neuve sea ice model LIM3.5: global and regional capabilities

    NASA Astrophysics Data System (ADS)

    Rousset, C.; Vancoppenolle, M.; Madec, G.; Fichefet, T.; Flavoni, S.; Barthélemy, A.; Benshila, R.; Chanut, J.; Levy, C.; Masson, S.; Vivier, F.

    2015-04-01

    We present the new 3.5 version of the Louvain-la-Neuve sea ice model (LIM) integrated in NEMO 3.6. The main novelty is the improvement of model robustness and versatility for a wide range of applications, from global to regional scales. Several modifications to the code were required. First, the time stepping scheme of the model was changed from parallel to sequential (ice dynamics first, then thermodynamics). Such a scheme enables to diagnose the different physical processes responsible for exchanges through the air-ice-ocean interfaces, as well as the online inspection of mass, heat and salt conservation properties of the code. In the course of these developments, several minor conservation leaks were found and fixed, so that LIM3.5 is exactly conservative. Second, lateral boundary conditions for regional ice-covered configurations have been implemented. To illustrate the new capabilities, two simulations are performed. One is a global simulation at a nominal 2° resolution forced by atmospheric climatologies and is found reasonably realistic although no specific tuning was done. The other is a regional simulation at 2 km resolution around the Svalbard Archipelago in the Arctic Ocean, with prescribed conditions at the four boundaries including tides. The simulation is able to resolve small-scale features and transient events such as the opening and closing of coastal polynyas. The ice mass budgets for both simulations are illustrated and mostly differ by the strength of ice formation in open water. LIM3.5 now forms a solid base for future scientific studies and model developments.

  7. A study of the ozone photochemistry in the upper stratosphere using LIMS data

    NASA Technical Reports Server (NTRS)

    Natarajan, M.; Callis, L. B.; Russell, J. M., III; Boughner, R. E.

    1985-01-01

    LIMS data at vernal equinox conditions are used to study the photochemistry of the upper stratosphere. The results indicate, and it has been recently reported, that with the use of recommended reaction rates, current models underestimate ozone mixing ratio by 20-40 percent. For ozone, good agreement with data is realized with the modification of six key reaction rates within the published limits of uncertainty. These modifications also yield better agreement with data for daytime NO2. Model results for other parameters such as the ratio HNO3/NO2, OH mixing ratio, and the temperature sensitivity of O3 are compared with data.

  8. Effects of polar stratospheric clouds in the Nimbus 7 LIMS Version 6 data set

    NASA Astrophysics Data System (ADS)

    Remsberg, Ellis; Harvey, V. Lynn

    2016-07-01

    The historic Limb Infrared Monitor of the Stratosphere (LIMS) measurements of 1978-1979 from the Nimbus 7 satellite were re-processed with Version 6 (V6) algorithms and archived in 2002. The V6 data set employs updated radiance registration methods, improved spectroscopic line parameters, and a common vertical resolution for all retrieved parameters. Retrieved profiles are spaced about every 1.6° of latitude along orbits and include the additional parameter of geopotential height. Profiles of O3 are sensitive to perturbations from emissions of polar stratospheric clouds (PSCs). This work presents results of implementing a first-order screening for effects of PSCs using simple algorithms based on vertical gradients of the O3 mixing ratio. Their occurrences are compared with the co-located, retrieved temperatures and related to the temperature thresholds needed for saturation of H2O and/or HNO3 vapor onto PSC particles. Observed daily locations where the major PSC screening criteria are satisfied are validated against PSCs observed with the Stratospheric Aerosol Monitor (SAM) II experiment also on Nimbus 7. Remnants of emissions from PSCs are characterized for O3 and HNO3 following the screening. PSCs may also impart a warm bias in the co-located LIMS temperatures, but by no more than 1-2 K at the altitudes of where effects of PSCs are a maximum in the ozone; thus, no PSC screening was applied to the V6 temperatures. Minimum temperatures vary between 187 and 194 K and often occur 1 to 2 km above where PSC effects are first identified in the ozone (most often between about 21 and 28 hPa). Those temperature-pressure values are consistent with conditions for the existence of nitric acid trihydrate (NAT) mixtures and to a lesser extent of super-cooled ternary solution (STS) droplets. A local, temporary uptake of HNO3 vapor of order 1-3 ppbv is indicated during mid-January for the 550 K surface. Seven-month time series of the distributions of LIMS O3 and HNO3 are shown

  9. Description of data on the Nimbus 7 LIMS map archive tape: Water vapor and nitrogen dioxide

    NASA Technical Reports Server (NTRS)

    Haggard, Kenneth V.; Marshall, B. T.; Kurzeja, Robert J.; Remsberg, Ellis E.; Russell, James M., III

    1988-01-01

    Described is the process by which the analysis of the Limb Infrared Monitor of the Stratosphere (LIMS) experiment data were used to produce estimates of synoptic maps of water vapor and nitrogen dioxide. In addition to a detailed description of the analysis procedure, also discussed are several interesting features in the data which are used to demonstrate how the analysis procedure produced the final maps and how one can estimate the uncertainties in the maps. In addition, features in the analysis are noted that would influence how one might use, or interpret, the results. These include subjects such as smoothing and the interpretation of wave components.

  10. Impact of the integration of a Maxwell-elastic-brittle rheology in NEMO-LIM3

    NASA Astrophysics Data System (ADS)

    Raulier, Jonathan; Fichefet, Thierry; Legat, Vincent; Weiss, Jérôme; Dansereau, Véronique

    2015-04-01

    Satellite observations of the Arctic sea ice show the existence of a dense mesh of leads constantly opening and closing over short time scales. Those leads are highly linked to the presence of linear kinematic features which are quasi linear patterns present in the strain field that stretch all across the Arctic basin. Current sea ice models fail to reproduce those linear kinematic features and the observed statistical distribution of deformation rate. In order to refine the physical representation of sea ice dynamics into sea ice models, a new approach has been adopted for the rheology of sea ice. This approach, based on a Maxwell elasto-brittle rheology, is being integrated in the NEMO-LIM3 global ocean-sea ice model (\\url{www.nemo-ocean.eu} ; \\url{www.elic.ucl.ac.be/lim}). In the present study, we examine the influence of the new rheology on the statistical characteristics of the simulated deformation rate and on the ability of the model to reproduce the existence of leads within the ice pack. We will also address the impact of the representation of leads on the fluxes between atmosphere and ocean.

  11. Description of data on the Nimbus 7 LIMS map archive tape: Ozone and nitric acid

    NASA Technical Reports Server (NTRS)

    Remsberg, E. E.; Kurzeja, R. J.; Haggard, K. V.; Russell, J. M., III; Gordley, L. L.

    1986-01-01

    The Nimbus 7 Limb Infrared Monitor of the Stratosphere (LIMS) data set has been processed into a Fourier coefficient representation with a Kalman filter algorithm applied to profile data at individual latitudes and pressure levels. The algorithm produces synoptic data at noon Greenwich Mean Time (GMT) from the asynoptic orbital profiles. This form of the data set is easy to use and is appropriate for time series analysis and further data manipulation and display. Ozone and nitric acid results are grouped together in this report because the LIMS vertical field of views (FOV's) and analysis characteristics for these species are similar. A comparison of the orbital input data with mixing ratios derived from Kalman filter coefficients indicates errors in mixing ratio of generally less than 5 percent, with 15 percent being a maximum error. The high quality of the mapped data was indicated by coherence of both the phases and the amplitudes of waves with latitude and pressure. Examples of the mapped fields are presented, and details are given concerning the importance of diurnal variations, the removal of polar stratospheric cloud signatures, and the interpretation of bias effects in the data near the tops of profiles.

  12. Description of data on the Nimbus 7 LIMS map archive tape: ozone and nitric acid

    SciTech Connect

    Remsberg, E.E.; Kurzeja, R.J.; Haggard, K.V.; Russell, J.M. III; Gordley, L.L.

    1986-12-01

    The Nimbus 7 Limb infrared monitor of the stratosphere (LIMS) data set has been processed into a Fourier coefficient representation with a Kalman filter algorithm applied to profile data at individual latitudes and pressure levels. The algorithm produces synoptic data at noon Greenwich Mean Time (GMT) from the asynoptic orbital profiles. This form of the data set is easy to use and is appropriate for time series analysis and further data manipulation and display. Ozone and nitric acid results are grouped together in this report because the LIMS vertical field of views (FOV's) and analysis characteristics for these species are similar. A comparison of the orbital input data with mixing ratios derived from Kalman filter coefficients indicates errors in mixing ratio of generally less than 5 percent, with 15 percent being a maximum error. The high quality of the mapped data was indicated by coherence of both the phases and the amplitudes of waves with latitude and pressure. Examples of the mapped fields are presented, and details are given concerning the importance of diurnal variations, the removal of polar stratospheric cloud signatures, and the interpretation of bias effects in the data near the tops of profiles.

  13. The Limb Infrared Monitor of the Stratosphere /LIMS/ experiment - Temperature and nitrogen dioxide results

    NASA Technical Reports Server (NTRS)

    Gille, J. C.; Russell, J. M.; Bailey, P. L.

    1981-01-01

    The Limb Infrared Monitor of the Stratosphere (LIMS) is a satellite-borne radiometer which scans across the earth's limb, measuring infrared radiation emitted by specific atmospheric gases in a number of broad channels. Instrumentation includes a folded telescope, which focuses the radiation from the limb onto an array of six mercury-cadmium-telluride detectors. A scan mirror within the telescope sweeps the fields of view across the horizon, with each up or down scan lasting 12 seconds. The measurements are inverted to yield distributions of temperature and nitrogen dioxide. The temperature is determined from two channels covering the intervals 595-739 per cm and 645-673 per cm of the 15 micron bands of CO2, with maximum signal to noise ratio of 500-1000. Nitrogen dioxide is determined from measurements in a channel from 1580-1613 per cm, with an S/N of approximately 100. Results show that the LIMS temperature and nitrogen dioxide data are characterized by high precision.

  14. LIMS Instrument Package (LIP) balloon experiment: Nimbus 7 satellite correlative temperature, ozone, water vapor, and nitric acid measurements

    NASA Technical Reports Server (NTRS)

    Lee, R. B., III; Gandrud, B. W.; Robbins, D. E.; Rossi, L. C.; Swann, N. R. W.

    1982-01-01

    The Limb Infrared Monitor of the Stratosphere (LIMS) LIP balloon experiment was used to obtain correlative temperature, ozone, water vapor, and nitric acid data at altitudes between 10 and 36 kilometers. The performance of the LIMS sensor flown on the Nimbus 7 Satellite was assessed. The LIP consists of the modified electrochemical concentration cell ozonesonde, the ultraviolet absorption photometric of ozone, the water vapor infrared radiometer sonde, the chemical absorption filter instrument for nitric acid vapor, and the infrared radiometer for nitric acid vapor. The limb instrument package (LIP), its correlative sensors, and the resulting data obtained from an engineering and four correlative flights are described.

  15. FluxTransgenics: a flexible LIMS-based tool for management of plant transformation experimental data

    PubMed Central

    2014-01-01

    Background The production and commercial release of Genetically Modified Organisms (GMOs) are currently the focus of important discussions. In order to guarantee the quality and reliability of their trials, companies and institutions working on this subject must adopt new approaches on management, organization and recording of laboratory conditions where field studies are performed. Computational systems for management and storage of laboratory data known as Laboratory Information Management Systems (LIMS) are essential tools to achieve this. Results In this work, we have used the SIGLa system – a workflow based LIMS as a framework to develop the FluxTransgenics system for a GMOs laboratory of Empresa Brasileira de Pesquisa Agropecuária (EMBRAPA) Maize and Sorghum (Sete Lagoas, MG - Brazil). A workflow representing all stages of the transgenic maize plants generation has been developed and uploaded in FluxTransgenics. This workflow models the activities involved in maize and sorghum transformation using the Agrobacterium tumefaciens method. By uploading this workflow in the SIGLa system we have created Fluxtransgenics, a complete LIMS for managing plant transformation data. Conclusions FluxTransgenics presents a solution for the management of the data produced by a laboratory of genetically modified plants that is efficient and supports different kinds of information. Its adoption will contribute to guarantee the quality of activities and products in the process of transgenic production and enforce the use of Good Laboratory Practices (GLP). The adoption of the transformation protocol associated to the use of FluxTransgenics has made it possible to increase productivity by at least 300%, increasing the efficiency of the experiments from between 0.5 and 1 percent to about 3%. This has been achieved by an increase in the number of experiments performed and a more accurate choice of parameters, all of which have been made possible because it became easier to

  16. LIMS data - Inferred stratospheric distribution of NOx and HOx trace constituents and the calculated odd nitrogen budget

    NASA Technical Reports Server (NTRS)

    Callis, L. B.; Russell, J. M.; Boughner, R. E.; Natarajan, M.

    1985-01-01

    LIMS, SAMS, SBUV and in-situ data have been used to infer species not measured but which are of photochemical interest, e.g., O(3P), O(1D), NO, N2O5, OH, HO2, ClO and HCl. (LIMS = limb infrared monitor of the stratosphere; SAMS = stratospheric and mesospheric sounder; and SBUV = solar backscattered ultraviolet instrument.) Production and loss of odd nitrogen have been calculated and estimates have been made of the odd nitrogen transport due to adiabatically driven circulation derived from LIMS data. Data used from LIMS include O3, NO2, HNO3, H2O and T. CH4 and N2O were taken from SAMS and the UV solar flux from the SBUV instrument. Species were inferred for periods in October, December, March and May. Results for December are discussed. Results indicate: (1) maximum stratospheric odd nitrogen levels of 25 ppbv; (2) evidence of odd nitrogen transport from the mesosphere appearing at 25 km in the wintertime polar latitudes; (3) the polar night build-up of high levels of N2O5 beginning after the autumnal equinox; and (4) the possibility of large downward fluxes of odd nitrogen into the troposphere during the winter at latitudes poleward of 60 degrees.

  17. Lmo4 and Other LIM domain only factors are necessary and sufficient for multiple retinal cell type development.

    PubMed

    Jin, Kangxin; Xiao, Dongchang; Andersen, Bogi; Xiang, Mengqing

    2016-08-01

    Understanding the molecular basis by which distinct cell types are specified is a central issue in retinogenesis and retinal disease development. Here we examined the role of LIM domain only 4 (Lmo4) in retinal development using both gain-of-function and loss-of-function approaches. By immunostaining, Lmo4 was found to be expressed in mouse retina from E10.5 to mature stages. Retroviral delivery of Lmo4 into retinal progenitor cells could promote the amacrine, bipolar and Müller cell fates at the expense of photoreceptors. It also inhibited the fate of early-born retinal ganglion cells. Using a dominant-negative form of Lmo4 which suppresses transcriptional activities of all LIM domain only factors, we demonstrated that LIM domain only factors are both necessary and sufficient for promoting amacrine and bipolar cell development, but not for the differentiation of ganglion, horizontal, Müller, or photoreceptor cells. Taken together, our study uncovers multiple roles of Lmo4 during retinal development and demonstrates the importance of LIM domain only factors in ensuring proper retinal cell specification and differentiation. © 2015 Wiley Periodicals, Inc. Develop Neurobiol 76: 900-915, 2016. PMID:26579872

  18. Development of quantitative laser ionization mass spectrometry (LIMS). Final report, 1 Aug 87-1 Jan 90

    SciTech Connect

    Odom, R.W.

    1991-06-04

    The objective of the research was to develop quantitative microanalysis methods for dielectric thin films using the laser ionization mass spectrometry (LIMS) technique. The research involved preparation of thin (5,000 A) films of SiO2, Al2O3, MgF2, TiO2, Cr2O3, Ta2O5, Si3N4, and ZrO2, and doping these films with ion implant impurities of 11B, 40Ca, 56Fe, 68Zn, 81Br, and 121Sb. Laser ionization mass spectrometry (LIMS), secondary ion mass spectrometry (SIMS) and Rutherford backscattering spectrometry (RBS) were performed on these films. The research demonstrated quantitative LIMS analysis down to detection levels of 10-100 ppm, and led to the development of (1) a compound thin film standards product line for the performing organization, (2) routine LIMS analytical methods, and (3) the manufacture of high speed preamplifiers for time-of-flight mass spectrometry (TOF-MS) techniques.

  19. On the quality of the Nimbus 7 LIMS version 6 ozone for studies of the middle atmosphere

    NASA Astrophysics Data System (ADS)

    Remsberg, Ellis; Lingenfelser, Gretchen; Natarajan, Murali; Gordley, Larry; Marshall, B. Thomas; Thompson, Earl

    2007-07-01

    The Nimbus 7 Limb Infrared Monitor of the Stratosphere (LIMS) radiance profile dataset of 1978/79 was reconditioned and reprocessed to Version 6 (V6) profiles of temperature and species that are improved significantly over those from Version 5 (V5). The LIMS V6 dataset was archived for public use in 2002. Improvements for its ozone include: (1) a more accurate accounting for instrument and spacecraft motion effects in the radiances, (2) the use of better spectroscopic line parameters for its ozone forward model, (3) retrievals of all its scans, (4) more accurate and compatible temperature versus pressure profiles (or T(p)) that are needed for the registration of the ozone radiances and for the removal of temperature effects from them, and (5) a better accounting for interfering species in the lower stratosphere. The retrieved V6 ozone profiles extend from near cloud top altitudes to about 80 km and from 64S to 84N latitude with better sampling along the orbit than for the V5 dataset. Calculated estimates of the single-profile precision and accuracy are provided; precision estimates based on the data themselves are of order 3% or better from 1 to 30 hPa. Estimates of total systematic error are hard to generalize because the separate sources of error may not all be of the same sign, and they depend somewhat on the atmospheric state. It is estimated that the accuracy of the V6 zonal mean ozone distribution is within ±9% from 50 10 hPa, improving to ±7% in the uppermost stratosphere. Simulation studies show that the LIMS T(p) retrievals are underestimating slightly the small amplitudes of the atmospheric temperature tides, which affect the retrieved day/night ozone differences. There are also small biases in the middle to lower stratosphere for the ascending versus descending node LIMS ozone, due principally to not accounting for the asymmetric weighting of its radiance within the tangent layer. The total accuracy for the LIMS ozone was assessed by comparing its

  20. Description of data on the Nimbus 7 LIMS map archive tape: Temperature and geopotential height

    NASA Technical Reports Server (NTRS)

    Haggard, K. V.; Remsberg, E. E.; Grose, W. L.; Russell, J. M., III; Marshall, B. T.; Lingenfelser, G.

    1986-01-01

    The process by which the analysis of the Limb Infared Monitor of the Stratosphere (LIMS) experiment data were used to produce estimates of synoptic maps of temperature and geopotential height is described. In addition to a detailed description of the analysis procedure, several interesting features in the data are discussed and these features are used to demonstrate how the analysis procedure produced the final maps and how one can estimate the uncertainties in the maps. In addition, features in the analysis are noted that would influence how one might use, or interpret, the results. These include subjects such as smoothing and the interpretation of wave components. While some suggestions are made for an improved analysis of the data, it is shown that, in general, the maps are an excellent estimation of the synoptic fields.

  1. A Lagrangian analysis of a sudden stratospheric warming - Comparison of a model simulation and LIMS observations

    NASA Technical Reports Server (NTRS)

    Pierce, R. B.; Remsberg, Ellis E.; Fairlie, T. D.; Blackshear, W. T.; Grose, William L.; Turner, Richard E.

    1992-01-01

    Lagrangian area diagnostics and trajectory techniques are used to investigate the radiative and dynamical characteristics of a spontaneous sudden warming which occurred during a 2-yr Langley Research Center model simulation. The ability of the Langley Research Center GCM to simulate the major features of the stratospheric circulation during such highly disturbed periods is illustrated by comparison of the simulated warming to the observed circulation during the LIMS observation period. The apparent sink of vortex area associated with Rossby wave-breaking accounts for the majority of the reduction of the size of the vortex and also acts to offset the radiatively driven increase in the area occupied by the 'surf zone'. Trajectory analysis of selected material lines substantiates the conclusions from the area diagnostics.

  2. Estimates of the stratospheric distribution of odd nitrogen from the LIMS data

    NASA Technical Reports Server (NTRS)

    Callis, L. B.; Russell, J. M., III; Natarajan, M.

    1985-01-01

    Reasonable lower-limit estimates of the latitude-altitude distribution of stratospheric NO(x) for December 1978 and March 1979 are presented. The lower-limit estimates are based on nighttime measurements of NO2 and HNO3 taken by the LIMS instrument aboard the NIMBUS 7 satellite. It is shown that the estimates do not depend upon model calculations or a priori knowledge of the nature of the stratospheric photochemical system. The results indicate that the measured sum of nighttime NO2 nd HNO3 is as high as 22.5 + or - ppbv and that atmospheric NO(x) levels may be as high as 26 + or - 4.5 ppbv at 37 km, which is larger than most calculated NO(x) levels.

  3. Improvements in the profiles and distributions of nitric acid and nitrogen dioxide with the LIMS version 6 dataset

    NASA Astrophysics Data System (ADS)

    Remsberg, E.; Natarajan, M.; Marshall, B. T.; Gordley, L. L.; Thompson, R. E.; Lingenfelser, G.

    2010-05-01

    The quality of the Nimbus 7 Limb Infrared Monitor of the Stratosphere (LIMS) nitric acid (HNO3) and nitrogen dioxide (NO2) profiles and distributions of 1978/1979 are described after their processing with an updated, Version 6 (V6) algorithm and subsequent archival in 2002. Estimates of the precision and accuracy of both of those species are developed and provided herein. The character of the V6 HNO3 profiles is relatively unchanged from that of the earlier LIMS Version 5 (V5) profiles, except in the upper stratosphere where the interfering effects of CO2 are accounted for better with V6. The accuracy of the retrieved V6 NO2 is also significantly better in the middle and upper stratosphere, due to improvements in its spectral line parameters and in the reduced biases for the accompanying V6 temperature and water vapor profiles. As a result of these important updates, there is better agreement with theoretical calculations for profiles of the HNO3/NO2 ratio, day-to-night NO2 ratio, and with estimates of the production of NO2 in the mesosphere and its descent to the upper stratosphere during polar night. In particular, the findings for middle and upper stratospheric NO2 should also be more compatible with those obtained from more recent satellite sensors because the effects of the spin-splitting of the NO2 lines are accounted for now with the LIMS V6 algorithm. The improved precisions and more frequent retrievals of the LIMS profiles along their orbit tracks provide for better continuity and detail in map analyses of these two species on pressure surfaces. It is judged that the chemical effects of the oxides of nitrogen on ozone can be studied quantitatively throughout the stratosphere with the LIMS V6 data.

  4. LIM Kinase, a Newly Identified Regulator of Presynaptic Remodeling by Rod Photoreceptors After Injury

    PubMed Central

    Wang, Weiwei; Townes-Anderson, Ellen

    2015-01-01

    Purpose Rod photoreceptors retract their axon terminals and develop neuritic sprouts in response to retinal detachment and reattachment, respectively. This study examines the role of LIM kinase (LIMK), a component of RhoA and Rac pathways, in the presynaptic structural remodeling of rod photoreceptors. Methods Phosphorylated LIMK (p-LIMK), the active form of LIMK, was examined in salamander retina with Western blot and confocal microscopy. Axon length within the first 7 hours and process growth after 3 days of culture were assessed in isolated rod photoreceptors treated with inhibitors of upstream regulators ROCK and p21-activated kinase (Pak) (Y27632 and IPA-3) and a direct LIMK inhibitor (BMS-5). Porcine retinal explants were also treated with BMS-5 and analyzed 24 hours after detachment. Because Ca2+ influx contributes to axonal retraction, L-type channels were blocked in some experiments with nicardipine. Results Phosphorylated LIMK is present in rod terminals during retraction and in newly formed processes. Axonal retraction over 7 hours was significantly reduced by inhibition of LIMK or its regulators, ROCK and Pak. Process growth was reduced by LIMK or Pak inhibition especially at the basal (axon-bearing) region of the rod cells. Combining Ca2+ channel and LIMK inhibition had no additional effect on retraction but did further inhibit sprouting after 3 days. In detached porcine retina, LIMK inhibition reduced rod axonal retraction and improved retinal morphology. Conclusions Thus structural remodeling, in the form of either axonal retraction or neuritic growth, requires LIMK activity. LIM kinase inhibition may have therapeutic potential for reducing pathologic rod terminal plasticity after retinal injury. PMID:26658506

  5. The integrated enstrophy budget of the winter stratosphere diagnosed from LIMS data

    NASA Technical Reports Server (NTRS)

    Schoeberl, M. R.; Smith, A. K.

    1985-01-01

    The quasi-geostrophic integrated enstrophy budget for the 1978 to 1979 winter has been analyzed from 10-0.1 mb using LIMS data. During January and late February periods a significant imbalance in the budget appears at 10mb. This imbalance is attributed to Rossby wave breaking. It is produced by the irreversible transfer of enstrophy to smaller scales not resolved by LIMS. The imbalance episodes correspond well to the appearance of Ertel vorticity filaments shown by McIntyre and Palmer (1984). From a seasonal viewpoint, the integrated enstrophy shows an average (although irregular) transfer from a zonal mean reservoir to waves which are then dissipated. On a shorter time scale the integrated enstrophy sloshes back and forth between the waves and mean flow in early winter; then, beginning with the January sudden warming, the total enstrophy is reduced more rapidly. Between 10 mb and 1 mb this reduction is more or less continuous until the end of February. However, in the mesosphere the total enstrophy decrease is very short lived, being quickly restored after the January warming. Even though the zonal mean integrated enstrophy is large, only about 10% can be utilized by the waves. The available integrated potential enstrophy is introduced, which is a better measure of how close the flow is to saturation by Rossby waves. The largest amount of available potential enstrophy in early January is at 1 mb with decreasing amounts above and below. Saturation of the flow by Rossby waves occurs below 1 mb only coincident with sudden warmings; however, at mesospheric heights the flow appears to be nearly saturated throughout the winter.

  6. Critical Roles of the LIM Domains of Lhx3 in Recruiting Coactivators to the Motor Neuron-Specifying Isl1-Lhx3 Complex.

    PubMed

    Seo, So Yeon; Lee, Bora; Lee, Seunghee

    2015-10-01

    During spinal cord development, the LIM domains of the LIM homeodomain factor Lhx3 bind to either the LIM cofactor nuclear LIM interactor (NLI) or another LIM homeodomain factor, Isl1, assembling the tetrameric V2 interneuron-specifying Lhx3 complex (2NLI:2Lhx3) or the hexameric motor neuron-specifying Isl1-Lhx3 complex (2NLI:2Isl1:2Lhx3). However, the detailed molecular basis by which the Lhx3-LIM domains contribute to motor neuron specification still remains poorly understood. Here, we show that the Lhx3-LIM domains are essential for recruiting transcriptional coactivators to the Isl1-Lhx3 complex. Using a yeast genetic screening system, we identify Lhx3 point mutants that bind to NLI but not Isl1. Accordingly, these mutants fail to assemble the Isl1-Lhx3 complex. However, their interaction with coactivators is relatively intact, and they are fully functional in the Lhx3 complex and V2 interneuron specification. Interestingly, when these Lhx3 mutants are directly fused to Isl1, their transcriptional activity in the Isl1-Lhx3 complex is restored. We further show that this restoration reflects an unexpected role of the Lhx3-LIM domains, likely together with Isl1, to form an interaction interface for coactivators. Our results suggest that the Lhx3-LIM domains play critical roles in transactivation of the Isl1-Lhx3 complex by not only directing the assembly of the Isl1-Lhx3 complex but also recruiting coactivators to the complex. PMID:26260513

  7. Critical Roles of the LIM Domains of Lhx3 in Recruiting Coactivators to the Motor Neuron-Specifying Isl1-Lhx3 Complex

    PubMed Central

    Seo, So Yeon; Lee, Bora

    2015-01-01

    During spinal cord development, the LIM domains of the LIM homeodomain factor Lhx3 bind to either the LIM cofactor nuclear LIM interactor (NLI) or another LIM homeodomain factor, Isl1, assembling the tetrameric V2 interneuron-specifying Lhx3 complex (2NLI:2Lhx3) or the hexameric motor neuron-specifying Isl1-Lhx3 complex (2NLI:2Isl1:2Lhx3). However, the detailed molecular basis by which the Lhx3-LIM domains contribute to motor neuron specification still remains poorly understood. Here, we show that the Lhx3-LIM domains are essential for recruiting transcriptional coactivators to the Isl1-Lhx3 complex. Using a yeast genetic screening system, we identify Lhx3 point mutants that bind to NLI but not Isl1. Accordingly, these mutants fail to assemble the Isl1-Lhx3 complex. However, their interaction with coactivators is relatively intact, and they are fully functional in the Lhx3 complex and V2 interneuron specification. Interestingly, when these Lhx3 mutants are directly fused to Isl1, their transcriptional activity in the Isl1-Lhx3 complex is restored. We further show that this restoration reflects an unexpected role of the Lhx3-LIM domains, likely together with Isl1, to form an interaction interface for coactivators. Our results suggest that the Lhx3-LIM domains play critical roles in transactivation of the Isl1-Lhx3 complex by not only directing the assembly of the Isl1-Lhx3 complex but also recruiting coactivators to the complex. PMID:26260513

  8. New snow thermodynamics for the Louvain-la-Neuve Sea Ice Model (LIM)

    NASA Astrophysics Data System (ADS)

    Lecomte, Olivier; Fichefet, Thierry

    2010-05-01

    The Louvain-la-Neuve sea Ice Model (LIM) is a three-dimensional global model for sea ice dynamics and thermodynamics that has been specifically designed for climate studies and that is fully coupled with the oceanic general circulation model OPA on the modelling platform NEMO. This study presents and assesses the skills of a new one-dimensional snow model developed for the thermodynamic component of LIM, by comparison with the former model thermodynamics and observations. Snow is a key element in sea ice physics and in the interactions between sea ice and atmosphere. Owing to its low thermal conductivity and hight albedo, the snow cover is a very efficient insulator and it contributes directly and indirectly to the sea ice mass balance. Given the high variability and heterogeneity of the snow cover above sea ice, it is necessary to represent different types of snow, depending on their characteristics. A multilayer approach has been chosen for the model, with time varying temperature, density and thermal conductivity for each layer. Vertical heat diffusion, surface and internal melt, precipitations, snow ice formation and a parameterisation for melt pond albedo are included in the model. The model has been validated at Ice Station POLarstern (ISPOL) in the western Weddell Sea during summer and at Point Barrow (Alaska) during winter. The new model simulates better temperature profiles, with an amount of good correlations between modelled and observed profiles increasing from 12% to 42% for the 2-layer and 6-layer configurations, respectively. Conductive fluxes and temperatures are highly sensitive to albedo and ocean heat flux during summer, and to the thermal conductivity parameterisation during winter. Ice ablation rate is quite insensitive to snow thermal conductivity in summer because almost all the variability at the surface is absorbed by snow, making the temperature gradient in the ice relatively small and steady. Nevertheless, during winter, when the air

  9. Validation of nitrogen dioxide results measured by the limb infrared monitor of the stratosphere (LIMS) experiment on NIMBUS 7

    NASA Technical Reports Server (NTRS)

    Russell, J. M., III; Remsberg, E. E.; Gille, J. C.; Bailey, P. L.; Gordley, L. L.; Drayson, S. R.; Fischer, H.; Girard, A.; Harries, J. E.; Evans, W. F. J.

    1984-01-01

    The validation of results from the nitrogen dioxide channel and the quality of the data are examined in connection with the LIMS experiment which ran from late October 1978 to late May 1979. Factors studied include: channel characteristics, experiment errors due to instrument and spacecraft effects, predicted and measured precision, predicted accuracy, and comparisons with correlative measurements made in a series of balloon underflights. Features such as profile shape and slope of the mixing ratio altitude distribution are in good agreement. The LIMS data also fall within the range of previous mixing ratio measurements and are consistent with model estimates. The calculated on-orbit precision is about 0.3 ppbv and the estimated accuracy from simulations is about 2 ppbv over the 3-10-mbar range. Accuracy is less at higher and lower pressure levels. These results provide the first day-night set of nitrogen dioxide measurements from space.

  10. Results from the LIMS experiment for the PMP-1 winter 1978/79. [Limb Infrared Monitor of the Stratosphere

    NASA Technical Reports Server (NTRS)

    Gille, J. C.; Bailey, P. L.; Lyjak, L. V.; Russell, J. M., III

    1982-01-01

    The results of a preliminary analysis of the data on the global distributions of temperature and trace constituents in the middle atmosphere (15-65 km) obtained with the limb IR monitor (LIMS) are presented. The 6-channel limb-scanning radiometer was sensitive to the 15 micron bands of CO2, as well as other wavelengths, and was launched on the Nimbus 7 satellite in 1978. The derived data were partially compared with Soviet rocketsonde data to track an evolution in a warming trend over Siberia. The morphology of the changes were attributed to stratospheric warming. The comparisons with rocketsonde data indicated that the LIMS data were suitable for the determination of planetary wave amplitudes and phases, heat and momentum transport, and Eliassen-Palm fluxes.

  11. Islamic Education Philosophy Development (Study Analysis on Ta'lim Al-Kitab Al-Zarnuji Muta'allim Works)

    ERIC Educational Resources Information Center

    Asrori, H. Achmad

    2016-01-01

    "Ta'lim Muta'allim" is one of the monumental works of Shaykh Tajuddin Nu'man ibn Ibrahim ibn al-Khalil al-Zarnuji, who lived in the 6th century H/13-14 M. The reason for writing this study ie: (1) it is very rich with the basic values of Islamic education, (2) the values are already widely practiced in the world of education, especially…

  12. Improvements in the profiles and distributions of nitric acid and nitrogen dioxide with the LIMS version 6 dataset

    NASA Astrophysics Data System (ADS)

    Remsberg, E.; Natarajan, M.; Marshall, T.; Gordley, L. L.; Thompson, R. E.; Lingenfelser, G.

    2010-02-01

    The quality of the Nimbus 7 Limb Infrared Monitor of the Stratosphere (LIMS) nitric acid (HNO3) and nitrogen dioxide (NO2) profiles and distributions of 1978/1979 is described after their processing with an updated, Version 6 (V6) algorithm and subsequent archival in 2002. Estimates of the precision and accuracy of both of those species are developed and provided herein. The character of the V6 HNO3 profiles is relatively unchanged from that of the earlier LIMS Version 5 (V5) profiles, except in the upper stratosphere where the interfering effects of CO2 are accounted for better with V6. The accuracy of the retrieved V6 NO2 is also significantly better in the middle and upper stratosphere, due to improvements in its spectral line parameters and in the reduced biases for the accompanying V6 temperature and water vapor profiles. As a result of these important updates, there is better agreement with theoretical calculations for profiles of the HNO3/NO2 ratio, day-to-night NO2 ratio, and with estimates of the production of NO2 in the mesosphere and its descent to the upper stratosphere during polar night. The improved precisions and more frequent retrievals of the profiles along the LIMS orbit tracks provide for better continuity and detail in map analyses of these two species on pressure surfaces. It is judged that the chemical effects of the oxides of nitrogen on ozone can be examined quantitatively throughout the stratosphere with the LIMS V6 data, and that the findings will be more compatible with those obtained from measurements of the same species from subsequent satellite sensors.

  13. Speciation of surface gold in pressure oxidized carbonaceous gold ores by TOF-SIMS and TOF-LIMS

    NASA Astrophysics Data System (ADS)

    Dimov, S. S.; Chryssoulis, S. L.; Sodhi, R. N.

    2003-01-01

    To the best of our knowledge, this is the first attempt ever to speciate gold preg-robbed by carbonaceous matter using a surface sensitive microbeam technique. This approach enables the direct determination of gold species sorbed on carbonaceous particulates thus providing a new tool in understanding the chemistry of gold sorption on carbon. The reasoning behind this effort was to study the detrimental effect chloride ions have on gold recovery by pressure oxidation of carbonaceous sulfide ores, a technology largely used by the mining industry. The characterization of the sorbed gold species involved three surface sensitive microbeam analytical techniques (TOF-SIMS, TOF-LIMS and XPS) providing confirmatory results for better accuracy. Optimum conditions for detection of gold compounds with minimum fragmentation by TOF-SIMS and TOF-LIMS mass spectrometers have been determined. A reference library of 16 major gold complexes with halogen, thiosulfate, cyanide and thiocyanate groups relevant to the gold recovery processes has been established. The most suitable of the microbeam techniques tested was found to be negative (-ve) ion TOF-LIMS, offering best sensitivity and a small analytical spot size.

  14. Centrosome movements in vivo correlate with specific neurite formation downstream of LIM homeodomain transcription factor activity.

    PubMed

    Andersen, Erica F; Halloran, Mary C

    2012-10-01

    Neurons must develop complex structure to form proper connections in the nervous system. The initiation of axons in defined locations on the cell body and their extension to synaptic targets are critical steps in neuronal morphogenesis, yet the mechanisms controlling axon formation in vivo are poorly understood. The centrosome has been implicated in multiple aspects of neuronal morphogenesis; however, its function in axon development is under debate. Conflicting results from studies of centrosome function in axonogenesis suggest that its role is context dependent and underscore the importance of studying centrosome function as neurons develop in their natural environment. Using live imaging of zebrafish Rohon-Beard (RB) sensory neurons in vivo, we discovered a spatiotemporal relationship between centrosome position and the formation of RB peripheral, but not central, axons. We tested centrosome function by laser ablation and found that centrosome disruption inhibited peripheral axon outgrowth. In addition, we show that centrosome position and motility are regulated by LIM homeodomain transcription factor activity, which is specifically required for the development of RB peripheral axons. Furthermore, we show a correlation between centrosome mislocalization and ectopic axon formation in bashful (laminin alpha 1) mutants. Thus, both intrinsic transcription factor activity and extracellular cues can influence centrosome position and axon formation in vivo. This study presents the first positive association between the centrosome and axon formation in vivo and suggests that the centrosome is important for differential neurite formation in neurons with complex axonal morphologies. PMID:22899847

  15. A manual for a Laboratory Information Management System (LIMS) for light stable isotopes

    USGS Publications Warehouse

    Coplen, Tyler B.

    1998-01-01

    The reliability and accuracy of isotopic data can be improved by utilizing database software to (i) store information about samples, (ii) store the results of mass spectrometric isotope-ratio analyses of samples, (iii) calculate analytical results using standardized algorithms stored in a database, (iv) normalize stable isotopic data to international scales using isotopic reference materials, and (v) generate multi-sheet paper templates for convenient sample loading of automated mass-spectrometer sample preparation manifolds. Such a database program is presented herein. Major benefits of this system include (i) an increase in laboratory efficiency, (ii) reduction in the use of paper, (iii) reduction in workload due to the elimination or reduction of retyping of data by laboratory personnel, and (iv) decreased errors in data reported to sample submitters. Such a database provides a complete record of when and how often laboratory reference materials have been analyzed and provides a record of what correction factors have been used through time. It provides an audit trail for stable isotope laboratories. Since the original publication of the manual for LIMS for Light Stable Isotopes, the isotopes 3 H, 3 He, and 14 C, and the chlorofluorocarbons (CFCs), CFC-11, CFC-12, and CFC-113, have been added to this program.

  16. A manual for a laboratory information management system (LIMS) for light stable isotopes

    USGS Publications Warehouse

    Coplen, Tyler B.

    1997-01-01

    The reliability and accuracy of isotopic data can be improved by utilizing database software to (i) store information about samples, (ii) store the results of mass spectrometric isotope-ratio analyses of samples, (iii) calculate analytical results using standardized algorithms stored in a database, (iv) normalize stable isotopic data to international scales using isotopic reference materials, and (v) generate multi-sheet paper templates for convenient sample loading of automated mass-spectrometer sample preparation manifolds. Such a database program is presented herein. Major benefits of this system include (i) an increase in laboratory efficiency, (ii) reduction in the use of paper, (iii) reduction in workload due to the elimination or reduction of retyping of data by laboratory personnel, and (iv) decreased errors in data reported to sample submitters. Such a database provides a complete record of when and how often laboratory reference materials have been analyzed and provides a record of what correction factors have been used through time. It provides an audit trail for stable isotope laboratories. Since the original publication of the manual for LIMS for Light Stable Isotopes, the isotopes 3 H, 3 He, and 14 C, and the chlorofluorocarbons (CFCs), CFC-11, CFC-12, and CFC-113, have been added to this program.

  17. LIM Kinase Inhibitor Pyr1 Reduces the Growth and Metastatic Load of Breast Cancers.

    PubMed

    Prunier, Chloé; Josserand, Véronique; Vollaire, Julien; Beerling, Evelyne; Petropoulos, Christos; Destaing, Olivier; Montemagno, Christopher; Hurbin, Amandine; Prudent, Renaud; de Koning, Leanne; Kapur, Reuben; Cohen, Pascale A; Albiges-Rizo, Corinne; Coll, Jean-Luc; van Rheenen, Jacco; Billaud, Marc; Lafanechère, Laurence

    2016-06-15

    LIM kinases (LIMK) are emerging targets for cancer therapy, and they function as network hubs to coordinate actin and microtubule dynamics. When LIMKs are inhibited, actin microfilaments are disorganized and microtubules are stabilized. Owing to their stabilizing effect on microtubules, LIMK inhibitors may provide a therapeutic strategy to treat taxane-resistant cancers. In this study, we investigated the effect of LIMK inhibition on breast tumor development and on paclitaxel-resistant tumors, using a novel selective LIMK inhibitor termed Pyr1. Treatment of breast cancer cells, including paclitaxel-resistant cells, blocked their invasion and proliferation in vitro and their growth in vivo in tumor xenograft assays. The tumor-invasive properties of Pyr1 were investigated in vivo by intravital microscopy of tumor xenografts. A striking change of cell morphology was observed with a rounded phenotype arising in a subpopulation of cells, while other cells remained elongated. Notably, although Pyr1 decreased the motility of elongated cells, it increased the motility of rounded cells in the tumor. Pyr1 administration prevented the growth of metastasis but not their spread. Overall, our results provided a preclinical proof of concept concerning how a small-molecule inhibitor of LIMK may offer a strategy to treat taxane-resistant breast tumors and metastases. Cancer Res; 76(12); 3541-52. ©2016 AACR. PMID:27216191

  18. Transcriptional profiling of GBM invasion genes identifies effective inhibitors of the LIM kinase-Cofilin pathway

    PubMed Central

    Golbourn, Brian; Bertrand, Kelsey C.; Luck, Amanda; Sabha, Nesrin; Smith, Christian A.; Byron, Sara; Zadeh, Gelareh; Croul, Sidney; Berens, Michael; Rutka, James T.

    2014-01-01

    Malignant gliomas are highly proliferative and invasive neoplasms where total surgical resection is often impossible and effective local radiation therapy difficult. Consequently, there is a need to develop a greater understanding of the molecular events driving invasion and to identify novel treatment targets. Using microarray analysis comparing normal brain samples and mesenchymal glioblastoma multiforme (GBM), we identified over 140 significant genes involved in cell migration and invasion. The cofilin (CFL) pathway, which disassembles actin filaments, was highly up-regulated compared to normal brain. Up-regulation of LIM domain kinase 1 and 2 (LIMK1/2), that phosphorylates and inactivates cofilin, was confirmed in an additional independent data set comparing normal brain to GBM. We identified and utilized two small molecule inhibitors BMS-5 and Cucurbitacin I directed against the cofilin regulating kinases, LIMK1 and LIMK2, to target this pathway. Significant decreases in cell viability were observed in glioma cells treated with BMS-5 and Cucurbitacin I, while no cytotoxic effects were seen in normal astrocytes that lack LIMK. BMS-5 and Cucurbitacin I promoted increased adhesion in GBM cells, and decreased migration and invasion. Collectively, these data suggest that use of LIMK inhibitors may provide a novel way to target the invasive machinery in GBM. PMID:25237832

  19. The LIM-homeodomain transcription factor Islet2a promotes angioblast migration.

    PubMed

    Lamont, Ryan E; Wu, Chang-Yi; Ryu, Jae-Ryeon; Vu, Wendy; Davari, Paniz; Sobering, Ryan E; Kennedy, Regan M; Munsie, Nicole M; Childs, Sarah J

    2016-06-15

    Angioblasts of the developing vascular system require many signaling inputs to initiate their migration, proliferation and differentiation into endothelial cells. What is less studied is which intrinsic cell factors interpret these extrinsic signals. Here, we show the Lim homeodomain transcription factor islet2a (isl2a) is expressed in the lateral posterior mesoderm prior to angioblast migration. isl2a deficient angioblasts show disorganized migration to the midline to form axial vessels and fail to spread around the tailbud of the embryo. Isl2a morphants have fewer vein cells and decreased vein marker expression. We demonstrate that isl2a is required cell autonomously in angioblasts to promote their incorporation into the vein, and is permissive for vein identity. Knockout of isl2a results in decreased migration and proliferation of angioblasts during intersegmental artery growth. Since Notch signaling controls both artery-vein identity and tip-stalk cell formation, we explored the interaction of isl2a and Notch. We find that isl2a expression is negatively regulated by Notch activity, and that isl2a positively regulates flt4, a VEGF-C receptor repressed by Notch during angiogenesis. Thus Isl2a may act as an intermediate between Notch signaling and genetic programs controlling angioblast number and migration, placing it as a novel transcriptional regulator of early angiogenesis. PMID:27126199

  20. LIM kinase inhibitors disrupt mitotic microtubule organization and impair tumor cell proliferation

    PubMed Central

    Mardilovich, Katerina; Baugh, Mark; Crighton, Diane; Kowalczyk, Dominika; Gabrielsen, Mads; Munro, June; Croft, Daniel R.; Lourenco, Filipe; James, Daniel; Kalna, Gabriella; McGarry, Lynn; Rath, Oliver; Shanks, Emma; Garnett, Mathew J.; McDermott, Ultan; Brookfield, Joanna; Charles, Mark; Hammonds, Tim; Olson, Michael F.

    2015-01-01

    The actin and microtubule cytoskeletons are critically important for cancer cell proliferation, and drugs that target microtubules are widely-used cancer therapies. However, their utility is compromised by toxicities due to dose and exposure. To overcome these issues, we characterized how inhibition of the actin and microtubule cytoskeleton regulatory LIM kinases could be used in drug combinations to increase efficacy. A previously-described LIMK inhibitor (LIMKi) induced dose-dependent microtubule alterations that resulted in significant mitotic defects, and increased the cytotoxic potency of microtubule polymerization inhibitors. By combining LIMKi with 366 compounds from the GSK Published Kinase Inhibitor Set, effective combinations were identified with kinase inhibitors including EGFR, p38 and Raf. These findings encouraged a drug discovery effort that led to development of CRT0105446 and CRT0105950, which potently block LIMK1 and LIMK2 activity in vitro, and inhibit cofilin phosphorylation and increase αTubulin acetylation in cells. CRT0105446 and CRT0105950 were screened against 656 cancer cell lines, and rhabdomyosarcoma, neuroblastoma and kidney cancer cells were identified as significantly sensitive to both LIMK inhibitors. These large-scale screens have identified effective LIMK inhibitor drug combinations and sensitive cancer types. In addition, the LIMK inhibitory compounds CRT0105446 and CRT0105950 will enable further development of LIMK-targeted cancer therapy. PMID:26540348

  1. Screensaver: an open source lab information management system (LIMS) for high throughput screening facilities

    PubMed Central

    2010-01-01

    Background Shared-usage high throughput screening (HTS) facilities are becoming more common in academe as large-scale small molecule and genome-scale RNAi screening strategies are adopted for basic research purposes. These shared facilities require a unique informatics infrastructure that must not only provide access to and analysis of screening data, but must also manage the administrative and technical challenges associated with conducting numerous, interleaved screening efforts run by multiple independent research groups. Results We have developed Screensaver, a free, open source, web-based lab information management system (LIMS), to address the informatics needs of our small molecule and RNAi screening facility. Screensaver supports the storage and comparison of screening data sets, as well as the management of information about screens, screeners, libraries, and laboratory work requests. To our knowledge, Screensaver is one of the first applications to support the storage and analysis of data from both genome-scale RNAi screening projects and small molecule screening projects. Conclusions The informatics and administrative needs of an HTS facility may be best managed by a single, integrated, web-accessible application such as Screensaver. Screensaver has proven useful in meeting the requirements of the ICCB-Longwood/NSRB Screening Facility at Harvard Medical School, and has provided similar benefits to other HTS facilities. PMID:20482787

  2. Palmitoylation of LIM Kinase-1 ensures spine-specific actin polymerization and morphological plasticity

    PubMed Central

    George, Joju; Soares, Cary; Montersino, Audrey; Beique, Jean-Claude; Thomas, Gareth M

    2015-01-01

    Precise regulation of the dendritic spine actin cytoskeleton is critical for neurodevelopment and neuronal plasticity, but how neurons spatially control actin dynamics is not well defined. Here, we identify direct palmitoylation of the actin regulator LIM kinase-1 (LIMK1) as a novel mechanism to control spine-specific actin dynamics. A conserved palmitoyl-motif is necessary and sufficient to target LIMK1 to spines and to anchor LIMK1 in spines. ShRNA knockdown/rescue experiments reveal that LIMK1 palmitoylation is essential for normal spine actin polymerization, for spine-specific structural plasticity and for long-term spine stability. Palmitoylation is critical for LIMK1 function because this modification not only controls LIMK1 targeting, but is also essential for LIMK1 activation by its membrane-localized upstream activator PAK. These novel roles for palmitoylation in the spatial control of actin dynamics and kinase signaling provide new insights into structural plasticity mechanisms and strengthen links between dendritic spine impairments and neuropathological conditions. DOI: http://dx.doi.org/10.7554/eLife.06327.001 PMID:25884247

  3. ROCK1 via LIM kinase regulates growth, maturation and actin based functions in mast cells

    PubMed Central

    Kapur, Reuben; Shi, Jianjian; Ghosh, Joydeep; Munugalavadla, Veerendra; Sims, Emily; Martin, Holly; Wei, Lei; Mali, Raghuveer Singh

    2016-01-01

    Understanding mast cell development is essential due to their critical role in regulating immunity and autoimmune diseases. Here, we show how Rho kinases (ROCK) regulate mast cell development and can function as therapeutic targets for treating allergic diseases. Rock1 deficiency results in delayed maturation of bone marrow derived mast cells (BMMCs) in response to IL-3 stimulation and reduced growth in response to stem cell factor (SCF) stimulation. Further, integrin-mediated adhesion and migration, and IgE-mediated degranulation are all impaired in Rock1-deficient BMMCs. To understand the mechanism behind altered mast cell development in Rock1−/− BMMCs, we analyzed the activation of ROCK and its downstream targets including LIM kinase (LIMK). We observed reduced activation of ROCK, LIMK, AKT and ERK1/2 in Rock1-deficient BMMCs in response to SCF stimulation. Further, loss of either Limk1 or Limk2 also demonstrated altered BMMC maturation and growth; combined deletion of both Limk1 and Limk2 resulted in further reduction in BMMC maturation and growth. In passive cutaneous anaphylaxis model, deficiency of Rock1 or treatment with ROCK inhibitor Fasudil protected mice against IgE-mediated challenge. Our results identify ROCK/LIMK pathway as a novel therapeutic target for treating allergic diseases involving mast cells. PMID:26943578

  4. Four and a Half LIM Domains 1b (Fhl1b) Is Essential for Regulating the Liver versus Pancreas Fate Decision and for β-Cell Regeneration.

    PubMed

    Xu, Jin; Cui, Jiaxi; Del Campo, Aranzazu; Shin, Chong Hyun

    2016-02-01

    The liver and pancreas originate from overlapping embryonic regions, and single-cell lineage tracing in zebrafish has shown that Bone morphogenetic protein 2b (Bmp2b) signaling is essential for determining the fate of bipotential hepatopancreatic progenitors towards the liver or pancreas. Despite its pivotal role, the gene regulatory networks functioning downstream of Bmp2b signaling in this process are poorly understood. We have identified four and a half LIM domains 1b (fhl1b), which is primarily expressed in the prospective liver anlage, as a novel target of Bmp2b signaling. fhl1b depletion compromised liver specification and enhanced induction of pancreatic cells from endodermal progenitors. Conversely, overexpression of fhl1b favored liver specification and inhibited induction of pancreatic cells. By single-cell lineage tracing, we showed that fhl1b depletion led lateral endodermal cells, destined to become liver cells, to become pancreatic cells. Reversely, when fhl1b was overexpressed, medially located endodermal cells, fated to differentiate into pancreatic and intestinal cells, contributed to the liver by directly or indirectly modulating the discrete levels of pdx1 expression in endodermal progenitors. Moreover, loss of fhl1b increased the regenerative capacity of β-cells by increasing pdx1 and neurod expression in the hepatopancreatic ductal system. Altogether, these data reveal novel and critical functions of Fhl1b in the hepatic versus pancreatic fate decision and in β-cell regeneration. PMID:26845333

  5. Four and a Half LIM Domains 1b (Fhl1b) Is Essential for Regulating the Liver versus Pancreas Fate Decision and for β-Cell Regeneration

    PubMed Central

    Xu, Jin; Cui, Jiaxi; Del Campo, Aranzazu; Shin, Chong Hyun

    2016-01-01

    The liver and pancreas originate from overlapping embryonic regions, and single-cell lineage tracing in zebrafish has shown that Bone morphogenetic protein 2b (Bmp2b) signaling is essential for determining the fate of bipotential hepatopancreatic progenitors towards the liver or pancreas. Despite its pivotal role, the gene regulatory networks functioning downstream of Bmp2b signaling in this process are poorly understood. We have identified four and a half LIM domains 1b (fhl1b), which is primarily expressed in the prospective liver anlage, as a novel target of Bmp2b signaling. fhl1b depletion compromised liver specification and enhanced induction of pancreatic cells from endodermal progenitors. Conversely, overexpression of fhl1b favored liver specification and inhibited induction of pancreatic cells. By single-cell lineage tracing, we showed that fhl1b depletion led lateral endodermal cells, destined to become liver cells, to become pancreatic cells. Reversely, when fhl1b was overexpressed, medially located endodermal cells, fated to differentiate into pancreatic and intestinal cells, contributed to the liver by directly or indirectly modulating the discrete levels of pdx1 expression in endodermal progenitors. Moreover, loss of fhl1b increased the regenerative capacity of β-cells by increasing pdx1 and neurod expression in the hepatopancreatic ductal system. Altogether, these data reveal novel and critical functions of Fhl1b in the hepatic versus pancreatic fate decision and in β-cell regeneration. PMID:26845333

  6. Expression of the Lhx genes apterous and lim1 in an errant polychaete: implications for bilaterian appendage evolution, neural development, and muscle diversification

    PubMed Central

    2013-01-01

    Background Arthropod and vertebrate appendages appear to have evolved via parallel co-option of a plesiomorphic gene regulatory network. Our previous work implies that annelids evolved unrelated appendage-forming mechanisms; we therefore found no support for homology of parapodia and arthropodia at the level of the whole appendage. We expand on that study here by asking whether expression of the LIM homeobox (Lhx) genes apterous and lim1 in the annelid Neanthes arenaceodentata supports homology of the dorsal branches as well as the proximodistal axes of parapodia and arthropodia. In addition, we explore whether the neural expression of apterous and lim1 in Neanthes supports the putative ancestral function of the Lhx gene family in regulating the differentiation and maintenance of neuronal subtypes. Results Both genes exhibit continuous expression in specific portions of the developing central nervous system, from hatching to at least the 13-chaetiger stage. For example, nerve cord expression occurs in segmentally iterated patterns consisting of diffuse sets of many lim1-positive cells and comparatively fewer, clustered pairs of apterous-positive cells. Additionally, continuous apterous expression is observed in presumed neurosecretory ganglia of the posterior brain, while lim1 is continuously expressed in stomatogastric ganglia of the anterior brain. apterous is also expressed in the jaw sacs, dorsal parapodial muscles, and a presumed pair of cephalic sensory organs, whereas lim1 is expressed in multiple pharyngeal ganglia, the segmental peripheral nervous system, neuropodial chaetal sac muscles, and parapodial ligules. Conclusions The early and persistent nervous system expression of apterous and lim1 in Neanthes juveniles supports conservation of Lhx function in bilaterian neural differentiation and maintenance. Our results also suggest that diversification of parapodial muscle precursors involves a complementary LIM code similar to those generating distinct

  7. FluxCTTX: A LIMS-based tool for management and analysis of cytotoxicity assays data

    PubMed Central

    2015-01-01

    Background Cytotoxicity assays have been used by researchers to screen for cytotoxicity in compound libraries. Researchers can either look for cytotoxic compounds or screen "hits" from initial high-throughput drug screens for unwanted cytotoxic effects before investing in their development as a pharmaceutical. These assays may be used as an alternative to animal experimentation and are becoming increasingly important in modern laboratories. However, the execution of these assays in large scale and different laboratories requires, among other things, the management of protocols, reagents, cell lines used as well as the data produced, which can be a challenge. The management of all this information is greatly improved by the utilization of computational tools to save time and guarantee quality. However, a tool that performs this task designed specifically for cytotoxicity assays is not yet available. Results In this work, we have used a workflow based LIMS -- the Flux system -- and the Together Workflow Editor as a framework to develop FluxCTTX, a tool for management of data from cytotoxicity assays performed at different laboratories. The main work is the development of a workflow, which represents all stages of the assay and has been developed and uploaded in Flux. This workflow models the activities of cytotoxicity assays performed as described in the OECD 129 Guidance Document. Conclusions FluxCTTX presents a solution for the management of the data produced by cytotoxicity assays performed at Interlaboratory comparisons. Its adoption will contribute to guarantee the quality of activities in the process of cytotoxicity tests and enforce the use of Good Laboratory Practices (GLP). Furthermore, the workflow developed is complete and can be adapted to other contexts and different tests for management of other types of data. PMID:26696462

  8. The Hawaiian Freshwater Algal Database (HfwADB): a laboratory LIMS and online biodiversity resource

    PubMed Central

    2012-01-01

    Background Biodiversity databases serve the important role of highlighting species-level diversity from defined geographical regions. Databases that are specially designed to accommodate the types of data gathered during regional surveys are valuable in allowing full data access and display to researchers not directly involved with the project, while serving as a Laboratory Information Management System (LIMS). The Hawaiian Freshwater Algal Database, or HfwADB, was modified from the Hawaiian Algal Database to showcase non-marine algal specimens collected from the Hawaiian Archipelago by accommodating the additional level of organization required for samples including multiple species. Description The Hawaiian Freshwater Algal Database is a comprehensive and searchable database containing photographs and micrographs of samples and collection sites, geo-referenced collecting information, taxonomic data and standardized DNA sequence data. All data for individual samples are linked through unique 10-digit accession numbers (“Isolate Accession”), the first five of which correspond to the collection site (“Environmental Accession”). Users can search online for sample information by accession number, various levels of taxonomy, habitat or collection site. HfwADB is hosted at the University of Hawaii, and was made publicly accessible in October 2011. At the present time the database houses data for over 2,825 samples of non-marine algae from 1,786 collection sites from the Hawaiian Archipelago. These samples include cyanobacteria, red and green algae and diatoms, as well as lesser representation from some other algal lineages. Conclusions HfwADB is a digital repository that acts as a Laboratory Information Management System for Hawaiian non-marine algal data. Users can interact with the repository through the web to view relevant habitat data (including geo-referenced collection locations) and download images of collection sites, specimen photographs and

  9. A role of the LIM-homeobox gene Lhx2 in the regulation of pituitary development

    PubMed Central

    Zhao, Yangu; Mailloux, Christina M.; Hermesz, Edit; Palkovits, Miklos; Westphal, Heiner

    2009-01-01

    The mammalian pituitary gland originates from two separate germinal tissues during embryonic development. The anterior and intermediate lobes of the pituitary are derived from Rathke's pouch, a pocket formed by an invagination of the oral ectoderm. The posterior lobe is derived from the infundibulum, which is formed by evagination of the neuroectoderm in the ventral diencephalon. Previous studies have shown that development of Rathke's pouch and the generation of distinct populations of hormone-producing endocrine cell lineages in the anterior/intermediate pituitary lobes is regulated by a number of transcription factors expressed in the pouch and by inductive signals from the ventral diencephalon/infundibulum. However, little is known about factors that regulate the development of the posterior pituitary lobe. In this study, we show that the LIM-homeobox gene Lhx2 is extensively expressed in the developing ventral diencephalon, including the infundibulum and the posterior lobe of the pituitary. Deletion of Lhx2 gene results in persistent cell proliferation, a complete failure of evagination of the neuroectoderm in the ventral diencephalon, and defects in the formation of the distinct morphological features of the infundibulum and the posterior pituitary lobe. Rathke's pouch is formed and endocrine cell lineages are generated in the anterior/intermediate pituitary lobes of the Lhx2 mutant. However, the shape and organization of the pouch and the anterior/intermediate pituitary lobes are severely altered due to the defects in development of the infundibulum and the posterior lobe. Our study thus reveals an essential role for Lhx2 in the regulation of posterior pituitary development and suggests a mechanism whereby development of the posterior lobe may affect the development of the anterior and intermediate lobes of the pituitary gland. PMID:19900438

  10. Gbx2 regulates thalamocortical axon guidance by modifying the LIM and Robo codes.

    PubMed

    Chatterjee, Mallika; Li, Kairong; Chen, Li; Maisano, Xu; Guo, Qiuxia; Gan, Lin; Li, James Y H

    2012-12-01

    Combinatorial expression of transcription factors forms transcriptional codes to confer neuronal identities and connectivity. However, how these intrinsic factors orchestrate the spatiotemporal expression of guidance molecules to dictate the responsiveness of axons to guidance cues is less understood. Thalamocortical axons (TCAs) represent the major input to the neocortex and modulate cognitive functions, consciousness and alertness. TCAs travel a long distance and make multiple target choices en route to the cortex. The homeodomain transcription factor Gbx2 is essential for TCA development, as loss of Gbx2 abolishes TCAs in mice. Using a novel TCA-specific reporter, we have discovered that thalamic axons are mostly misrouted to the ventral midbrain and dorsal midline of the diencephalon in Gbx2-deficient mice. Furthermore, conditionally deleting Gbx2 at different embryonic stages has revealed a sustained role of Gbx2 in regulating TCA navigation and targeting. Using explant culture and mosaic analyses, we demonstrate that Gbx2 controls the intrinsic responsiveness of TCAs to guidance cues. The guidance defects of Gbx2-deficient TCAs are associated with abnormal expression of guidance receptors Robo1 and Robo2. Finally, we demonstrate that Gbx2 controls Robo expression by regulating LIM-domain transcription factors through three different mechanisms: Gbx2 and Lhx2 compete for binding to the Lmo3 promoter and exert opposing effects on its transcription; repressing Lmo3 by Gbx2 is essential for Lhx2 activity to induce Robo2; and Gbx2 represses Lhx9 transcription, which in turn induces Robo1. Our findings illustrate the transcriptional control of differential expression of Robo1 and Robo2, which may play an important role in establishing the topography of TCAs. PMID:23136391

  11. The LIM Homeodomain Factor Lhx2 Is Required for Hypothalamic Tanycyte Specification and Differentiation

    PubMed Central

    Salvatierra, Juan; Lee, Daniel A.; Zibetti, Cristina; Duran-Moreno, Maria; Yoo, Sooyeon; Newman, Elizabeth A.; Wang, Hong; Bedont, Joseph L.; de Melo, Jimmy; Miranda-Angulo, Ana L.; Gil-Perotin, Sara; Garcia-Verdugo, Jose Manuel

    2014-01-01

    Hypothalamic tanycytes, a radial glial-like ependymal cell population that expresses numerous genes selectively enriched in embryonic hypothalamic progenitors and adult neural stem cells, have recently been observed to serve as a source of adult-born neurons in the mammalian brain. The genetic mechanisms that regulate the specification and maintenance of tanycyte identity are unknown, but are critical for understanding how these cells can act as adult neural progenitor cells. We observe that LIM (Lin-11, Isl-1, Mec-3)-homeodomain gene Lhx2 is selectively expressed in hypothalamic progenitor cells and tanycytes. To test the function of Lhx2 in tanycyte development, we used an intersectional genetic strategy to conditionally delete Lhx2 in posteroventral hypothalamic neuroepithelium, both embryonically and postnatally. We observed that tanycyte development was severely disrupted when Lhx2 function was ablated during embryonic development. Lhx2-deficient tanycytes lost expression of tanycyte-specific genes, such as Rax, while also displaying ectopic expression of genes specific to cuboid ependymal cells, such as Rarres2. Ultrastructural analysis revealed that mutant tanycytes exhibited a hybrid identity, retaining radial morphology while becoming multiciliated. In contrast, postnatal loss of function of Lhx2 resulted only in loss of expression of tanycyte-specific genes. Using chromatin immunoprecipitation, we further showed that Lhx2 directly regulated expression of Rax, an essential homeodomain factor for tanycyte development. This study identifies Lhx2 as a key intrinsic regulator of tanycyte differentiation, sustaining Rax-dependent activation of tanycyte-specific genes while also inhibiting expression of ependymal cell-specific genes. These findings provide key insights into the transcriptional regulatory network specifying this still poorly characterized cell type. PMID:25505333

  12. LIM-homeobox gene Lhx5 is required for normal development of Cajal-Retzius cells.

    PubMed

    Miquelajáuregui, Amaya; Varela-Echavarría, Alfredo; Ceci, M Laura; García-Moreno, Fernando; Ricaño, Itzel; Hoang, Kimmi; Frade-Pérez, Daniela; Portera-Cailliau, Carlos; Tamariz, Elisa; De Carlos, Juan A; Westphal, Heiner; Zhao, Yangu

    2010-08-01

    Cajal-Retzius (C-R) cells play important roles in the lamination of the mammalian cortex via reelin secretion. The genetic mechanisms underlying the development of these neurons have just begun to be unraveled. Here, we show that two closely related LIM-homeobox genes Lhx1 and Lhx5 are expressed in reelin+ cells in various regions in the mouse telencephalon at or adjacent to sites where the C-R cells are generated, including the cortical hem, the mantle region of the septal/retrobulbar area, and the ventral pallium. Whereas Lhx5 is expressed in all of these reelin-expressing domains, Lhx1 is preferentially expressed in the septal area and in a continuous domain spanning from lateral olfactory region to caudomedial territories. Genetic ablation of Lhx5 results in decreased reelin+ and p73+ cells in the neocortical anlage, in the cortical hem, and in the septal, olfactory, and caudomedial telencephalic regions. The overall reduction in number of C-R cells in Lhx5 mutants is accompanied by formation of ectopic reelin+ cell clusters at the caudal telencephalon. Based on differential expression of molecular markers and by fluorescent cell tracing in cultured embryos, we located the origin of reelin+ ectopic cell clusters at the caudomedial telencephalic region. We also confirmed the existence of a normal migration stream of reelin+ cells from the caudomedial area to telencephalic olfactory territories in wild-type embryos. These results reveal a complex role for Lhx5 in regulating the development and normal distribution of C-R cells in the developing forebrain. PMID:20685998

  13. Transcription of follicle-stimulating hormone subunit genes is modulated by porcine LIM homeobox transcription factors, LHX2 and LHX3

    PubMed Central

    YOSHIDA, Saishu; KATO, Takako; NISHIMURA, Naoto; KANNO, Naoko; CHEN, Mo; UEHARU, Hiroki; NISHIHARA, Hiroto; KATO, Yukio

    2016-01-01

    The LIM-homeobox transcription factors LHX2 and LHX3s (LHX3a and LHX3b) are thought to be involved in regulating the pituitary glycoprotein hormone subunit genes Cga and Fshβ. These two factors show considerable differences in their amino acid sequences for DNA binding and protein-protein interactions and in their vital function in pituitary development. Hence, we compared the DNA binding properties and transcriptional activities of Cga and Fshβ between LHX2 and LHX3s. A gel mobility shift assay for approximately 1.1 kb upstream of Cga and 2.0 kb upstream of Fshβ varied in binding profiles between LHX2 and LHX3s. DNase I footprinting revealed DNA binding sites in 8 regions of the Cga promoter for LHX2 and LHX3s with small differences in the binding range and strength. In the Fshβ promoter, 14 binding sites were identified for LHX2 and LHX3, respectively. There were alternative binding sites to either gene in addition to similar differences observed in the Cga promoter. The transcriptional activities of LHX2 and LHX3s according to a reporter assay showed cell-type dependent activity with repression in the pituitary gonadotrope lineage LβT2 cells and stimulation in Chinese hamster ovary lineage CHO cells. Reactivity of LHX2 and LHX3s was observed in all regions, and differences were observed in the 5'-upstream region of Fshβ. However, immunohistochemistry showed that LHX2 resides in a small number of gonadotropes in contrast to LHX3. Thus, LHX3 mainly controls Cga and Fshβ expression. PMID:26853788

  14. Transcription of follicle-stimulating hormone subunit genes is modulated by porcine LIM homeobox transcription factors, LHX2 and LHX3.

    PubMed

    Yoshida, Saishu; Kato, Takako; Nishimura, Naoto; Kanno, Naoko; Chen, Mo; Ueharu, Hiroki; Nishihara, Hiroto; Kato, Yukio

    2016-06-17

    The LIM-homeobox transcription factors LHX2 and LHX3s (LHX3a and LHX3b) are thought to be involved in regulating the pituitary glycoprotein hormone subunit genes Cga and Fshβ. These two factors show considerable differences in their amino acid sequences for DNA binding and protein-protein interactions and in their vital function in pituitary development. Hence, we compared the DNA binding properties and transcriptional activities of Cga and Fshβ between LHX2 and LHX3s. A gel mobility shift assay for approximately 1.1 kb upstream of Cga and 2.0 kb upstream of Fshβ varied in binding profiles between LHX2 and LHX3s. DNase I footprinting revealed DNA binding sites in 8 regions of the Cga promoter for LHX2 and LHX3s with small differences in the binding range and strength. In the Fshβ promoter, 14 binding sites were identified for LHX2 and LHX3, respectively. There were alternative binding sites to either gene in addition to similar differences observed in the Cga promoter. The transcriptional activities of LHX2 and LHX3s according to a reporter assay showed cell-type dependent activity with repression in the pituitary gonadotrope lineage LβT2 cells and stimulation in Chinese hamster ovary lineage CHO cells. Reactivity of LHX2 and LHX3s was observed in all regions, and differences were observed in the 5'-upstream region of Fshβ. However, immunohistochemistry showed that LHX2 resides in a small number of gonadotropes in contrast to LHX3. Thus, LHX3 mainly controls Cga and Fshβ expression. PMID:26853788

  15. Lim homeobox genes in the Ctenophore Mnemiopsis leidyi: the evolution of neural cell type specification

    PubMed Central

    2012-01-01

    Background Nervous systems are thought to be important to the evolutionary success and diversification of metazoans, yet little is known about the origin of simple nervous systems at the base of the animal tree. Recent data suggest that ctenophores, a group of macroscopic pelagic marine invertebrates, are the most ancient group of animals that possess a definitive nervous system consisting of a distributed nerve net and an apical statocyst. This study reports on details of the evolution of the neural cell type specifying transcription factor family of LIM homeobox containing genes (Lhx), which have highly conserved functions in neural specification in bilaterian animals. Results Using next generation sequencing, the first draft of the genome of the ctenophore Mnemiopsis leidyi has been generated. The Lhx genes in all animals are represented by seven subfamilies (Lhx1/5, Lhx3/4, Lmx, Islet, Lhx2/9, Lhx6/8, and LMO) of which four were found to be represented in the ctenophore lineage (Lhx1/5, Lhx3/4, Lmx, and Islet). Interestingly, the ctenophore Lhx gene complement is more similar to the sponge complement (sponges do not possess neurons) than to either the cnidarian-bilaterian or placozoan Lhx complements. Using whole mount in situ hybridization, the Lhx gene expression patterns were examined and found to be expressed around the blastopore and in cells that give rise to the apical organ and putative neural sensory cells. Conclusion This research gives us a first look at neural cell type specification in the ctenophore M. leidyi. Within M. leidyi, Lhx genes are expressed in overlapping domains within proposed neural cellular and sensory cell territories. These data suggest that Lhx genes likely played a conserved role in the patterning of sensory cells in the ancestor of sponges and ctenophores, and may provide a link to the expression of Lhx orthologs in sponge larval photoreceptive cells. Lhx genes were later co-opted into patterning more diversified complements of

  16. Effect of the HITRAN 92 spectral data on the retrieval of NO2 mixing ratios from Nimbus 7 LIMS

    NASA Technical Reports Server (NTRS)

    Remsburg, Ellis E.; Bhatt, Praful P.; Eckman, Richard S.; Gordley, Larry L.; Russell, James M., III; Siskind, David E.

    1994-01-01

    To ensure spectral consistency when comparing Nimbus 7 Limb Infrared Monitor of the Stratosphere Experiment (LIMS) NO2 distributions with those from Atmosphere Trace Molecule Spectroscopy (ATMOS) and Upper Atmosphere Research Satellite Experiments (UARS), 1 day (May 5, 1979) of LIMS measurements were reprocessed using the NO2 line list on the HITRAN 92 tape compiled by the Airforce Geophysics Laboratory (AFGL). The revised NO2 mixing ratios are smaller by up to 20%. The decrease is not constant with height, latitude, or time of day but depends on the absolute amount of NO2 in the profile, as a result of a change in the degree of saturation for the strong NO2 spectral lines. The revised NO2 agrees better with correlative measurements and with NO2 distributions from the Stratospheric Aerosol and Gas Experiment (SAGE) and Halogen Occultation Experiment (HALOE) satellite experiments but not with those from ATMOS 85. Profiles of the day/night ratio of revised NO2 are now larger near 5 hPa. There is also some improvement between observed and modeled ozone in the upper stratosphere, when the revised nighttime NO2 profile is used as the estimate of NO(y) for the model calculations.

  17. Spatial-frequency response of the Limb Infrared Monitor of the Stratosphere /LIMS/ radiometer. [on Nimbus 7 satellite

    NASA Technical Reports Server (NTRS)

    Wilson, R. G.; Jalink, A., Jr.; Kahlbaum, W. M., Jr.

    1979-01-01

    The Limb Infrared Monitor of the Stratosphere (LIMS), an experiment on the Nimbus-7 satellite, is designed to scan the earth's limb vertically and measure spectral emission profiles of trace atmospheric gases believed to be important in processes controlling the stratospheric ozone distribution. Experiment objectives are reviewed and several analyses and measurements are described which were performed to determine the adequacy of the system for satisfying these objectives. Adequate spatial-frequency response for all the spectral channels is required to provide, through inversion of the measured limb-radiance profiles, important information about the temperature, structure, and composition of the atmosphere. From the LIMS design-model data, the MTF was calculated for the optical system, the detector field mask, the electronics, and the overall system, for each channel. The signal output performance of the instrument was predicted from the system MTF data model input radiance data for each channel. MTF measurements made on the flight sensor confirmed the analytical results. The predictions indicate that the instrument can satisfy the basic measurement objectives of the experiment.

  18. The mean ozone profile and its temperature sensitivity in the upper stratosphere and lower mesosphere - An analysis of LIMS observations

    NASA Technical Reports Server (NTRS)

    Froidevaux, Lucien; Allen, Mark; Berman, Stanley; Daughton, Alan

    1989-01-01

    Multiple simultaneous LIMS (Limb Infrared Monitor of the Stratosphere) observations of both O3 and temperature T in the upper stratosphere and lower mesosphere are analyzed to derive O3-T correlations for comparisons with model calculations. The zonally averaged O3 profile is compared with results from a simplified photochemical model that assumes O3 to be in a photochemical steady state. The model profile is systematically lower than the observed profile. Key parameters are identified in which changes can systematically increase the O3 profile in both the stratosphere and the mesosphere. The LIMS-derived values for the sensitivity of O3 to changes in T are compared with equilibrium model calculations which include the T-driven opacity feedback effect on photodissociation rate constants. The theoretical O3 response to temperature perturbations is investigated. It is shown how the O3-T sensitivity coefficient is affected by zonal and vertical advection terms as well as by the photochemical coupling between O3 and T.

  19. A guide for the laboratory information management system (LIMS) for light stable isotopes--Versions 7 and 8

    USGS Publications Warehouse

    Coplen, Tyler B.

    2000-01-01

    The reliability and accuracy of isotopic data can be improved by utilizing database software to (i) store information about samples, (ii) store the results of mass spectrometric isotope-ratio analyses of samples, (iii) calculate analytical results using standardized algorithms stored in a database, (iv) normalize stable isotopic data to international scales using isotopic reference materials, and (v) generate multi-sheet paper templates for convenient sample loading of automated mass-spectrometer sample preparation manifolds. Such a database program, the Laboratory Information Management System (LIMS) for Light Stable Isotopes, is presented herein. Major benefits of this system include (i) a dramatic improvement in quality assurance, (ii) an increase in laboratory efficiency, (iii) a reduction in workload due to the elimination or reduction of retyping of data by laboratory personnel, and (iv) a decrease in errors in data reported to sample submitters. Such a database provides a complete record of when and how often laboratory reference materials have been analyzed and provides a record of what correction factors have been used through time. It provides an audit trail for laboratories. LIMS for Light Stable Isotopes is available for both Microsoft Office 97 Professional and Microsoft Office 2000 Professional as versions 7 and 8, respectively. Both source code (mdb file) and precompiled executable files (mde) are available. Numerous improvements have been made for continuous flow isotopic analysis in this version (specifically 7.13 for Microsoft Access 97 and 8.13 for Microsoft Access 2000). It is much easier to import isotopic results from Finnigan ISODAT worksheets, even worksheets on which corrections for amount of sample (linearity corrections) have been added. The capability to determine blank corrections using isotope mass balance from analyses of elemental analyzer samples has been added. It is now possible to calculate and apply drift corrections to isotopic

  20. Molecular cloning of LIM homeodomain transcription factor Lhx2 as a transcription factor of porcine follicle-stimulating hormone beta subunit (FSHβ) gene.

    PubMed

    Kato, Takako; Ishikawa, Akio; Yoshida, Saishu; Sano, Yoshiya; Kitahara, Kousuke; Nakayama, Michie; Susa, Takao; Kato, Yukio

    2012-01-01

    We cloned the LIM-homeodomain protein LHX2 as a transcription factor for the porcine follicle-stimulating hormone β subunit gene (Fshβ) by the Yeast One-Hybrid Cloning System using the upstream region of -852/-746 bases (b) from the transcription start site, called Fd2, as a bait sequence. The reporter assay in LβT2 and CHO cells revealed the presence of an LHX2-responsive region other than Fd2. A potential LHX2 binding sequence was confirmed as AATTAAT containing a consensus homeodomain binding core sequence AATT by Systematic Evolution of Ligands by Exponential Enrichment analysis. DNase I footprinting demonstrated three AATTAAT sequences located at regions -835/-829, -818/-812 and -806/-800 b in the Fd2 region and 12 binding sites in the distal and proximal regions mostly containing an AATT-core sequence. RT-PCR analysis of Lhx2 expression during porcine fetal and postnatal pituitary development showed a gradual increase from fetal day (f) 40 to postnatal day (p) 8 followed by a slight decrease to p230, suggesting that LHX2 may play its role largely in the late fetal and postnatal periods. The analyses of Lhx2 expression in pituitary tumor-derived cell lines showed their expressions in cell lines including αT31, LβT2 and others. Since LHX2 was previously identified as a transcription factor for Cga and the in vitro experiments in the present study suggested that LHX2 regulated the expression of Fshβ, it is possible that LHX2 controls the synthesis of FSH at the transcription level. PMID:22134063

  1. LIM homeobox transcription factor Lhx2 inhibits skeletal muscle differentiation in part via transcriptional activation of Msx1 and Msx2.

    PubMed

    Kodaka, Yusaku; Tanaka, Kiyoko; Kitajima, Kenji; Tanegashima, Kosuke; Matsuda, Ryoichi; Hara, Takahiko

    2015-02-15

    LIM homeobox transcription factor Lhx2 is known to be an important regulator of neuronal development, homeostasis of hair follicle stem cells, and self-renewal of hematopoietic stem cells; however, its function in skeletal muscle development is poorly understood. In this study, we found that overexpression of Lhx2 completely inhibits the myotube-forming capacity of C2C12 cells and primary myoblasts. The muscle dedifferentiation factors Msx1 and Msx2 were strongly induced by the Lhx2 overexpression. Short interfering RNA-mediated knockdown of Lhx2 in the developing limb buds of mouse embryos resulted in a reduction in Msx1 and Msx2 mRNA levels, suggesting that they are downstream target genes of Lhx2. We found two Lhx2 consensus-binding sites in the -2097 to -1189 genomic region of Msx1 and two additional sites in the -536 to +73 genomic region of Msx2. These sequences were shown by luciferase reporter assay to be essential for Lhx2-mediated transcriptional activation. Moreover, electrophoretic mobility shift assays and chromatin immunoprecipitation assays showed that Lhx2 is present in chromatin DNA complexes bound to the enhancer regions of the Msx1 and Msx2 genes. These data demonstrate that Msx1 and Msx2 are direct transcriptional targets of Lhx2. In addition, overexpression of Lhx2 significantly enhanced the mRNA levels of bone morphogenetic protein 4 and transforming growth factor beta family genes. We propose that Lhx2 is involved in the early stage of skeletal muscle development by inducing multiple differentiation inhibitory factors. PMID:25460335

  2. Recording information on protein complexes in an information management system

    PubMed Central

    Savitsky, Marc; Diprose, Jonathan M.; Morris, Chris; Griffiths, Susanne L.; Daniel, Edward; Lin, Bill; Daenke, Susan; Bishop, Benjamin; Siebold, Christian; Wilson, Keith S.; Blake, Richard; Stuart, David I.; Esnouf, Robert M.

    2011-01-01

    The Protein Information Management System (PiMS) is a laboratory information management system (LIMS) designed for use with the production of proteins in a research environment. The software is distributed under the CCP4 licence, and so is available free of charge to academic laboratories. Like most LIMS, the underlying PiMS data model originally had no support for protein–protein complexes. To support the SPINE2-Complexes project the developers have extended PiMS to meet these requirements. The modifications to PiMS, described here, include data model changes, additional protocols, some user interface changes and functionality to detect when an experiment may have formed a complex. Example data are shown for the production of a crystal of a protein complex. Integration with SPINE2-Complexes Target Tracker application is also described. PMID:21605682

  3. Protein

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Proteins are the major structural and functional components of all cells in the body. They are macromolecules that comprise 1 or more chains of amino acids that vary in their sequence and length and are folded into specific 3-dimensional structures. The sizes and conformations of proteins, therefor...

  4. Proteins.

    ERIC Educational Resources Information Center

    Doolittle, Russell F.

    1985-01-01

    Examines proteins which give rise to structure and, by virtue of selective binding to other molecules, make genes. Binding sites, amino acids, protein evolution, and molecular paleontology are discussed. Work with encoding segments of deoxyribonucleic acid (exons) and noncoding stretches (introns) provides new information for hypotheses. (DH)

  5. The global distribution and variability of stratospheric constituents measured by LIMS. [Limb Infrared Monitor of the Stratosphere

    NASA Technical Reports Server (NTRS)

    Russell, J. M., III

    1984-01-01

    The purpose of the Nimbus 7 LIMS experiment was to sound the composition and structure of the upper atmosphere and provide data for study of photochemistry, radiation, and dynamics processes. Vertical profiles were measured of temperature and ozone (O3) over the 10-km to 65-km range and water vapor, nitrogen dioxide, and nitric acid over the 10-km to 50-km range. Latitude coverage extended from 64 deg S to 84 deg N. Several general features of the atmosphere have emerged from data analyzes thus far. Nitrogen dioxide exhibits rapid latitudinal variations in winter and shows hemispheric asymmetry with generally higher vertical column amount in the summer hemisphere. HNO3 data show that this gas is highly variable with altitude, latitude, and season. Smallest mixing ratios occur in the tropics, and the largest values occur in the high latitude winter hemisphere. The results show that O3, NO2, and HNO3 are strongly affected during a stratospheric warming.

  6. In vivo imaging of cell behaviors and F-actin reveals LIM-HD transcription factor regulation of peripheral versus central sensory axon development

    PubMed Central

    2011-01-01

    Background Development of specific neuronal morphology requires precise control over cell motility processes, including axon formation, outgrowth and branching. Dynamic remodeling of the filamentous actin (F-actin) cytoskeleton is critical for these processes; however, little is known about the mechanisms controlling motile axon behaviors and F-actin dynamics in vivo. Neuronal structure is specified in part by intrinsic transcription factor activity, yet the molecular and cellular steps between transcription and axon behavior are not well understood. Zebrafish Rohon-Beard (RB) sensory neurons have a unique morphology, with central axons that extend in the spinal cord and a peripheral axon that innervates the skin. LIM homeodomain (LIM-HD) transcription factor activity is required for formation of peripheral RB axons. To understand how neuronal morphogenesis is controlled in vivo and how LIM-HD transcription factor activity differentially regulates peripheral versus central axons, we used live imaging of axon behavior and F-actin distribution in vivo. Results We used an F-actin biosensor containing the actin-binding domain of utrophin to characterize actin rearrangements during specific developmental processes in vivo, including axon initiation, consolidation and branching. We found that peripheral axons initiate from a specific cellular compartment and that F-actin accumulation and protrusive activity precede peripheral axon initiation. Moreover, disruption of LIM-HD transcriptional activity has different effects on the motility of peripheral versus central axons; it inhibits peripheral axon initiation, growth and branching, while increasing the growth rate of central axons. Our imaging revealed that LIM-HD transcription factor activity is not required for F-actin based protrusive activity or F-actin accumulation during peripheral axon initiation, but can affect positioning of F-actin accumulation and axon formation. Conclusion Our ability to image the dynamics of

  7. LIM-homeodomain transcription factor Awh is a key component activating all three fibroin genes, fibH, fibL and fhx, in the silk gland of the silkworm, Bombyx mori.

    PubMed

    Kimoto, Mai; Tsubota, Takuya; Uchino, Keiro; Sezutsu, Hideki; Takiya, Shigeharu

    2015-01-01

    In the silkworm Bombyx mori, three fibroin genes, fibroin-heavy-chain (fibH), fibroin-light-chain (fibL) and fibrohexamerin (fhx), are coexpressed only in the posterior silk gland (PSG) cells, while the sericin genes encoding silk glue proteins are expressed in the middle silk gland (MSG) cells. Silk gland factor-2 (SGF-2) is a PSG-specific activator complex of fibH, composed of a LIM-homeodomain protein, Awh, and its cofactors, Ldb and Lcaf. We investigated whether SGF-2 can activate other fibroin genes using transgenic silkworms. The genes for Ldb and Lcaf were expressed ubiquitously in various tissues, while the gene for Awh was expressed strictly specific in PSG of the wild type silkworms. Misexpression of Awh in transgenic silkworms induced ectopic expression of fibL and fhx as well as fibH in MSG. Coincidently with the induction of fibL and fhx by Awh, binding of SGF-2 to the promoter of fibL and fhx was detected in vitro, and SGF-2 binds directly to the fhx core promoter. Ectopic expression of the fibroin genes was observed at high levels in the middle part of MSG. Moreover, fibL and fhx were induced in the anterior silk gland (ASG) of the transgenic silkworms, but fibH was not. These results indicate that Awh is a key activator of all three fibroin genes, and the activity is probably regulated in conjunction with additional factors. PMID:25449130

  8. Complete genome sequence of Cellulophaga lytica type strain (LIM-21T)

    SciTech Connect

    Pati, Amrita; Abt, Birte; Teshima, Hazuki; Nolan, Matt; Lapidus, Alla L.; Lucas, Susan; Hammon, Nancy; Deshpande, Shweta; Cheng, Jan-Fang; Tapia, Roxanne; Han, Cliff; Goodwin, Lynne A.; Pitluck, Sam; Liolios, Konstantinos; Pagani, Ioanna; Mavromatis, K; Ovchinnikova, Galina; Chen, Amy; Palaniappan, Krishna; Land, Miriam L; Hauser, Loren John; Jeffries, Cynthia; Detter, J. Chris; Brambilla, Evelyne-Marie; Kannan, K. Palani; Rohde, Manfred; Spring, Stefan; Goker, Markus; Woyke, Tanja; Bristow, James; Eisen, Jonathan; Markowitz, Victor; Hugenholtz, Philip; Kyrpides, Nikos C; Klenk, Hans-Peter; Ivanova, N

    2011-01-01

    Cellulophaga lytica (Lewin 1969) Johansen et al. 1999 is the type species of the genus Cellulophaga, which belongs to the family Flavobacteriaceae within the phylum 'Bacteroidetes' and was isolated from marine beach mud in Limon, Costa Rica. The species is of biotechnological interest because its members produce a wide range of extracellular enzymes capable of degrading proteins and polysaccharides. After the genome sequence of Cellulophaga algicola this is the second completed genome sequence of a member of the genus Cellulophaga. The 3,765,936 bp long genome with its 3,303 protein-coding and 55 RNA genes consists of one circular chromosome and is a part of the Genomic Encyclopedia of Bacteria and Archaea project.

  9. Satellite observations of upper atmosphere O3 and HNO3 from the Limb Infrared Monitor of the Stratosphere /LIMS/ experiment on Nimbus 7

    NASA Technical Reports Server (NTRS)

    Russell, J. M., III; Remsberg, E. E.; Gille, J. C.; Gordley, L. L.

    1981-01-01

    The Limb Infrared Monitor of the Stratosphere (LIMS) experiment, launched on-board the Nimbus satellite, was designed to measure both day and night limb radiance which is used to infer upper atmosphere temperature profiles and chemical compound concentrations important in the study of ozone chemistry. The LIMS instrument is a six channel, limb scanning, thermal radiometer which uses HgCdTe detectors, cooled to approximately 64 K. Measurements include ozone, water vapor, nitrogen dioxide, and nitric acid. The ozone and nitric acid channels are centered at approximately 9.6 microns and 11.3 microns, respectively. Results from the channels are described with an emphasis on data validation. The O3 and HNO3 channels have demonstrated high measurement precision.

  10. Conformational flexibility of the oncogenic protein LMO2 primes the formation of the multi-protein transcription complex

    PubMed Central

    Sewell, H.; Tanaka, T.; Omari, K. El; Mancini, E. J.; Cruz, A.; Fernandez-Fuentes, N.; Chambers, J.; Rabbitts, T. H.

    2014-01-01

    LMO2 was discovered via chromosomal translocations in T-cell leukaemia and shown normally to be essential for haematopoiesis. LMO2 is made up of two LIM only domains (thus it is a LIM-only protein) and forms a bridge in a multi-protein complex. We have studied the mechanism of formation of this complex using a single domain antibody fragment that inhibits LMO2 by sequestering it in a non-functional form. The crystal structure of LMO2 with this antibody fragment has been solved revealing a conformational difference in the positioning and angle between the two LIM domains compared with its normal binding. This contortion occurs by bending at a central helical region of LMO2. This is a unique mechanism for inhibiting an intracellular protein function and the structural contusion implies a model in which newly synthesized, intrinsically disordered LMO2 binds to a partner protein nucleating further interactions and suggests approaches for therapeutic targeting of LMO2. PMID:24407558

  11. Surface phase diagram and stability of (001) and (111) LiM n2O4 spinel oxides

    NASA Astrophysics Data System (ADS)

    Kim, Soo; Aykol, Muratahan; Wolverton, C.

    2015-09-01

    The (001) and (111) surface structures of the LiM n2O4 (LMO) spinel are examined using density functional theory (DFT) calculations within the generalized gradient approximation (GGA )+U approach. In order to clarify discrepancies in the literature for previous DFT calculations of these surfaces, we first carefully study the effects of surface termination/reconstruction, slab construction, and relaxation schemes, as well as magnetic ordering and U values for Mn on the calculated surface energies of LMO. We explain these discrepancies and show that the relaxation scheme and surface reconstruction play the key role in determining the relative stability of (001) and (111) surfaces. We have further analyzed the thermodynamic stability of LMO surfaces as a function of oxygen and lithium chemical potentials. We have found that the ratio of (001) to (111) surface energies is ˜1.09 up to the oxygen chemical potential corresponding to ˜800 K . This ratio favors the formation of truncated-octahedron shaped LMO particles dominated by Li-terminated reconstructed (111) facets with no surface Mn, which can help suppress Mn dissolution. Higher temperatures or closed systems (with no exchange of material with surroundings) favor the formation of (001) dominated particles. The observation of a wide spectrum of polyhedral shapes between (001)- and (111)-dominated LMO particles in experiments can be explained by the narrow range of surface energies and their sensitivity to synthesis conditions.

  12. Interactions between Arctic sea ice drift, concentration and thickness modeled by NEMO-LIM3 at different resolutions

    NASA Astrophysics Data System (ADS)

    Docquier, David; Massonnet, François; Raulier, Jonathan; Lecomte, Olivier; Fichefet, Thierry

    2016-04-01

    Sea ice concentration and thickness have substantially decreased in the Arctic since the beginning of the satellite era. As a result, mechanical strength has decreased allowing more fracturing and leading to increased sea ice drift. However, recent studies have highlighted that the interplay between sea ice thermodynamics and dynamics is poorly represented in contemporary global climate model (GCM) simulations. Thus, the considerable inter-model spread in terms of future sea ice extent projections could be reduced by better understanding the interactions between drift, concentration and thickness. This study focuses on the results coming from the global coupled ocean-sea ice model NEMO-LIM3 between 1979 and 2012. Three different simulations are forced by the Drakkar Forcing Set (DFS) 5.2 and run on the global tripolar ORCA grid at spatial resolutions of 0.25, 1° and 2°. The relation between modeled sea ice drift, concentration and thickness is further analyzed, compared to observations and discussed in the framework of the above-mentioned poor representation. It is proposed as a process-based metric for evaluating model performance. This study forms part of the EU Horizon 2020 PRIMAVERA project aiming at developing a new generation of advanced and well-evaluated high-resolution GCMs.

  13. Validation of temperature retrievals obtained by the Limb Infrared Monitor of the Stratosphere (LIMS) experiment on Nimbus 7

    NASA Technical Reports Server (NTRS)

    Gille, J. C.; Bailey, P. L.; Lyjak, L. V.; Beck, S. A.; Russell, J. M., III; Remsberg, E. E.; Gordley, L. L.; Lienesch, J. H.; Planet, W. G.; House, F. B.

    1984-01-01

    An outline is given of LIMS temperature determinations (as a function of pressure) from measurements in two channels covering portions of the 15-micrometer band of carbon dioxide. The known sources of error from the radiometer and data reduction are used to estimate the systematic and random errors expected of the results. Observational determinations of the complete end-to-end precision are obtained by computing the standard deviation of six sequential temperature retrievals in regions where the atmosphere is horizontally uniform. This yields values of 0.2 to 0.6 K, in reasonable agreement with the estimates. A correction for horizontal gradients in the atmosphere leads to a large reduction in the differences between the stratospheric temperatures determined on the ascending and descending portions of the orbit. The temperatures agree in the mean with radiosondes and rocketsondes to within 1-2 K in most regions below 1 mbar. Several interesting, previously unseen features included cold regions in the mid-latitude mesosphere and wavelike vertical variations in the tropics.

  14. Implications of the stratospheric water vapor distribution as determined from the Nimbus 7 LIMS experiment. [Limb Infrared Monitor of Stratosphere

    NASA Technical Reports Server (NTRS)

    Remsberg, E. E.; Russell, J. M., III; Gordley, L. L.; Gille, J. C.; Bailey, P. L.

    1984-01-01

    The LIMS experiment on Nimbus 7 has provided new results on the stratospheric water vapor distribution. The data show (1) a latitudinal gradient with mixing ratios that increase by a factor of 2 from equator to + or - 60 degrees at 50 mb, (2) most of the time there is a fairly uniform mixing ratio of 5 ppmv at high latitudes, but more variation exists during winter, (3) a well-developed hygropause at low to midlatitudes of the lower stratosphere, (4) a source region of water vapor in the upper stratospehere to lower mesosphere that is consistent with methane oxidation chemistry, at least within the uncertainties of the data, (5) an apparent zonal mean H2O distribution that is consistent with the circulation proposed by Brewer in 1949, and (6) a zonal mean distribution in the lower stratosphere that is consistent with the idea of quasi-isentropic transport by eddies in the meridional direction. Limits to the use of the data in the refinement of our understanding of the stratospheric water vapor budget are noted.

  15. High resolution seismic imaging of faults beneath Limón Bay, northern Panama Canal, Republic of Panama

    NASA Astrophysics Data System (ADS)

    Pratt, Thomas L.; Holmes, Mark; Schweig, Eugene S.; Gomberg, Joan; Cowan, Hugh A.

    2003-06-01

    High-resolution seismic reflection profiles from Limón Bay, Republic of Panama, were acquired as part of a seismic hazard investigation of the northern Panama Canal region. The seismic profiles image gently west and northwest dipping strata of upper Miocene Gatún Formation, unconformably overlain by a thin (<20 m) sequence of Holocene muds. Numerous faults, which have northeast trends where they can be correlated between seismic profiles, break the upper Miocene strata. Some of the faults have normal displacement, but on many faults, the amount and type of displacement cannot be determined. The age of displacement is constrained to be Late Miocene or younger, and regional geologic considerations suggest Pliocene movement. The faults may be part of a more extensive set of north- to northeast-trending faults and fractures in the canal region of central Panama. Low topography and the faults in the canal area may be the result of the modern regional stress field, bending of the Isthmus of Panama, shearing in eastern Panama, or minor deformation of the Panama Block above the Caribbean subduction zone. For seismic hazard analysis of the northern canal area, these faults led us to include a source zone of shallow faults proximal to northern canal facilities.

  16. The LIM-only transcription factor LMO2 determines tumorigenic and angiogenic traits in glioma stem cells.

    PubMed

    Kim, S-H; Kim, E-J; Hitomi, M; Oh, S-Y; Jin, X; Jeon, H-M; Beck, S; Jin, X; Kim, J-K; Park, C G; Chang, S-Y; Yin, J; Kim, T; Jeon, Y-J; Song, J; Lim, Y C; Lathia, J D; Nakano, I; Kim, H

    2015-09-01

    Glioblastomas (GBMs) maintain their cellular heterogeneity with glioma stem cells (GSCs) producing a variety of tumor cell types. Here we interrogated the oncogenic roles of Lim domain only 2 (LMO2) in GBM and GSCs in mice and human. High expression of LMO2 was found in human patient-derived GSCs compared with the differentiated progeny cells. LMO2 is required for GSC proliferation both in vitro and in vivo, as shRNA-mediated LMO2 silencing attenuated tumor growth derived from human GSCs. Further, LMO2 is sufficient to induce stem cell characteristics (stemness) in mouse premalignant astrocytes, as forced LMO2 expression facilitated in vitro and in vivo growth of astrocytes derived from Ink4a/Arf null mice and acquisition of GSC phenotypes. A subset of mouse and human GSCs converted into vascular endothelial-like tumor cells both in vitro and in vivo, which phenotype was attenuated by LMO2 silencing and promoted by LMO2 overexpression. Mechanistically, the action of LMO2 for induction of glioma stemness is mediated by transcriptional regulation of Jagged1 resulting in activation of the Notch pathway, whereas LMO2 directly occupies the promoter regions of the VE-cadherin gene for a gain of endothelial cellular phenotype. Subsequently, selective ablation of human GSC-derived VE-cadherin-expressing cells attenuated vascular formation in mouse intracranial tumors, thereby significantly prolonging mouse survival. Clinically, LMO2 expression was elevated in GBM tissues and inversely correlated with prognosis of GBM patients. Taken together, our findings describe novel dual roles of LMO2 to induce tumorigenesis and angiogenesis, and provide potential therapeutic targets in GBMs. PMID:25721045

  17. Mitochondrial Translocation of High Mobility Group Box 1 Facilitates LIM Kinase 2-Mediated Programmed Necrotic Neuronal Death.

    PubMed

    Hyun, Hye-Won; Ko, Ah-Reum; Kang, Tae-Cheon

    2016-01-01

    High mobility group box 1 (HMGB1) acts a signaling molecule regulating a wide range of inflammatory responses in extracellular space. HMGB1 also stabilizes nucleosomal structure and facilitates gene transcription. Under pathophysiological conditions, nuclear HMGB1 is immediately transported to the cytoplasm through chromosome region maintenance 1 (CRM1). Recently, we have reported that up-regulation of LIM kinase 2 (LIMK2) expression induces HMGB1 export from neuronal nuclei during status epilepticus (SE)-induced programmed neuronal necrosis in the rat hippocampus. Thus, we investigated whether HMGB1 involves LIMK2-mediated programmed neuronal necrosis, but such role is not reported. In the present study, SE was induced by pilocarpine in rats that were intracerebroventricularly infused with saline, control siRNA, LIMK2 siRNA or leptomycin B (LMB, a CRM1 inhibitor) prior to SE induction. Thereafter, we performed Fluoro-Jade B staining, western blots and immunohistochemical studies. LIMK2 knockdown effectively attenuated SE-induced neuronal death and HMGB1 import into mitochondria accompanied by inhibiting nuclear HMGB1 release and abnormal mitochondrial elongation. LMB alleviated SE-induced neuronal death and nuclear HMGB1 release. However, LMB did not prevent mitochondrial elongation induced by SE, but inhibited the HMGB1 import into mitochondria. The efficacy of LMB was less effective to attenuate SE-induced neuronal death than that of LIMK2 siRNA. These findings indicate that nuclear HMGB1 release and the subsequent mitochondrial import may facilitate and deteriorate programmed necrotic neuronal deaths. The present data suggest that the nuclear HMGB1 release via CRM1 may be a potential therapeutic target for the programmed necrotic neuronal death induced by SE. PMID:27147971

  18. Mitochondrial Translocation of High Mobility Group Box 1 Facilitates LIM Kinase 2-Mediated Programmed Necrotic Neuronal Death

    PubMed Central

    Hyun, Hye-Won; Ko, Ah-Reum; Kang, Tae-Cheon

    2016-01-01

    High mobility group box 1 (HMGB1) acts a signaling molecule regulating a wide range of inflammatory responses in extracellular space. HMGB1 also stabilizes nucleosomal structure and facilitates gene transcription. Under pathophysiological conditions, nuclear HMGB1 is immediately transported to the cytoplasm through chromosome region maintenance 1 (CRM1). Recently, we have reported that up-regulation of LIM kinase 2 (LIMK2) expression induces HMGB1 export from neuronal nuclei during status epilepticus (SE)-induced programmed neuronal necrosis in the rat hippocampus. Thus, we investigated whether HMGB1 involves LIMK2-mediated programmed neuronal necrosis, but such role is not reported. In the present study, SE was induced by pilocarpine in rats that were intracerebroventricularly infused with saline, control siRNA, LIMK2 siRNA or leptomycin B (LMB, a CRM1 inhibitor) prior to SE induction. Thereafter, we performed Fluoro-Jade B staining, western blots and immunohistochemical studies. LIMK2 knockdown effectively attenuated SE-induced neuronal death and HMGB1 import into mitochondria accompanied by inhibiting nuclear HMGB1 release and abnormal mitochondrial elongation. LMB alleviated SE-induced neuronal death and nuclear HMGB1 release. However, LMB did not prevent mitochondrial elongation induced by SE, but inhibited the HMGB1 import into mitochondria. The efficacy of LMB was less effective to attenuate SE-induced neuronal death than that of LIMK2 siRNA. These findings indicate that nuclear HMGB1 release and the subsequent mitochondrial import may facilitate and deteriorate programmed necrotic neuronal deaths. The present data suggest that the nuclear HMGB1 release via CRM1 may be a potential therapeutic target for the programmed necrotic neuronal death induced by SE. PMID:27147971

  19. Effects of different agricultural systems on soil quality in Northern Limón province, Costa Rica.

    PubMed

    Cornwell, Emma

    2014-09-01

    Conversion of native rainforest ecosystems in Limón Province of Costa Rica to banana and pineapple monoculture has led to reductions in biodiversity and soil quality. Agroforestry management of cacao (Theobroma cacao) is an alternative system that may maintain the agricultural livelihood of the region while more closely mimicking native ecosystems. This study compared physical, biological and chemical soil quality indicators of a cacao plantation under organic agroforestry management with banana, pineapple, and pasture systems; a native forest nearby served as a control. For bulk density and earthworm analysis, 18 samples were collected between March and April 2012 from each ecosystem paired with 18 samples from the cacao. Cacao had a lower bulk density than banana and pineapple monocultures, but greater than the forest (p < 0.05). Cacao also hosted a greater number and mass of earthworms than banana and pineapple (p < 0.05), but similar to forest and pasture. For soil chemical characteristics, three composite samples were collected in March 2012 from each agroecosystem paired with three samples from the cacao plantation. Forest and pineapple ecosystems had the lowest pH, cation exchange capacity, and exchangeable nutrient cations, while cacao had the greatest (p < 0.05). Total nutrient levels of P and N were slightly greater in banana, pineapple and pasture than in cacao; probably related to addition of chemical fertilizer and manure from cattle grazing. Forest and cacao also had greater %C, than other ecosystems, which is directly related to soil organic matter content (p < 0.0001). Overall, cacao had more favorable physical, biological and chemical soil characteristics than banana and pineapple monocultures, while trends were less conclusive compared to the pastureland. While organic cacao was inferior to native forest in some soil characteristics such as bulk density and organic carbon, its soil quality did best mimic that of the native forest. This supports

  20. Demonstration in vivo of the role of Arabidopsis PLIM2 actin-binding proteins during pollination.

    PubMed

    Sudo, Keisuke; Park, Jong-In; Sakazono, Satomi; Masuko-Suzuki, Hiromi; Osaka, Masaaki; Kawagishi, Mizuho; Fujita, Kotomi; Maruoka, Mayumi; Nanjo, Hikaru; Suzuki, Go; Suwabe, Keita; Watanabe, Masao

    2013-01-01

    In plant reproduction, pollination is the initial key process in bringing together the male and female gametophytes. When a pollen grain lands on the surface of the stigma, information is exchanged between the pollen and stigmatic cell to determine whether the pollen grain will be accepted or rejected. If it is accepted, the stigmatic papilla cell supplies water and other resources to the pollen for germination and pollen tube elongation. Cellular processes involving actin are essential for pollen germination and tube growth, and actin-binding proteins regulate these processes by interacting with actin filaments to assemble cytoskeletal structures and actin networks. LIM proteins, which belong to a subfamily of cysteine-rich proteins, are a family of actin-binding proteins in plants, and are considered to be important for formation of the actin cytoskeleton and maintenance of its dynamics. Although the physiological and biochemical characteristics of LIMs have been elucidated in vitro in a variety of cell types, their exact role in pollen germination and pollen tube growth during pollination remained unclear. In this manuscript, we focus on the pollen-specific LIM proteins, AtPLIM2a and AtPLIM2c, and define their biological function during pollination in Arabidopsis thaliana. The atplim2a/atplim2c double knockdown RNAi plants showed a reduced pollen germination, approximately one-fifth of wild type, and slower pollen tube growth in the pistil, that is 80.4 μm/hr compared to 140.8 μm/hr in wild type. These defects led to an occasional unfertilized ovule at the bottom of the silique in RNAi plants. Our data provide direct evidence of the biological function of LIM proteins during pollination as actin-binding proteins, modulating cytoskeletal structures and actin networks, and their consequent importance in seed production. PMID:24694391

  1. LIM-kinase 2 induces formation of stress fibres, focal adhesions and membrane blebs, dependent on its activation by Rho-associated kinase-catalysed phosphorylation at threonine-505.

    PubMed Central

    Amano, T; Tanabe, K; Eto, T; Narumiya, S; Mizuno, K

    2001-01-01

    LIM-kinase 1 and 2 (LIMK1 and LIMK2) phosphorylate cofilin and induce actin cytoskeletal reorganization. LIMK1 is activated by Rho-associated, coiled-coil-forming protein kinase (ROCK) and p21-activated kinase 1 (PAK1), but activation mechanisms and cellular functions of LIMK2 have remained to be determined. We report here that LIMK1 and LIMK2 phosphorylate both cofilin and actin-depolymerizing factor (ADF) specifically at Ser-3 and exhibit partially distinct substrate specificity when tested using site-directed cofilin mutants as substrates. We also show that LIMK2 is activated by ROCK by phosphorylation at Thr-505 within the activation loop. Wild-type LIMK2, but not its mutant (T505V) with replacement of Thr-505 by Val, was activated by ROCK in vitro and in vivo. LIMK2 mutants with replacement of Thr-505 by one or two Glu residues (T505E or T505EE) increased the kinase activity about 3.6-fold but were not further activated by ROCK. When expressed in HeLa cells, wild-type LIMK2, but not the T505V mutant, induced the formation of stress fibres, focal adhesions and membrane blebs. Furthermore, inhibitors of Rho and ROCK significantly suppressed LIMK2-induced stress fibres and membrane blebs. These results suggest that LIMK2 functions downstream of the Rho-ROCK signalling pathway and plays a role in reorganization of actin filaments and membrane structures, by phosphorylating cofilin/ADF proteins. PMID:11171090

  2. LIMS (Limb Infrared Monitor of the Stratosphere) observation of traveling planetary waves and potential vorticity advection in the stratosphere and mesosphere

    SciTech Connect

    Dunkerton, T.J. )

    1991-02-20

    Eastward and westward traveling waves were observed by the Nimbus 7 Limb Infrared Monitor of the Stratosphere (LIMS) during the northern winter 1978-1979. Eastward waves were prevalent in early winter and were involved in a minor Canadian warming in December 1978. A large westward traveling wave, as described by previous authors, was observed in January 1979 during a series of minor warmings. By comparing these two events, it is shown that in both cases the superposition of traveling and quasi-stationary waves led to constructive interference that was responsible for the warmings. However, there was significant asymmetry between eastward and westward traveling components. Eastward disturbances were confined mostly within the polar vortex, whereas quasi-stationary and westward traveling components propagated to the vortex periphery and beyond, into the tropics and mid-latitude mesosphere. This behavior is consistent with Rossby wave propagation and indicates that the location and magnitude of planetary wave breaking is sensitive to the frequency spectrum entering the middle atmosphere. However, this asymmetry is also a signature of the nonlinear critical layer as it projects onto the frequency spectrum. Both interpretations are shown to be valid during wave events observed by LIMS. A local Eulerian analysis of potential vorticity (PV) transport indicates that adiabatic, geostrophic advection by the resolvable scales of motion explains qualitatively (but not quantitatively) the observed potential vorticity tendencies in the LIMS northern hemisphere winter. In particular, calculated advection explains the eastward rotation of the main vortex, intrusion of low PV air into the polar cap, and formation of high PV filaments at the vortex periphery.

  3. Heterogenic Final Cell Cycle by Chicken Retinal Lim1 Horizontal Progenitor Cells Leads to Heteroploid Cells with a Remaining Replicated Genome

    PubMed Central

    Shirazi Fard, Shahrzad; Jarrin, Miguel; Boije, Henrik; Fillon, Valerie; All-Eriksson, Charlotta; Hallböök, Finn

    2013-01-01

    Retinal progenitor cells undergo apical mitoses during the process of interkinetic nuclear migration and newly generated post-mitotic neurons migrate to their prospective retinal layer. Whereas this is valid for most types of retinal neurons, chicken horizontal cells are generated by delayed non-apical mitoses from dedicated progenitors. The regulation of such final cell cycle is not well understood and we have studied how Lim1 expressing horizontal progenitor cells (HPCs) exit the cell cycle. We have used markers for S- and G2/M-phase in combination with markers for cell cycle regulators Rb1, cyclin B1, cdc25C and p27Kip1 to characterise the final cell cycle of HPCs. The results show that Lim1+ HPCs are heterogenic with regards to when and during what phase they leave the final cell cycle. Not all horizontal cells were generated by a non-apical (basal) mitosis; instead, the HPCs exhibited three different behaviours during the final cell cycle. Thirty-five percent of the Lim1+ horizontal cells was estimated to be generated by non-apical mitoses. The other horizontal cells were either generated by an interkinetic nuclear migration with an apical mitosis or by a cell cycle with an S-phase that was not followed by any mitosis. Such cells remain with replicated DNA and may be regarded as somatic heteroploids. The observed heterogeneity of the final cell cycle was also seen in the expression of Rb1, cyclin B1, cdc25C and p27Kip1. Phosphorylated Rb1-Ser608 was restricted to the Lim1+ cells that entered S-phase while cyclin B1 and cdc25C were exclusively expressed in HPCs having a basal mitosis. Only HPCs that leave the cell cycle after an apical mitosis expressed p27Kip1. We speculate that the cell cycle heterogeneity with formation of heteroploid cells may present a cellular context that contributes to the suggested propensity of these cells to generate cancer when the retinoblastoma gene is mutated. PMID:23527113

  4. Neotectonics and paleoseismology of the Limón and pedro miguel faults in Panamá: earthquake hazard to the Panamá canal

    USGS Publications Warehouse

    Rockwell, Thomas; Gath, Edon; Gonzalez, Tania; Madden, Chris; Verdugo, Danielle; Lippincott, Caitlin; Dawson, Tim; Owen, Lewis A.; Fuchs, Markus; Cadena, Ana; Williams, Pat; Weldon, Elise; Franceschi, Pastora

    2010-01-01

    We present new geologic, tectonic geomorphic, and geochronologic data on the slip rate, timing, and size of past surface ruptures for the right-lateral Lim??n and Pedro Miguel faults in central Panam??. These faults are part of a system of conjugate faults that accommodate the internal deformation of Panam?? resulting from the ongoing collision of Central and South America. There have been at least three surface ruptures on the Lim??n fault in the past 950-1400 years, with the most recent during the past 365 years. Displacement in this young event is at least 1.2 m (based on trenching) and may be 1.6-2 m (based on small channel offsets). Awell-preserved 4.2 m offset suggests that the penultimate event also sustained significant displacement. The Holocene slip rate has averaged about 6 mm=yr, based on a 30-m offset terrace riser incised into a 5-ka abandoned channel. The Pedro Miguel fault has sustained three surface ruptures in the past 1600 years, the most recent being the 2 May 1621 earthquake that partially destroyed Panam?? Viejo. At least 2.1 m of slip occurred in this event near the Canal, with geomorphic offsets suggesting 2.5-3 m. The historic Camino de Cruces is offset 2.8 m, indicating multimeter displacement over at least 20 km of fault length. Channel offsets of 100-400 m, together with a climate-induced incision model, suggest a Late Quaternary slip rate of about 5 mm=yr, which is consistent with the paleoseismic results. Comparison of the timing of surface ruptures between the Lim??n and Pedro Miguel faults suggests that large earthquakes may rupture both faults with 2-3 m of displacement for over 40 km, such as is likely in earthquakes in the M 7 range. Altogether, our observations indicate that the Lim??n and Pedro Miguel faults represent a significant seismic hazard to central Panam?? and, specifically, to the Canal and Panam?? City.

  5. An overview of wave-mean flow interactions during the winter of 1978-79 derived from LIMS observations. [Limb Infrared Monitor of Stratosphere

    NASA Technical Reports Server (NTRS)

    Gille, J. C.; Lyjak, L. V.

    1984-01-01

    Gradient winds, Eliassen-Palm (EP) fluxes and flux divergences, and the squared refractive index for planetary waves have been calculated from mapped data from the Limb Infrared Monitor of the Stratosphere (LIMS) experiment on Nimbus 7. The changes in the zonal mean atmospheric state, from early winter through 3 disturbances, is described. Convergence or divergence of the EP fluxes clearly produces changes in the zonal mean wind. The steering of the waves by the refractive index structure is not as clear on a daily basis.

  6. Laboratory information management system for membrane protein structure initiative--from gene to crystal.

    PubMed

    Troshin, Petr V; Morris, Chris; Prince, Stephen M; Papiz, Miroslav Z

    2008-12-01

    Membrane Protein Structure Initiative (MPSI) exploits laboratory competencies to work collaboratively and distribute work among the different sites. This is possible as protein structure determination requires a series of steps, starting with target selection, through cloning, expression, purification, crystallization and finally structure determination. Distributed sites create a unique set of challenges for integrating and passing on information on the progress of targets. This role is played by the Protein Information Management System (PIMS), which is a laboratory information management system (LIMS), serving as a hub for MPSI, allowing collaborative structural proteomics to be carried out in a distributed fashion. It holds key information on the progress of cloning, expression, purification and crystallization of proteins. PIMS is employed to track the status of protein targets and to manage constructs, primers, experiments, protocols, sample locations and their detailed histories: thus playing a key role in MPSI data exchange. It also serves as the centre of a federation of interoperable information resources such as local laboratory information systems and international archival resources, like PDB or NCBI. During the challenging task of PIMS integration, within the MPSI, we discovered a number of prerequisites for successful PIMS integration. In this article we share our experiences and provide invaluable insights into the process of LIMS adaptation. This information should be of interest to partners who are thinking about using LIMS as a data centre for their collaborative efforts. PMID:18991141

  7. LIMS (Limb Infrared Monitor of the Stratosphere) observation of traveling planetary waves and potential vorticity advection in the stratosphere and mesosphere

    NASA Technical Reports Server (NTRS)

    Dunkerton, Timothy J.

    1991-01-01

    Eastward and westward traveling waves were observed by the Nimbus 7 Limb Infrared Monitor of the Stratosphere (LIMS) during the northern winter 1978-1979. Eastward waves were prevalent in early winter and were involved in a minor Canadian warming in December 1978. A large westward traveling wave, as described by previous authors, was observed in January 1979 during a series of minor warmings. By comparing these two events, it is shown that in both cases the superposition of traveling and quasi-stationary waves led to constructive interference that was responsible for the warmings. However, there was significant asymmetry between eastward and westward traveling components. A local Eulerian analysis of potential vorticity (PV) transport indicates that adiabatic, geostrophic advection by the resolvable scales of motion explains qualitatively (but not quantitatively) the observed potential vorticity tendencies in the LIMS Northern Hemisphere winter. In particular, calculated advection explains the eastward rotation of the main vortex, intrusion of low PV air into the polar cap, and formation of high PV filaments at the vortex periphery.

  8. The focal adhesion protein PINCH-1 associates with EPLIN at integrin adhesion sites

    PubMed Central

    Karaköse, Esra; Geiger, Tamar; Flynn, Kevin; Lorenz-Baath, Katrin; Zent, Roy; Mann, Matthias; Fässler, Reinhard

    2015-01-01

    ABSTRACT PINCH-1 is a LIM-only domain protein that forms a ternary complex with integrin-linked kinase (ILK) and parvin (to form the IPP complex) downstream of integrins. Here, we demonstrate that PINCH-1 (also known as Lims1) gene ablation in the epidermis of mice caused epidermal detachment from the basement membrane, epidermal hyperthickening and progressive hair loss. PINCH-1-deficient keratinocytes also displayed profound adhesion, spreading and migration defects in vitro that were substantially more severe than those of ILK-deficient keratinocytes indicating that PINCH-1 also exerts functions in an ILK-independent manner. By isolating the PINCH-1 interactome, the LIM-domain-containing and actin-binding protein epithelial protein lost in neoplasm (EPLIN, also known as LIMA1) was identified as a new PINCH-1-associated protein. EPLIN localized, in a PINCH-1-dependent manner, to integrin adhesion sites of keratinocytes in vivo and in vitro and its depletion severely attenuated keratinocyte spreading and migration on collagen and fibronectin without affecting PINCH-1 levels in focal adhesions. Given that the low PINCH-1 levels in ILK-deficient keratinocytes were sufficient to recruit EPLIN to integrin adhesions, our findings suggest that PINCH-1 regulates integrin-mediated adhesion of keratinocytes through the interactions with ILK as well as EPLIN. PMID:25609703

  9. The Protein Information Management System (PiMS): a generic tool for any structural biology research laboratory.

    PubMed

    Morris, Chris; Pajon, Anne; Griffiths, Susanne L; Daniel, Ed; Savitsky, Marc; Lin, Bill; Diprose, Jonathan M; da Silva, Alan Wilter; Pilicheva, Katya; Troshin, Peter; van Niekerk, Johannes; Isaacs, Neil; Naismith, James; Nave, Colin; Blake, Richard; Wilson, Keith S; Stuart, David I; Henrick, Kim; Esnouf, Robert M

    2011-04-01

    The techniques used in protein production and structural biology have been developing rapidly, but techniques for recording the laboratory information produced have not kept pace. One approach is the development of laboratory information-management systems (LIMS), which typically use a relational database schema to model and store results from a laboratory workflow. The underlying philosophy and implementation of the Protein Information Management System (PiMS), a LIMS development specifically targeted at the flexible and unpredictable workflows of protein-production research laboratories of all scales, is described. PiMS is a web-based Java application that uses either Postgres or Oracle as the underlying relational database-management system. PiMS is available under a free licence to all academic laboratories either for local installation or for use as a managed service. PMID:21460443

  10. The Protein Information Management System (PiMS): a generic tool for any structural biology research laboratory

    PubMed Central

    Morris, Chris; Pajon, Anne; Griffiths, Susanne L.; Daniel, Ed; Savitsky, Marc; Lin, Bill; Diprose, Jonathan M.; Wilter da Silva, Alan; Pilicheva, Katya; Troshin, Peter; van Niekerk, Johannes; Isaacs, Neil; Naismith, James; Nave, Colin; Blake, Richard; Wilson, Keith S.; Stuart, David I.; Henrick, Kim; Esnouf, Robert M.

    2011-01-01

    The techniques used in protein production and structural biology have been developing rapidly, but techniques for recording the laboratory information produced have not kept pace. One approach is the development of laboratory information-management systems (LIMS), which typically use a relational database schema to model and store results from a laboratory workflow. The underlying philosophy and implementation of the Protein Information Management System (PiMS), a LIMS development specifically targeted at the flexible and unpredictable workflows of protein-production research laboratories of all scales, is described. PiMS is a web-based Java application that uses either Postgres or Oracle as the underlying relational database-management system. PiMS is available under a free licence to all academic laboratories either for local installation or for use as a managed service. PMID:21460443

  11. Deep sequencing of RNA from three different extracellular vesicle (EV) subtypes released from the human LIM1863 colon cancer cell line uncovers distinct miRNA-enrichment signatures.

    PubMed

    Ji, Hong; Chen, Maoshan; Greening, David W; He, Weifeng; Rai, Alin; Zhang, Wenwei; Simpson, Richard J

    2014-01-01

    Secreted microRNAs (miRNAs) enclosed within extracellular vesicles (EVs) play a pivotal role in intercellular communication by regulating recipient cell gene expression and affecting target cell function. Here, we report the isolation of three distinct EV subtypes from the human colon carcinoma cell line LIM1863--shed microvesicles (sMVs) and two exosome populations (immunoaffinity isolated A33-exosomes and EpCAM-exosomes). Deep sequencing of miRNA libraries prepared from parental LIM1863 cells/derived EV subtype RNA yielded 254 miRNA identifications, of which 63 are selectively enriched in the EVs--miR-19a/b-3p, miR-378a/c/d, and miR-577 and members of the let-7 and miR-8 families being the most prominent. Let-7a-3p*, let-7f-1-3p*, miR-451a, miR-574-5p*, miR-4454 and miR-7641 are common to all EV subtypes, and 6 miRNAs (miR-320a/b/c/d, miR-221-3p, and miR-200c-3p) discern LIM1863 exosomes from sMVs; miR-98-5p was selectively represented only in sMVs. Notably, A33-Exos contained the largest number (32) of exclusively-enriched miRNAs; 14 of these miRNAs have not been reported in the context of CRC tissue/biofluid analyses and warrant further examination as potential diagnostic markers of CRC. Surprisingly, miRNA passenger strands (star miRNAs) for miR-3613-3p*, -362-3p*, -625-3p*, -6842-3p* were the dominant strand in A33-Exos, the converse to that observed in parental cells. This finding suggests miRNA biogenesis may be interlinked with endosomal/exosomal processing. PMID:25330373

  12. The variability of stratospheric and mesospheric NO2 in the polar winter night observed by LIMS. [Limb Infrared Monitor of Stratosphere

    NASA Technical Reports Server (NTRS)

    Russell, M. J., III; Remsberg, E. E.; Callis, L. B.; Solomon, S.; Gordley, L. L.

    1984-01-01

    The LIMS experiment sounded the upper atmosphere from late October 1978 to late May 1979 and provided vertical profiles of atmospheric temperature, 03, H2O, HNO3, and NO2. Radiance averaging was used before retrieval to measure the altitude distribution of NO2 over the altitude range from the lower stratosphere into the mesosphere. Observations in the polar winter night region northward of about 70 deg N reveal NO2 levels near 175 ppbv at about 70 km, and they show a significant longitudinal variability (factor of 4 to 7). A definite temporal trend exists, showing a buildup of mesospheric and stratospheric NO2 during the polar night and a subsequent slowing of the increase of decline after sunlight returns, depending on altitude. The data represent the first experimental evidence that the thermosphere is an NO(x) source for the mesosphere and stratosphere.

  13. Homeodomain proteins: an update.

    PubMed

    Bürglin, Thomas R; Affolter, Markus

    2016-06-01

    Here, we provide an update of our review on homeobox genes that we wrote together with Walter Gehring in 1994. Since then, comprehensive surveys of homeobox genes have become possible due to genome sequencing projects. Using the 103 Drosophila homeobox genes as example, we present an updated classification. In animals, there are 16 major classes, ANTP, PRD, PRD-LIKE, POU, HNF, CUT (with four subclasses: ONECUT, CUX, SATB, and CMP), LIM, ZF, CERS, PROS, SIX/SO, plus the TALE superclass with the classes IRO, MKX, TGIF, PBC, and MEIS. In plants, there are 11 major classes, i.e., HD-ZIP (with four subclasses: I to IV), WOX, NDX, PHD, PLINC, LD, DDT, SAWADEE, PINTOX, and the two TALE classes KNOX and BEL. Most of these classes encode additional domains apart from the homeodomain. Numerous insights have been obtained in the last two decades into how homeodomain proteins bind to DNA and increase their specificity by interacting with other proteins to regulate cell- and tissue-specific gene expression. Not only protein-DNA base pair contacts are important for proper target selection; recent experiments also reveal that the shape of the DNA plays a role in specificity. Using selected examples, we highlight different mechanisms of homeodomain protein-DNA interaction. The PRD class of homeobox genes was of special interest to Walter Gehring in the last two decades. The PRD class comprises six families in Bilateria, and tinkers with four different motifs, i.e., the PAIRED domain, the Groucho-interacting motif EH1 (aka Octapeptide or TN), the homeodomain, and the OAR motif. Homologs of the co-repressor protein Groucho are also present in plants (TOPLESS), where they have been shown to interact with small amphipathic motives (EAR), and in yeast (TUP1), where we find an EH1-like motif in MATα2. PMID:26464018

  14. Epigenetic repression of PDZ-LIM domain-containing protein 2 promotes ovarian cancer via NOS2-derived nitric oxide signaling

    PubMed Central

    Lau, Wayne Bond; Lau, Bonnie; Luo, Zhongyue; Lin, Qiao; Yang, Huiliang; Xuan, Yu; Yi, Tao; Zhao, Xia; Wei, Yuquan

    2016-01-01

    Ovarian cancer constitutes one of the most lethal gynaecological malignancies worldwide and currently no satisfactory therapeutic approaches have been established. Therefore, elucidation of molecular mechanisms to develop targeted therapy of ovarian cancer is crucial. PDLIM2 is critical to promote ubiquitination of nuclear p65 and thus its role in inflammation has been highlighted recently. We demonstrate that PDLIM2 is decreased in both ovarian high-grade serous carcinoma and in various human ovarian cancer cell lines compared with normal ovary tissues and human ovarian surface epithelial cells (HOSE). Further functional analysis revealed that PDLIM2 is epigenetically repressed in ovarian cancer development and inhibition of PDLIM2 promoted ovarian cancer growth both in vivo and in vitro via NOS2-derived nitric oxide signaling, leading to recruitment of M2 type macrophages. These results suggest that PDLIM2 might be involved in ovarian cancer pathogenesis, which could serve as a promising therapeutic target for ovarian cancer patients. PMID:26593252

  15. Neogene-quaternary Ostracoda and paleoenvironments, of the Limón basin, Costa Rica, and Bocas del Toro basin, Panama

    USGS Publications Warehouse

    Borne, P.F.; Cronin, T. M.; Hazel, J.E.

    1999-01-01

    Tropical marine ostracodes from Neogene and Quaternary sediments of the Central American Caribbean region have been the subject of biostratigraphic, ecological, taxonomic, and evolutionary studies. As part of the Panama Paleontology Project (PPP), Neogene and Quaternary ostracodes are being studied from the Central American region. The overall goal of this research is to evaluate the impact of the emergence of the Central American Isthmus as a land barrier between the Caribbean/tropical Atlantic and the Pacific oceans on marine ostracode biodiversity and the oceanic environments in which extant ostracodes evolved. Due to the ecological specificity of many living tropical ostracode species, they are ideally suited for reconstructing paleoenvironments on the basis of their occurrence in fossil assemblages, which in turn can lead to a better understanding of the tropical climatic and tectonic history of Central America. The principal aims of this chapter are: (a) to document the composition of the ostracode assemblages from the Limón Basin of Costa Rica and the Bocas del Toro Basin of Panama, two areas yielding extensive ma rine ostracode assemblages; (b) to describe the environments of deposition within these basins; and (c) to document the stratigraphic distribution of potentially agediagnostic ostracode species in the Limón and Bocas del Toro basins in order to enhance their use in Central American biostratigraphy. A secondary, but none-the-less important goal is to assemble a database on the distribution of modem ostracode species in the Caribbean and adjacent areas as a basis for comparison with fossil assemblages. Although the ecological, biostratigraphic and paleoenvironmental conclusions presented here will improve as additional material is studied, these fossil and modem ostracode databases constitute the foundation for future evolutionary and geochernical studies of tropical Caribbean and eastern Pacific Ocean ostracodes. Moreover, we present here evidence

  16. Human p53 with a mutation in codone 273 facilitates the amplification of the gene dhfr in rat-1 and LIM1215 cells

    SciTech Connect

    Il`inskaya, G.V.; Sokova, O.I.; Kopnin, B.P.

    1995-05-01

    The effect of the expression of the exogenous human mutant p53 (Arg {yields} His in codone 273) on the amplification rate of the gene dhfr in permissive Rat-1 and LIM1215 cells was studied. It was shown that introduction of a retroviral construct with p53His273 resulted in the accumulation of methotrexate-resistant variants with an increased number of dhfr copies in populations of recipient cells. Luria-Delbruck fluctuation analysis revealed a four- to six-fold increase in the rate of appearance of new methotrexate-resistant cells. Chromosomal analysis demonstrated an extrachromosomal location of amplified DNA in cells containing p53His273, as was the case for control sublines. The data obtained indicate that modifications of p53 may induce gene amplification not only via removing the proliferation block of cells with amplified genes in selective medium, but also via some other mechanisms that seem to increase the chromosomal recombination rate. 24 refs., 3 figs., 3 tabs.

  17. Li mobility in Nasicon-type materials LiM2(PO4)3, M = Ge, Ti, Sn, Zr and Hf, followed by 7Li NMR spectroscopy.

    PubMed

    Arbi, K; París, M A; Sanz, J

    2011-10-21

    Lithium mobility in LiM(2)(PO(4))(3) compounds, M = Ge and Sn, has been investigated by (7)Li Nuclear Magnetic Resonance (NMR) spectroscopy, and deduced information compared with that reported previously in Ti, Zr and Hf members of the series in the temperature range 100-500 K. From the analysis of (7)Li NMR quadrupole interactions (C(Q) and η parameters), spin-spin T(2)(-1) and spin-lattice T(1)(-1) relaxation rates, structural sites occupancy and mobility of lithium have been deduced. Below 250 K, Li ions are preferentially located at M(1) sites in rhombohedral phases, but occupy intermediate M(12) sites between M(1) and M(2) sites in triclinic ones. In high-temperature rhombohedral phases, a superionic state is achieved when residence times at M(1) and M(12) sites become similar and correlation effects on Li motion decrease. This state can be obtained by large order-disorder transformations in rhombohedral phases or by sharp first order transitions in triclinic ones. The presence of two relaxation mechanisms in T(1)(-1) plots of rhombohedral phases has been associated with departures of conductivity from the Arrhenius behavior. Long term mobility of lithium is discussed in terms of the cation vacancy distribution along conduction paths. PMID:21897945

  18. Synthesis, crystal structures and optical properties of two congruent-melting isotypic diphosphates: LiM3P2O7 (M=Na, K)

    NASA Astrophysics Data System (ADS)

    Shi, Yunjing; Wang, Ying; Pan, Shilie; Yang, Zhihua; Dong, Xiaoyu; Wu, Hongping; Zhang, Min; Cao, Jian; Zhou, Zhongxiang

    2013-01-01

    Two new isotypic diphosphates, LiNa3P2O7 (1) and LiK3P2O7 (2), have been synthesized by conventional solid-state reaction. The single-crystal X-ray structural analyses have shown that they crystallize in the orthorhombic space group C2221 (No. 20) with the unit cells: a=5.4966(2) Å, b=9.1365(4) Å, c=12.2764(5) Å for compound 1 and a=6.0373(14) Å, b=9.339(2) Å, c=13.292(3) Å for compound 2. The LiM3P2O7 (M=Na, K) consist of two-dimensional [LiP2O7]3- layers, which are composed by LiO4 tetrahedral and diphosphate groups, the Na or K atoms are filled in the interlayers and balance the charge. Second harmonic generation (SHG) on powder samples have been measured using Kurtz and Perry techniques. Thermal analyses, IR spectroscopy, UV-vis-NIR diffuse reflectance spectra, and band structure calculations are performed on the reported compounds.

  19. Relationship between the initial rate of protein aggregation and the lag period for amorphous aggregation.

    PubMed

    Borzova, Vera A; Markossian, Kira A; Kurganov, Boris I

    2014-07-01

    Lag period is an inherent characteristic of the kinetic curves registered for protein aggregation. The appearance of a lag period is connected with the nucleation stage and the stages of the formation of folding or unfolding intermediates prone to aggregation (for example, the stage of protein unfolding under stress conditions). Discovering the kinetic regularities essential for elucidation of the protein aggregation mechanism comprises deducing the relationship between the lag period and aggregation rate. Fändrich proposed the following equation connecting the duration of the lag phase (tlag) and the aggregate growth rate (kg) in the amyloid fibrillation: kg=const/tlag. To establish the relationship between the initial rate of protein aggregation (v) and the lag period (t0) in the case of amorphous aggregation, the kinetics of dithithreitol-induced aggregation of holo-α-lactalbumin from bovine milk was studied (0.1M Na-phosphate buffer, pH 6.8; 37°C). The order of aggregation with respect to protein (n) was calculated from the dependence of the initial rate of protein aggregation on the α-lactalbumin concentration (n=5.3). The following equation connecting v and t0 has been proposed: v(1/n)=const/(t0-t0,lim), where t0,lim is the limiting value of t0 at high concentrations of the protein. PMID:24794200

  20. The four and a half LIM domains 2 (FHL2) regulates ovarian granulosa cell tumor progression via controlling AKT1 transcription

    PubMed Central

    Hua, G; He, C; Lv, X; Fan, L; Wang, C; Remmenga, S W; Rodabaugh, K J; Yang, L; Lele, S M; Yang, P; Karpf, A R; Davis, J S; Wang, C

    2016-01-01

    The four and a half LIM domains 2 (FHL2) has been shown to play important roles in the regulation of cell proliferation, survival, adhesion, motility and signal transduction in a cell type and tissue-dependent manner. However, the function of FHL2 in ovarian physiology and pathology is unclear. The aim of this study was to determine the role and functional mechanism of FHL2 in the progression of ovarian granulosa cell tumors (GCTs). Immunohistochemical analysis indicated that FHL2 was overexpressed in GCT tissues. Cellular localization of FHL2 in GCT cells was cell cycle dependent. Knockdown of FHL2 suppressed GCT cell growth, reduced cell viability and inhibited cell migration. Consistently, ectopic expression of FHL2 in GCT cells with very low endogenous FHL2 promoted cell growth, improved cell viability and enhance cell migration. Importantly, overexpression of FHL2 promoted GCT progression in vivo. Mechanistic studies indicated that FHL2 regulates AKT1 gene expression in vitro and in vivo. Knockdown of FHL2 or AKT1 in GCT cell lines induced very similar phenotypes. Ectopic expression of constitutively active AKT1 rescued FHL2 knockdown-induced arrest of GCT cell growth and reduction of GCT cell viability, suggesting that FHL2 regulates GCT cell growth and viability through controlling AKT1 expression. Finally, co-immunoprecipitation and chromatin immunoprecipitation analyses indicated that FHL2 functions as a co-activator of NFκB and AP-1 to regulate AKT1 gene transcription. In conclusion, results from the present study indicate that FHL2 exerts its oncogenic action in GCT cells via controlling AKT1 gene expression. FHL2 is a promising target for the development of novel drugs against ovarian granulosa cell tumor. PMID:27415427

  1. The four and a half LIM domains 2 (FHL2) regulates ovarian granulosa cell tumor progression via controlling AKT1 transcription.

    PubMed

    Hua, G; He, C; Lv, X; Fan, L; Wang, C; Remmenga, S W; Rodabaugh, K J; Yang, L; Lele, S M; Yang, P; Karpf, A R; Davis, J S; Wang, C

    2016-01-01

    The four and a half LIM domains 2 (FHL2) has been shown to play important roles in the regulation of cell proliferation, survival, adhesion, motility and signal transduction in a cell type and tissue-dependent manner. However, the function of FHL2 in ovarian physiology and pathology is unclear. The aim of this study was to determine the role and functional mechanism of FHL2 in the progression of ovarian granulosa cell tumors (GCTs). Immunohistochemical analysis indicated that FHL2 was overexpressed in GCT tissues. Cellular localization of FHL2 in GCT cells was cell cycle dependent. Knockdown of FHL2 suppressed GCT cell growth, reduced cell viability and inhibited cell migration. Consistently, ectopic expression of FHL2 in GCT cells with very low endogenous FHL2 promoted cell growth, improved cell viability and enhance cell migration. Importantly, overexpression of FHL2 promoted GCT progression in vivo. Mechanistic studies indicated that FHL2 regulates AKT1 gene expression in vitro and in vivo. Knockdown of FHL2 or AKT1 in GCT cell lines induced very similar phenotypes. Ectopic expression of constitutively active AKT1 rescued FHL2 knockdown-induced arrest of GCT cell growth and reduction of GCT cell viability, suggesting that FHL2 regulates GCT cell growth and viability through controlling AKT1 expression. Finally, co-immunoprecipitation and chromatin immunoprecipitation analyses indicated that FHL2 functions as a co-activator of NFκB and AP-1 to regulate AKT1 gene transcription. In conclusion, results from the present study indicate that FHL2 exerts its oncogenic action in GCT cells via controlling AKT1 gene expression. FHL2 is a promising target for the development of novel drugs against ovarian granulosa cell tumor. PMID:27415427

  2. A LIM-homeobox gene is required for differentiation of Wnt-expressing cells at the posterior end of the planarian body.

    PubMed

    Hayashi, Tetsutaro; Motoishi, Minako; Yazawa, Shigenobu; Itomi, Kazu; Tanegashima, Chiharu; Nishimura, Osamu; Agata, Kiyokazu; Tarui, Hiroshi

    2011-09-01

    Planarians have high regenerative ability, which is dependent on pluripotent adult somatic stem cells called neoblasts. Recently, canonical Wnt/β-catenin signaling was shown to be required for posterior specification, and Hedgehog signaling was shown to control anterior-posterior polarity via activation of the Djwnt1/P-1 gene at the posterior end of planarians. Thus, various signaling molecules play an important role in planarian stem cell regulation. However, the molecular mechanisms directly involved in stem cell differentiation have remained unclear. Here, we demonstrate that one of the planarian LIM-homeobox genes, Djislet, is required for the differentiation of Djwnt1/P-1-expressing cells from stem cells at the posterior end. RNA interference (RNAi)-treated planarians of Djislet [Djislet(RNAi)] show a tail-less phenotype. Thus, we speculated that Djislet might be involved in activation of the Wnt signaling pathway in the posterior blastema. When we carefully examined the expression pattern of Djwnt1/P-1 by quantitative real-time PCR during posterior regeneration, we found two phases of Djwnt1/P-1 expression: the first phase was detected in the differentiated cells in the old tissue in the early stage of regeneration and then a second phase was observed in the cells derived from stem cells in the posterior blastema. Interestingly, Djislet is expressed in stem cell-derived DjPiwiA- and Djwnt1/P-1-expressing cells, and Djislet(RNAi) only perturbed the second phase. Thus, we propose that Djislet might act to trigger the differentiation of cells expressing Djwnt1/P-1 from stem cells. PMID:21828095

  3. Facilitation of Endosomal Recycling by an IRG Protein Homolog Maintains Apical Tubule Structure in Caenorhabditis elegans.

    PubMed

    Grussendorf, Kelly A; Trezza, Christopher J; Salem, Alexander T; Al-Hashimi, Hikmat; Mattingly, Brendan C; Kampmeyer, Drew E; Khan, Liakot A; Hall, David H; Göbel, Verena; Ackley, Brian D; Buechner, Matthew

    2016-08-01

    Determination of luminal diameter is critical to the function of small single-celled tubes. A series of EXC proteins, including EXC-1, prevent swelling of the tubular excretory canals in Caenorhabditis elegans In this study, cloning of exc-1 reveals it to encode a homolog of mammalian IRG proteins, which play roles in immune response and autophagy and are associated with Crohn's disease. Mutants in exc-1 accumulate early endosomes, lack recycling endosomes, and exhibit abnormal apical cytoskeletal structure in regions of enlarged tubules. EXC-1 interacts genetically with two other EXC proteins that also affect endosomal trafficking. In yeast two-hybrid assays, wild-type and putative constitutively active EXC-1 binds to the LIM-domain protein EXC-9, whose homolog, cysteine-rich intestinal protein, is enriched in mammalian intestine. These results suggest a model for IRG function in forming and maintaining apical tubule structure via regulation of endosomal recycling. PMID:27334269

  4. Three-dimensional structure determination of proteins related to human health in their functional context at The Israel Structural Proteomics Center (ISPC). This paper was presented at ICCBM10.

    PubMed

    Albeck, Shira; Burstein, Yigal; Dym, Orly; Jacobovitch, Yossi; Levi, Nurit; Meged, Ran; Michael, Yigal; Peleg, Yoav; Prilusky, Jaime; Schreiber, Gideon; Silman, Israel; Unger, Tamar; Sussman, Joel L

    2005-10-01

    The principal goal of the Israel Structural Proteomics Center (ISPC) is to determine the structures of proteins related to human health in their functional context. Emphasis is on the solution of structures of proteins complexed with their natural partner proteins and/or with DNA. To date, the ISPC has solved the structures of 14 proteins, including two protein complexes. It has adopted automated high-throughput (HTP) cloning and expression techniques and is now expressing in Escherichia coli, Pichia pastoris and baculovirus, and in a cell-free E. coli system. Protein expression in E. coli is the primary system of choice in which different parameters are tested in parallel. Much effort is being devoted to development of automated refolding of proteins expressed as inclusion bodies in E. coli. The current procedure utilizes tagged proteins from which the tag can subsequently be removed by TEV protease, thus permitting streamlined purification of a large number of samples. Robotic protein crystallization screens and optimization utilize both the batch method under oil and vapour diffusion. In order to record and organize the data accumulated by the ISPC, a laboratory information-management system (LIMS) has been developed which facilitates data monitoring and analysis. This permits optimization of conditions at all stages of protein production and structure determination. A set of bioinformatics tools, which are implemented in our LIMS, is utilized to analyze each target. PMID:16204888

  5. LASP1, a Newly Identified Melanocytic Protein with a Possible Role in Melanin Release, but Not in Melanoma Progression

    PubMed Central

    Houben, Roland; Grunewald, Thomas G. P.; Goebeler, Matthias; Butt, Elke

    2015-01-01

    The LIM and SH3 protein 1 (LASP1) is a focal adhesion protein. Its expression is increased in many malignant tumors. However, little is known about the physiological role of the protein. In the present study, we investigated the expression and function of LASP1 in normal skin, melanocytic nevi and malignant melanoma. In normal skin, a distinct LASP1 expression is visible only in the basal epidermal layer while in nevi LASP1 protein is detected in all melanocytes. Melanoma exhibit no increase in LASP1 mRNA compared to normal skin. In melanocytes, the protein is bound to dynamin and mainly localized at late melanosomes along the edges and at the tips of the cell. Knockdown of LASP1 results in increased melanin concentration in the cells. Collectively, we identified LASP1 as a hitherto unknown protein in melanocytes and as novel partner of dynamin in the physiological process of membrane constriction and melanosome vesicle release. PMID:26061439

  6. Chemical analysis of solid materials by a LIMS instrument designed for space research: 2D elemental imaging, sub-nm depth profiling and molecular surface analysis

    NASA Astrophysics Data System (ADS)

    Moreno-García, Pavel; Grimaudo, Valentine; Riedo, Andreas; Neuland, Maike B.; Tulej, Marek; Broekmann, Peter; Wurz, Peter

    2016-04-01

    Direct quantitative chemical analysis with high lateral and vertical resolution of solid materials is of prime importance for the development of a wide variety of research fields, including e.g., astrobiology, archeology, mineralogy, electronics, among many others. Nowadays, studies carried out by complementary state-of-the-art analytical techniques such as Auger Electron Spectroscopy (AES), X-ray Photoelectron Spectroscopy (XPS), Secondary Ion Mass Spectrometry (SIMS), Glow Discharge Time-of-Flight Mass Spectrometry (GD-TOF-MS) or Laser Ablation Inductively Coupled Plasma Mass Spectrometry (LA-ICP-MS) provide extensive insight into the chemical composition and allow for a deep understanding of processes that might have fashioned the outmost layers of an analyte due to its interaction with the surrounding environment. Nonetheless, these investigations typically employ equipment that is not suitable for implementation on spacecraft, where requirements concerning weight, size and power consumption are very strict. In recent years Laser Ablation/Ionization Mass Spectrometry (LIMS) has re-emerged as a powerful analytical technique suitable not only for laboratory but also for space applications.[1-3] Its improved performance and measurement capabilities result from the use of cutting edge ultra-short femtosecond laser sources, improved vacuum technology and fast electronics. Because of its ultimate compactness, simplicity and robustness it has already proven to be a very suitable analytical tool for elemental and isotope investigations in space research.[4] In this contribution we demonstrate extended capabilities of our LMS instrument by means of three case studies: i) 2D chemical imaging performed on an Allende meteorite sample,[5] ii) depth profiling with unprecedented sub-nm vertical resolution on Cu electrodeposited interconnects[6,7] and iii) preliminary molecular desorption of polymers without assistance of matrix or functionalized substrates.[8] On the whole

  7. Protein open-access liquid chromatography/mass spectrometry.

    PubMed

    White, Wendy L; Wagner, Craig D; Hall, John T; Chaney, Erin E; George, Bindu; Hofmann, Karen; Miller, Luke A D; Williams, Jon D

    2005-01-01

    Each year increasing numbers of proteins are submitted for routine characterization by liquid chromatography/mass spectrometry (LC/MS). This paper reports a solution that transforms routine LC/MS analysis of proteins into a fully automated process that significantly reduces analyst intervention. The solution developed, protein open-access (OA) LC/MS, consists of web-enabled sample submission and registration, automated data processing, data interpretation, and report generation. Sample submissions and results are recorded in a LIMS that utilizes an Oracle database. The protein sequence is captured during the sample submission process, stored in the database, and utilized to determine the theoretical protein molecular weight. This calculated mass is used to set the parameters for transformation of the mass-to-charge spectra to the mass domain and evaluate the presence or absence of the desired protein. Three protein OA-LC/MS instruments have been deployed in our facility to support protein characterization, purification, and modification efforts. PMID:15609371

  8. LIMS for Lasers 2015 for achieving long-term accuracy and precision of δ2H, δ17O, and δ18O of waters using laser absorption spectrometry

    USGS Publications Warehouse

    Coplen, Tyler B.; Wassenaar, Leonard I

    2015-01-01

    Although laser absorption spectrometry (LAS) instrumentation is easy to use, its incorporation into laboratory operations is not easy, owing to extensive offline manipulation of comma-separated-values files for outlier detection, between-sample memory correction, nonlinearity (δ-variation with water amount) correction, drift correction, normalization to VSMOW-SLAP scales, and difficulty in performing long-term QA/QC audits. METHODS: A Microsoft Access relational-database application, LIMS (Laboratory Information Management System) for Lasers 2015, was developed. It automates LAS data corrections and manages clients, projects, samples, instrument-sample lists, and triple-isotope (δ(17) O, δ(18) O, and δ(2) H values) instrumental data for liquid-water samples. It enables users to (1) graphically evaluate sample injections for variable water yields and high isotope-delta variance; (2) correct for between-sample carryover, instrumental drift, and δ nonlinearity; and (3) normalize final results to VSMOW-SLAP scales. RESULTS: Cost-free LIMS for Lasers 2015 enables users to obtain improved δ(17) O, δ(18) O, and δ(2) H values with liquid-water LAS instruments, even those with under-performing syringes. For example, LAS δ(2) HVSMOW measurements of USGS50 Lake Kyoga (Uganda) water using an under-performing syringe having ±10 % variation in water concentration gave +31.7 ± 1.6 ‰ (2-σ standard deviation), compared with the reference value of +32.8 ± 0.4 ‰, after correction for variation in δ value with water concentration, between-sample memory, and normalization to the VSMOW-SLAP scale. CONCLUSIONS: LIMS for Lasers 2015 enables users to create systematic, well-founded instrument templates, import δ(2) H, δ(17) O, and δ(18) O results, evaluate performance with automatic graphical plots, correct for δ nonlinearity due to variable water concentration, correct for between-sample memory, adjust for drift, perform VSMOW-SLAP normalization, and

  9. The Protein Information Management System (PiMS): a generic tool for any structural biology research laboratory

    SciTech Connect

    Morris, Chris; Pajon, Anne; Griffiths, Susanne L.; Daniel, Ed; Savitsky, Marc; Lin, Bill; Diprose, Jonathan M.; Wilter da Silva, Alan; Pilicheva, Katya; Troshin, Peter; Niekerk, Johannes van; Isaacs, Neil; Naismith, James; Nave, Colin; Blake, Richard; Wilson, Keith S.; Stuart, David I.; Henrick, Kim; Esnouf, Robert M.

    2011-04-01

    The Protein Information Management System (PiMS) is described together with a discussion of how its features make it well suited to laboratories of all sizes. The techniques used in protein production and structural biology have been developing rapidly, but techniques for recording the laboratory information produced have not kept pace. One approach is the development of laboratory information-management systems (LIMS), which typically use a relational database schema to model and store results from a laboratory workflow. The underlying philosophy and implementation of the Protein Information Management System (PiMS), a LIMS development specifically targeted at the flexible and unpredictable workflows of protein-production research laboratories of all scales, is described. PiMS is a web-based Java application that uses either Postgres or Oracle as the underlying relational database-management system. PiMS is available under a free licence to all academic laboratories either for local installation or for use as a managed service.

  10. Nuclear translocation of the cytoskeleton-associated protein, smALP, upon induction of skeletal muscle differentiation

    SciTech Connect

    Cambier, Linda; Pomies, Pascal

    2011-06-17

    Highlights: {yields} The cytoskeleton-associated protein, smALP, is expressed in differentiated skeletal muscle. {yields} smALP is translocated from the cytoplasm to the nucleus of C2C12 myoblasts upon induction of myogenesis. {yields} The differentiation-dependent nuclear translocation of smALP occurs in parallel with the nuclear accumulation of myogenin. {yields} The LIM domain of smALP is essential for the nuclear accumulation of the protein. {yields} smALP might act in the nucleus to control some critical aspect of the muscle differentiation process. -- Abstract: The skALP isoform has been shown to play a critical role in actin organization and anchorage within the Z-discs of skeletal muscles, but no data is available on the function of the smALP isoform in skeletal muscle cells. Here, we show that upon induction of differentiation a nuclear translocation of smALP from the cytoplasm to the nucleus of C2C12 myoblasts, concomitant to an up-regulation of the protein expression, occurs in parallel with the nuclear accumulation of myogenin. Moreover, we demonstrate that the LIM domain of smALP is essential for the nuclear translocation of the protein.

  11. Kinematic 3-D Retro-Modeling of an Orogenic Bend in the South Limón Fold-and-Thrust Belt, Eastern Costa Rica: Prediction of the Incremental Internal Strain Distribution

    NASA Astrophysics Data System (ADS)

    Brandes, Christian; Tanner, David C.; Winsemann, Jutta

    2016-03-01

    The South Limón fold-and-thrust belt, in the back-arc area of southern Costa Rica, is characterized by a 90° curvature of the strike of the thrust planes and is therefore a natural laboratory for the analysis of curved orogens. The analysis of curved fold-and-thrust belts is a challenge because of the varying structural orientations within the belt. Based on seismic reflection lines, we created a 3-D subsurface model containing three major thrust faults and three stratigraphic horizons. 3-D kinematic retro-deformation modeling was carried out to analyze the spatial evolution of the fold-and-thrust belt. The maximum amount of displacement on each of the faults is (from hinterland to foreland); thrust 1: 800 m; thrust 2: 600 m; thrust 3: 250 m. The model was restored sequentially to its pre-deformational state. The strain history of the stratigraphic horizons in the model was calculated at every step. This shows that the internal strain pattern has an abrupt change at the orogenic bend. Contractional strain occurs in the forelimbs of the hanging-wall anticlines, while a zone of dilative strain spreads from the anticline crests to the backlimbs. The modeling shows that a NNE-directed transport direction best explains the structural evolution of the bend. This would require a left-lateral strike-slip zone in the North to compensate for the movement and thereby decoupling the South Limón fold-and-thrust belt from northern Costa Rica. Therefore, our modeling supports the presence of the Trans-Isthmic fault system, at least during the Plio-Pleistocene.

  12. Early embryonic expression of a LIM-homeobox gene Cs-lhx3 is downstream of beta-catenin and responsible for the endoderm differentiation in Ciona savignyi embryos.

    PubMed

    Satou, Y; Imai, K S; Satoh, N

    2001-09-01

    In early Ciona embryos, nuclear accumulation of beta-catenin is most probably the first step of endodermal cell specification. If beta-catenin is mis- and/or overexpressed, presumptive notochord cells and epidermal cells change their fates into endodermal cells, whereas if beta-catenin nuclear localization is downregulated by the overexpression of cadherin, the endoderm differentiation is suppressed, accompanied with the differentiation of extra epidermal cells ( Imai, K., Takada, N., Satoh, N. and Satou, Y. (2000) Development 127, 3009-3020). Subtractive hybridization screens of mRNAs between beta-catenin overexpressed embryos and cadherin overexpressed embryos were conducted to identify potential beta-catenin target genes that are responsible for endoderm differentiation in Ciona savignyi embryos. We found that a LIM-homeobox gene (Cs-lhx3), an otx homolog (Cs-otx) and an NK-2 class gene (Cs-ttf1) were among beta-catenin downstream genes. In situ hybridization signals for early zygotic expression of Cs-lhx3 were evident only in the presumptive endodermal cells as early as the 32-cell stage, those of Cs-otx in the mesoendodermal cells at the 32-cell stage and those of Cs-ttf1 in the endodermal cells at the 64-cell stage. Later, Cs-lhx3 was expressed again in a set of neuronal cells in the tailbud embryo, while Cs-otx was expressed in the anterior nervous system of the embryo. Expression of all three genes was upregulated in beta-catenin overexpressed embryos and downregulated in cadherin overexpressed embryos. Injection of morpholino oligonucleotides against Cs-otx did not affect the embryonic endoderm differentiation, although the formation of the central nervous system was suppressed. Injection of Cs-ttf1 morpholino oligonucleotides also failed to suppress the endoderm differentiation, although injection of its synthetic mRNAs resulted in ectopic development of endoderm differentiation marker alkaline phosphatase. By contrast, injection of Cs-lhx3 morpholino

  13. Analyzing protein-protein interactions from affinity purification-mass spectrometry data with SAINT

    PubMed Central

    Choi, Hyungwon; Liu, Guomin; Mellacheruvu, Dattatreya; Tyers, Mike; Gingras, Anne-Claude; Nesvizhskii, Alexey I.

    2012-01-01

    Significance Analysis of INTeractome (SAINT) is a software package for scoring protein-protein interactions based on label-free quantitative proteomics data (e.g. spectral count or intensity) in affinity purification – mass spectrometry (AP-MS) experiments. SAINT allows bench scientists to select bona fide interactions and remove non-specific interactions in an unbiased manner. However, there is no `one-size-fits-all' statistical model for every dataset, since the experimental design varies across studies. Key variables include the number of baits, the number of biological replicates per bait, and control purifications. Here we give a detailed account of input data format, control data, selection of high confidence interactions, and visualization of filtered data. We explain additional options for customizing the statistical model for optimal filtering in specific datasets. We also discuss a graphical user interface of SAINT in connection to the LIMS system ProHits which can be installed as a virtual machine on Mac OSX or PC Windows computers. PMID:22948729

  14. Cysteine-rich intestinal protein binds zinc during transmucosal zinc transport.

    PubMed Central

    Hempe, J M; Cousins, R J

    1991-01-01

    The mechanism of zinc absorption has not been delineated, but kinetic studies show that both passive and carrier-mediated processes are involved. We have identified a low molecular mass zinc-binding protein in the soluble fraction of rat intestinal mucosa that could function as an intracellular zinc carrier. The protein was not detected in liver or pancreas, suggesting a role specific to the intestine. The protein binds zinc during transmucosal zinc transport and shows signs of saturation at higher luminal zinc concentrations, characteristics consistent with a role in carrier-mediated zinc absorption. Microsequence analysis of the protein purified by gel-filtration HPLC and SDS/PAGE showed complete identity within the first 41 N-terminal amino acids with the deduced protein sequence of cysteine-rich intestinal protein [Birkenmeier, E. H. & Gordon, J. I. (1986) Proc. Natl. Acad. Sci. USA 83, 2516-2520]. These investigators showed that the gene for this protein is developmentally regulated in neonates during the suckling period, conserved in many vertebrate species, and predominantly expressed in the small intestine. Cysteine-rich intestinal protein contains a recently identified conserved sequence of histidine and cysteine residues, the LIM motif, which our results suggest confers metal-binding properties that are important for zinc transport and/or functions of this micronutrient. Images PMID:1946385

  15. Cysteine-rich intestinal protein binds zinc during transmucosal zinc transport

    SciTech Connect

    Hempe, J.M.; Cousins, R.J. )

    1991-11-01

    The mechanism of zinc absorption has not been delineated, but kinetic studies show that both passive and carrier-mediated processes are involved. The authors have identified a low molecular mass zinc-binding protein in the soluble fraction of rat intestinal mucosa that could function as an intracellular zinc carrier. The protein was not detected in liver or pancreas, suggesting a role specific to the intestine. The protein binds zinc during transmucosal zinc transport and shows signs of saturation at higher luminal zinc concentrations, characteristics consistent with a role in carrier-mediated zinc absorption. Microsequence analysis of the protein purified by gel-filtration HPCL and SDS/PAGE showed complete identity within the first 41 N-terminal amino acids with the deduced protein sequence of cysteine-rich intestinal protein. These investigators showed that the gene for this protein is developmentally regulated in neonates during the suckling period, conserved in many vertebrate species, and predominantly expressed in the small intestine. Cysteine-rich intestinal protein contains a recently identified conserved sequence of histidine and cysteine residues, the LIM motif, which our results suggest confers metal-binding properties that are important for zinc transport and/or functions of this micronutrient.

  16. The netrin receptor UNC-40/DCC stimulates axon attraction and outgrowth through enabled and, in parallel, Rac and UNC-115/AbLIM.

    PubMed

    Gitai, Zemer; Yu, Timothy W; Lundquist, Erik A; Tessier-Lavigne, Marc; Bargmann, Cornelia I

    2003-01-01

    Netrins promote axon outgrowth and turning through DCC/UNC-40 receptors. To characterize Netrin signaling, we generated a gain-of-function UNC-40 molecule, MYR::UNC-40. MYR::UNC-40 causes axon guidance defects, excess axon branching, and excessive axon and cell body outgrowth. These defects are suppressed by loss-of-function mutations in ced-10 (a Rac GTPase), unc-34 (an Enabled homolog), and unc-115 (a putative actin binding protein). ced-10, unc-34, and unc-115 also function in endogenous unc-40 signaling. Our results indicate that Enabled functions in axonal attraction as well as axon repulsion. UNC-40 has two conserved cytoplasmic motifs that mediate distinct downstream pathways: CED-10, UNC-115, and the UNC-40 P2 motif act in one pathway, and UNC-34 and the UNC-40 P1 motif act in the other. Thus, UNC-40 might act as a scaffold to deliver several independent signals to the actin cytoskeleton. PMID:12526772

  17. Protein Condensation

    NASA Astrophysics Data System (ADS)

    Gunton, James D.; Shiryayev, Andrey; Pagan, Daniel L.

    2007-09-01

    Preface; 1. Introduction; 2. Globular protein structure; 3. Experimental methods; 4. Thermodynamics and statistical mechanics; 5. Protein-protein interactions; 6. Theoretical studies of equilibrium; 7. Nucleation theory; 8. Experimental studies of nucleation; 9. Lysozyme; 10. Some other globular proteins; 11. Membrane proteins; 12. Crystallins and cataracts; 13. Sickle hemoglobin and sickle cell anemia; 14, Alzheimer's disease; Index.

  18. Protein Condensation

    NASA Astrophysics Data System (ADS)

    Gunton, James D.; Shiryayev, Andrey; Pagan, Daniel L.

    2014-07-01

    Preface; 1. Introduction; 2. Globular protein structure; 3. Experimental methods; 4. Thermodynamics and statistical mechanics; 5. Protein-protein interactions; 6. Theoretical studies of equilibrium; 7. Nucleation theory; 8. Experimental studies of nucleation; 9. Lysozyme; 10. Some other globular proteins; 11. Membrane proteins; 12. Crystallins and cataracts; 13. Sickle hemoglobin and sickle cell anemia; 14, Alzheimer's disease; Index.

  19. Total protein

    MedlinePlus

    The total protein test measures the total amount of two classes of proteins found in the fluid portion of your ... nutritional problems, kidney disease or liver disease . If total protein is abnormal, you will need to have more ...

  20. Storage Proteins

    PubMed Central

    Fujiwara, Toru; Nambara, Eiji; Yamagishi, Kazutoshi; Goto, Derek B.; Naito, Satoshi

    2002-01-01

    Plants accumulate storage substances such as starch, lipids and proteins in certain phases of development. Storage proteins accumulate in both vegetative and reproductive tissues and serve as a reservoir to be used in later stages of plant development. The accumulation of storage protein is thus beneficial for the survival of plants. Storage proteins are also an important source of dietary plant proteins. Here, we summarize the genome organization and regulation of gene expression of storage protein genes in Arabidopsis. PMID:22303197

  1. Dietary Proteins

    MedlinePlus

    ... grains and beans. Proteins from meat and other animal products are complete proteins. This means they supply all of the amino acids the body can't make on its own. Most plant proteins are incomplete. You should eat different types of plant proteins every day to get ...

  2. Protein Analysis

    NASA Astrophysics Data System (ADS)

    Chang, Sam K. C.

    Proteins are an abundant component in all cells, and almost all except storage proteins are important for biological functions and cell structure. Food proteins are very complex. Many have been purified and characterized. Proteins vary in molecular mass, ranging from approximately 5000 to more than a million Daltons. They are composed of elements including hydrogen, carbon, nitrogen, oxygen, and sulfur. Twenty α-amino acids are the building blocks of proteins; the amino acid residues in a protein are linked by peptide bonds. Nitrogen is the most distinguishing element present in proteins. However, nitrogen content in various food proteins ranges from 13.4 to 19.1% (1) due to the variation in the specific amino acid composition of proteins. Generally, proteins rich in basic amino acids contain more nitrogen.

  3. The Nimbus 7 LIMS map archival tape

    NASA Technical Reports Server (NTRS)

    Haggard, K. V.; Remsberg, E. E.; Russell, J. M., III

    1986-01-01

    The development of the Langley Map Archival Tape (LAMAT) is described. The role of the Nimbus 7 orbit characteristics in the analysis and interpretation of LAMAT data is discussed. The pressure ranges and heights for the LAMAT components are given and the synoptic fields for each pressure and latitude were modeled as Fourier series in longitude with time-dependent coefficients. Examples of LAMAT temperature fields are presented and the accuracies of the LAMAT fields are evaluated. The fit between the Langley inverted profile archival tape (LAIPAT) and LAMAT is examined by comparing averaged zonal mean fields and standard deviations. Consideration is given to diurnal variations of LAIPAT data, and the time and spatial smoothing of the data.

  4. Total protein

    MedlinePlus

    ... page: //medlineplus.gov/ency/article/003483.htm Total protein To use the sharing features on this page, please enable JavaScript. The total protein test measures the total amount of two classes ...

  5. Whey Protein

    MedlinePlus

    ... shows that taking whey protein in combination with strength training increases lean body mass, strength, and muscle size. ... grams/kg of whey protein in combination with strength training for 6-10 weeks. For HIV/AIDS-related ...

  6. Protein Microarrays

    NASA Astrophysics Data System (ADS)

    Ricard-Blum, S.

    Proteins are key actors in the life of the cell, involved in many physiological and pathological processes. Since variations in the expression of messenger RNA are not systematically correlated with variations in the protein levels, the latter better reflect the way a cell functions. Protein microarrays thus supply complementary information to DNA chips. They are used in particular to analyse protein expression profiles, to detect proteins within complex biological media, and to study protein-protein interactions, which give information about the functions of those proteins [3-9]. They have the same advantages as DNA microarrays for high-throughput analysis, miniaturisation, and the possibility of automation. Section 18.1 gives a brief overview of proteins. Following this, Sect. 18.2 describes how protein microarrays can be made on flat supports, explaining how proteins can be produced and immobilised on a solid support, and discussing the different kinds of substrate and detection method. Section 18.3 discusses the particular format of protein microarrays in suspension. The diversity of protein microarrays and their applications are then reported in Sect. 18.4, with applications to therapeutics (protein-drug interactions) and diagnostics. The prospects for future developments of protein microarrays are then outlined in the conclusion. The bibliography provides an extensive list of reviews and detailed references for those readers who wish to go further in this area. Indeed, the aim of the present chapter is not to give an exhaustive or detailed analysis of the state of the art, but rather to provide the reader with the basic elements needed to understand how proteins are designed and used.

  7. Protein Structure

    ERIC Educational Resources Information Center

    Asmus, Elaine Garbarino

    2007-01-01

    Individual students model specific amino acids and then, through dehydration synthesis, a class of students models a protein. The students clearly learn amino acid structure, primary, secondary, tertiary, and quaternary structure in proteins and the nature of the bonds maintaining a protein's shape. This activity is fun, concrete, inexpensive and…

  8. Protein folds and protein folding

    PubMed Central

    Schaeffer, R. Dustin; Daggett, Valerie

    2011-01-01

    The classification of protein folds is necessarily based on the structural elements that distinguish domains. Classification of protein domains consists of two problems: the partition of structures into domains and the classification of domains into sets of similar structures (or folds). Although similar topologies may arise by convergent evolution, the similarity of their respective folding pathways is unknown. The discovery and the characterization of the majority of protein folds will be followed by a similar enumeration of available protein folding pathways. Consequently, understanding the intricacies of structural domains is necessary to understanding their collective folding pathways. We review the current state of the art in the field of protein domain classification and discuss methods for the systematic and comprehensive study of protein folding across protein fold space via atomistic molecular dynamics simulation. Finally, we discuss our large-scale Dynameomics project, which includes simulations of representatives of all autonomous protein folds. PMID:21051320

  9. IBR5 Modulates Temperature-Dependent, R Protein CHS3-Mediated Defense Responses in Arabidopsis

    PubMed Central

    Bao, Fei; Ao, Kevin; Zhang, Xiaoyan; Zhang, Yuelin; Yang, Shuhua

    2015-01-01

    Plant responses to low temperature are tightly associated with defense responses. We previously characterized the chilling-sensitive mutant chs3-1 resulting from the activation of the Toll and interleukin 1 receptor-nucleotide binding-leucine-rich repeat (TIR-NB-LRR)-type resistance (R) protein harboring a C-terminal LIM (Lin-11, Isl-1 and Mec-3 domains) domain. Here we report the identification of a suppressor of chs3, ibr5-7 (indole-3-butyric acid response 5), which largely suppresses chilling-activated defense responses. IBR5 encodes a putative dual-specificity protein phosphatase. The accumulation of CHS3 protein at chilling temperatures is inhibited by the IBR5 mutation. Moreover, chs3-conferred defense phenotypes were synergistically suppressed by mutations in HSP90 and IBR5. Further analysis showed that IBR5, with holdase activity, physically associates with CHS3, HSP90 and SGT1b (Suppressor of the G2 allele of skp1) to form a complex that protects CHS3. In addition to the positive role of IBR5 in regulating CHS3, IBR5 is also involved in defense responses mediated by R genes, including SNC1 (Suppressor of npr1-1, Constitutive 1), RPS4 (Resistance to P. syringae 4) and RPM1 (Resistance to Pseudomonas syringae pv. maculicola 1). Thus, the results of the present study reveal a role for IBR5 in the regulation of multiple R protein-mediated defense responses. PMID:26451844

  10. Protein Dynamics

    NASA Astrophysics Data System (ADS)

    Frauenfelder, Hans

    2011-03-01

    Proteins combine properties of solids, liquids, and glasses. Schrödinger anticipated the main features of biomolecules long ago by stating that they had to be solid-like, but able to assume many different conformations. Indeed proteins can assume a gigantic number of conformational substates with the same primary sequence but different conformations. The different substates are described as craters in a very-high-dimensional energy landscape. The energy landscape is organized in a hierarchy of tiers, craters within craters within craters. Protein motions are pictured as transition between substates - jumps from crater to crater. Initially we assumed that these jumps were controlled by internal barriers between substates, but experiments have shown that nature selected a different approach. Proteins are surrounded by one to two layers of water and are embedded in a bulk solvent. Structural motions of the protein are controlled by the alpha fluctuations in the solvent surrounding the protein. Some internal motions most likely involving side chains are controlled electrostatically by beta fluctuations in the hydration shell. The dynamics of proteins is consequently dominated by the environment (H. Frauenfelder et al. PNAS 106, 5129 (2009). One can speculate that this organization permits exchange of information among biomolecules. The energy landscape is not just organized into two tiers, alpha and beta, but cryogenic experiments have revealed more tiers and protein more properties similar to that of glasses. While proteins function at ambient temperatures, cryogenic studies are necessary to understand the physics relevant for biology.

  11. Interfacial Protein-Protein Associations

    PubMed Central

    Langdon, Blake B.; Kastantin, Mark; Walder, Robert; Schwartz, Daniel K.

    2014-01-01

    While traditional models of protein adsorption focus primarily on direct protein-surface interactions, recent findings suggest that protein-protein interactions may play a central role. Using high-throughput intermolecular resonance energy transfer (RET) tracking, we directly observed dynamic, protein-protein associations of bovine serum albumin on poly(ethylene glycol) modified surfaces. The associations were heterogeneous and reversible, and associating molecules resided on the surface for longer times. The appearance of three distinct RET states suggested a spatially heterogeneous surface – with areas of high protein density (i.e. strongly-interacting clusters) coexisting with mobile monomers. Distinct association states exhibited characteristic behavior, i.e. partial-RET (monomer-monomer) associations were shorter-lived than complete-RET (protein-cluster) associations. While the fractional surface area covered by regions with high protein density (i.e. clusters) increased with increasing concentration, the distribution of contact times between monomers and clusters was independent of solution concentration, suggesting that associations were a local phenomenon, and independent of the global surface coverage. PMID:24274729

  12. Noc2, a putative zinc finger protein involved in exocytosis in endocrine cells.

    PubMed

    Kotake, K; Ozaki, N; Mizuta, M; Sekiya, S; Inagaki, N; Seino, S

    1997-11-21

    We have cloned a cDNA encoding a novel protein of 302 amino acids (designated Noc2, no C2 domain) that has 40.7% amino acid identity with and 77.9% similarity to the N-terminal region of rabphilin-3A, a target molecule of Rab3A. However, unlike rabphilin-3A, Noc2 lacks two C2 domains that are thought to interact with Ca2+ and phospholipids. Noc2 is expressed predominantly in endocrine tissues and hormone-secreting cell lines and at very low levels in brain. Immunoblot analysis of subcellular fractions of the insulin-secreting cell line MIN6 and immunocytochemistry reveal that Noc2 is a 38-kDa protein present in the cytoplasm. Overexpression of Noc2 in PC12 cells cotransfected with growth hormone enhances high K+-induced growth hormone secretion. Screening a mouse embryonic cDNA library with the yeast two-hybrid system shows that Noc2 interacts with the LIM domain-containing protein zyxin, a component of the cytoskeleton, and this interaction is further confirmed by the coimmunoprecipitation experiment. Accordingly, Noc2 is probably involved in regulated exocytosis in endocrine cells by interacting with the cytoskeleton. PMID:9367993

  13. Comparison of the Ventricle Muscle Proteome between Patients with Rheumatic Heart Disease and Controls with Mitral Valve Prolapse: HSP 60 May Be a Specific Protein in RHD

    PubMed Central

    Zheng, Dawei; Xu, Limin; Sun, Lebo; Feng, Qiang; Wang, Zishan; Shao, Guofeng; Ni, Yiming

    2014-01-01

    Objective. Rheumatic heart disease (RHD) is a serious autoimmune heart disease. The present study was aimed at identifying the differentially expressed proteins between patients with RHD and controls with mitral valve prolapse. Methods. Nine patients with RHD and nine controls with mitral valve prolapsed were enrolled for this study. Two-dimensional difference in-gel electrophoresis (2D-DIGE) and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) were performed. Results. A total of 39 protein spots with differential expressions were identified between the two groups (P < 0.05, Average Ratio > 1.2 or Average Ratio < −1.2) and four upregulated proteins (including heat shock protein 60 (HSP 60), desmin, PDZ and LIM domain protein 1, and proteasome subunit alpha type-1) and three downregulated proteins (including tropomyosin alpha-1 chain, malate dehydrogenase, and chaperone activity of bc1 complex homolog) were determined. Conclusion. These seven proteins, especially HSP 60, may serve as potential biomarkers for the diagnosis of RHD and provide evidence to explain the mechanisms of this complex disease in the future. PMID:24738046

  14. Whey Protein

    MedlinePlus

    ... intolerance, for replacing or supplementing milk-based infant formulas, and for reversing weight loss and increasing glutathione ( ... allergic reactions compared to infants who receive standard formula. However, taking why protein might not be helpful ...

  15. Extensive surface protein profiles of extracellular vesicles from cancer cells may provide diagnostic signatures from blood samples

    PubMed Central

    Belov, Larissa; Matic, Kieran J.; Hallal, Susannah; Best, O. Giles; Mulligan, Stephen P.; Christopherson, Richard I.

    2016-01-01

    Extracellular vesicles (EV) are membranous particles (30–1,000 nm in diameter) secreted by cells. Important biological functions have been attributed to 2 subsets of EV, the exosomes (bud from endosomal membranes) and the microvesicles (MV; bud from plasma membranes). Since both types of particles contain surface proteins derived from their cell of origin, their detection in blood may enable diagnosis and prognosis of disease. We have used an antibody microarray (DotScan) to compare the surface protein profiles of live cancer cells with those of their EV, based on their binding patterns to immobilized antibodies. Initially, EV derived from the cancer cell lines, LIM1215 (colorectal cancer) and MEC1 (B-cell chronic lymphocytic leukaemia; CLL), were used for assay optimization. Biotinylated antibodies specific for EpCAM (CD326) and CD19, respectively, were used to detect captured particles by enhanced chemiluminescence. Subsequently, this approach was used to profile CD19+ EV from the plasma of CLL patients. These EV expressed a subset (~40%) of the proteins detected on CLL cells from the same patients: moderate or high levels of CD5, CD19, CD31, CD44, CD55, CD62L, CD82, HLA-A,B,C, HLA-DR; low levels of CD21, CD49c, CD63. None of these proteins was detected on EV from the plasma of age- and gender-matched healthy individuals. PMID:27086589

  16. Extensive surface protein profiles of extracellular vesicles from cancer cells may provide diagnostic signatures from blood samples.

    PubMed

    Belov, Larissa; Matic, Kieran J; Hallal, Susannah; Best, O Giles; Mulligan, Stephen P; Christopherson, Richard I

    2016-01-01

    Extracellular vesicles (EV) are membranous particles (30-1,000 nm in diameter) secreted by cells. Important biological functions have been attributed to 2 subsets of EV, the exosomes (bud from endosomal membranes) and the microvesicles (MV; bud from plasma membranes). Since both types of particles contain surface proteins derived from their cell of origin, their detection in blood may enable diagnosis and prognosis of disease. We have used an antibody microarray (DotScan) to compare the surface protein profiles of live cancer cells with those of their EV, based on their binding patterns to immobilized antibodies. Initially, EV derived from the cancer cell lines, LIM1215 (colorectal cancer) and MEC1 (B-cell chronic lymphocytic leukaemia; CLL), were used for assay optimization. Biotinylated antibodies specific for EpCAM (CD326) and CD19, respectively, were used to detect captured particles by enhanced chemiluminescence. Subsequently, this approach was used to profile CD19(+) EV from the plasma of CLL patients. These EV expressed a subset (~40%) of the proteins detected on CLL cells from the same patients: moderate or high levels of CD5, CD19, CD31, CD44, CD55, CD62L, CD82, HLA-A,B,C, HLA-DR; low levels of CD21, CD49c, CD63. None of these proteins was detected on EV from the plasma of age- and gender-matched healthy individuals. PMID:27086589

  17. Designed protein-protein association.

    PubMed

    Grueninger, Dirk; Treiber, Nora; Ziegler, Mathias O P; Koetter, Jochen W A; Schulze, Monika-Sarah; Schulz, Georg E

    2008-01-11

    The analysis of natural contact interfaces between protein subunits and between proteins has disclosed some general rules governing their association. We have applied these rules to produce a number of novel assemblies, demonstrating that a given protein can be engineered to form contacts at various points of its surface. Symmetry plays an important role because it defines the multiplicity of a designed contact and therefore the number of required mutations. Some of the proteins needed only a single side-chain alteration in order to associate to a higher-order complex. The mobility of the buried side chains has to be taken into account. Four assemblies have been structurally elucidated. Comparisons between the designed contacts and the results will provide useful guidelines for the development of future architectures. PMID:18187656

  18. ZASP Interacts with the Mechanosensing Protein Ankrd2 and p53 in the Signalling Network of Striated Muscle

    PubMed Central

    Martinelli, Valentina C.; Kyle, W. Buck; Kojic, Snezana; Vitulo, Nicola; Li, Zhaohui; Belgrano, Anna; Maiuri, Paolo; Banks, Lawrence; Vatta, Matteo; Valle, Giorgio; Faulkner, Georgine

    2014-01-01

    ZASP is a cytoskeletal PDZ-LIM protein predominantly expressed in striated muscle. It forms multiprotein complexes and plays a pivotal role in the structural integrity of sarcomeres. Mutations in the ZASP protein are associated with myofibrillar myopathy, left ventricular non-compaction and dilated cardiomyopathy. The ablation of its murine homologue Cypher results in neonatal lethality. ZASP has several alternatively spliced isoforms, in this paper we clarify the nomenclature of its human isoforms as well as their dynamics and expression pattern in striated muscle. Interaction is demonstrated between ZASP and two new binding partners both of which have roles in signalling, regulation of gene expression and muscle differentiation; the mechanosensing protein Ankrd2 and the tumour suppressor protein p53. These proteins and ZASP form a triple complex that appears to facilitate poly-SUMOylation of p53. We also show the importance of two of its functional domains, the ZM-motif and the PDZ domain. The PDZ domain can bind directly to both Ankrd2 and p53 indicating that there is no competition between it and p53 for the same binding site on Ankrd2. However there is competition for this binding site between p53 and a region of the ZASP protein lacking the PDZ domain, but containing the ZM-motif. ZASP is negative regulator of p53 in transactivation experiments with the p53-responsive promoters, MDM2 and BAX. Mutations in the ZASP ZM-motif induce modification in protein turnover. In fact, two mutants, A165V and A171T, were not able to bind Ankrd2 and bound only poorly to alpha-actinin2. This is important since the A165V mutation is responsible for zaspopathy, a well characterized autosomal dominant distal myopathy. Although the mechanism by which this mutant causes disease is still unknown, this is the first indication of how a ZASP disease associated mutant protein differs from that of the wild type ZASP protein. PMID:24647531

  19. Protein Crystallization

    NASA Technical Reports Server (NTRS)

    Chernov, Alexander A.

    2005-01-01

    Nucleation, growth and perfection of protein crystals will be overviewed along with crystal mechanical properties. The knowledge is based on experiments using optical and force crystals behave similar to inorganic crystals, though with a difference in orders of magnitude in growing parameters. For example, the low incorporation rate of large biomolecules requires up to 100 times larger supersaturation to grow protein, rather than inorganic crystals. Nucleation is often poorly reproducible, partly because of turbulence accompanying the mixing of precipitant with protein solution. Light scattering reveals fluctuations of molecular cluster size, its growth, surface energies and increased clustering as protein ages. Growth most often occurs layer-by-layer resulting in faceted crystals. New molecular layer on crystal face is terminated by a step where molecular incorporation occurs. Quantitative data on the incorporation rate will be discussed. Rounded crystals with molecularly disordered interfaces will be explained. Defects in crystals compromise the x-ray diffraction resolution crucially needed to find the 3D atomic structure of biomolecules. The defects are immobile so that birth defects stay forever. All lattice defects known for inorganics are revealed in protein crystals. Contribution of molecular conformations to lattice disorder is important, but not studied. This contribution may be enhanced by stress field from other defects. Homologous impurities (e.g., dimers, acetylated molecules) are trapped more willingly by a growing crystal than foreign protein impurities. The trapped impurities induce internal stress eliminated in crystals exceeding a critical size (part of mni for ferritin, lysozyme). Lesser impurities are trapped from stagnant, as compared to the flowing, solution. Freezing may induce much more defects unless quickly amorphysizing intracrystalline water.

  20. Bacteriophage protein-protein interactions.

    PubMed

    Häuser, Roman; Blasche, Sonja; Dokland, Terje; Haggård-Ljungquist, Elisabeth; von Brunn, Albrecht; Salas, Margarita; Casjens, Sherwood; Molineux, Ian; Uetz, Peter

    2012-01-01

    Bacteriophages T7, λ, P22, and P2/P4 (from Escherichia coli), as well as ϕ29 (from Bacillus subtilis), are among the best-studied bacterial viruses. This chapter summarizes published protein interaction data of intraviral protein interactions, as well as known phage-host protein interactions of these phages retrieved from the literature. We also review the published results of comprehensive protein interaction analyses of Pneumococcus phages Dp-1 and Cp-1, as well as coliphages λ and T7. For example, the ≈55 proteins encoded by the T7 genome are connected by ≈43 interactions with another ≈15 between the phage and its host. The chapter compiles published interactions for the well-studied phages λ (33 intra-phage/22 phage-host), P22 (38/9), P2/P4 (14/3), and ϕ29 (20/2). We discuss whether different interaction patterns reflect different phage lifestyles or whether they may be artifacts of sampling. Phages that infect the same host can interact with different host target proteins, as exemplified by E. coli phage λ and T7. Despite decades of intensive investigation, only a fraction of these phage interactomes are known. Technical limitations and a lack of depth in many studies explain the gaps in our knowledge. Strategies to complete current interactome maps are described. Although limited space precludes detailed overviews of phage molecular biology, this compilation will allow future studies to put interaction data into the context of phage biology. PMID:22748812

  1. Recombinant protein production technology

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Recombinant protein production is an important technology for antibody production, biochemical activity study, and structural determination during the post-genomic era. Limiting factors in recombinant protein production include low-level protein expression, protein precipitation, and loss of protein...

  2. CPNA-1, a copine domain protein, is located at integrin adhesion sites and is required for myofilament stability in Caenorhabditis elegans.

    PubMed

    Warner, Adam; Xiong, Ge; Qadota, Hiroshi; Rogalski, Teresa; Vogl, A Wayne; Moerman, Donald G; Benian, Guy M

    2013-03-01

    We identify cpna-1 (F31D5.3) as a novel essential muscle gene in the nematode Caenorhabditis elegans. Antibodies specific to copine domain protein atypical-1 (CPNA-1), as well as a yellow fluorescent protein translational fusion, are localized to integrin attachment sites (M-lines and dense bodies) in the body-wall muscle of C. elegans. CPNA-1 contains an N-terminal predicted transmembrane domain and a C-terminal copine domain and binds to the M-line/dense body protein PAT-6 (actopaxin) and the M-line proteins UNC-89 (obscurin), LIM-9 (FHL), SCPL-1 (SCP), and UNC-96. Proper CPNA-1 localization is dependent upon PAT-6 in embryonic and adult muscle. Nematodes lacking cpna-1 arrest elongation at the twofold stage of embryogenesis and display disruption of the myofilament lattice. The thick-filament component myosin heavy chain MYO-3 and the M-line component UNC-89 are initially localized properly in cpna-1-null embryos. However, in these embryos, when contraction begins, MYO-3 and UNC-89 become mislocalized into large foci and animals die. We propose that CPNA-1 acts as a linker between an integrin-associated protein, PAT-6, and membrane-distal components of integrin adhesion complexes in the muscle of C. elegans. PMID:23283987

  3. Analysis of tarantula skeletal muscle protein sequences and identification of transcriptional isoforms

    PubMed Central

    Zhu, Jingui; Sun, Yongqiao; Zhao, Fa-Qing; Yu, Jun; Craig, Roger; Hu, Songnian

    2009-01-01

    Background Tarantula has been used as a model system for studying skeletal muscle structure and function, yet data on the genes expressed in tarantula muscle are lacking. Results We constructed a cDNA library from Aphonopelma sp. (Tarantula) skeletal muscle and got 2507 high-quality 5'ESTs (expressed sequence tags) from randomly picked clones. EST analysis showed 305 unigenes, among which 81 had more than 2 ESTs. Twenty abundant unigenes had matches to skeletal muscle-related genes including actin, myosin, tropomyosin, troponin-I, T and C, paramyosin, muscle LIM protein, muscle protein 20, a-actinin and tandem Ig/Fn motifs (found in giant sarcomere-related proteins). Matches to myosin light chain kinase and calponin were also identified. These results support the existence of both actin-linked and myosin-linked regulation in tarantula skeletal muscle. We have predicted full-length as well as partial cDNA sequences both experimentally and computationally for myosin heavy and light chains, actin, tropomyosin, and troponin-I, T and C, and have deduced the putative peptides. A preliminary analysis of the structural and functional properties was also carried out. Sequence similarities suggested multiple isoforms of most myofibrillar proteins, supporting the generality of multiple isoforms known from previous muscle sequence studies. This may be related to a mix of muscle fiber types. Conclusion The present study serves as a basis for defining the transcriptome of tarantula skeletal muscle, for future in vitro expression of tarantula proteins, and for interpreting structural and functional observations in this model species. PMID:19298669

  4. Protein inference: A protein quantification perspective.

    PubMed

    He, Zengyou; Huang, Ting; Liu, Xiaoqing; Zhu, Peijun; Teng, Ben; Deng, Shengchun

    2016-08-01

    In mass spectrometry-based shotgun proteomics, protein quantification and protein identification are two major computational problems. To quantify the protein abundance, a list of proteins must be firstly inferred from the raw data. Then the relative or absolute protein abundance is estimated with quantification methods, such as spectral counting. Until now, most researchers have been dealing with these two processes separately. In fact, the protein inference problem can be regarded as a special protein quantification problem in the sense that truly present proteins are those proteins whose abundance values are not zero. Some recent published papers have conceptually discussed this possibility. However, there is still a lack of rigorous experimental studies to test this hypothesis. In this paper, we investigate the feasibility of using protein quantification methods to solve the protein inference problem. Protein inference methods aim to determine whether each candidate protein is present in the sample or not. Protein quantification methods estimate the abundance value of each inferred protein. Naturally, the abundance value of an absent protein should be zero. Thus, we argue that the protein inference problem can be viewed as a special protein quantification problem in which one protein is considered to be present if its abundance is not zero. Based on this idea, our paper tries to use three simple protein quantification methods to solve the protein inference problem effectively. The experimental results on six data sets show that these three methods are competitive with previous protein inference algorithms. This demonstrates that it is plausible to model the protein inference problem as a special protein quantification task, which opens the door of devising more effective protein inference algorithms from a quantification perspective. The source codes of our methods are available at: http://code.google.com/p/protein-inference/. PMID:26935399

  5. Homologous Transcription Factors DUX4 and DUX4c Associate with Cytoplasmic Proteins during Muscle Differentiation

    PubMed Central

    Ansseau, Eugénie; Matteotti, Christel; Yip, Cassandre; Liu, Jian; Leroy, Baptiste; Hubeau, Céline; Gerbaux, Cécile; Cloet, Samuel; Wauters, Armelle; Zorbo, Sabrina; Meyer, Pierre; Pirson, Isabelle; Laoudj-Chenivesse, Dalila; Wattiez, Ruddy; Harper, Scott Q.; Belayew, Alexandra; Coppée, Frédérique

    2016-01-01

    Hundreds of double homeobox (DUX) genes map within 3.3-kb repeated elements dispersed in the human genome and encode DNA-binding proteins. Among these, we identified DUX4, a potent transcription factor that causes facioscapulohumeral muscular dystrophy (FSHD). In the present study, we performed yeast two-hybrid screens and protein co-purifications with HaloTag-DUX fusions or GST-DUX4 pull-down to identify protein partners of DUX4, DUX4c (which is identical to DUX4 except for the end of the carboxyl terminal domain) and DUX1 (which is limited to the double homeodomain). Unexpectedly, we identified and validated (by co-immunoprecipitation, GST pull-down, co-immunofluorescence and in situ Proximal Ligation Assay) the interaction of DUX4, DUX4c and DUX1 with type III intermediate filament protein desmin in the cytoplasm and at the nuclear periphery. Desmin filaments link adjacent sarcomere at the Z-discs, connect them to sarcolemma proteins and interact with mitochondria. These intermediate filament also contact the nuclear lamina and contribute to positioning of the nuclei. Another Z-disc protein, LMCD1 that contains a LIM domain was also validated as a DUX4 partner. The functionality of DUX4 or DUX4c interactions with cytoplasmic proteins is underscored by the cytoplasmic detection of DUX4/DUX4c upon myoblast fusion. In addition, we identified and validated (by co-immunoprecipitation, co-immunofluorescence and in situ Proximal Ligation Assay) as DUX4/4c partners several RNA-binding proteins such as C1QBP, SRSF9, RBM3, FUS/TLS and SFPQ that are involved in mRNA splicing and translation. FUS and SFPQ are nuclear proteins, however their cytoplasmic translocation was reported in neuronal cells where they associated with ribonucleoparticles (RNPs). Several other validated or identified DUX4/DUX4c partners are also contained in mRNP granules, and the co-localizations with cytoplasmic DAPI-positive spots is in keeping with such an association. Large muscle RNPs were

  6. Homologous Transcription Factors DUX4 and DUX4c Associate with Cytoplasmic Proteins during Muscle Differentiation.

    PubMed

    Ansseau, Eugénie; Eidahl, Jocelyn O; Lancelot, Céline; Tassin, Alexandra; Matteotti, Christel; Yip, Cassandre; Liu, Jian; Leroy, Baptiste; Hubeau, Céline; Gerbaux, Cécile; Cloet, Samuel; Wauters, Armelle; Zorbo, Sabrina; Meyer, Pierre; Pirson, Isabelle; Laoudj-Chenivesse, Dalila; Wattiez, Ruddy; Harper, Scott Q; Belayew, Alexandra; Coppée, Frédérique

    2016-01-01

    Hundreds of double homeobox (DUX) genes map within 3.3-kb repeated elements dispersed in the human genome and encode DNA-binding proteins. Among these, we identified DUX4, a potent transcription factor that causes facioscapulohumeral muscular dystrophy (FSHD). In the present study, we performed yeast two-hybrid screens and protein co-purifications with HaloTag-DUX fusions or GST-DUX4 pull-down to identify protein partners of DUX4, DUX4c (which is identical to DUX4 except for the end of the carboxyl terminal domain) and DUX1 (which is limited to the double homeodomain). Unexpectedly, we identified and validated (by co-immunoprecipitation, GST pull-down, co-immunofluorescence and in situ Proximal Ligation Assay) the interaction of DUX4, DUX4c and DUX1 with type III intermediate filament protein desmin in the cytoplasm and at the nuclear periphery. Desmin filaments link adjacent sarcomere at the Z-discs, connect them to sarcolemma proteins and interact with mitochondria. These intermediate filament also contact the nuclear lamina and contribute to positioning of the nuclei. Another Z-disc protein, LMCD1 that contains a LIM domain was also validated as a DUX4 partner. The functionality of DUX4 or DUX4c interactions with cytoplasmic proteins is underscored by the cytoplasmic detection of DUX4/DUX4c upon myoblast fusion. In addition, we identified and validated (by co-immunoprecipitation, co-immunofluorescence and in situ Proximal Ligation Assay) as DUX4/4c partners several RNA-binding proteins such as C1QBP, SRSF9, RBM3, FUS/TLS and SFPQ that are involved in mRNA splicing and translation. FUS and SFPQ are nuclear proteins, however their cytoplasmic translocation was reported in neuronal cells where they associated with ribonucleoparticles (RNPs). Several other validated or identified DUX4/DUX4c partners are also contained in mRNP granules, and the co-localizations with cytoplasmic DAPI-positive spots is in keeping with such an association. Large muscle RNPs were

  7. Interfacing protein lysine acetylation and protein phosphorylation

    PubMed Central

    Tran, Hue T.; Uhrig, R. Glen; Nimick, Mhairi; Moorhead, Greg B.

    2012-01-01

    Recognition that different protein covalent modifications can operate in concert to regulate a single protein has forced us to re-think the relationship between amino acid side chain modifications and protein function. Results presented by Tran et al. 2012 demonstrate the association of a protein phosphatase (PP2A) with a histone/lysine deacetylase (HDA14) on plant microtubules along with a histone/lysine acetyltransferase (ELP3). This finding reveals a regulatory interface between two prevalent covalent protein modifications, protein phosphorylation and acetylation, emphasizing the integrated complexity of post-translational protein regulation found in nature. PMID:22827947

  8. Comparative analysis of zinc finger proteins involved in plant disease resistance.

    PubMed

    Gupta, Santosh Kumar; Rai, Amit Kumar; Kanwar, Shamsher Singh; Sharma, Tilak R

    2012-01-01

    A meta-analysis was performed to understand the role of zinc finger domains in proteins of resistance (R) genes cloned from different crops. We analyzed protein sequences of seventy R genes of various crops in which twenty six proteins were found to have zinc finger domains along with nucleotide binding sites - leucine rice repeats (NBS-LRR) domains. We identified thirty four zinc finger domains in the R proteins of nine crops and were grouped into 19 types of zinc fingers. The size of individual zinc finger domain within the R genes varied from 11 to 84 amino acids, whereas the size of proteins containing these domains varied from 263 to 1305 amino acids. The biophysical analysis revealed that molecular weight of Pi54 zinc finger was lowest whereas the highest one was found in rice Pib zinc finger named as Transposes Transcription Factor (TTF). The instability (R(2) =0.95) and the aliphatic (R(2) =0.94) indices profile of zinc finger domains follows the polynomial distribution pattern. The pairwise identity analysis showed that the Lin11, Isl-1 & Mec-3 (LIM) zinc finger domain of rice blast resistance protein pi21 have 12.3% similarity with the nuclear transcription factor, X-box binding-like 1 (NFX) type zinc finger domain of Pi54 protein. For the first time, we reported that Pi54 (Pi-k(h)-Tetep), a rice blast resistance (R) protein have a small zinc finger domain of NFX type located on the C-terminal in between NBS and LRR domains of the R-protein. Compositional analysis depicted by the helical wheel diagram revealed the presence of a hydrophobic region within this domain which might help in exposing the LRR region for a possible R-Avr interaction. This domain is unique among all other cloned plant disease resistance genes and might play an important role in broad-spectrum nature of rice blast resistance gene Pi54. PMID:22916136

  9. Comparative Analysis of Zinc Finger Proteins Involved in Plant Disease Resistance

    PubMed Central

    Gupta, Santosh Kumar; Rai, Amit Kumar; Kanwar, Shamsher Singh; Sharma, Tilak R.

    2012-01-01

    A meta-analysis was performed to understand the role of zinc finger domains in proteins of resistance (R) genes cloned from different crops. We analyzed protein sequences of seventy R genes of various crops in which twenty six proteins were found to have zinc finger domains along with nucleotide binding sites - leucine rice repeats (NBS-LRR) domains. We identified thirty four zinc finger domains in the R proteins of nine crops and were grouped into 19 types of zinc fingers. The size of individual zinc finger domain within the R genes varied from 11 to 84 amino acids, whereas the size of proteins containing these domains varied from 263 to 1305 amino acids. The biophysical analysis revealed that molecular weight of Pi54 zinc finger was lowest whereas the highest one was found in rice Pib zinc finger named as Transposes Transcription Factor (TTF). The instability (R2 = 0.95) and the aliphatic (R2 = 0.94) indices profile of zinc finger domains follows the polynomial distribution pattern. The pairwise identity analysis showed that the Lin11, Isl-1 & Mec-3 (LIM) zinc finger domain of rice blast resistance protein pi21 have 12.3% similarity with the nuclear transcription factor, X-box binding-like 1 (NFX) type zinc finger domain of Pi54 protein. For the first time, we reported that Pi54 (Pi-kh-Tetep), a rice blast resistance (R) protein have a small zinc finger domain of NFX type located on the C-terminal in between NBS and LRR domains of the R-protein. Compositional analysis depicted by the helical wheel diagram revealed the presence of a hydrophobic region within this domain which might help in exposing the LRR region for a possible R-Avr interaction. This domain is unique among all other cloned plant disease resistance genes and might play an important role in broad-spectrum nature of rice blast resistance gene Pi54. PMID:22916136

  10. Actin turnover-dependent fast dissociation of capping protein in the dendritic nucleation actin network: evidence of frequent filament severing.

    PubMed

    Miyoshi, Takushi; Tsuji, Takahiro; Higashida, Chiharu; Hertzog, Maud; Fujita, Akiko; Narumiya, Shuh; Scita, Giorgio; Watanabe, Naoki

    2006-12-18

    Actin forms the dendritic nucleation network and undergoes rapid polymerization-depolymerization cycles in lamellipodia. To elucidate the mechanism of actin disassembly, we characterized molecular kinetics of the major filament end-binding proteins Arp2/3 complex and capping protein (CP) using single-molecule speckle microscopy. We have determined the dissociation rates of Arp2/3 and CP as 0.048 and 0.58 s(-1), respectively, in lamellipodia of live XTC fibroblasts. This CP dissociation rate is three orders of magnitude faster than in vitro. CP dissociates slower from actin stress fibers than from the lamellipodial actin network, suggesting that CP dissociation correlates with actin filament dynamics. We found that jasplakinolide, an actin depolymerization inhibitor, rapidly blocked the fast CP dissociation in cells. Consistently, the coexpression of LIM kinase prolonged CP speckle lifetime in lamellipodia. These results suggest that cofilin-mediated actin disassembly triggers CP dissociation from actin filaments. We predict that filament severing and end-to-end annealing might take place fairly frequently in the dendritic nucleation actin arrays. PMID:17178911

  11. Length, protein protein interactions, and complexity

    NASA Astrophysics Data System (ADS)

    Tan, Taison; Frenkel, Daan; Gupta, Vishal; Deem, Michael W.

    2005-05-01

    The evolutionary reason for the increase in gene length from archaea to prokaryotes to eukaryotes observed in large-scale genome sequencing efforts has been unclear. We propose here that the increasing complexity of protein-protein interactions has driven the selection of longer proteins, as they are more able to distinguish among a larger number of distinct interactions due to their greater average surface area. Annotated protein sequences available from the SWISS-PROT database were analyzed for 13 eukaryotes, eight bacteria, and two archaea species. The number of subcellular locations to which each protein is associated is used as a measure of the number of interactions to which a protein participates. Two databases of yeast protein-protein interactions were used as another measure of the number of interactions to which each S. cerevisiae protein participates. Protein length is shown to correlate with both number of subcellular locations to which a protein is associated and number of interactions as measured by yeast two-hybrid experiments. Protein length is also shown to correlate with the probability that the protein is encoded by an essential gene. Interestingly, average protein length and number of subcellular locations are not significantly different between all human proteins and protein targets of known, marketed drugs. Increased protein length appears to be a significant mechanism by which the increasing complexity of protein-protein interaction networks is accommodated within the natural evolution of species. Consideration of protein length may be a valuable tool in drug design, one that predicts different strategies for inhibiting interactions in aberrant and normal pathways.

  12. EDITORIAL: Precision proteins Precision proteins

    NASA Astrophysics Data System (ADS)

    Demming, Anna

    2010-06-01

    Since the birth of modern day medicine, during the times of Hippocrates in ancient Greece, the profession has developed from the rudimentary classification of disease into a rigorous science with an inspiring capability to treat and cure. Scientific methodology has distilled clinical diagnostic tools from the early arts of prognosis, which used to rely as much on revelation and prophecy, as intuition and judgement [1]. Over the past decade, research into the interactions between proteins and nanosystems has provided some ingenious and apt techniques for delving into the intricacies of anatomical systems. In vivo biosensing has emerged as a vibrant field of research, as much of medical diagnosis relies on the detection of substances or an imbalance in the chemicals in the body. The inherent properties of nanoscale structures, such as cantilevers, make them well suited to biosensing applications that demand the detection of molecules at very low concentrations. Measurable deflections in cantilevers functionalised with antibodies provide quantitative indicators of the presence of specific antigens when the two react. Such developments have roused mounting interest in the interactions of proteins with nanostructures, such as carbon nanotubes [3], which have demonstrated great potential as generic biomarkers. Plasmonic properties are also being exploited in sensing applications, such as the molecular sentinel recently devised by researchers in the US. The device uses the plasmonic properties of a silver nanoparticle linked to a Raman labelled hairpin DNA probe to signal changes in the probe geometry resulting from interactions with substances in the environment. Success stories so far include the detection of two specific genes associated with breast cancer [4]. A greater understanding of how RNA interference regulates gene expression has highlighted the potential of using this natural process as another agent for combating disease in personalized medicine. However, the

  13. EDITORIAL: Precision proteins Precision proteins

    NASA Astrophysics Data System (ADS)

    Demming, Anna

    2010-06-01

    Since the birth of modern day medicine, during the times of Hippocrates in ancient Greece, the profession has developed from the rudimentary classification of disease into a rigorous science with an inspiring capability to treat and cure. Scientific methodology has distilled clinical diagnostic tools from the early arts of prognosis, which used to rely as much on revelation and prophecy, as intuition and judgement [1]. Over the past decade, research into the interactions between proteins and nanosystems has provided some ingenious and apt techniques for delving into the intricacies of anatomical systems. In vivo biosensing has emerged as a vibrant field of research, as much of medical diagnosis relies on the detection of substances or an imbalance in the chemicals in the body. The inherent properties of nanoscale structures, such as cantilevers, make them well suited to biosensing applications that demand the detection of molecules at very low concentrations. Measurable deflections in cantilevers functionalised with antibodies provide quantitative indicators of the presence of specific antigens when the two react. Such developments have roused mounting interest in the interactions of proteins with nanostructures, such as carbon nanotubes [3], which have demonstrated great potential as generic biomarkers. Plasmonic properties are also being exploited in sensing applications, such as the molecular sentinel recently devised by researchers in the US. The device uses the plasmonic properties of a silver nanoparticle linked to a Raman labelled hairpin DNA probe to signal changes in the probe geometry resulting from interactions with substances in the environment. Success stories so far include the detection of two specific genes associated with breast cancer [4]. A greater understanding of how RNA interference regulates gene expression has highlighted the potential of using this natural process as another agent for combating disease in personalized medicine. However, the

  14. Shotgun protein sequencing.

    SciTech Connect

    Faulon, Jean-Loup Michel; Heffelfinger, Grant S.

    2009-06-01

    A novel experimental and computational technique based on multiple enzymatic digestion of a protein or protein mixture that reconstructs protein sequences from sequences of overlapping peptides is described in this SAND report. This approach, analogous to shotgun sequencing of DNA, is to be used to sequence alternative spliced proteins, to identify post-translational modifications, and to sequence genetically engineered proteins.

  15. Protein Crystal Based Nanomaterials

    NASA Technical Reports Server (NTRS)

    Bell, Jeffrey A.; VanRoey, Patrick

    2001-01-01

    This is the final report on a NASA Grant. It concerns a description of work done, which includes: (1) Protein crystals cross-linked to form fibers; (2) Engineering of protein to favor crystallization; (3) Better knowledge-based potentials for protein-protein contacts; (4) Simulation of protein crystallization.

  16. Protein folding, protein homeostasis, and cancer

    PubMed Central

    Van Drie, John H.

    2011-01-01

    Proteins fold into their functional 3-dimensional structures from a linear amino acid sequence. In vitro this process is spontaneous; while in vivo it is orchestrated by a specialized set of proteins, called chaperones. Protein folding is an ongoing cellular process, as cellular proteins constantly undergo synthesis and degradation. Here emerging links between this process and cancer are reviewed. This perspective both yields insights into the current struggle to develop novel cancer chemotherapeutics and has implications for future chemotherapy discovery. PMID:21272445

  17. Split-Protein Systems: Beyond Binary Protein-Protein Interactions

    PubMed Central

    Shekhawat, Sujan S.; Ghosh, Indraneel

    2011-01-01

    It has been estimated that 650,000 protein-protein interactions exist in the human interactome [1], a subset of all possible macromolecular partnerships that dictate life. Thus there is a continued need for the development of sensitive and user-friendly methods for cataloguing biomacromolecules in complex environments and for detecting their interactions, modifications, and cellular location. Such methods also allow for establishing differences in the interactome between a normal and diseased cellular state and for quantifying the outcome of therapeutic intervention. A promising approach for deconvoluting the role of macromolecular partnerships is split-protein reassembly, also called protein fragment complementation. This approach relies on the appropriate fragmentation of protein reporters, such as the green fluorescent protein or firefly luciferase, which when attached to possible interacting partners can reassemble and regain function, thereby confirming the partnership. Split-protein methods have been effectively utilized for detecting protein-protein interactions in cell-free systems, E. coli, yeast, mammalian cells, plants, and live animals. Herein, we present recent advances in engineering split-protein systems that allow for the rapid detection of ternary protein complexes, small molecule inhibitors, as well as a variety of macromolecules including nucleic acids, poly(ADP) ribose, and iron sulfur clusters. We also present advances that combine split-protein systems with chemical inducers of dimerization strategies that allow for regulating the activity of orthogonal split-proteases as well as aid in identifying enzyme inhibitors. Finally, we discuss autoinhibition strategies leading to turn-on sensors as well as future directions in split-protein methodology including possible therapeutic approaches. PMID:22070901

  18. Split-protein systems: beyond binary protein-protein interactions.

    PubMed

    Shekhawat, Sujan S; Ghosh, Indraneel

    2011-12-01

    It has been estimated that 650,000 protein-protein interactions exist in the human interactome (Stumpf et al., 2008), a subset of all possible macromolecular partnerships that dictate life. Thus there is a continued need for the development of sensitive and user-friendly methods for cataloguing biomacromolecules in complex environments and for detecting their interactions, modifications, and cellular location. Such methods also allow for establishing differences in the interactome between a normal and diseased cellular state and for quantifying the outcome of therapeutic intervention. A promising approach for deconvoluting the role of macromolecular partnerships is split-protein reassembly, also called protein fragment complementation. This approach relies on the appropriate fragmentation of protein reporters, such as the green fluorescent protein or firefly luciferase, which when attached to possible interacting partners can reassemble and regain function, thereby confirming the partnership. Split-protein methods have been effectively utilized for detecting protein-protein interactions in cell-free systems, Escherichia coli, yeast, mammalian cells, plants, and live animals. Herein, we present recent advances in engineering split-protein systems that allow for the rapid detection of ternary protein complexes, small molecule inhibitors, as well as a variety of macromolecules including nucleic acids, poly(ADP) ribose, and iron sulfur clusters. We also present advances that combine split-protein systems with chemical inducers of dimerization strategies that allow for regulating the activity of orthogonal split-proteases as well as aid in identifying enzyme inhibitors. Finally, we discuss autoinhibition strategies leading to turn-on sensors as well as future directions in split-protein methodology including possible therapeutic approaches. PMID:22070901

  19. Protein in diet

    MedlinePlus

    ... basic structure of protein is a chain of amino acids. You need protein in your diet to help ... Protein foods are broken down into parts called amino acids during digestion. The human body needs a number ...

  20. Protein-losing enteropathy

    MedlinePlus

    ... this page: //medlineplus.gov/ency/article/007338.htm Protein-losing enteropathy To use the sharing features on this page, please enable JavaScript. Protein-losing enteropathy is an abnormal loss of protein ...

  1. Protein electrophoresis - serum

    MedlinePlus

    ... digestive tract to absorb proteins ( protein-losing enteropathy ) Malnutrition Kidney disorder called nephrotic syndrome Scarring of the ... may indicate: Abnormally low level of LDL cholesterol Malnutrition Increased gamma globulin proteins may indicate: Bone marrow ...

  2. Domains mediate protein-protein interactions and nucleate protein assemblies.

    PubMed

    Costa, S; Cesareni, G

    2008-01-01

    Cell physiology is governed by an intricate mesh of physical and functional links among proteins, nucleic acids and other metabolites. The recent information flood coming from large-scale genomic and proteomic approaches allows us to foresee the possibility of compiling an exhaustive list of the molecules present within a cell, enriched with quantitative information on concentration and cellular localization. Moreover, several high-throughput experimental and computational techniques have been devised to map all the protein interactions occurring in a living cell. So far, such maps have been drawn as graphs where nodes represent proteins and edges represent interactions. However, this representation does not take into account the intrinsically modular nature of proteins and thus fails in providing an effective description of the determinants of binding. Since proteins are composed of domains that often confer on proteins their binding capabilities, a more informative description of the interaction network would detail, for each pair of interacting proteins in the network, which domains mediate the binding. Understanding how protein domains combine to mediate protein interactions would allow one to add important features to the protein interaction network, making it possible to discriminate between simultaneously occurring and mutually exclusive interactions. This objective can be achieved by experimentally characterizing domain recognition specificity or by analyzing the frequency of co-occurring domains in proteins that do interact. Such approaches allow gaining insights on the topology of complexes with unknown three-dimensional structure, thus opening the prospect of adopting a more rational strategy in developing drugs designed to selectively target specific protein interactions. PMID:18491061

  3. Drugging Membrane Protein Interactions.

    PubMed

    Yin, Hang; Flynn, Aaron D

    2016-07-11

    The majority of therapeutics target membrane proteins, accessible on the surface of cells, to alter cellular signaling. Cells use membrane proteins to transduce signals into cells, transport ions and molecules, bind cells to a surface or substrate, and catalyze reactions. Newly devised technologies allow us to drug conventionally "undruggable" regions of membrane proteins, enabling modulation of protein-protein, protein-lipid, and protein-nucleic acid interactions. In this review, we survey the state of the art of high-throughput screening and rational design in drug discovery, and we evaluate the advances in biological understanding and technological capacity that will drive pharmacotherapy forward against unorthodox membrane protein targets. PMID:26863923

  4. Protein sensing with engineered protein nanopores*

    PubMed Central

    Mohammad, Mohammad M.; Movileanu, Liviu

    2013-01-01

    The use of nanopores is a powerful new frontier in single-molecule sciences. Nanopores have been used effectively in exploring various biophysical features of small polypeptides and proteins, such as their folding state and structure, ligand interactions, and enzymatic activity. In particular, the α-hemolysin protein pore (αHL) has been used extensively for the detection, characterization and analysis of polypeptides, because this protein nanopore is highly robust, versatile and tractable under various experimental conditions. Inspired by the mechanisms of protein translocation across the outer membrane translocases of mitochondria, we have shown the ability to use nanopore-probe techniques in controlling a single protein using engineered αHL pores. Here, we provide a detailed protocol for the preparation of αHL protein nanopores. Moreover, we demonstrate that placing attractive electrostatic traps is instrumental in tackling single-molecule stochastic sensing of folded proteins. PMID:22528256

  5. Nanotechnologies in protein microarrays.

    PubMed

    Krizkova, Sona; Heger, Zbynek; Zalewska, Marta; Moulick, Amitava; Adam, Vojtech; Kizek, Rene

    2015-01-01

    Protein microarray technology became an important research tool for study and detection of proteins, protein-protein interactions and a number of other applications. The utilization of nanoparticle-based materials and nanotechnology-based techniques for immobilization allows us not only to extend the surface for biomolecule immobilization resulting in enhanced substrate binding properties, decreased background signals and enhanced reporter systems for more sensitive assays. Generally in contemporarily developed microarray systems, multiple nanotechnology-based techniques are combined. In this review, applications of nanoparticles and nanotechnologies in creating protein microarrays, proteins immobilization and detection are summarized. We anticipate that advanced nanotechnologies can be exploited to expand promising fields of proteins identification, monitoring of protein-protein or drug-protein interactions, or proteins structures. PMID:26039143

  6. Detoxified lipooligosaccharide from nontypeable Haemophilus influenzae conjugated to proteins confers protection against otitis media in chinchillas.

    PubMed Central

    Gu, X X; Sun, J; Jin, S; Barenkamp, S J; Lim, D J; Robbins, J B; Battey, J

    1997-01-01

    Detoxified-lipooligosaccharide (dLOS)-protein conjugates from nontypeable Haemophilus influenzae (NTHi) elicited a significant rise of anti-LOS antibodies with bactericidal activity in rabbits (X.-X. Gu, C.-M. Tsai, T. Ueyama, S. J. Barenkamp, J. B. Robbins, and D. J. Lim, Infect. Immun. 64:4047-4053, 1996). In this study, we evaluated whether vaccination with the conjugates would protect against NTHi otitis media in chinchillas. Fifty-eight chinchillas received three subcutaneous or intramuscular injections of dLOS-conjugated tetanus toxoid, dLOS-conjugated high-molecular-weight proteins from NTHi, or saline (control) in Freund's adjuvant and then were challenged by intrabullar inoculation with 140 CFU of NTHi. All vaccinated animals responded with elevated serum titers of anti-LOS antibody, and 49% (19 of 39) demonstrated bactericidal activity against the homologous strain. Otitis media with culture-positive NTHi effusions developed in all 19 controls and 56% (22 of 39) of the vaccinated animals during a period of 21 days (P < 0.001). Bacterial counts of the middle ear effusions were lower in the vaccine groups than in the controls (P < 0.01). The incidences of infection in the unchallenged ear or inner ear were 26 or 28% in the vaccine groups and 53 or 58% in the controls (P < 0.05). The signs of infection observed by otoscopy were less severe in the vaccine groups than in the controls. There was no significant difference between the two vaccine groups. These data indicate that active immunization with LOS-based conjugates reduces the incidence of NTHi-induced otitis media. PMID:9353024

  7. PREFACE: Protein protein interactions: principles and predictions

    NASA Astrophysics Data System (ADS)

    Nussinov, Ruth; Tsai, Chung-Jung

    2005-06-01

    Proteins are the `workhorses' of the cell. Their roles span functions as diverse as being molecular machines and signalling. They carry out catalytic reactions, transport, form viral capsids, traverse membranes and form regulated channels, transmit information from DNA to RNA, making possible the synthesis of new proteins, and they are responsible for the degradation of unnecessary proteins and nucleic acids. They are the vehicles of the immune response and are responsible for viral entry into the cell. Given their importance, considerable effort has been centered on the prediction of protein function. A prime way to do this is through identification of binding partners. If the function of at least one of the components with which the protein interacts is known, that should let us assign its function(s) and the pathway(s) in which it plays a role. This holds since the vast majority of their chores in the living cell involve protein-protein interactions. Hence, through the intricate network of these interactions we can map cellular pathways, their interconnectivities and their dynamic regulation. Their identification is at the heart of functional genomics; their prediction is crucial for drug discovery. Knowledge of the pathway, its topology, length, and dynamics may provide useful information for forecasting side effects. The goal of predicting protein-protein interactions is daunting. Some associations are obligatory, others are continuously forming and dissociating. In principle, from the physical standpoint, any two proteins can interact, but under what conditions and at which strength? The principles of protein-protein interactions are general: the non-covalent interactions of two proteins are largely the outcome of the hydrophobic effect, which drives the interactions. In addition, hydrogen bonds and electrostatic interactions play important roles. Thus, many of the interactions observed in vitro are the outcome of experimental overexpression. Protein disorder

  8. Protein sequence comparison and protein evolution

    SciTech Connect

    Pearson, W.R.

    1995-12-31

    This tutorial was one of eight tutorials selected to be presented at the Third International Conference on Intelligent Systems for Molecular Biology which was held in the United Kingdom from July 16 to 19, 1995. This tutorial examines how the information conserved during the evolution of a protein molecule can be used to infer reliably homology, and thus a shared proteinfold and possibly a shared active site or function. The authors start by reviewing a geological/evolutionary time scale. Next they look at the evolution of several protein families. During the tutorial, these families will be used to demonstrate that homologous protein ancestry can be inferred with confidence. They also examine different modes of protein evolution and consider some hypotheses that have been presented to explain the very earliest events in protein evolution. The next part of the tutorial will examine the technical aspects of protein sequence comparison. Both optimal and heuristic algorithms and their associated parameters that are used to characterize protein sequence similarities are discussed. Perhaps more importantly, they survey the statistics of local similarity scores, and how these statistics can both be used to improve the selectivity of a search and to evaluate the significance of a match. They them examine distantly related members of three protein families, the serine proteases, the glutathione transferases, and the G-protein-coupled receptors (GCRs). Finally, the discuss how sequence similarity can be used to examine internal repeated or mosaic structures in proteins.

  9. Sorghum and millet proteins

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Sorghum and millet proteins are an important source of dietary protein for significant numbers of people living throughout Africa and parts of Asia. Compared to other food proteins, such as those found in milk, eggs and wheat, little is known about the functionality of sorghum and millet proteins. ...

  10. Whey protein fractionation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Concentrated whey protein products from cheese whey, such as whey protein concentrate (WPC) and whey protein isolate (WPI), contain more than seven different types of proteins: alpha-lactalbumin (alpha-LA), beta-lactoglobulin (beta-LG), bovine serum albumin (BSA), immunoglobulins (Igs), lactoferrin ...

  11. Protein in diet

    MedlinePlus

    ... protein. The basic structure of protein is a chain of amino acids. You need protein in your diet to help your body repair cells and make new ones. Protein is also important for growth and development in children, teens, and pregnant women.

  12. Techniques in protein methylation.

    PubMed

    Lee, Jaeho; Cheng, Donghang; Bedford, Mark T

    2004-01-01

    Proteins can be methylated on the side-chain nitrogens of arginine and lysine residues or on carboxy-termini. Protein methylation is a way of subtly changing the primary sequence of a peptide so that it can encode more information. This common posttranslational modification is implicated in the regulation of a variety of processes including protein trafficking, transcription and protein-protein interactions. In this chapter, we will use the arginine methyltransferases to illustrate different approaches that have been developed to assess protein methylation. Both in vivo and in vitro methylation techniques are described, and the use of small molecule inhibitors of protein methylation will be demonstrated. PMID:15173617

  13. Biochemical Approaches for Discovering Protein-Protein Interactions

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Protein-protein interactions or protein complexes are indigenous to nearly all cellular processes, ranging from metabolism to structure. Elucidating both individual protein associations and complex protein interaction networks, while challenging, is an essential goal of functional genomics. For ex...

  14. Urine Protein and Urine Protein to Creatinine Ratio

    MedlinePlus

    ... limited. Home Visit Global Sites Search Help? Urine Protein and Urine Protein to Creatinine Ratio Share this page: Was this page helpful? Also known as: 24-Hour Urine Protein; Urine Total Protein; Urine Protein to Creatinine Ratio; ...

  15. [Protein expression and purification].

    PubMed

    Růčková, E; Müller, P; Vojtěšek, B

    2014-01-01

    Production of recombinant proteins is essential for many applications in both basic research and also in medicine, where recombinant proteins are used as pharmaceuticals. This review summarizes procedures involved in recombinant protein expression and purification, including molecular cloning of target genes into expression vectors, selection of the appropriate expression system, and protein purification techniques. Recombinant DNA technology allows protein engineering to modify protein stability, activity and function or to facilitate protein purification by affinity tag fusions. A wide range of cloning systems enabling fast and effective design of expression vectors is currently available. A first choice of protein expression system is usually the bacteria Escherichia coli. The main advantages of this prokaryotic expression system are low cost and simplicity; on the other hand this system is often unsuitable for production of complex mammalian proteins. Protein expression mediated by eukaryotic cells (yeast, insect and mammalian cells) usually produces properly folded and posttranslationally modified proteins. How-ever, cultivation of insect and, especially, mammalian cells is time consuming and expensive. Affinity tagged recombinant proteins are purified efficiently using affinity chromatography. An affinity tag is a protein or peptide that mediates specific binding to a chromatography column, unbound proteins are removed during a washing step and pure protein is subsequently eluted. PMID:24945544

  16. Protein- protein interaction detection system using fluorescent protein microdomains

    DOEpatents

    Waldo, Geoffrey S.; Cabantous, Stephanie

    2010-02-23

    The invention provides a protein labeling and interaction detection system based on engineered fragments of fluorescent and chromophoric proteins that require fused interacting polypeptides to drive the association of the fragments, and further are soluble and stable, and do not change the solubility of polypeptides to which they are fused. In one embodiment, a test protein X is fused to a sixteen amino acid fragment of GFP (.beta.-strand 10, amino acids 198-214), engineered to not perturb fusion protein solubility. A second test protein Y is fused to a sixteen amino acid fragment of GFP (.beta.-strand 11, amino acids 215-230), engineered to not perturb fusion protein solubility. When X and Y interact, they bring the GFP strands into proximity, and are detected by complementation with a third GFP fragment consisting of GFP amino acids 1-198 (strands 1-9). When GFP strands 10 and 11 are held together by interaction of protein X and Y, they spontaneous association with GFP strands 1-9, resulting in structural complementation, folding, and concomitant GFP fluorescence.

  17. Designing Fluorinated Proteins.

    PubMed

    Marsh, E N G

    2016-01-01

    As methods to incorporate noncanonical amino acid residues into proteins have become more powerful, interest in their use to modify the physical and biological properties of proteins and enzymes has increased. This chapter discusses the use of highly fluorinated analogs of hydrophobic amino acids, for example, hexafluoroleucine, in protein design. In particular, fluorinated residues have proven to be generally effective in increasing the thermodynamic stability of proteins. The chapter provides an overview of the different fluorinated amino acids that have been used in protein design and the various methods available for producing fluorinated proteins. It discusses model proteins systems into which highly fluorinated amino acids have been introduced and the reasons why fluorinated residues are generally stabilizing, with particular reference to thermodynamic and structural studies from our laboratory. Lastly, details of the methodology we have developed to measure the thermodynamic stability of oligomeric fluorinated proteins are presented, as this may be generally applicable to many proteins. PMID:27586337

  18. Surface Mediated Protein Disaggregation

    NASA Astrophysics Data System (ADS)

    Radhakrishna, Mithun; Kumar, Sanat K.

    2014-03-01

    Preventing protein aggregation is of both biological and industrial importance. Biologically these aggregates are known to cause amyloid type diseases like Alzheimer's and Parkinson's disease. Protein aggregation leads to reduced activity of the enzymes in industrial applications. Inter-protein interactions between the hydrophobic residues of the protein are known to be the major driving force for protein aggregation. In the current paper we show how surface chemistry and curvature can be tuned to mitigate these inter-protein interactions. Our results calculated in the framework of the Hydrophobic-Polar (HP) lattice model show that, inter-protein interactions can be drastically reduced by increasing the surface hydrophobicity to a critical value corresponding to the adsorption transition of the protein. At this value of surface hydrophobicity, proteins lose inter-protein contacts to gain surface contacts and thus the surface helps in reducing the inter-protein interactions. Further, we show that the adsorption of the proteins inside hydrophobic pores of optimal sizes are most efficient both in reducing inter-protein contacts and simultaneously retaining most of the native-contacts due to strong protein-surface interactions coupled with stabilization due to the confinement. Department of Energy (Grant No DE-FG02-11ER46811).

  19. PINT: Protein-protein Interactions Thermodynamic Database.

    PubMed

    Kumar, M D Shaji; Gromiha, M Michael

    2006-01-01

    The first release of Protein-protein Interactions Thermodynamic Database (PINT) contains >1500 data of several thermodynamic parameters along with sequence and structural information, experimental conditions and literature information. Each entry contains numerical data for the free energy change, dissociation constant, association constant, enthalpy change, heat capacity change and so on of the interacting proteins upon binding, which are important for understanding the mechanism of protein-protein interactions. PINT also includes the name and source of the proteins involved in binding, their Protein Information Resource, SWISS-PROT and Protein Data Bank (PDB) codes, secondary structure and solvent accessibility of residues at mutant positions, measuring methods, experimental conditions, such as buffers, ions and additives, and literature information. A WWW interface facilitates users to search data based on various conditions, feasibility to select the terms for output and different sorting options. Further, PINT is cross-linked with other related databases, PIR, SWISS-PROT, PDB and NCBI PUBMED literature database. The database is freely available at http://www.bioinfodatabase.com/pint/index.html. PMID:16381844

  20. Rho-associated protein kinase 1 (ROCK1) is increased in Alzheimer's disease and ROCK1 depletion reduces amyloid-β levels in brain.

    PubMed

    Henderson, Benjamin W; Gentry, Erik G; Rush, Travis; Troncoso, Juan C; Thambisetty, Madhav; Montine, Thomas J; Herskowitz, Jeremy H

    2016-08-01

    Alzheimer's disease (AD) is the leading cause of dementia and mitigating amyloid-β (Aβ) levels may serve as a rational therapeutic avenue to slow AD progression. Pharmacologic inhibition of the Rho-associated protein kinases (ROCK1 and ROCK2) is proposed to curb Aβ levels, and mechanisms that underlie ROCK2's effects on Aβ production are defined. How ROCK1 affects Aβ generation remains a critical barrier. Here, we report that ROCK1 protein levels were elevated in mild cognitive impairment due to AD (MCI) and AD brains compared to controls. Aβ42 oligomers marginally increased ROCK1 and ROCK2 protein levels in neurons but strongly induced phosphorylation of Lim kinase 1 (LIMK1), suggesting that Aβ42 activates ROCKs. RNAi depletion of ROCK1 or ROCK2 suppressed endogenous Aβ40 production in neurons, and Aβ40 levels were reduced in brains of ROCK1 heterozygous knock-out mice compared to wild-type littermate controls. ROCK1 knockdown decreased amyloid precursor protein (APP), and treatment with bafilomycin accumulated APP levels in neurons depleted of ROCK1. These observations suggest that reduction of ROCK1 diminishes Aβ levels by enhancing APP protein degradation. Collectively, these findings support the hypothesis that both ROCK1 and ROCK2 are therapeutic targets to combat Aβ production in AD. Mitigating amyloid-β (Aβ) levels is a rational strategy for Alzheimer's disease (AD) treatment, however, therapeutic targets with clinically available drugs are lacking. We hypothesize that Aβ accumulation in mild cognitive impairment because of AD (MCI) and AD activates the RhoA/ROCK pathway which in turn fuels production of Aβ. Escalation of this cycle over the course of many years may contribute to the buildup of amyloid pathology in MCI and/or AD. PMID:27246255

  1. LIM Homeobox Domain 2 Is Required for Corneal Epithelial Homeostasis

    PubMed Central

    Sartaj, Rachel; Chee, Ru‐ik; Yang, Jing; Wan, Pengxia; Liu, Aihong; Guaiquil, Victor; Fuchs, Elaine

    2016-01-01

    Abstract The cornea requires constant epithelial renewal to maintain clarity for appropriate vision. A subset of stem cells residing at the limbus is primarily responsible for maintaining corneal epithelium homeostasis. Trauma and disease may lead to stem cell deficiency and therapeutic targeting to replenish the stemness capacity has been stalled by the lack of reliable corneal epithelial stem cell markers. Here we identified the location of Lhx2 in mice (mLhx2) cornea and conjunctival tissue using an Lhx2eGFP reporter model and in human tissues (hLHX2). Lhx2 localized to the basal cells of central cornea, the conjunctiva and the entire limbal epithelium in humans and mice. To ascribe a functional role we generated Lhx2 conditional knockout (cKO) mice and the phenotypic effects in corneas were analyzed by slit lamp microscopy, in cell‐based assays and in a model of corneal epithelium debridement. Immunodetection on corneal sections were used to visualize conjunctivalization, a sign of limbal barrier failure. Lhx2cKO mice produced reduced body hair and spontaneous epithelial defects in the cornea that included neovascularization, perforation with formation of scar tissue and opacification. Cell based assays showed that Lhx2cKO derived corneal epithelial cells have a significantly lower capacity to form colonies over time and delayed wound‐healing recovery when compared to wildtype cells. Repeated corneal epithelial wounding resulted in decreased re‐epithelialization and multiple cornea lesions in Lhx2cKO mice compared to normal recovery seen in wildtype mice. We conclude that Lhx2 is required for maintenance of the corneal epithelial cell compartment and the limbal barrier. Stem Cells 2016;34:493–503 PMID:26661907

  2. LIM Homeobox Domain 2 Is Required for Corneal Epithelial Homeostasis.

    PubMed

    Sartaj, Rachel; Chee, Ru-ik; Yang, Jing; Wan, Pengxia; Liu, Aihong; Guaiquil, Victor; Fuchs, Elaine; Rosenblatt, Mark I

    2016-02-01

    The cornea requires constant epithelial renewal to maintain clarity for appropriate vision. A subset of stem cells residing at the limbus is primarily responsible for maintaining corneal epithelium homeostasis. Trauma and disease may lead to stem cell deficiency and therapeutic targeting to replenish the stemness capacity has been stalled by the lack of reliable corneal epithelial stem cell markers. Here we identified the location of Lhx2 in mice (mLhx2) cornea and conjunctival tissue using an Lhx2eGFP reporter model and in human tissues (hLHX2). Lhx2 localized to the basal cells of central cornea, the conjunctiva and the entire limbal epithelium in humans and mice. To ascribe a functional role we generated Lhx2 conditional knockout (cKO) mice and the phenotypic effects in corneas were analyzed by slit lamp microscopy, in cell-based assays and in a model of corneal epithelium debridement. Immunodetection on corneal sections were used to visualize conjunctivalization, a sign of limbal barrier failure. Lhx2cKO mice produced reduced body hair and spontaneous epithelial defects in the cornea that included neovascularization, perforation with formation of scar tissue and opacification. Cell based assays showed that Lhx2cKO derived corneal epithelial cells have a significantly lower capacity to form colonies over time and delayed wound-healing recovery when compared to wildtype cells. Repeated corneal epithelial wounding resulted in decreased re-epithelialization and multiple cornea lesions in Lhx2cKO mice compared to normal recovery seen in wildtype mice. We conclude that Lhx2 is required for maintenance of the corneal epithelial cell compartment and the limbal barrier. PMID:26661907

  3. The Limb Infrared Monitor of the Stratosphere (LIMS) experiment

    NASA Technical Reports Server (NTRS)

    Russell, J. M.; Gille, J. C.

    1978-01-01

    The Limb Infrared Monitor of the Stratosphere is used to obtain vertical profiles and maps of temperature and the concentration of ozone, water vapor, nitrogen dioxide, and nitric acid for the region of the stratosphere bounded by the upper troposphere and the lower mesosphere.

  4. Transport of trace species deduced from LIMS observations

    NASA Technical Reports Server (NTRS)

    Gille, J. C.; Lyjak, L. V.; Smith, A. K.; Bailey, P. L.; Massie, S. T.; Smythe, C. M.

    1985-01-01

    One of the critical problems in the middle atmosphere at this time is understanding the interaction between atmospheric motions and photochemistry, and the way in which these two processes maintain the distributions of photochemically active species such as ozone. Recent work has shown that a great deal can be learned by considering the zonally averaged distributions and transports, i.e., working in the latitude-altitude plane. This 2-dimensional approach has been used in the past in Eulerian formation. Recent work (DUNKERTON, 1978) has shown that formulating the transport problems in terms of the residual mean circulation, which has similarity to the Lagrangian mean motion, results in a more physical picture of the way in which the transport takes place. Here, data from the Limb Infrared Monitor of the Stratosphere (LIMB) experiment are used to investigate how well the residual mean formulation works with real data, in the sense of asking how well the predicted ozone changes match those in regions of the lower stratosphere in which chemical changes may be neglected. This allows a test of the approximations that are made, and of the strengths and weaknesses of this approach to transport calculation.

  5. Miniature LIMS System for In Situ Detection of Biosignatures

    NASA Astrophysics Data System (ADS)

    Riedo, A.; Tulej, M.; Neuland, M. B.; Wurz, P.

    2016-05-01

    The current measurement capabilities of our miniature Laser Ablation Ionization Mass Spectrometer for sensitive and quantitative in situ chemical analyses (element, isotope and molecular) of solids on planetary surfaces will be presented.

  6. DNA mimicry by proteins.

    PubMed

    Dryden, D T F; Tock, M R

    2006-04-01

    It has been discovered recently, via structural and biophysical analyses, that proteins can mimic DNA structures in order to inhibit proteins that would normally bind to DNA. Mimicry of the phosphate backbone of DNA, the hydrogen-bonding properties of the nucleotide bases and the bending and twisting of the DNA double helix are all present in the mimics discovered to date. These mimics target a range of proteins and enzymes such as DNA restriction enzymes, DNA repair enzymes, DNA gyrase and nucleosomal and nucleoid-associated proteins. The unusual properties of these protein DNA mimics may provide a foundation for the design of targeted inhibitors of DNA-binding proteins. PMID:16545103

  7. Development and optimization of a cell-based assay for the selection of synthetic compounds that potentiate bone morphogenetic protein-2 activity.

    PubMed

    Okada, Motohiro; Sangadala, Sreedhara; Liu, Yunshan; Yoshida, Munehito; Reddy, Boojala Vijay B; Titus, Louisa; Boden, Scott D

    2009-12-01

    The requirement of large amounts of the recombinant human bone morphogenetic protein-2 (BMP-2) produces a huge translational barrier for its routine clinical use due to high cost. This leads to an urgent need to develop alternative methods to lower costs and/or increase efficacies for using BMP-2. In this study, we describe the development and optimization of a cell-based assay that is sensitive, reproducible, and reliable in identifying reagents that potentiate the effects of BMP-2 in inducing transdifferentiation of C2C12 myoblasts into the osteoblastic phenotype. The assay is based on a BMP-responsive Smad1-driven luciferase reporter gene. LIM mineralization protein-1 (LMP-1) is a novel intracellular LIM domain protein that has been shown by our group to enhance cellular responsiveness to BMP-2. Our previous report elucidated that the binding of LMP-1 with the WW2 domain in Smad ubiquitin regulatory factor-1 (Smurf1) rescues the osteogenic Smads from degradation. Here, using the optimized cell-based assay, we first evaluated the activity of the recombinantly prepared proteins, LMP-1, and its mutant (LMP-1DeltaSmurf1) that lacks the Smurf1-WW2 domain-binding motif. Both the wild type and the mutant proteins were engineered to contain an 11-amino acid HIV-TAT protein derived membrane transduction domain to aid the cellular delivery of recombinant proteins. The cell-based reporter assay confirmed that LMP-1 potentiates the BMP-induced stimulation of C2C12 cells towards the osteoblastic phenotype. The potentiating effect of LMP-1 was significantly reduced when a specific-motif known to interact with Smurf1 was mutated. We validated the results obtained in the reporter assay by also monitoring the expression of mRNA for osteocalcin and alkaline phosphatase (ALP) which is widely accepted osteoblast differentiation marker genes. Finally, we provide further confirmation of our results by measuring the activity of alkaline phosphatase in support of the accuracy and

  8. Physics of protein motility and motor proteins

    NASA Astrophysics Data System (ADS)

    Kolomeisky, Anatoly B.

    2013-09-01

    Motor proteins are enzymatic molecules that transform chemical energy into mechanical motion and work. They are critically important for supporting various cellular activities and functions. In the last 15 years significant progress in understanding the functioning of motor proteins has been achieved due to revolutionary breakthroughs in single-molecule experimental techniques and strong advances in theoretical modelling. However, microscopic mechanisms of protein motility are still not well explained, and the collective efforts of many scientists are needed in order to solve these complex problems. In this special section the reader will find the latest advances on the difficult road to mapping motor proteins dynamics in various systems. Recent experimental developments have allowed researchers to monitor and to influence the activity of single motor proteins with a high spatial and temporal resolution. It has stimulated significant theoretical efforts to understand the non-equilibrium nature of protein motility phenomena. The latest results from all these advances are presented and discussed in this special section. We would like to thank the scientists from all over the world who have reported their latest research results for this special section. We are also grateful to the staff and editors of Journal of Physics: Condensed Matter for their invaluable help in handling all the administrative and refereeing activities. The field of motor proteins and protein motility is fast moving, and we hope that this collection of articles will be a useful source of information in this highly interdisciplinary area. Physics of protein motility and motor proteins contents Physics of protein motility and motor proteinsAnatoly B Kolomeisky Identification of unique interactions between the flexible linker and the RecA-like domains of DEAD-box helicase Mss116 Yuan Zhang, Mirkó Palla, Andrew Sun and Jung-Chi Liao The load dependence of the physical properties of a molecular motor

  9. Protein C blood test

    MedlinePlus

    ... a normal substance in the body that prevents blood clotting. A blood test can be done to see ... history of blood clots. Protein C helps control blood clotting. A lack of this protein or problem with ...

  10. Protein S blood test

    MedlinePlus

    ... a normal substance in your body that prevents blood clotting. A blood test can be done to see ... family history of blood clots. Protein S helps control blood clotting. A lack of this protein or problem with ...

  11. Protein electrophoresis - urine

    MedlinePlus

    ... nephropathy Kidney failure Multiple myeloma Nephrotic syndrome Acute urinary tract infection Risks There are no risks associated with this ... Primary amyloidosis Protein in diet Protein urine test Urinary tract infection - adults Update Date 5/29/2014 Updated by: ...

  12. [Protein-losing enteropathy].

    PubMed

    Amiot, A

    2015-07-01

    Protein-losing enteropathy is a rare syndrome of gastrointestinal protein loss. The primary causes can be classified into lymphatic leakage due to increased interstitial pressure and increased leakage of protein-rich fluids due to erosive or non-erosive gastrointestinal disorders. The diagnosis of protein-losing enteropathy should be considered in patients with chronic diarrhea and peripheral oedema. The diagnosis of protein-losing enteropathy is most commonly based on the determination of fecal alpha-1 antitrypsin clearance. Most protein-losing enteropathy cases are the result of either lymphatic obstruction or a variety of gastrointestinal disorders and cardiac diseases, while primary intestinal lymphangiectasia (Waldmann's disease) is less common. Treatment of protein-losing enteropathy targets the underlying disease but also includes dietary modification, such as high-protein and low-fat diet along with medium-chain triglyceride supplementation. PMID:25618488

  13. Learning about Proteins

    MedlinePlus

    ... body, and protecting you from disease. All About Amino Acids When you eat foods that contain protein, the ... called amino (say: uh-MEE-no) acids. The amino acids then can be reused to make the proteins ...

  14. Hydrodynamic effects in proteins

    NASA Astrophysics Data System (ADS)

    Szymczak, Piotr; Cieplak, Marek

    2011-01-01

    Experimental and numerical results pertaining to flow-induced effects in proteins are reviewed. Special emphasis is placed on shear-induced unfolding and on the role of solvent mediated hydrodynamic interactions in the conformational transitions in proteins.

  15. Hydrodynamic effects in proteins.

    PubMed

    Szymczak, Piotr; Cieplak, Marek

    2011-01-26

    Experimental and numerical results pertaining to flow-induced effects in proteins are reviewed. Special emphasis is placed on shear-induced unfolding and on the role of solvent mediated hydrodynamic interactions in the conformational transitions in proteins. PMID:21406855

  16. Understanding protein folding: small proteins in silico.

    PubMed

    Zimmermann, Olav; Hansmann, Ulrich H E

    2008-01-01

    Recent improvements in methodology and increased computer power now allow atomistic computer simulations of protein folding. We briefly review several advanced Monte Carlo algorithms that have contributed to this development. Details of folding simulations of three designed mini proteins are shown. Adding global translations and rotations has allowed us to handle multiple chains and to simulate the aggregation of six beta-amyloid fragments. In a different line of research we have developed several algorithms to predict local features from sequence. In an outlook we sketch how such biasing could extend the application spectrum of Monte Carlo simulations to structure prediction of larger proteins. PMID:18036571

  17. Imaging Protein-protein Interactions in vivo

    PubMed Central

    Seegar, Tom; Barton, William

    2010-01-01

    Protein-protein interactions are a hallmark of all essential cellular processes. However, many of these interactions are transient, or energetically weak, preventing their identification and analysis through traditional biochemical methods such as co-immunoprecipitation. In this regard, the genetically encodable fluorescent proteins (GFP, RFP, etc.) and their associated overlapping fluorescence spectrum have revolutionized our ability to monitor weak interactions in vivo using Förster resonance energy transfer (FRET)1-3. Here, we detail our use of a FRET-based proximity assay for monitoring receptor-receptor interactions on the endothelial cell surface. PMID:20972411

  18. CSF myelin basic protein

    MedlinePlus

    CSF myelin basic protein is a test to measure the level of myelin basic protein (MBP) in the cerebrospinal fluid (CSF). The CSF ... less than 4 ng/mL of myelin basic protein in the CSF. Normal value ranges may vary ...

  19. Modeling Protein Domain Function

    ERIC Educational Resources Information Center

    Baker, William P.; Jones, Carleton "Buck"; Hull, Elizabeth

    2007-01-01

    This simple but effective laboratory exercise helps students understand the concept of protein domain function. They use foam beads, Styrofoam craft balls, and pipe cleaners to explore how domains within protein active sites interact to form a functional protein. The activity allows students to gain content mastery and an understanding of the…

  20. Palmitoylation of Hedgehog proteins.

    PubMed

    Buglino, John A; Resh, Marilyn D

    2012-01-01

    Hedgehog (Hh) proteins are secreted signaling proteins that contain amide-linked palmitate at the N-terminus and cholesterol at the C-terminus. Palmitoylation of Hh proteins is critical for effective long- and short-range signaling. The palmitoylation reaction occurs during transit of Hh through the secretory pathway, most likely in the lumen of the ER. Attachment of palmitate to Hh proteins is independent of cholesterol modification and autoprocessing and is catalyzed by Hhat (Hedgehog acyltransferase). Hhat is a member of the membrane bound O-acyltransferase (MBOAT) family, a subgroup of multipass membrane proteins that catalyze transfer of fatty acyl groups to lipids and proteins. Several classes of secreted proteins have recently been shown to be substrates for MBOAT acyltransferases, including Hh proteins and Spitz (palmitoylated by Hhat), Wg/Wnt proteins (modified with palmitate and/or palmitoleate by Porcupine) and ghrelin (octanoylated by ghrelin O-acyltransferase). These findings highlight protein fatty acylation as a mechanism that not only influences membrane binding of intracellular proteins but also regulates the signaling range and efficacy of secreted proteins. PMID:22391306

  1. Protein electrophoresis - serum

    MedlinePlus

    Normal value ranges are: Total protein: 6.4 to 8.3 g/dL (grams per deciliter) Albumin: 3.5 to 5.0 g/dL Alpha-1 ... Decreased total protein may indicate: Abnormal loss of protein from the digestive tract or the inability of the digestive tract ...

  2. CSF total protein

    MedlinePlus

    CSF total protein is a test to determine the amount of protein in your spinal fluid, also called cerebrospinal fluid (CSF). ... The normal protein range varies from lab to lab, but is typically about 15 to 60 mg/dL. Note: mg/dL = ...

  3. Modeling Protein Self Assembly

    ERIC Educational Resources Information Center

    Baker, William P.; Jones, Carleton Buck; Hull, Elizabeth

    2004-01-01

    Understanding the structure and function of proteins is an important part of the standards-based science curriculum. Proteins serve vital roles within the cell and malfunctions in protein self assembly are implicated in degenerative diseases. Experience indicates that this topic is a difficult one for many students. We have found that the concept…

  4. Texturized dairy proteins

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Dairy proteins are amenable to structural modifications induced by high temperature, shear and moisture; in particular, whey proteins can change conformation to new unfolded states. The change in protein state is a basis for creating new foods. The dairy products, nonfat dried milk (NDM), whey prote...

  5. Destabilized bioluminescent proteins

    DOEpatents

    Allen, Michael S.; Rakesh, Gupta; Gary, Sayler S.

    2007-07-31

    Purified nucleic acids, vectors and cells containing a gene cassette encoding at least one modified bioluminescent protein, wherein the modification includes the addition of a peptide sequence. The duration of bioluminescence emitted by the modified bioluminescent protein is shorter than the duration of bioluminescence emitted by an unmodified form of the bioluminescent protein.

  6. Protein - Which is Best?

    PubMed

    Hoffman, Jay R; Falvo, Michael J

    2004-09-01

    Protein intake that exceeds the recommended daily allowance is widely accepted for both endurance and power athletes. However, considering the variety of proteins that are available much less is known concerning the benefits of consuming one protein versus another. The purpose of this paper is to identify and analyze key factors in order to make responsible recommendations to both the general and athletic populations. Evaluation of a protein is fundamental in determining its appropriateness in the human diet. Proteins that are of inferior content and digestibility are important to recognize and restrict or limit in the diet. Similarly, such knowledge will provide an ability to identify proteins that provide the greatest benefit and should be consumed. The various techniques utilized to rate protein will be discussed. Traditionally, sources of dietary protein are seen as either being of animal or vegetable origin. Animal sources provide a complete source of protein (i.e. containing all essential amino acids), whereas vegetable sources generally lack one or more of the essential amino acids. Animal sources of dietary protein, despite providing a complete protein and numerous vitamins and minerals, have some health professionals concerned about the amount of saturated fat common in these foods compared to vegetable sources. The advent of processing techniques has shifted some of this attention and ignited the sports supplement marketplace with derivative products such as whey, casein and soy. Individually, these products vary in quality and applicability to certain populations. The benefits that these particular proteins possess are discussed. In addition, the impact that elevated protein consumption has on health and safety issues (i.e. bone health, renal function) are also reviewed. Key PointsHigher protein needs are seen in athletic populations.Animal proteins is an important source of protein, however potential health concerns do exist from a diet of protein

  7. Protein crystallization with paper

    NASA Astrophysics Data System (ADS)

    Matsuoka, Miki; Kakinouchi, Keisuke; Adachi, Hiroaki; Maruyama, Mihoko; Sugiyama, Shigeru; Sano, Satoshi; Yoshikawa, Hiroshi Y.; Takahashi, Yoshinori; Yoshimura, Masashi; Matsumura, Hiroyoshi; Murakami, Satoshi; Inoue, Tsuyoshi; Mori, Yusuke; Takano, Kazufumi

    2016-05-01

    We developed a new protein crystallization method that incorporates paper. A small piece of paper, such as facial tissue or KimWipes, was added to a drop of protein solution in the traditional sitting drop vapor diffusion technique, and protein crystals grew by incorporating paper. By this method, we achieved the growth of protein crystals with reducing osmotic shock. Because the technique is very simple and the materials are easy to obtain, this method will come into wide use for protein crystallization. In the future, it could be applied to nanoliter-scale crystallization screening on a paper sheet such as in inkjet printing.

  8. Highly thermostable fluorescent proteins

    DOEpatents

    Bradbury, Andrew M.; Waldo, Geoffrey S.; Kiss, Csaba

    2011-03-22

    Thermostable fluorescent proteins (TSFPs), methods for generating these and other stability-enhanced proteins, polynucleotides encoding such proteins, and assays and method for using the TSFPs and TSFP-encoding nucleic acid molecules are provided. The TSFPs of the invention show extremely enhanced levels of stability and thermotolerance. In one case, for example, a TSFP of the invention is so stable it can be heated to 99.degree. C. for short periods of time without denaturing, and retains 85% of its fluorescence when heated to 80.degree. C. for several minutes. The invention also provides a method for generating stability-enhanced variants of a protein, including but not limited to fluorescent proteins.

  9. Highly thermostable fluorescent proteins

    DOEpatents

    Bradbury, Andrew M.; Waldo, Geoffrey S.; Kiss, Csaba

    2012-05-01

    Thermostable fluorescent proteins (TSFPs), methods for generating these and other stability-enhanced proteins, polynucleotides encoding such proteins, and assays and method for using the TSFPs and TSFP-encoding nucleic acid molecules are provided. The TSFPs of the invention show extremely enhanced levels of stability and thermotolerance. In one case, for example, a TSFP of the invention is so stable it can be heated to 99.degree. C. for short periods of time without denaturing, and retains 85% of its fluorescence when heated to 80.degree. C. for several minutes. The invention also provides a method for generating stability-enhanced variants of a protein, including but not limited to fluorescent proteins.

  10. Highly thermostable fluorescent proteins

    DOEpatents

    Bradbury, Andrew M.; Waldo, Geoffrey S.; Kiss, Csaba

    2011-11-29

    Thermostable fluorescent proteins (TSFPs), methods for generating these and other stability-enhanced proteins, polynucleotides encoding such proteins, and assays and method for using the TSFPs and TSFP-encoding nucleic acid molecules are provided. The TSFPs of the invention show extremely enhanced levels of stability and thermotolerance. In one case, for example, a TSFP of the invention is so stable it can be heated to 99.degree. C. for short periods of time without denaturing, and retains 85% of its fluorescence when heated to 80.degree. C. for several minutes. The invention also provides a method for generating stability-enhanced variants of a protein, including but not limited to fluorescent proteins.

  11. Selective Precipitation of Proteins.

    PubMed

    Matulis, Daumantas

    2016-01-01

    Selective precipitation of proteins can be used as a bulk method to recover the majority of proteins from a crude lysate, as a selective method to fractionate a subset of proteins from a protein solution, or as a very specific method to recover a single protein of interest from a purification step. This unit describes a number of methods suitable for selective precipitation. In each of the protocols that are outlined, the physical or chemical basis of the precipitation process, the parameters that can be varied for optimization, and the basic steps for developing an optimized precipitation are described. PMID:26836410

  12. Forces Stabilizing Proteins

    PubMed Central

    Pace, C. Nick; Scholtz, J. Martin; Grimsley, Gerald R.

    2014-01-01

    The goal of this article is to summarize what has been learned about the major forces stabilizing proteins since the late 1980s when site-directed mutagenesis became possible. The following conclusions are derived from experimental studies of hydrophobic and hydrogen bonding variants. 1. Based on studies of 138 hydrophobic interaction variants in 11 proteins, burying a –CH2– group on folding contributes 1.1 ± 0.5 kcal/mol to protein stability. 2. The burial of nonpolar side chains contributes to protein stability in two ways: first, a term that depends on the removal of the side chains from water and, more importantly, the enhanced London dispersion forces that result from the tight packing in the protein interior. 3. Based on studies of 151 hydrogen bonding variants in 15 proteins, forming a hydrogen bond on folding contributes 1.1 ± 0.8 kcal/mol to protein stability. 4. The contribution of hydrogen bonds to protein stability is strongly context dependent. 5. Hydrogen bonds by side chains and peptide groups make similar contributions to protein stability. 6. Polar group burial can make a favorable contribution to protein stability even if the polar group is not hydrogen bonded. 7. Hydrophobic interactions and hydrogen bonds both make large contributions to protein stability. PMID:24846139

  13. Mechanism of protein decarbonylation.

    PubMed

    Wong, Chi-Ming; Marcocci, Lucia; Das, Dividutta; Wang, Xinhong; Luo, Haibei; Zungu-Edmondson, Makhosazane; Suzuki, Yuichiro J

    2013-12-01

    Ligand/receptor stimulation of cells promotes protein carbonylation that is followed by the decarbonylation process, which might involve thiol-dependent reduction (C.M. Wong et al., Circ. Res. 102:301-318; 2008). This study further investigated the properties of this protein decarbonylation mechanism. We found that the thiol-mediated reduction of protein carbonyls is dependent on heat-labile biologic components. Cysteine and glutathione were efficient substrates for decarbonylation. Thiols decreased the protein carbonyl content, as detected by 2,4-dinitrophenylhydrazine, but not the levels of malondialdehyde or 4-hydroxynonenal protein adducts. Mass spectrometry identified proteins that undergo thiol-dependent decarbonylation, which include peroxiredoxins. Peroxiredoxin-2 and -6 were carbonylated and subsequently decarbonylated in response to the ligand/receptor stimulation of cells. siRNA knockdown of glutaredoxin inhibited the decarbonylation of peroxiredoxin. These results strengthen the concept that thiol-dependent decarbonylation defines the kinetics of protein carbonylation signaling. PMID:24044890

  14. Pigment-protein complexes

    SciTech Connect

    Siegelman, H W

    1980-01-01

    The photosynthetically-active pigment protein complexes of procaryotes and eucaryotes include chlorophyll proteins, carotenochlorophyll proteins, and biliproteins. They are either integral components or attached to photosynthetic membranes. Detergents are frequently required to solubilize the pigment-protein complexes. The membrane localization and detergent solubilization strongly suggest that the pigment-protein complexes are bound to the membranes by hydrophobic interactions. Hydrophobic interactions of proteins are characterized by an increase in entropy. Their bonding energy is directly related to temperature and ionic strength. Hydrophobic-interaction chromatography, a relatively new separation procedure, can furnish an important method for the purification of pigment-protein complexes. Phycobilisome purification and properties provide an example of the need to maintain hydrophobic interactions to preserve structure and function.

  15. Protein solubility modeling

    NASA Technical Reports Server (NTRS)

    Agena, S. M.; Pusey, M. L.; Bogle, I. D.

    1999-01-01

    A thermodynamic framework (UNIQUAC model with temperature dependent parameters) is applied to model the salt-induced protein crystallization equilibrium, i.e., protein solubility. The framework introduces a term for the solubility product describing protein transfer between the liquid and solid phase and a term for the solution behavior describing deviation from ideal solution. Protein solubility is modeled as a function of salt concentration and temperature for a four-component system consisting of a protein, pseudo solvent (water and buffer), cation, and anion (salt). Two different systems, lysozyme with sodium chloride and concanavalin A with ammonium sulfate, are investigated. Comparison of the modeled and experimental protein solubility data results in an average root mean square deviation of 5.8%, demonstrating that the model closely follows the experimental behavior. Model calculations and model parameters are reviewed to examine the model and protein crystallization process. Copyright 1999 John Wiley & Sons, Inc.

  16. Protein kinesis: The dynamics of protein trafficking and stability

    SciTech Connect

    1995-12-31

    The purpose of this conference is to provide a multidisciplinary forum for exchange of state-of-the-art information on protein kinesis. This volume contains abstracts of papers in the following areas: protein folding and modification in the endoplasmic reticulum; protein trafficking; protein translocation and folding; protein degradation; polarity; nuclear trafficking; membrane dynamics; and protein import into organelles.

  17. Phage display of proteins.

    PubMed

    Kościelska, K; Kiczak, L; Kasztura, M; Wesołowska, O; Otlewski, J

    1998-01-01

    In recent years the phage display approach has become an increasingly popular method in protein research. This method enables the presentation of large peptide and protein libraries on the surface of phage particles from which molecules of desired functional property(ies) can be rapidly selected. The great advantage of this method is a direct linkage between an observed phenotype and encapsulated genotype, which allows fast determination of selected sequences. The phage display approach is a powerful tool in generating highly potent biomolecules, including: search for specific antibodies, determining enzyme specificity, exploring protein-protein and protein-DNA interactions, minimizing proteins, introducing new functions into different protein scaffolds, and searching sequence space of protein folding. In this article many examples are given to illustrate that this technique can be used in different fields of protein science. The phage display has a potential of the natural evolution and its possibilities are far beyond rational prediction. Assuming that we can design the selection agents and conditions we should be able to engineer any desired protein function or feature. PMID:9918498

  18. Energy design for protein-protein interactions

    PubMed Central

    Ravikant, D. V. S.; Elber, Ron

    2011-01-01

    Proteins bind to other proteins efficiently and specifically to carry on many cell functions such as signaling, activation, transport, enzymatic reactions, and more. To determine the geometry and strength of binding of a protein pair, an energy function is required. An algorithm to design an optimal energy function, based on empirical data of protein complexes, is proposed and applied. Emphasis is made on negative design in which incorrect geometries are presented to the algorithm that learns to avoid them. For the docking problem the search for plausible geometries can be performed exhaustively. The possible geometries of the complex are generated on a grid with the help of a fast Fourier transform algorithm. A novel formulation of negative design makes it possible to investigate iteratively hundreds of millions of negative examples while monotonically improving the quality of the potential. Experimental structures for 640 protein complexes are used to generate positive and negative examples for learning parameters. The algorithm designed in this work finds the correct binding structure as the lowest energy minimum in 318 cases of the 640 examples. Further benchmarks on independent sets confirm the significant capacity of the scoring function to recognize correct modes of interactions. PMID:21842951

  19. Modeling Protein Expression and Protein Signaling Pathways

    PubMed Central

    Telesca, Donatello; Müller, Peter; Kornblau, Steven M.; Suchard, Marc A.; Ji, Yuan

    2015-01-01

    High-throughput functional proteomic technologies provide a way to quantify the expression of proteins of interest. Statistical inference centers on identifying the activation state of proteins and their patterns of molecular interaction formalized as dependence structure. Inference on dependence structure is particularly important when proteins are selected because they are part of a common molecular pathway. In that case, inference on dependence structure reveals properties of the underlying pathway. We propose a probability model that represents molecular interactions at the level of hidden binary latent variables that can be interpreted as indicators for active versus inactive states of the proteins. The proposed approach exploits available expert knowledge about the target pathway to define an informative prior on the hidden conditional dependence structure. An important feature of this prior is that it provides an instrument to explicitly anchor the model space to a set of interactions of interest, favoring a local search approach to model determination. We apply our model to reverse-phase protein array data from a study on acute myeloid leukemia. Our inference identifies relevant subpathways in relation to the unfolding of the biological process under study. PMID:26246646

  20. Protein-protein docking with backbone flexibility.

    PubMed

    Wang, Chu; Bradley, Philip; Baker, David

    2007-10-19

    Computational protein-protein docking methods currently can create models with atomic accuracy for protein complexes provided that the conformational changes upon association are restricted to the side chains. However, it remains very challenging to account for backbone conformational changes during docking, and most current methods inherently keep monomer backbones rigid for algorithmic simplicity and computational efficiency. Here we present a reformulation of the Rosetta docking method that incorporates explicit backbone flexibility in protein-protein docking. The new method is based on a "fold-tree" representation of the molecular system, which seamlessly integrates internal torsional degrees of freedom and rigid-body degrees of freedom. Problems with internal flexible regions ranging from one or more loops or hinge regions to all of one or both partners can be readily treated using appropriately constructed fold trees. The explicit treatment of backbone flexibility improves both sampling in the vicinity of the native docked conformation and the energetic discrimination between near-native and incorrect models. PMID:17825317

  1. Energy design for protein-protein interactions

    NASA Astrophysics Data System (ADS)

    Ravikant, D. V. S.; Elber, Ron

    2011-08-01

    Proteins bind to other proteins efficiently and specifically to carry on many cell functions such as signaling, activation, transport, enzymatic reactions, and more. To determine the geometry and strength of binding of a protein pair, an energy function is required. An algorithm to design an optimal energy function, based on empirical data of protein complexes, is proposed and applied. Emphasis is made on negative design in which incorrect geometries are presented to the algorithm that learns to avoid them. For the docking problem the search for plausible geometries can be performed exhaustively. The possible geometries of the complex are generated on a grid with the help of a fast Fourier transform algorithm. A novel formulation of negative design makes it possible to investigate iteratively hundreds of millions of negative examples while monotonically improving the quality of the potential. Experimental structures for 640 protein complexes are used to generate positive and negative examples for learning parameters. The algorithm designed in this work finds the correct binding structure as the lowest energy minimum in 318 cases of the 640 examples. Further benchmarks on independent sets confirm the significant capacity of the scoring function to recognize correct modes of interactions.

  2. Mechanisms Regulating Protein Localization.

    PubMed

    Bauer, Nicholas C; Doetsch, Paul W; Corbett, Anita H

    2015-10-01

    Cellular functions are dictated by protein content and activity. There are numerous strategies to regulate proteins varying from modulating gene expression to post-translational modifications. One commonly used mode of regulation in eukaryotes is targeted localization. By specifically redirecting the localization of a pool of existing protein, cells can achieve rapid changes in local protein function. Eukaryotic cells have evolved elegant targeting pathways to direct proteins to the appropriate cellular location or locations. Here, we provide a general overview of these localization pathways, with a focus on nuclear and mitochondrial transport, and present a survey of the evolutionarily conserved regulatory strategies identified thus far. We end with a description of several specific examples of proteins that exploit localization as an important mode of regulation. PMID:26172624

  3. Electrophoretic separation of proteins.

    PubMed

    Chakavarti, Bulbul; Chakavarti, Deb

    2008-01-01

    Electrophoresis is used to separate complex mixtures of proteins (e.g., from cells, subcellular fractions, column fractions, or immunoprecipitates), to investigate subunit compositions, and to verify homogeneity of protein samples. It can also serve to purify proteins for use in further applications. In polyacrylamide gel electrophoresis, proteins migrate in response to an electrical field through pores in a polyacrylamide gel matrix; pore size decreases with increasing acrylamide concentration. The combination of pore size and protein charge, size, and shape determines the migration rate of the protein. In this unit, the standard Laemmli method is described for discontinuous gel electrophoresis under denaturing conditions, i.e., in the presence of sodium dodecyl sulfate (SDS). PMID:19066548

  4. Outer membrane protein purification.

    PubMed

    Arigita, C; Jiskoot, W; Graaf, M R; Kersten, G F

    2001-01-01

    The major outer membrane proteins (OMPs) from Neisseria meningitidis, which are expressed at high levels, are subdivided in five classes based on molecular weight (1,2) (see Table 1). Table 1 Major Meningococcal Outer-Membrane Proteins Outer-membrane proteins Name Molecular maass Function/characteristics Class 1 PorA 44-47 kDa Porin Class 2/3 PorB 37-42 kDa Porin Class 4 Rmp Reductionmodifiableprotein, unknown Class 5 Opa 26-30 kDa Adhesion,opacity protein Opc 25 kDa Invasion, opacity protein Iron-regulated proteins Mirp 37 kDa Iron acquisition (?);majoriron-regulatedprotein FrpB 70 kDa Ferric enterobactin receptor (also FetA) Adapted from ref. (1). PMID:21336748

  5. Biofilm Matrix Proteins

    PubMed Central

    Fong, Jiunn N. C.; Yildiz, Fitnat H.

    2015-01-01

    Proteinaceous components of the biofilm matrix include secreted extracellular proteins, cell surface adhesins and protein subunits of cell appendages such as flagella and pili. Biofilm matrix proteins play diverse roles in biofilm formation and dissolution. They are involved in attaching cells to surfaces, stabilizing the biofilm matrix via interactions with exopolysaccharide and nucleic acid components, developing three-dimensional biofilm architectures, and dissolving biofilm matrix via enzymatic degradation of polysaccharides, proteins, and nucleic acids. In this chapter, we will review functions of matrix proteins in a selected set of microorganisms, studies of the matrix proteomes of Vibrio cholerae and Pseudomonas aeruginosa, and roles of outer membrane vesicles and of nucleoid-binding proteins in biofilm formation. PMID:26104709

  6. Principles of Flexible Protein-Protein Docking

    PubMed Central

    Andrusier, Nelly; Mashiach, Efrat; Nussinov, Ruth; Wolfson, Haim J.

    2008-01-01

    Treating flexibility in molecular docking is a major challenge in cell biology research. Here we describe the background and the principles of existing flexible protein-protein docking methods, focusing on the algorithms and their rational. We describe how protein flexibility is treated in different stages of the docking process: in the preprocessing stage, rigid and flexible parts are identified and their possible conformations are modeled. This preprocessing provides information for the subsequent docking and refinement stages. In the docking stage, an ensemble of pre-generated conformations or the identified rigid domains may be docked separately. In the refinement stage, small-scale movements of the backbone and side-chains are modeled and the binding orientation is improved by rigid-body adjustments. For clarity of presentation, we divide the different methods into categories. This should allow the reader to focus on the most suitable method for a particular docking problem. PMID:18655061

  7. Antimicrobial proteins: From old proteins, new tricks.

    PubMed

    Smith, Valerie J; Dyrynda, Elisabeth A

    2015-12-01

    This review describes the main types of antimicrobial peptides (AMPs) synthesised by crustaceans, primarily those identified in shrimp, crayfish, crab and lobster. It includes an overview of their range of microbicidal activities and the current landscape of our understanding of their gene expression patterns in different body tissues. It further summarises how their expression might change following various types of immune challenges. The review further considers proteins or protein fragments from crustaceans that have antimicrobial properties but are more usually associated with other biological functions, or are derived from such proteins. It discusses how these unconventional AMPs might be generated at, or delivered to, sites of infection and how they might contribute to crustacean host defence in vivo. It also highlights recent work that is starting to reveal the extent of multi-functionality displayed by some decapod AMPs, particularly their participation in other aspects of host protection. Examples of such activities include proteinase inhibition, phagocytosis, antiviral activity and haematopoiesis. PMID:26320628

  8. Elastic proteins and elastomeric protein alloys.

    PubMed

    Aghaei-Ghareh-Bolagh, Behnaz; Mithieux, Suzanne M; Weiss, Anthony S

    2016-06-01

    The elastomeric proteins elastin and resilin have been used extensively in the fabrication of biomaterials for tissue engineering applications due to their unique mechanical and biological properties. Tropoelastin is the soluble monomer component of elastin. Tropoelastin and resilin are both highly elastic with high resilience, substantial extensibility, high durability and low energy loss, which makes them excellent candidates for the fabrication of elastic tissues that demand regular and repetitive movement like the skin, lung, blood vessels, muscles and vocal folds. Combinations of these proteins with silk fibroin further enhance their biomechanical and biological properties leading to a new class of protein alloy materials with versatile properties. In this review, the properties of tropoelastin-based and resilin-based biomaterials with and without silk are described in concert with examples of their applications in tissue engineering. PMID:26780495

  9. Protein Crystal Quality Studies

    NASA Technical Reports Server (NTRS)

    1998-01-01

    Eddie Snell, Post-Doctoral Fellow the National Research Council (NRC) uses a reciprocal space mapping diffractometer for macromolecular crystal quality studies. The diffractometer is used in mapping the structure of macromolecules such as proteins to determine their structure and thus understand how they function with other proteins in the body. This is one of several analytical tools used on proteins crystallized on Earth and in space experiments. Photo credit: NASA/Marshall Space Flight Center (MSFC)

  10. Protein oxidation and peroxidation.

    PubMed

    Davies, Michael J

    2016-04-01

    Proteins are major targets for radicals and two-electron oxidants in biological systems due to their abundance and high rate constants for reaction. With highly reactive radicals damage occurs at multiple side-chain and backbone sites. Less reactive species show greater selectivity with regard to the residues targeted and their spatial location. Modification can result in increased side-chain hydrophilicity, side-chain and backbone fragmentation, aggregation via covalent cross-linking or hydrophobic interactions, protein unfolding and altered conformation, altered interactions with biological partners and modified turnover. In the presence of O2, high yields of peroxyl radicals and peroxides (protein peroxidation) are formed; the latter account for up to 70% of the initial oxidant flux. Protein peroxides can oxidize both proteins and other targets. One-electron reduction results in additional radicals and chain reactions with alcohols and carbonyls as major products; the latter are commonly used markers of protein damage. Direct oxidation of cysteine (and less commonly) methionine residues is a major reaction; this is typically faster than with H2O2, and results in altered protein activity and function. Unlike H2O2, which is rapidly removed by protective enzymes, protein peroxides are only slowly removed, and catabolism is a major fate. Although turnover of modified proteins by proteasomal and lysosomal enzymes, and other proteases (e.g. mitochondrial Lon), can be efficient, protein hydroperoxides inhibit these pathways and this may contribute to the accumulation of modified proteins in cells. Available evidence supports an association between protein oxidation and multiple human pathologies, but whether this link is causal remains to be established. PMID:27026395

  11. Protein oxidation and peroxidation

    PubMed Central

    Davies, Michael J.

    2016-01-01

    Proteins are major targets for radicals and two-electron oxidants in biological systems due to their abundance and high rate constants for reaction. With highly reactive radicals damage occurs at multiple side-chain and backbone sites. Less reactive species show greater selectivity with regard to the residues targeted and their spatial location. Modification can result in increased side-chain hydrophilicity, side-chain and backbone fragmentation, aggregation via covalent cross-linking or hydrophobic interactions, protein unfolding and altered conformation, altered interactions with biological partners and modified turnover. In the presence of O2, high yields of peroxyl radicals and peroxides (protein peroxidation) are formed; the latter account for up to 70% of the initial oxidant flux. Protein peroxides can oxidize both proteins and other targets. One-electron reduction results in additional radicals and chain reactions with alcohols and carbonyls as major products; the latter are commonly used markers of protein damage. Direct oxidation of cysteine (and less commonly) methionine residues is a major reaction; this is typically faster than with H2O2, and results in altered protein activity and function. Unlike H2O2, which is rapidly removed by protective enzymes, protein peroxides are only slowly removed, and catabolism is a major fate. Although turnover of modified proteins by proteasomal and lysosomal enzymes, and other proteases (e.g. mitochondrial Lon), can be efficient, protein hydroperoxides inhibit these pathways and this may contribute to the accumulation of modified proteins in cells. Available evidence supports an association between protein oxidation and multiple human pathologies, but whether this link is causal remains to be established. PMID:27026395

  12. Computer Models of Proteins

    NASA Technical Reports Server (NTRS)

    2000-01-01

    Dr. Marc Pusey (seated) and Dr. Craig Kundrot use computers to analyze x-ray maps and generate three-dimensional models of protein structures. With this information, scientists at Marshall Space Flight Center can learn how proteins are made and how they work. The computer screen depicts a proten structure as a ball-and-stick model. Other models depict the actual volume occupied by the atoms, or the ribbon-like structures that are crucial to a protein's function.

  13. Pressure cryocooling protein crystals

    DOEpatents

    Kim, Chae Un; Gruner, Sol M.

    2011-10-04

    Preparation of cryocooled protein crystal is provided by use of helium pressurizing and cryocooling to obtain cryocooled protein crystal allowing collection of high resolution data and by heavier noble gas (krypton or xenon) binding followed by helium pressurizing and cryocooling to obtain cryocooled protein crystal for collection of high resolution data and SAD phasing simultaneously. The helium pressurizing is carried out on crystal coated to prevent dehydration or on crystal grown in aqueous solution in a capillary.

  14. The Non-native Helical Intermediate State May Accumulate at Low pH in the Folding and Aggregation Landscape of the Intestinal Fatty Acid Binding Protein.

    PubMed

    Sarkar-Banerjee, Suparna; Chowdhury, Sourav; Paul, Simanta Sarani; Dutta, Debashis; Ghosh, Anisa; Chattopadhyay, Krishnananda

    2016-08-16

    There has been widespread interest in studying early intermediate states and their roles in protein folding. The interest in intermediate states has been further emphasized in the recent literature because of their implications for protein aggregation. Unfortunately, direct kinetic characterization of intermediates has been difficult because of the limited time resolutions offered by the kinetic techniques and the heterogeneity of the folding and aggregation landscape. Even in equilibrium experiments, the characterization of intermediate states could be difficult because (a) their populations in equilibrium could be low and/or (b) they lack any specific biochemical or biophysical signatures for their identification. In this paper, we have used fluorescence correlation spectroscopy to study the nature of a low-pH intermediate state of the intestinal fatty acid binding protein, a small protein with predominantly β-sheet structure. Our results have shown that the pH 3 intermediate diffuses faster than the folded protein and has strong helix forming propensity. These behaviors support Lim's hypothesis according to which even an entirely β-sheet protein would form helical bundles at the early stage. Using dynamic light scattering and thioflavin T binding measurements, we have observed that the pH 3 intermediate is prone to aggregation. We believe that early helix formation is the result of a local effect, which originates from the interaction of the neighboring amino acids around the hydrophobic core residues. This early intermediate reorganizes subsequently, and this structural reorganization is initiated by the destabilizing interactions induced by the distant residues, unfavorable entropic costs, and steric constraints of the hydrophobic side chains. Mutational analyses show further that the increase in the hydrophobicity in the hydrophobic core region increases the population of the α-helical intermediate, enhancing the aggregation propensity of the protein

  15. Protein-protein interactions as drug targets.

    PubMed

    Skwarczynska, Malgorzata; Ottmann, Christian

    2015-10-01

    Modulation of protein-protein interactions (PPIs) is becoming increasingly important in drug discovery and chemical biology. While a few years ago this 'target class' was deemed to be largely undruggable an impressing number of publications and success stories now show that targeting PPIs with small, drug-like molecules indeed is a feasible approach. Here, we summarize the current state of small-molecule inhibition and stabilization of PPIs and review the active molecules from a structural and medicinal chemistry angle, especially focusing on the key examples of iNOS, LFA-1 and 14-3-3. PMID:26510391

  16. Biomolecular membrane protein crystallization

    NASA Astrophysics Data System (ADS)

    Reddy Bolla, Jani; Su, Chih-Chia; Yu, Edward W.

    2012-07-01

    Integral membrane proteins comprise approximately 30% of the sequenced genomes, and there is an immediate need for their high-resolution structural information. Currently, the most reliable approach to obtain these structures is X-ray crystallography. However, obtaining crystals of membrane proteins that diffract to high resolution appears to be quite challenging, and remains a major obstacle in structural determination. This brief review summarizes a variety of methodologies for use in crystallizing these membrane proteins. Hopefully, by introducing the available methods, techniques, and providing a general understanding of membrane proteins, a rational decision can be made about now to crystallize these complex materials.

  17. Self assembling proteins

    DOEpatents

    Yeates, Todd O.; Padilla, Jennifer; Colovos, Chris

    2004-06-29

    Novel fusion proteins capable of self-assembling into regular structures, as well as nucleic acids encoding the same, are provided. The subject fusion proteins comprise at least two oligomerization domains rigidly linked together, e.g. through an alpha helical linking group. Also provided are regular structures comprising a plurality of self-assembled fusion proteins of the subject invention, and methods for producing the same. The subject fusion proteins find use in the preparation of a variety of nanostructures, where such structures include: cages, shells, double-layer rings, two-dimensional layers, three-dimensional crystals, filaments, and tubes.

  18. Consensus protein design.

    PubMed

    Porebski, Benjamin T; Buckle, Ashley M

    2016-07-01

    A popular and successful strategy in semi-rational design of protein stability is the use of evolutionary information encapsulated in homologous protein sequences. Consensus design is based on the hypothesis that at a given position, the respective consensus amino acid contributes more than average to the stability of the protein than non-conserved amino acids. Here, we review the consensus design approach, its theoretical underpinnings, successes, limitations and challenges, as well as providing a detailed guide to its application in protein engineering. PMID:27274091

  19. Prediction of protein-protein interactions based on protein-protein correlation using least squares regression.

    PubMed

    Huang, De-Shuang; Zhang, Lei; Han, Kyungsook; Deng, Suping; Yang, Kai; Zhang, Hongbo

    2014-01-01

    In order to transform protein sequences into the feature vectors, several works have been done, such as computing auto covariance (AC), conjoint triad (CT), local descriptor (LD), moran autocorrelation (MA), normalized moreaubroto autocorrelation (NMB) and so on. In this paper, we shall adopt these transformation methods to encode the proteins, respectively, where AC, CT, LD, MA and NMB are all represented by '+' in a unified manner. A new method, i.e. the combination of least squares regression with '+' (abbreviated as LSR(+)), will be introduced for encoding a protein-protein correlation-based feature representation and an interacting protein pair. Thus there are totally five different combinations for LSR(+), i.e. LSRAC, LSRCT, LSRLD, LSRMA and LSRNMB. As a result, we combined a support vector machine (SVM) approach with LSR(+) to predict protein-protein interactions (PPI) and PPI networks. The proposed method has been applied on four datasets, i.e. Saaccharomyces cerevisiae, Escherichia coli, Homo sapiens and Caenorhabditis elegans. The experimental results demonstrate that all LSR(+) methods outperform many existing representative algorithms. Therefore, LSR(+) is a powerful tool to characterize the protein-protein correlations and to infer PPI, whilst keeping high performance on prediction of PPI networks. PMID:25059329

  20. Human Mitochondrial Protein Database

    National Institute of Standards and Technology Data Gateway

    SRD 131 Human Mitochondrial Protein Database (Web, free access)   The Human Mitochondrial Protein Database (HMPDb) provides comprehensive data on mitochondrial and human nuclear encoded proteins involved in mitochondrial biogenesis and function. This database consolidates information from SwissProt, LocusLink, Protein Data Bank (PDB), GenBank, Genome Database (GDB), Online Mendelian Inheritance in Man (OMIM), Human Mitochondrial Genome Database (mtDB), MITOMAP, Neuromuscular Disease Center and Human 2-D PAGE Databases. This database is intended as a tool not only to aid in studying the mitochondrion but in studying the associated diseases.

  1. PIC: Protein Interactions Calculator.

    PubMed

    Tina, K G; Bhadra, R; Srinivasan, N

    2007-07-01

    Interactions within a protein structure and interactions between proteins in an assembly are essential considerations in understanding molecular basis of stability and functions of proteins and their complexes. There are several weak and strong interactions that render stability to a protein structure or an assembly. Protein Interactions Calculator (PIC) is a server which, given the coordinate set of 3D structure of a protein or an assembly, computes various interactions such as disulphide bonds, interactions between hydrophobic residues, ionic interactions, hydrogen bonds, aromatic-aromatic interactions, aromatic-sulphur interactions and cation-pi interactions within a protein or between proteins in a complex. Interactions are calculated on the basis of standard, published criteria. The identified interactions between residues can be visualized using a RasMol and Jmol interface. The advantage with PIC server is the easy availability of inter-residue interaction calculations in a single site. It also determines the accessible surface area and residue-depth, which is the distance of a residue from the surface of the protein. User can also recognize specific kind of interactions, such as apolar-apolar residue interactions or ionic interactions, that are formed between buried or exposed residues or near the surface or deep inside. PMID:17584791

  2. Glycolipid transfer proteins

    PubMed Central

    Brown, Rhoderick E.; Mattjus, Peter

    2007-01-01

    Glycolipid transfer proteins (GLTPs) are small (24 kD), soluble, ubiquitous proteins characterized by their ability to accelerate the intermembrane transfer of glycolipids in vitro. GLTP specificity encompasses both sphingoid- and glycerol-based glycolipids, but with a strict requirement that the initial sugar residue be beta-linked to the hydrophobic lipid backbone. The 3D protein structures of GLTP reveal liganded structures with unique lipid binding modes. The biochemical properties of GLTP action at the membrane surface have been studied rather comprehensively, but the biological role of GLTP remains enigmatic. What is clear is that GLTP differs distinctly from other known glycolipid-binding proteins, such as nonspecific lipid transfer proteins, lysosomal sphingolipid activator proteins, lectins, lung surfactant proteins as well as other lipid binding/transfer proteins. Based on the unique conformational architecture that targets GLTP to membranes and enables glycolipid binding, GLTP is now considered the prototypical and founding member of a new protein superfamily in eukaryotes. PMID:17320476

  3. Engineering therapeutic protein disaggregases.

    PubMed

    Shorter, James

    2016-05-15

    Therapeutic agents are urgently required to cure several common and fatal neurodegenerative disorders caused by protein misfolding and aggregation, including amyotrophic lateral sclerosis (ALS), Parkinson's disease (PD), and Alzheimer's disease (AD). Protein disaggregases that reverse protein misfolding and restore proteins to native structure, function, and localization could mitigate neurodegeneration by simultaneously reversing 1) any toxic gain of function of the misfolded form and 2) any loss of function due to misfolding. Potentiated variants of Hsp104, a hexameric AAA+ ATPase and protein disaggregase from yeast, have been engineered to robustly disaggregate misfolded proteins connected with ALS (e.g., TDP-43 and FUS) and PD (e.g., α-synuclein). However, Hsp104 has no metazoan homologue. Metazoa possess protein disaggregase systems distinct from Hsp104, including Hsp110, Hsp70, and Hsp40, as well as HtrA1, which might be harnessed to reverse deleterious protein misfolding. Nevertheless, vicissitudes of aging, environment, or genetics conspire to negate these disaggregase systems in neurodegenerative disease. Thus, engineering potentiated human protein disaggregases or isolating small-molecule enhancers of their activity could yield transformative therapeutics for ALS, PD, and AD. PMID:27255695

  4. Cellulose synthase interacting protein

    PubMed Central

    Somerville, Chris

    2010-01-01

    Cellulose is the most abundant biopolymer on earth. The great abundance of cellulose places it at the forefront as a primary source of biomass for renewable biofuels. However, the knowledge of how plant cells make cellulose remains very rudimentary. Cellulose microfibrils are synthesized at the plasma membrane by hexameric protein complexes, also known as cellulose synthase complexes. The only known components of cellulose synthase complexes are cellulose synthase (CESA) proteins until the recent identification of a novel component. CSI1, which encodes CESA interacting protein 1 (CSI1) in Arabidopsis. CSI1, as the first non-CESA proteins associated with cellulose synthase complexes, opens up many opportunities. PMID:21150290

  5. Consensus protein design

    PubMed Central

    Porebski, Benjamin T.; Buckle, Ashley M.

    2016-01-01

    A popular and successful strategy in semi-rational design of protein stability is the use of evolutionary information encapsulated in homologous protein sequences. Consensus design is based on the hypothesis that at a given position, the respective consensus amino acid contributes more than average to the stability of the protein than non-conserved amino acids. Here, we review the consensus design approach, its theoretical underpinnings, successes, limitations and challenges, as well as providing a detailed guide to its application in protein engineering. PMID:27274091

  6. Acanthamoeba castellanii STAT protein.

    PubMed

    Kicinska, Anna; Leluk, Jacek; Jarmuszkiewicz, Wieslawa

    2014-01-01

    STAT (signal transducers and activators of transcription) proteins are one of the important mediators of phosphotyrosine-regulated signaling in metazoan cells. We described the presence of STAT protein in a unicellular, free-living amoebae with a simple life cycle, Acanthamoeba castellanii. A. castellanii is the only, studied to date, Amoebozoan that does not belong to Mycetozoa but possesses STATs. A sequence of the A. castellanii STAT protein includes domains similar to those of the Dictyostelium STAT proteins: a coiled coil (characteristic for Dictyostelium STAT coiled coil), a STAT DNA-binding domain and a Src-homology domain. The search for protein sequences homologous to A. castellanii STAT revealed 17 additional sequences from lower eukaryotes. Interestingly, all of these sequences come from Amoebozoa organisms that belong to either Mycetozoa (slime molds) or Centramoebida. We showed that there are four separated clades within the slime mold STAT proteins. The A. castellanii STAT protein branches next to a group of STATc proteins from Mycetozoa. We also demonstrate that Amoebozoa form a distinct monophyletic lineage within the STAT protein world that is well separated from the other groups. PMID:25338074

  7. Engineering therapeutic protein disaggregases

    PubMed Central

    Shorter, James

    2016-01-01

    Therapeutic agents are urgently required to cure several common and fatal neurodegenerative disorders caused by protein misfolding and aggregation, including amyotrophic lateral sclerosis (ALS), Parkinson’s disease (PD), and Alzheimer’s disease (AD). Protein disaggregases that reverse protein misfolding and restore proteins to native structure, function, and localization could mitigate neurodegeneration by simultaneously reversing 1) any toxic gain of function of the misfolded form and 2) any loss of function due to misfolding. Potentiated variants of Hsp104, a hexameric AAA+ ATPase and protein disaggregase from yeast, have been engineered to robustly disaggregate misfolded proteins connected with ALS (e.g., TDP-43 and FUS) and PD (e.g., α-synuclein). However, Hsp104 has no metazoan homologue. Metazoa possess protein disaggregase systems distinct from Hsp104, including Hsp110, Hsp70, and Hsp40, as well as HtrA1, which might be harnessed to reverse deleterious protein misfolding. Nevertheless, vicissitudes of aging, environment, or genetics conspire to negate these disaggregase systems in neurodegenerative disease. Thus, engineering potentiated human protein disaggregases or isolating small-molecule enhancers of their activity could yield transformative therapeutics for ALS, PD, and AD. PMID:27255695

  8. Ultrafiltration of pegylated proteins

    NASA Astrophysics Data System (ADS)

    Molek, Jessica R.

    There is considerable clinical interest in the use of "second-generation" therapeutics produced by conjugation of a native protein with various polymers including polyethylene glycol (PEG). PEG--protein conjugates, so-called PEGylated proteins, can exhibit enhanced stability, half-life, and bioavailability. One of the challenges in the commercial production of PEGylated proteins is the purification required to remove unreacted polymer, native protein, and in many cases PEGylated proteins with nonoptimal degrees of conjugation. The overall objective of this thesis was to examine the use of ultrafiltration for the purification of PEGylated proteins. This included: (1) analysis of size-based separation of PEGylated proteins using conventional ultrafiltration membranes, (2) use of electrically-charged membranes to exploit differences in electrostatic interactions, and (3) examination of the effects of PEGylation on protein fouling. The experimental results were analyzed using appropriate theoretical models, with the underlying physical properties of the PEGylated proteins evaluated using size exclusion chromatography, capillary electrophoresis, dynamic light scattering, and reverse phase chromatography. PEGylated proteins were produced by covalent attachment of activated PEG to a protein via primary amines on the lysine residues. A simple model was developed for the reaction kinetics, which was used to explore the effect of reaction conditions and mode of operation on the distribution of PEGylated products. The effective size of the PEGylated proteins was evaluated using size exclusion chromatography, with appropriate correlations developed for the size in terms of the molecular weight of the native protein and attached PEG. The electrophoretic mobility of the PEGylated proteins were evaluated by capillary electrophoresis with the data in good agreement with a simple model accounting for the increase in protein size and the reduction in the number of protonated amine

  9. Protein metabolism and requirements.

    PubMed

    Biolo, Gianni

    2013-01-01

    Skeletal muscle adaptation to critical illness includes insulin resistance, accelerated proteolysis, and increased release of glutamine and the other amino acids. Such amino acid efflux from skeletal muscle provides precursors for protein synthesis and energy fuel to the liver and to the rapidly dividing cells of the intestinal mucosa and the immune system. From these adaptation mechanisms, severe muscle wasting, glutamine depletion, and hyperglycemia, with increased patient morbidity and mortality, may ensue. Protein/amino acid nutrition, through either enteral or parenteral routes, plays a pivotal role in treatment of metabolic abnormalities in critical illness. In contrast to energy requirement, which can be accurately assessed by indirect calorimetry, methods to determine individual protein/amino acid needs are not currently available. In critical illness, a decreased ability of protein/amino acid intake to promote body protein synthesis is defined as anabolic resistance. This abnormality leads to increased protein/amino acid requirement and relative inefficiency of nutritional interventions. In addition to stress mediators, immobility and physical inactivity are key determinants of anabolic resistance. The development of mobility protocols in the intensive care unit should be encouraged to enhance the efficacy of nutrition. In critical illness, protein/amino acid requirement has been defined as the intake level associated with the lowest rate of catabolism. The optimal protein-sparing effects in patients receiving adequate energy are achieved when protein/amino acids are administered at rates between 1.3 and 1.5 g/kg/day. Extra glutamine supplementation is required in conditions of severe systemic inflammatory response. Protein requirement increases during hypocaloric feeding and in patients with acute renal failure on continuous renal replacement therapy. Evidence suggests that receiving adequate protein/amino acid intake may be more important than achieving

  10. Binding Efficiency of Protein-Protein Complexes

    PubMed Central

    Day, Eric S.; Cote, Shaun M.; Whitty, Adrian

    2012-01-01

    We examine the relationship between binding affinity and interface size for reversible protein-protein interactions (PPI), using cytokines from the tumor necrosis factor (TNF) superfamily and their receptors as a test case. Using surface plasmon resonance, we measured single-site binding affinities for the large receptor TNFR1 binding to its ligands TNFα (KD = 1.4 ± 0.4 nM) and lymphotoxin-α (KD = 50 ± 10 nM), and also for the small receptor Fn14 binding to TWEAK (KD = 70 ± 10 nM). We additionally assembled data for all other TNF/TNFR family complexes for which reliable single site binding affinities have been reported. We used these values to calculate the binding efficiency – defined as binding energy per Å2 of surface area buried at the contact interface – for the nine of these complexes for which co-crystal structures are available, and compared the results to those for a set of 144 protein-protein complexes with published affinity values. The results show that the most efficient PPI complexes generate ~20 cal.mol−1/Å2 of binding energy. A minimum contact area of ~500 Å2 is required for a stable complex, required to generate sufficient interaction energy to pay the entropic cost of co-localizing two proteins from 1 M solution. The most compact and efficient TNF/TNFR complex was BAFF/BR3, which achieved ~80% of the maximum achievable binding efficiency. Other small receptors also gave high binding efficiencies, while the larger receptors generated only 44-49% of this limit despite interacting primarily through just a single small domain. The results provide new insight into how much binding energy can be generated by a PPI interface of a given size, and establish a quantitative method to predict how large a natural or engineered contact interface must be to achieve a given level of binding affinity. PMID:23088250

  11. Protein Attachment on Nanodiamonds.

    PubMed

    Lin, Chung-Lun; Lin, Cheng-Huang; Chang, Huan-Cheng; Su, Meng-Chih

    2015-07-16

    A recent advance in nanotechnology is the scale-up production of small and nonaggregated diamond nanoparticles suitable for biological applications. Using detonation nanodiamonds (NDs) with an average diameter of ∼4 nm as the adsorbents, we have studied the static attachment of three proteins (myoglobin, bovine serum albumin, and insulin) onto the nanoparticles by optical spectroscopy, mass spectrometry, and dynamic light scattering, and electrophoretic zeta potential measurements. Results show that the protein surface coverage is predominantly determined by the competition between protein-protein and protein-ND interactions, giving each protein a unique and characteristic structural configuration in its own complex. Specifically, both myoglobin and bovine serum albumin show a Langmuir-type adsorption behavior, forming 1:1 complexes at saturation, whereas insulin folds into a tightly bound multimer before adsorption. The markedly different adsorption patterns appear to be independent of the protein concentration and are closely related to the affinity of the individual proteins for the NDs. The present study provides a fundamental understanding for the use of NDs as a platform for nanomedical drug delivery. PMID:25815400

  12. Poxviral Ankyrin Proteins

    PubMed Central

    Herbert, Michael H.; Squire, Christopher J.; Mercer, Andrew A

    2015-01-01

    Multiple repeats of the ankyrin motif (ANK) are ubiquitous throughout the kingdoms of life but are absent from most viruses. The main exception to this is the poxvirus family, and specifically the chordopoxviruses, with ANK repeat proteins present in all but three species from separate genera. The poxviral ANK repeat proteins belong to distinct orthologue groups spread over different species, and align well with the phylogeny of their genera. This distribution throughout the chordopoxviruses indicates these proteins were present in an ancestral vertebrate poxvirus, and have since undergone numerous duplication events. Most poxviral ANK repeat proteins contain an unusual topology of multiple ANK motifs starting at the N-terminus with a C-terminal poxviral homologue of the cellular F-box enabling interaction with the cellular SCF ubiquitin ligase complex. The subtle variations between ANK repeat proteins of individual poxviruses suggest an array of different substrates may be bound by these protein-protein interaction domains and, via the F-box, potentially directed to cellular ubiquitination pathways and possible degradation. Known interaction partners of several of these proteins indicate that the NF-κB coordinated anti-viral response is a key target, whilst some poxviral ANK repeat domains also have an F-box independent affect on viral host-range. PMID:25690795

  13. Proteins and Amino Acids

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Proteins are the most abundant substances in living organisms and cells. All proteins are constructed from the same twenty amino acids that are linked together by covalent bonds. Shorter chains of two or more amino acids can be linked by covalent bonds to form polypeptides. There are twenty amino...

  14. Proteins and glasses

    SciTech Connect

    Frauenfelder, H.

    1997-12-31

    The structure, the energy landscape, and the dynamics of proteins and glasses are similar. Both types of systems display characteristic nonexponential time dependencies of relaxation phenomena. Experiments suggest that both, proteins and glasses, are heterogeneous and that this fact causes the observed time dependence. This result is discussed in terms of the rough energy landscape characteristic of complex systems.

  15. Synthesis of Lipidated Proteins.

    PubMed

    Mejuch, Tom; Waldmann, Herbert

    2016-08-17

    Protein lipidation is one of the major post-translational modifications (PTM) of proteins. The attachment of the lipid moiety frequently determines the localization and the function of the lipoproteins. Lipidated proteins participate in many essential biological processes in eukaryotic cells, including vesicular trafficking, signal transduction, and regulation of the immune response. Malfunction of these cellular processes usually leads to various diseases such as cancer. Understanding the mechanism of cellular signaling and identifying the protein-protein and protein-lipid interactions in which the lipoproteins are involved is a crucial task. To achieve these goals, fully functional lipidated proteins are required. However, access to lipoproteins by means of standard expression is often rather limited. Therefore, semisynthetic methods, involving the synthesis of lipidated peptides and their subsequent chemoselective ligation to yield full-length lipoproteins, were developed. In this Review we summarize the commonly used methods for lipoprotein synthesis and the development of the corresponding chemoselective ligation techniques. Several key studies involving full-length semisynthetic lipidated Ras, Rheb, and LC3 proteins are presented. PMID:27444727

  16. The AVIT protein family

    PubMed Central

    Kaser, Alexandra; Winklmayr, Martina; Lepperdinger, Günther; Kreil, Günther

    2003-01-01

    Homologues of a protein originally isolated from snake venom and frog skin secretions are present in many vertebrate species. They contain 80–90 amino acids, 10 of which are cysteines with identical spacing. Various names have been given to these proteins, such as mamba intestinal protein 1 (MIT1), Bv8 (Bombina variegata molecular mass ∼8 kDa), prokineticins and endocrine-gland vascular endothelial growth factor (EG-VEGF). Their amino-terminal sequences are identical, and so we propose that the sequence of their first four residues, AVIT, is used as a name for this family. From a comparison of the sequences, two types of AVIT proteins can be discerned. These proteins seem to be distributed widely in mammalian tissues and are known to bind to G-protein-coupled receptors. Members of this family have been shown to stimulate contraction of the guinea pig ileum, to cause hyperalgesia after injection into rats and to be active as specific growth factors. Moreover, the messenger RNA level of one of these AVIT proteins changes rhythmically in the region of the brain known as the suprachiasmatic nucleus. This shows that members of this new family of small proteins are involved in diverse biological processes. PMID:12728244

  17. Protein Kinases and Addiction

    PubMed Central

    Lee, Anna M.; Messing, Robert O.

    2011-01-01

    Although drugs of abuse have different chemical structures and interact with different protein targets, all appear to usurp common neuronal systems that regulate reward and motivation. Addiction is a complex disease that is thought to involve drug-induced changes in synaptic plasticity due to alterations in cell signaling, gene transcription, and protein synthesis. Recent evidence suggests that drugs of abuse interact with and change a common network of signaling pathways that include a subset of specific protein kinases. The best studied of these kinases are reviewed here and include extracellular signal-regulated kinase, cAMP-dependent protein kinase, cyclin-dependent protein kinase 5, protein kinase C, calcium/calmodulin-dependent protein kinase II, and Fyn tyrosine kinase. These kinases have been implicated in various aspects of drug addiction including acute drug effects, drug self-administration, withdrawal, reinforcement, sensitization, and tolerance. Identifying protein kinase substrates and signaling pathways that contribute to the addicted state may provide novel approaches for new pharma-cotherapies to treat drug addiction. PMID:18991950

  18. Loss of mitochondrial DNA-encoded protein ND1 results in disruption of complex I biogenesis during early stages of assembly.

    PubMed

    Lim, Sze Chern; Hroudová, Jana; Van Bergen, Nicole J; Lopez Sanchez, M Isabel G; Trounce, Ian A; McKenzie, Matthew

    2016-06-01

    Mitochondrial complex I (NADH:ubiquinone oxidoreductase) must be assembled precisely from 45 protein subunits for it to function correctly. One of its mitochondrial DNA (mtDNA) encoded subunits, ND1, is incorporated during the early stages of complex I assembly. However, little is known about how mutations in ND1 affect this assembly process. We found that in human 143B cybrid cells carrying a homoplasmic MT-ND1 mutation, ND1 protein could not be translated. As a result, the early stages of complex I assembly were disrupted, with mature complex I undetectable and complex I-linked respiration severely reduced to 2.0% of control levels. Interestingly, complex IV (ferrocytochrome c:oxygen oxidoreductase) steady-state levels were also reduced to 40.3%, possibly due to its diminished stability in the absence of respiratory supercomplex formation. This was in comparison with 143B cybrid controls (that contained wild-type mtDNA on the same nuclear background), which exhibited normal complex I, complex IV, and supercomplex assembly. We conclude that the loss of ND1 stalls complex I assembly during the early stages of its biogenesis, which not only results in the loss of mature complex I but also disrupts the stability of complex IV and the respiratory supercomplex to cause mitochondrial dysfunction.-Lim, S. C., Hroudová, J., Van Bergen, N. J., Lopez Sanchez, M. I. G., Trounce, I. A., McKenzie, M. Loss of mitochondrial DNA-encoded protein ND1 results in disruption of complex I biogenesis during early stages of assembly. PMID:26929434

  19. Drugging Membrane Protein Interactions

    PubMed Central

    Yin, Hang; Flynn, Aaron D.

    2016-01-01

    The majority of therapeutics target membrane proteins, accessible on the surface of cells, to alter cellular signaling. Cells use membrane proteins to transduce signals into cells, transport ions and molecules, bind the cell to a surface or substrate, and catalyze reactions. Newly devised technologies allow us to drug conventionally “undruggable” regions of membrane proteins, enabling modulation of protein–protein, protein–lipid, and protein–nucleic acid interactions. In this review, we survey the state of the art in high-throughput screening and rational design in drug discovery, and we evaluate the advances in biological understanding and technological capacity that will drive pharmacotherapy forward against unorthodox membrane protein targets. PMID:26863923

  20. Manipulating and Visualizing Proteins

    SciTech Connect

    Simon, Horst D.

    2003-12-05

    ProteinShop Gives Researchers a Hands-On Tool for Manipulating, Visualizing Protein Structures. The Human Genome Project and other biological research efforts are creating an avalanche of new data about the chemical makeup and genetic codes of living organisms. But in order to make sense of this raw data, researchers need software tools which let them explore and model data in a more intuitive fashion. With this in mind, researchers at Lawrence Berkeley National Laboratory and the University of California, Davis, have developed ProteinShop, a visualization and modeling program which allows researchers to manipulate protein structures with pinpoint control, guided in large part by their own biological and experimental instincts. Biologists have spent the last half century trying to unravel the ''protein folding problem,'' which refers to the way chains of amino acids physically fold themselves into three-dimensional proteins. This final shape, which resembles a crumpled ribbon or piece of origami, is what determines how the protein functions and translates genetic information. Understanding and modeling this geometrically complex formation is no easy matter. ProteinShop takes a given sequence of amino acids and uses visualization guides to help generate predictions about the secondary structures, identifying alpha helices and flat beta strands, and the coil regions that bind them. Once secondary structures are in place, researchers can twist and turn these pre-configurations until they come up with a number of possible tertiary structure conformations. In turn, these are fed into a computationally intensive optimization procedure that tries to find the final, three-dimensional protein structure. Most importantly, ProteinShop allows users to add human knowledge and intuition to the protein structure prediction process, thus bypassing bad configurations that would otherwise be fruitless for optimization. This saves compute cycles and accelerates the entire process, so

  1. Proteins, fluctuations and complexity

    SciTech Connect

    Frauenfelder, Hans; Chen, Guo; Fenimore, Paul W

    2008-01-01

    Glasses, supercooled liquids, and proteins share common properties, in particular the existence of two different types of fluctuations, {alpha} and {beta}. While the effect of the {alpha} fluctuations on proteins has been known for a few years, the effect of {beta} fluctuations has not been understood. By comparing neutron scattering data on the protein myoglobin with the {beta} fluctuations in the hydration shell measured by dielectric spectroscopy we show that the internal protein motions are slaved to these fluctuations. We also show that there is no 'dynamic transition' in proteins near 200 K. The rapid increase in the mean square displacement with temperature in many neutron scattering experiments is quantitatively predicted by the {beta} fluctuations in the hydration shell.

  2. Structures of membrane proteins

    PubMed Central

    Vinothkumar, Kutti R.; Henderson, Richard

    2010-01-01

    In reviewing the structures of membrane proteins determined up to the end of 2009, we present in words and pictures the most informative examples from each family. We group the structures together according to their function and architecture to provide an overview of the major principles and variations on the most common themes. The first structures, determined 20 years ago, were those of naturally abundant proteins with limited conformational variability, and each membrane protein structure determined was a major landmark. With the advent of complete genome sequences and efficient expression systems, there has been an explosion in the rate of membrane protein structure determination, with many classes represented. New structures are published every month and more than 150 unique membrane protein structures have been determined. This review analyses the reasons for this success, discusses the challenges that still lie ahead, and presents a concise summary of the key achievements with illustrated examples selected from each class. PMID:20667175

  3. Protein sequence databases.

    PubMed

    Apweiler, Rolf; Bairoch, Amos; Wu, Cathy H

    2004-02-01

    A variety of protein sequence databases exist, ranging from simple sequence repositories, which store data with little or no manual intervention in the creation of the records, to expertly curated universal databases that cover all species and in which the original sequence data are enhanced by the manual addition of further information in each sequence record. As the focus of researchers moves from the genome to the proteins encoded by it, these databases will play an even more important role as central comprehensive resources of protein information. Several the leading protein sequence databases are discussed here, with special emphasis on the databases now provided by the Universal Protein Knowledgebase (UniProt) consortium. PMID:15036160

  4. Proteins in unexpected locations.

    PubMed Central

    Smalheiser, N R

    1996-01-01

    Members of all classes of proteins--cytoskeletal components, secreted growth factors, glycolytic enzymes, kinases, transcription factors, chaperones, transmembrane proteins, and extracellular matrix proteins--have been identified in cellular compartments other than their conventional sites of action. Some of these proteins are expressed as distinct compartment-specific isoforms, have novel mechanisms for intercompartmental translocation, have distinct endogenous biological actions within each compartment, and are regulated in a compartment-specific manner as a function of physiologic state. The possibility that many, if not most, proteins have distinct roles in more than one cellular compartment has implications for the evolution of cell organization and may be important for understanding pathological conditions such as Alzheimer's disease and cancer. PMID:8862516

  5. New York-Structural GenomiX Research Consortium (NYXGXRC): a Large Scale Center for the Protein Structure Initiative

    SciTech Connect

    Bonanno,J.; Almo, S.; Bresnick, A.; Chance, M.; Fiser, A.; Swaminathan, S.; Jiang, J.; Studier, F.; Shapiro, L.; et al.

    2005-01-01

    Structural GenomiX, Inc. (SGX), four New York area institutions, and two University of California schools have formed the New York Structural GenomiX Research Consortium (NYSGXRC), an industrial/academic Research Consortium that exploits individual core competencies to support all aspects of the NIH-NIGMS funded Protein Structure Initiative (PSI), including protein family classification and target selection, generation of protein for biophysical analyses, sample preparation for structural studies, structure determination and analyses, and dissemination of results. At the end of the PSI Pilot Study Phase (PSI-1), the NYSGXRC will be capable of producing 100-200 experimentally determined protein structures annually. All Consortium activities can be scaled to increase production capacity significantly during the Production Phase of the PSI (PSI-2). The Consortium utilizes both centralized and de-centralized production teams with clearly defined deliverables and hand-off procedures that are supported by a web-based target/sample tracking system (SGX Laboratory Information Data Management System, LIMS, and NYSGXRC Internal Consortium Experimental Database, ICE-DB). Consortium management is provided by an Executive Committee, which is composed of the PI and all Co-PIs. Progress to date is tracked on a publicly available Consortium web site (http://www.nysgxrc.org) and all DNA/protein reagents and experimental protocols are distributed freely from the New York City Area institutions. In addition to meeting the requirements of the Pilot Study Phase and preparing for the Production Phase of the PSI, the NYSGXRC aims to develop modular technologies that are transferable to structural biology laboratories in both academe and industry. The NYSGXRC PI and Co-PIs intend the PSI to have a transforming effect on the disciplines of X-ray crystallography and NMR spectroscopy of biological macromolecules. Working with other PSI-funded Centers, the NYSGXRC seeks to create the

  6. Protein crystal growth

    NASA Technical Reports Server (NTRS)

    Bugg, Charles E.

    1993-01-01

    Proteins account for 50% or more of the dry weight of most living systems and play a crucial role in virtually all biological processes. Since the specific functions of essentially all biological molecules are determined by their three-dimensional structures, it is obvious that a detailed understanding of the structural makeup of a protein is essential to any systematic research pertaining to it. At the present time, protein crystallography has no substitute, it is the only technique available for elucidating the atomic arrangements within complicated biological molecules. Most macromolecules are extremely difficult to crystallize, and many otherwise exciting and promising projects have terminated at the crystal growth stage. There is a pressing need to better understand protein crystal growth, and to develop new techniques that can be used to enhance the size and quality of protein crystals. There are several aspects of microgravity that might be exploited to enhance protein crystal growth. The major factor that might be expected to alter crystal growth processes in space is the elimination of density-driven convective flow. Another factor that can be readily controlled in the absence of gravity is the sedimentation of growing crystal in a gravitational field. Another potential advantage of microgravity for protein crystal growth is the option of doing containerless crystal growth. One can readily understand why the microgravity environment established by Earth-orbiting vehicles is perceived to offer unique opportunities for the protein crystallographer. The near term objectives of the Protein Crystal Growth in a Microgravity Environment (PCG/ME) project is to continue to improve the techniques, procedures, and hardware systems used to grow protein crystals in Earth orbit.

  7. The centrality of cancer proteins in human protein-protein interaction network: a revisit.

    PubMed

    Xiong, Wei; Xie, Luyu; Zhou, Shuigeng; Liu, Hui; Guan, Jihong

    2014-01-01

    Topological analysis of protein-protein interaction (PPI) networks has been widely applied to the investigation on cancer mechanisms. However, there is still a debate on whether cancer proteins exhibit more topological centrality compared to the other proteins in the human PPI network. To resolve this debate, we first identified four sets of human proteins, and then mapped these proteins into the yeast PPI network by homologous genes. Finally, we compared these proteins' properties in human and yeast PPI networks. Experiments over two real datasets demonstrated that cancer proteins tend to have higher degree and smaller clustering coefficient than non-cancer proteins. Experimental results also validated that cancer proteins have larger betweenness centrality compared to the other proteins on the STRING dataset. However, on the BioGRID dataset, the average betweenness centrality of cancer proteins is larger than that of disease and control proteins, but smaller than that of essential proteins. PMID:24878726

  8. Protein Regulation in Signal Transduction.

    PubMed

    Lee, Michael J; Yaffe, Michael B

    2016-01-01

    SUMMARYCells must respond to a diverse, complex, and ever-changing mix of signals, using a fairly limited set of parts. Changes in protein level, protein localization, protein activity, and protein-protein interactions are critical aspects of signal transduction, allowing cells to respond highly specifically to a nearly limitless set of cues and also to vary the sensitivity, duration, and dynamics of the response. Signal-dependent changes in levels of gene expression and protein synthesis play an important role in regulation of protein levels, whereas posttranslational modifications of proteins regulate their degradation, localization, and functional interactions. Protein ubiquitylation, for example, can direct proteins to the proteasome for degradation or provide a signal that regulates their interactions and/or location within the cell. Similarly, protein phosphorylation by specific kinases is a key mechanism for augmenting protein activity and relaying signals to other proteins that possess domains that recognize the phosphorylated residues. PMID:27252361

  9. Bacterial Ice Crystal Controlling Proteins

    PubMed Central

    Lorv, Janet S. H.; Rose, David R.; Glick, Bernard R.

    2014-01-01

    Across the world, many ice active bacteria utilize ice crystal controlling proteins for aid in freezing tolerance at subzero temperatures. Ice crystal controlling proteins include both antifreeze and ice nucleation proteins. Antifreeze proteins minimize freezing damage by inhibiting growth of large ice crystals, while ice nucleation proteins induce formation of embryonic ice crystals. Although both protein classes have differing functions, these proteins use the same ice binding mechanisms. Rather than direct binding, it is probable that these protein classes create an ice surface prior to ice crystal surface adsorption. Function is differentiated by molecular size of the protein. This paper reviews the similar and different aspects of bacterial antifreeze and ice nucleation proteins, the role of these proteins in freezing tolerance, prevalence of these proteins in psychrophiles, and current mechanisms of protein-ice interactions. PMID:24579057

  10. Protein Binding Pocket Dynamics.

    PubMed

    Stank, Antonia; Kokh, Daria B; Fuller, Jonathan C; Wade, Rebecca C

    2016-05-17

    The dynamics of protein binding pockets are crucial for their interaction specificity. Structural flexibility allows proteins to adapt to their individual molecular binding partners and facilitates the binding process. This implies the necessity to consider protein internal motion in determining and predicting binding properties and in designing new binders. Although accounting for protein dynamics presents a challenge for computational approaches, it expands the structural and physicochemical space for compound design and thus offers the prospect of improved binding specificity and selectivity. A cavity on the surface or in the interior of a protein that possesses suitable properties for binding a ligand is usually referred to as a binding pocket. The set of amino acid residues around a binding pocket determines its physicochemical characteristics and, together with its shape and location in a protein, defines its functionality. Residues outside the binding site can also have a long-range effect on the properties of the binding pocket. Cavities with similar functionalities are often conserved across protein families. For example, enzyme active sites are usually concave surfaces that present amino acid residues in a suitable configuration for binding low molecular weight compounds. Macromolecular binding pockets, on the other hand, are located on the protein surface and are often shallower. The mobility of proteins allows the opening, closing, and adaptation of binding pockets to regulate binding processes and specific protein functionalities. For example, channels and tunnels can exist permanently or transiently to transport compounds to and from a binding site. The influence of protein flexibility on binding pockets can vary from small changes to an already existent pocket to the formation of a completely new pocket. Here, we review recent developments in computational methods to detect and define binding pockets and to study pocket dynamics. We introduce five

  11. PSC: protein surface classification

    PubMed Central

    Tseng, Yan Yuan; Li, Wen-Hsiung

    2012-01-01

    We recently proposed to classify proteins by their functional surfaces. Using the structural attributes of functional surfaces, we inferred the pairwise relationships of proteins and constructed an expandable database of protein surface classification (PSC). As the functional surface(s) of a protein is the local region where the protein performs its function, our classification may reflect the functional relationships among proteins. Currently, PSC contains a library of 1974 surface types that include 25 857 functional surfaces identified from 24 170 bound structures. The search tool in PSC empowers users to explore related surfaces that share similar local structures and core functions. Each functional surface is characterized by structural attributes, which are geometric, physicochemical or evolutionary features. The attributes have been normalized as descriptors and integrated to produce a profile for each functional surface in PSC. In addition, binding ligands are recorded for comparisons among homologs. PSC allows users to exploit related binding surfaces to reveal the changes in functionally important residues on homologs that have led to functional divergence during evolution. The substitutions at the key residues of a spatial pattern may determine the functional evolution of a protein. In PSC (http://pocket.uchicago.edu/psc/), a pool of changes in residues on similar functional surfaces is provided. PMID:22669905

  12. Structure Prediction of Protein Complexes

    NASA Astrophysics Data System (ADS)

    Pierce, Brian; Weng, Zhiping

    Protein-protein interactions are critical for biological function. They directly and indirectly influence the biological systems of which they are a part. Antibodies bind with antigens to detect and stop viruses and other infectious agents. Cell signaling is performed in many cases through the interactions between proteins. Many diseases involve protein-protein interactions on some level, including cancer and prion diseases.

  13. (PCG) Protein Crystal Growth Canavalin

    NASA Technical Reports Server (NTRS)

    1989-01-01

    (PCG) Protein Crystal Growth Canavalin. The major storage protein of leguminous plants and a major source of dietary protein for humans and domestic animals. It is studied in efforts to enhance nutritional value of proteins through protein engineerings. It is isolated from Jack Bean because of it's potential as a nutritional substance. Principal Investigator on STS-26 was Alex McPherson.

  14. Protein Crystal Quality Studies

    NASA Technical Reports Server (NTRS)

    1998-01-01

    Eddie Snell (standing), Post-Doctoral Fellow the National Research Council (NRC),and Marc Pusey of Marshall Space Flight Center (MSFC) use a reciprocal space mapping diffractometer for marcromolecular crystal quality studies. The diffractometer is used in mapping the structure of marcromolecules such as proteins to determine their structure and thus understand how they function with other proteins in the body. This is one of several analytical tools used on proteins crystalized on Earth and in space experiments. Photo credit: NASA/Marshall Space Flight Center (MSFC)

  15. Protein Crystal Malic Enzyme

    NASA Technical Reports Server (NTRS)

    1992-01-01

    Malic Enzyme is a target protein for drug design because it is a key protein in the life cycle of intestinal parasites. After 2 years of effort on Earth, investigators were unable to produce any crystals that were of high enough quality and for this reason the structure of this important protein could not be determined. Crystals obtained from one STS-50 were of superior quality allowing the structure to be determined. This is just one example why access to space is so vital for these studies. Principal Investigator is Larry DeLucas.

  16. Piezoelectric allostery of protein

    NASA Astrophysics Data System (ADS)

    Ohnuki, Jun; Sato, Takato; Takano, Mitsunori

    2016-07-01

    Allostery is indispensable for a protein to work, where a locally applied stimulus is transmitted to a distant part of the molecule. While the allostery due to chemical stimuli such as ligand binding has long been studied, the growing interest in mechanobiology prompts the study of the mechanically stimulated allostery, the physical mechanism of which has not been established. By molecular dynamics simulation of a motor protein myosin, we found that a locally applied mechanical stimulus induces electrostatic potential change at distant regions, just like the piezoelectricity. This novel allosteric mechanism, "piezoelectric allostery", should be of particularly high value for mechanosensor/transducer proteins.

  17. Emerging fluorescent protein technologies.

    PubMed

    Enterina, Jhon Ralph; Wu, Lanshi; Campbell, Robert E

    2015-08-01

    Fluorescent proteins (FPs), such as the Aequorea jellyfish green FP (GFP), are firmly established as fundamental tools that enable a wide variety of biological studies. Specifically, FPs can serve as versatile genetically encoded markers for tracking proteins, organelles, or whole cells, and as the basis for construction of biosensors that can be used to visualize a growing array of biochemical events in cells and tissues. In this review we will focus on emerging applications of FPs that represent unprecedented new directions for the field. These emerging applications include new strategies for using FPs in biosensing applications, and innovative ways of using FPs to manipulate protein function or gene expression. PMID:26043278

  18. Piezoelectric allostery of protein.

    PubMed

    Ohnuki, Jun; Sato, Takato; Takano, Mitsunori

    2016-07-01

    Allostery is indispensable for a protein to work, where a locally applied stimulus is transmitted to a distant part of the molecule. While the allostery due to chemical stimuli such as ligand binding has long been studied, the growing interest in mechanobiology prompts the study of the mechanically stimulated allostery, the physical mechanism of which has not been established. By molecular dynamics simulation of a motor protein myosin, we found that a locally applied mechanical stimulus induces electrostatic potential change at distant regions, just like the piezoelectricity. This novel allosteric mechanism, "piezoelectric allostery", should be of particularly high value for mechanosensor/transducer proteins. PMID:27575163

  19. Protein crystallography prescreen kit

    DOEpatents

    Segelke, Brent W.; Krupka, Heike I.; Rupp, Bernhard

    2007-10-02

    A kit for prescreening protein concentration for crystallization includes a multiplicity of vials, a multiplicity of pre-selected reagents, and a multiplicity of sample plates. The reagents and a corresponding multiplicity of samples of the protein in solutions of varying concentrations are placed on sample plates. The sample plates containing the reagents and samples are incubated. After incubation the sample plates are examined to determine which of the sample concentrations are too low and which the sample concentrations are too high. The sample concentrations that are optimal for protein crystallization are selected and used.

  20. Protein crystallography prescreen kit

    DOEpatents

    Segelke, Brent W.; Krupka, Heike I.; Rupp, Bernhard

    2005-07-12

    A kit for prescreening protein concentration for crystallization includes a multiplicity of vials, a multiplicity of pre-selected reagents, and a multiplicity of sample plates. The reagents and a corresponding multiplicity of samples of the protein in solutions of varying concentrations are placed on sample plates. The sample plates containing the reagents and samples are incubated. After incubation the sample plates are examined to determine which of the sample concentrations are too low and which the sample concentrations are too high. The sample concentrations that are optimal for protein crystallization are selected and used.

  1. Evolution of proteins.

    NASA Technical Reports Server (NTRS)

    Dayhoff, M. O.

    1971-01-01

    The amino acid sequences of proteins from living organisms are dealt with. The structure of proteins is first discussed; the variation in this structure from one biological group to another is illustrated by the first halves of the sequences of cytochrome c, and a phylogenetic tree is derived from the cytochrome c data. The relative geological times associated with the events of this tree are discussed. Errors which occur in the duplication of cells during the evolutionary process are examined. Particular attention is given to evolution of mutant proteins, globins, ferredoxin, and transfer ribonucleic acids (tRNA's). Finally, a general outline of biological evolution is presented.

  2. Protein based Block Copolymers

    PubMed Central

    Rabotyagova, Olena S.; Cebe, Peggy; Kaplan, David L.

    2011-01-01

    Advances in genetic engineering have led to the synthesis of protein-based block copolymers with control of chemistry and molecular weight, resulting in unique physical and biological properties. The benefits from incorporating peptide blocks into copolymer designs arise from the fundamental properties of proteins to adopt ordered conformations and to undergo self-assembly, providing control over structure formation at various length scales when compared to conventional block copolymers. This review covers the synthesis, structure, assembly, properties, and applications of protein-based block copolymers. PMID:21235251

  3. A Bayesian Estimator of Protein-Protein Association Probabilities

    SciTech Connect

    Gilmore, Jason M.; Auberry, Deanna L.; Sharp, Julia L.; White, Amanda M.; Anderson, Kevin K.; Daly, Don S.

    2008-07-01

    The Bayesian Estimator of Protein-Protein Association Probabilities (BEPro3) is a software tool for estimating probabilities of protein-protein association between bait and prey protein pairs using data from multiple-bait, multiple-replicate, protein pull-down LC-MS assay experiments. BEPro3 is open source software that runs on both Windows XP and Mac OS 10.4 or newer versions, and is freely available from http://www.pnl.gov/statistics/BEPro3.

  4. Interactive protein manipulation

    SciTech Connect

    SNCrivelli@lbl.gov

    2003-07-01

    We describe an interactive visualization and modeling program for the creation of protein structures ''from scratch''. The input to our program is an amino acid sequence -decoded from a gene- and a sequence of predicted secondary structure types for each amino acid-provided by external structure prediction programs. Our program can be used in the set-up phase of a protein structure prediction process; the structures created with it serve as input for a subsequent global internal energy minimization, or another method of protein structure prediction. Our program supports basic visualization methods for protein structures, interactive manipulation based on inverse kinematics, and visualization guides to aid a user in creating ''good'' initial structures.

  5. Protein Colloidal Aggregation Project

    NASA Technical Reports Server (NTRS)

    Oliva-Buisson, Yvette J. (Compiler)

    2014-01-01

    To investigate the pathways and kinetics of protein aggregation to allow accurate predictive modeling of the process and evaluation of potential inhibitors to prevalent diseases including cataract formation, chronic traumatic encephalopathy, Alzheimer's Disease, Parkinson's Disease and others.

  6. Engineered Proteins for Bioelectrochemistry

    NASA Astrophysics Data System (ADS)

    Akram, Muhammad Safwan; Rehman, Jawad Ur; Hall, Elizabeth A. H.

    2014-06-01

    It is only in the past two decades that excellent protein engineering tools have begun to meet parallel advances in materials chemistry, nanofabrication, and electronics. This is revealing scenarios from which synthetic enzymes can emerge, which were previously impossible, as well as interfaces with novel electrode materials. That means the control of the protein structure, electron transport pathway, and electrode surface can usher us into a new era of bioelectrochemistry. This article reviews the principle of electron transfer (ET) and considers how its application at the electrode, within the protein, and at a redox group is directing key advances in the understanding of protein structure to create systems that exhibit better efficiency and unique bioelectrochemistry.

  7. Protein Model Database

    SciTech Connect

    Fidelis, K; Adzhubej, A; Kryshtafovych, A; Daniluk, P

    2005-02-23

    The phenomenal success of the genome sequencing projects reveals the power of completeness in revolutionizing biological science. Currently it is possible to sequence entire organisms at a time, allowing for a systemic rather than fractional view of their organization and the various genome-encoded functions. There is an international plan to move towards a similar goal in the area of protein structure. This will not be achieved by experiment alone, but rather by a combination of efforts in crystallography, NMR spectroscopy, and computational modeling. Only a small fraction of structures are expected to be identified experimentally, the remainder to be modeled. Presently there is no organized infrastructure to critically evaluate and present these data to the biological community. The goal of the Protein Model Database project is to create such infrastructure, including (1) public database of theoretically derived protein structures; (2) reliable annotation of protein model quality, (3) novel structure analysis tools, and (4) access to the highest quality modeling techniques available.

  8. The Pentapeptide Repeat Proteins

    SciTech Connect

    Vetting,M.; Hegde, S.; Fajardo, J.; Fiser, A.; Roderick, S.; Takiff, H.; Blanchard, J.

    2006-01-01

    The Pentapeptide Repeat Protein (PRP) family has over 500 members in the prokaryotic and eukaryotic kingdoms. These proteins are composed of, or contain domains composed of, tandemly repeated amino acid sequences with a consensus sequence of [S, T,A, V][D, N][L, F]-[S, T,R][G]. The biochemical function of the vast majority of PRP family members is unknown. The three-dimensional structure of the first member of the PRP family was determined for the fluoroquinolone resistance protein (MfpA) from Mycobacterium tuberculosis. The structure revealed that the pentapeptide repeats encode the folding of a novel right-handed quadrilateral {beta}-helix. MfpA binds to DNA gyrase and inhibits its activity. The rod-shaped, dimeric protein exhibits remarkable size, shape and electrostatic similarity to DNA.

  9. Untying knots in proteins.

    PubMed

    Sułkowska, Joanna I; Sułkowski, Piotr; Szymczak, Piotr; Cieplak, Marek

    2010-10-13

    A shoelace can be readily untied by pulling its ends rather than its loops. Attempting to untie a native knot in a protein can also succeed or fail depending on where one pulls. However, thermal fluctuations induced by the surrounding water affect conformations stochastically and may add to the uncertainty of the outcome. When the protein is pulled by the termini, the knot can only get tightened, and any attempt at untying results in failure. We show that, by pulling specific amino acids, one may easily retract a terminal segment of the backbone from the knotting loop and untangle the knot. At still other amino acids, the outcome of pulling can go either way. We study the dependence of the untying probability on the way the protein is grasped, the pulling speed, and the temperature. Elucidation of the mechanisms underlying this dependence is critical for a successful experimental realization of protein knot untying. PMID:20857930

  10. Membrane Protein Prediction Methods

    PubMed Central

    Punta, Marco; Forrest, Lucy R.; Bigelow, Henry; Kernytsky, Andrew; Liu, Jinfeng; Rost, Burkhard

    2007-01-01

    We survey computational approaches that tackle membrane protein structure and function prediction. While describing the main ideas that have led to the development of the most relevant and novel methods, we also discuss pitfalls, provide practical hints and highlight the challenges that remain. The methods covered include: sequence alignment, motif search, functional residue identification, transmembrane segment and protein topology predictions, homology and ab initio modeling. Overall, predictions of functional and structural features of membrane proteins are improving, although progress is hampered by the limited amount of high-resolution experimental information available. While predictions of transmembrane segments and protein topology rank among the most accurate methods in computational biology, more attention and effort will be required in the future to ameliorate database search, homology and ab initio modeling. PMID:17367718

  11. Bence-Jones protein - quantitative

    MedlinePlus

    Immunoglobulin light chains - urine; Urine Bence-Jones protein ... Bence-Jones proteins are a part of regular antibodies called light chains. These proteins are not normally in urine. Sometimes, when ...

  12. Protein Nitrogen Determination

    NASA Astrophysics Data System (ADS)

    Nielsen, S. Suzanne

    The protein content of foods can be determined by numerous methods. The Kjeldahl method and the nitrogen combustion (Dumas) method for protein analysis are based on nitrogen determination. Both methods are official for the purposes of nutrition labeling of foods. While the Kjeldahl method has been used widely for over a hundred years, the recent availability of automated instrumentation for the Dumas method in many cases is replacing use of the Kjeldahl method.

  13. The Malignant Protein Puzzle.

    PubMed

    Walker, Lary C; Jucker, Mathias

    2016-01-01

    When most people hear the words malignant and brain, cancer immediately comes to mind. But our authors argue that proteins can be malignant too, and can spread harmfully through the brain in neurodegenerative diseases that include Alzheimer's, Parkinson's, CTE, and ALS. Studying how proteins such as PrP, amyloid beta, tau, and others aggregate and spread, and kill brain cells, represents a crucial new frontier in neuroscience. PMID:27408676

  14. Recombinant Collagenlike Proteins

    NASA Technical Reports Server (NTRS)

    Fertala, Andzej

    2007-01-01

    A group of collagenlike recombinant proteins containing high densities of biologically active sites has been invented. The method used to express these proteins is similar to a method of expressing recombinant procollagens and collagens described in U. S. Patent 5,593,859, "Synthesis of human procollagens and collagens in recombinant DNA systems." Customized collagenous proteins are needed for biomedical applications. In particular, fibrillar collagens are attractive for production of matrices needed for tissue engineering and drug delivery. Prior to this invention, there was no way of producing customized collagenous proteins for these and other applications. Heretofore, collagenous proteins have been produced by use of such biological systems as yeasts, bacteria, and transgenic animals and plants. These products are normal collagens that can also be extracted from such sources as tendons, bones, and hides. These products cannot be made to consist only of biologically active, specific amino acid sequences that may be needed for specific applications. Prior to this invention, it had been established that fibrillar collagens consist of domains that are responsible for such processes as interaction with cells, binding of growth factors, and interaction with a number of structural proteins present in the extracellular matrix. A normal collagen consists of a sequence of domains that can be represented by a corresponding sequence of labels, e.g., D1D2D3D4. A collagenlike protein of the present invention contains regions of collagen II that contain multiples of a single domain (e.g., D1D1D1D1 or D4D4D4D4) chosen for its specific biological activity. By virtue of the multiplicity of the chosen domain, the density of sites having that specific biological activity is greater than it is in a normal collagen. A collagenlike protein according to this invention can thus be made to have properties that are necessary for tissue engineering.

  15. Protein conducting nanopores

    NASA Astrophysics Data System (ADS)

    Harsman, Anke; Krüger, Vivien; Bartsch, Philipp; Honigmann, Alf; Schmidt, Oliver; Rao, Sanjana; Meisinger, Christof; Wagner, Richard

    2010-11-01

    About 50% of the cellular proteins have to be transported into or across cellular membranes. This transport is an essential step in the protein biosynthesis. In eukaryotic cells secretory proteins are transported into the endoplasmic reticulum before they are transported in vesicles to the plasma membrane. Almost all proteins of the endosymbiotic organelles chloroplasts and mitochondria are synthesized on cytosolic ribosomes and posttranslationally imported. Genetic, biochemical and biophysical approaches led to rather detailed knowledge on the composition of the translocon-complexes which catalyze the membrane transport of the preproteins. Comprehensive concepts on the targeting and membrane transport of polypeptides emerged, however little detail on the molecular nature and mechanisms of the protein translocation channels comprising nanopores has been achieved. In this paper we will highlight recent developments of the diverse protein translocation systems and focus particularly on the common biophysical properties and functions of the protein conducting nanopores. We also provide a first analysis of the interaction between the genuine protein conducting nanopore Tom40SC as well as a mutant Tom40SC (\\mathrm {S}_{54} \\to E ) containing an additional negative charge at the channel vestibule and one of its native substrates, CoxIV, a mitochondrial targeting peptide. The polypeptide induced a voltage-dependent increase in the frequency of channel closure of Tom40SC corresponding to a voltage-dependent association rate, which was even more pronounced for the Tom40SC S54E mutant. The corresponding dwelltime reflecting association/transport of the peptide could be determined with \\bar {t}_{\\mathrm {off}} \\cong 1.1 ms for the wildtype, whereas the mutant Tom40SC S54E displayed a biphasic dwelltime distribution (\\bar {t}_{\\mathrm {off}}^1 \\cong 0.4 ms \\bar {t}_{\\mathrm {off}}^2 \\cong 4.6 ms).

  16. Cotton and Protein Interactions

    SciTech Connect

    Goheen, Steven C.; Edwards, J. V.; Rayburn, Alfred R.; Gaither, Kari A.; Castro, Nathan J.

    2006-06-30

    The adsorbent properties of important wound fluid proteins and cotton cellulose are reviewed. This review focuses on the adsorption of albumin to cotton-based wound dressings and some chemically modified derivatives targeted for chronic wounds. Adsorption of elastase in the presence of albumin was examined as a model to understand the interactive properties of these wound fluid components with cotton fibers. In the chronic non-healing wound, elastase appears to be over-expressed, and it digests tissue and growth factors, interfering with the normal healing process. Albumin is the most prevalent protein in wound fluid, and in highly to moderately exudative wounds, it may bind significantly to the fibers of wound dressings. Thus, the relative binding properties of both elastase and albumin to wound dressing fibers are of interest in the design of more effective wound dressings. The present work examines the binding of albumin to two different derivatives of cotton, and quantifies the elastase binding to the same derivatives following exposure of albumin to the fiber surface. An HPLC adsorption technique was employed coupled with a colorimetric enzyme assay to quantify the relative binding properties of albumin and elastase to cotton. The results of wound protein binding are discussed in relation to the porosity and surface chemistry interactions of cotton and wound proteins. Studies are directed to understanding the implications of protein adsorption phenomena in terms of fiber-protein models that have implications for rationally designing dressings for chronic wounds.

  17. Stretching to Understand Proteins

    NASA Astrophysics Data System (ADS)

    Cieplak, Marek

    2007-03-01

    Mechanical stretching of single proteins has been studied experimentally for about 50 proteins yielding a variety of force patterns and values of the peak forces. We have performed a theoretical survey of 7749 proteins of known native structure and map out the landscape of possible dynamical behaviors unders stretching at constant speed. The model used is constructed based on the native geometry. It is solved by methods of molecular dynamics and validated by comparing the theoretical predictions to experimental results. We characterize the distribution of peak forces and on correlations with the system size and with the structure classification as characterized by the CATH scheme. We identify proteins with the biggest forces and show that they belong to few topology classes. We determine which protein segments act as mechanical clamps and show that, in most cases, they correspond to long stretches of parallel beta-strands, but other mechanisms are also possible. We then consider stretching by fluid flows. We show that unfolding induced by a uniform flow shows a richer behavior than that in the force clamp. The dynamics of unfolding is found to depend strongly on the selection of the amino acid, usually one of the termini, which is anchored. These features offer potentially wider diagnostic tools to investigate structure of proteins compared to experiments based on the atomic force microscopy.

  18. Fast protein folding kinetics

    PubMed Central

    Gelman, Hannah; Gruebele, Martin

    2014-01-01

    Fast folding proteins have been a major focus of computational and experimental study because they are accessible to both techniques: they are small and fast enough to be reasonably simulated with current computational power, but have dynamics slow enough to be observed with specially developed experimental techniques. This coupled study of fast folding proteins has provided insight into the mechanisms which allow some proteins to find their native conformation well less than 1 ms and has uncovered examples of theoretically predicted phenomena such as downhill folding. The study of fast folders also informs our understanding of even “slow” folding processes: fast folders are small, relatively simple protein domains and the principles that govern their folding also govern the folding of more complex systems. This review summarizes the major theoretical and experimental techniques used to study fast folding proteins and provides an overview of the major findings of fast folding research. Finally, we examine the themes that have emerged from studying fast folders and briefly summarize their application to protein folding in general as well as some work that is left to do. PMID:24641816

  19. Use of protein-protein interactions in affinity chromatography.

    PubMed

    Muronetz, V I; Sholukh, M; Korpela, T

    2001-10-30

    Biospecific recognition between proteins is a phenomenon that can be exploited for designing affinity-chromatographic purification systems for proteins. In principle, the approach is straightforward, and there are usually many alternative ways, since a protein can be always found which binds specifically enough to the desired protein. Routine immunoaffinity chromatography utilizes the recognition of antigenic epitopes by antibodies. However, forces involved in protein-protein interactions as well the forces keeping the three-dimensional structures of proteins intact are complicated, and proteins are easily unfolded by various factors with unpredictable results. Because of this and because of the generally high association strength between proteins, the correct adjustment of binding forces between an immobilized protein and the protein to be purified as well as the release of bound proteins in biologically active form from affinity complexes are the main problem. Affinity systems involving interactions like enzyme-enzyme, subunit-oligomer, protein-antibody, protein-chaperone and the specific features involved in each case are presented as examples. This article also aims to sketch prospects for further development of the use of protein-protein interactions for the purification of proteins. PMID:11694271

  20. Protein crystal growth in space

    NASA Technical Reports Server (NTRS)

    Bugg, C. E.; Clifford, D. W.

    1987-01-01

    The advantages of protein crystallization in space, and the applications of protein crystallography to drug design, protein engineering, and the design of synthetic vaccines are examined. The steps involved in using protein crystallography to determine the three-dimensional structure of a protein are discussed. The growth chamber design and the hand-held apparatus developed for protein crystal growth by vapor diffusion techniques (hanging-drop method) are described; the experimental data from the four Shuttle missions are utilized to develop hardware for protein crystal growth in space and to evaluate the effects of gravity on protein crystal growth.

  1. Multifunctional protein: cardiac ankyrin repeat protein*

    PubMed Central

    Zhang, Na; Xie, Xiao-jie; Wang, Jian-an

    2016-01-01

    Cardiac ankyrin repeat protein (CARP) not only serves as an important component of muscle sarcomere in the cytoplasm, but also acts as a transcription co-factor in the nucleus. Previous studies have demonstrated that CARP is up-regulated in some cardiovascular disorders and muscle diseases; however, its role in these diseases remains controversial now. In this review, we will discuss the continued progress in the research related to CARP, including its discovery, structure, and the role it plays in cardiac development and heart diseases. PMID:27143260

  2. Bioinformatics and Moonlighting Proteins.

    PubMed

    Hernández, Sergio; Franco, Luís; Calvo, Alejandra; Ferragut, Gabriela; Hermoso, Antoni; Amela, Isaac; Gómez, Antonio; Querol, Enrique; Cedano, Juan

    2015-01-01

    Multitasking or moonlighting is the capability of some proteins to execute two or more biochemical functions. Usually, moonlighting proteins are experimentally revealed by serendipity. For this reason, it would be helpful that Bioinformatics could predict this multifunctionality, especially because of the large amounts of sequences from genome projects. In the present work, we analyze and describe several approaches that use sequences, structures, interactomics, and current bioinformatics algorithms and programs to try to overcome this problem. Among these approaches are (a) remote homology searches using Psi-Blast, (b) detection of functional motifs and domains, (c) analysis of data from protein-protein interaction databases (PPIs), (d) match the query protein sequence to 3D databases (i.e., algorithms as PISITE), and (e) mutation correlation analysis between amino acids by algorithms as MISTIC. Programs designed to identify functional motif/domains detect mainly the canonical function but usually fail in the detection of the moonlighting one, Pfam and ProDom being the best methods. Remote homology search by Psi-Blast combined with data from interactomics databases (PPIs) has the best performance. Structural information and mutation correlation analysis can help us to map the functional sites. Mutation correlation analysis can only be used in very specific situations - it requires the existence of multialigned family protein sequences - but can suggest how the evolutionary process of second function acquisition took place. The multitasking protein database MultitaskProtDB (http://wallace.uab.es/multitask/), previously published by our group, has been used as a benchmark for the all of the analyses. PMID:26157797

  3. Self-Assembling Protein Microarrays

    NASA Astrophysics Data System (ADS)

    Ramachandran, Niroshan; Hainsworth, Eugenie; Bhullar, Bhupinder; Eisenstein, Samuel; Rosen, Benjamin; Lau, Albert Y.; C. Walter, Johannes; LaBaer, Joshua

    2004-07-01

    Protein microarrays provide a powerful tool for the study of protein function. However, they are not widely used, in part because of the challenges in producing proteins to spot on the arrays. We generated protein microarrays by printing complementary DNAs onto glass slides and then translating target proteins with mammalian reticulocyte lysate. Epitope tags fused to the proteins allowed them to be immobilized in situ. This obviated the need to purify proteins, avoided protein stability problems during storage, and captured sufficient protein for functional studies. We used the technology to map pairwise interactions among 29 human DNA replication initiation proteins, recapitulate the regulation of Cdt1 binding to select replication proteins, and map its geminin-binding domain.

  4. Purine inhibitors of protein kinases, G proteins and polymerases

    DOEpatents

    Gray, Nathanael S.; Schultz, Peter; Kim, Sung-Hou; Meijer, Laurent

    2001-07-03

    The present invention relates to purine analogs that inhibit, inter alia, protein kinases, G-proteins and polymerases. In addition, the present invention relates to methods of using such purine analogs to inhibit protein kinases, G-proteins, polymerases and other cellular processes and to treat cellular proliferative diseases.

  5. Benchtop Detection of Proteins

    NASA Technical Reports Server (NTRS)

    Scardelletti, Maximilian C.; Varaljay, Vanessa

    2007-01-01

    A process, and a benchtop-scale apparatus for implementing the process, have been developed to detect proteins associated with specific microbes in water. The process and apparatus may also be useful for detection of proteins in other, more complex liquids. There may be numerous potential applications, including monitoring lakes and streams for contamination, testing of blood and other bodily fluids in medical laboratories, and testing for microbial contamination of liquids in restaurants and industrial food-processing facilities. A sample can be prepared and analyzed by use of this process and apparatus within minutes, whereas an equivalent analysis performed by use of other processes and equipment can often take hours to days. The process begins with the conjugation of near-infrared-fluorescent dyes to antibodies that are specific to a particular protein. Initially, the research has focused on using near-infrared dyes to detect antigens or associated proteins in solution, which has proven successful vs. microbial cells, and streamlining the technique in use for surface protein detection on microbes would theoretically render similar results. However, it is noted that additional work is needed to transition protein-based techniques to microbial cell detection. Consequently, multiple such dye/antibody pairs could be prepared to enable detection of multiple selected microbial species, using a different dye for each species. When excited by near-infrared light of a suitable wavelength, each dye fluoresces at a unique longer wavelength that differs from those of the other dyes, enabling discrimination among the various species. In initial tests, the dye/antibody pairs are mixed into a solution suspected of containing the selected proteins, causing the binding of the dye/antibody pairs to such suspect proteins that may be present. The solution is then run through a microcentrifuge that includes a membrane that acts as a filter in that it retains the dye/antibody/protein

  6. Solid State NMR and Protein-Protein Interactions in Membranes

    PubMed Central

    Miao, Yimin; Cross, Timothy A.

    2013-01-01

    Solid state NMR spectroscopy has evolved rapidly in recent years into an excellent tool for the characterization of membrane proteins and their complexes. In the past few years it has also become clear that the structure of membrane proteins, especially helical membrane proteins is determined, in part, by the membrane environment. Therefore, the modeling of this environment by a liquid crystalline lipid bilayer for solid state NMR has generated a unique tool for the characterization of native conformational states, local and global dynamics, and high resolution structure for these proteins. Protein-protein interactions can also benefit from this solid state NMR capability to characterize membrane proteins in a native-like environment. These complexes take the form of oligomeric structures and hetero-protein interactions both with water soluble proteins and other membrane proteins. PMID:24034903

  7. Solid state NMR and protein-protein interactions in membranes.

    PubMed

    Miao, Yimin; Cross, Timothy A

    2013-12-01

    Solid state NMR spectroscopy has evolved rapidly in recent years into an excellent tool for the characterization of membrane proteins and their complexes. In the past few years it has also become clear that the structure of membrane proteins, especially helical membrane proteins is determined, in part, by the membrane environment. Therefore, the modeling of this environment by a liquid crystalline lipid bilayer for solid state NMR has generated a unique tool for the characterization of native conformational states, local and global dynamics, and high-resolution structure for these proteins. Protein-protein interactions can also benefit from this solid state NMR capability to characterize membrane proteins in a native-like environment. These complexes take the form of oligomeric structures and hetero-protein interactions both with water-soluble proteins and other membrane proteins. PMID:24034903

  8. The detection of DNA-binding proteins by protein blotting.

    PubMed Central

    Bowen, B; Steinberg, J; Laemmli, U K; Weintraub, H

    1980-01-01

    A method, called "protein blotting," for the detection of DNA-binding proteins is described. Proteins are separated on an SDA-polyacrylamide gel. The gel is sandwiched between 2 nitrocellulose filters and the proteins allowed to diffuse out of the gel and onto the filters. The proteins are tightly bound to each filter, producing a replica of the original gel pattern. The replica is used to detect DNA-binding proteins, RNA-binding proteins or histone-binding proteins by incubation of the filter with [32P]DNA, [125I]RNA, or [125I] histone. Evidence is also presented that specific protein-DNA interactions may be detected by this technique; under appropriate conditions, the lac repressor binds only to DNA containing the lac operator. Strategies for the detection of specific protein-DNA interactions are discussed. Images PMID:6243775

  9. Histophilus somni Surface Proteins.

    PubMed

    Corbeil, Lynette B

    2016-01-01

    The pathogen surface is usually the first site of interaction with the host. Histophilus somni was earlier thought to only have an outer membrane on its surface. Now it is known that the surface is composed of many virulence factors, including outer membrane proteins, lipooligosaccharide or endotoxin, a fibrillar network, and an exopolysaccharide. Outer membrane blebs, endotoxin, the fibrillar network, and the exopolysaccharide are also shed from the surface. This review will focus on the surface proteins of this pathogen that may colonize the mucosal surface of ruminants as a commensal or may cause pneumonia, septicemia, myocarditis, thrombotic meningoencephalitis, arthritis, and/or abortion. The major outer membrane protein has been well studied. Since its size and epitopes vary from strain to strain, it may be useful for typing strains. Iron-regulated OMPs have also received much attention because of their role in iron uptake for in vivo growth of H. somni. Other OMPs may be protective, based on passive immunization with monospecific antibodies and active immunization experiments. The surface and shed fibrillar network has been shown to be an immunoglobulin-binding protein in that it binds bovine IgG2 by the Fc portion. Two repeat domains (DR1 and DR2) have cytotoxic Fic motifs. Vaccine studies with recombinant DR2 are promising. Studies of the bacterial genome as well as comparison of surface proteins of different strains from the various H. somni syndromes and carrier states will be discussed and have provided much insight into pathogenesis and protection. PMID:26728061

  10. Plant protein kinase substrates identification using protein microarrays.

    PubMed

    Ma, Shisong; Dinesh-Kumar, Savithramma P

    2015-01-01

    Protein kinases regulate signaling pathways by phosphorylating their targets. They play critical roles in plant signaling networks. Although many important protein kinases have been identified in plants, their substrates are largely unknown. We have developed and produced plant protein microarrays with more than 15,000 purified plant proteins. Here, we describe a detailed protocol to use these microarrays to identify plant protein kinase substrates via in vitro phosphorylation assays on these arrays. PMID:25930701

  11. How Many Protein-Protein Interactions Types Exist in Nature?

    PubMed Central

    Mitra, Pralay; Zhang, Yang

    2012-01-01

    Protein quaternary structure universe” refers to the ensemble of all protein-protein complexes across all organisms in nature. The number of quaternary folds thus corresponds to the number of ways proteins physically interact with other proteins. This study focuses on answering two basic questions: Whether the number of protein-protein interactions is limited and, if yes, how many different quaternary folds exist in nature. By all-to-all sequence and structure comparisons, we grouped the protein complexes in the protein data bank (PDB) into 3,629 families and 1,761 folds. A statistical model was introduced to obtain the quantitative relation between the numbers of quaternary families and quaternary folds in nature. The total number of possible protein-protein interactions was estimated around 4,000, which indicates that the current protein repository contains only 42% of quaternary folds in nature and a full coverage needs approximately a quarter century of experimental effort. The results have important implications to the protein complex structural modeling and the structure genomics of protein-protein interactions. PMID:22719985

  12. How many protein-protein interactions types exist in nature?

    PubMed

    Garma, Leonardo; Mukherjee, Srayanta; Mitra, Pralay; Zhang, Yang

    2012-01-01

    "Protein quaternary structure universe" refers to the ensemble of all protein-protein complexes across all organisms in nature. The number of quaternary folds thus corresponds to the number of ways proteins physically interact with other proteins. This study focuses on answering two basic questions: Whether the number of protein-protein interactions is limited and, if yes, how many different quaternary folds exist in nature. By all-to-all sequence and structure comparisons, we grouped the protein complexes in the protein data bank (PDB) into 3,629 families and 1,761 folds. A statistical model was introduced to obtain the quantitative relation between the numbers of quaternary families and quaternary folds in nature. The total number of possible protein-protein interactions was estimated around 4,000, which indicates that the current protein repository contains only 42% of quaternary folds in nature and a full coverage needs approximately a quarter century of experimental effort. The results have important implications to the protein complex structural modeling and the structure genomics of protein-protein interactions. PMID:22719985

  13. Computational drug design targeting protein-protein interactions.

    PubMed

    Bienstock, Rachelle J

    2012-01-01

    Novel discoveries in molecular disease pathways within the cell, combined with increasing information regarding protein binding partners has lead to a new approach in drug discovery. There is interest in designing drugs to modulate protein-protein interactions as opposed to solely targeting the catalytic active site within a single enzyme or protein. There are many challenges in this new approach to drug discovery, particularly since the protein-protein interface has a larger surface area, can comprise a discontinuous epitope, and is more amorphous and less well defined than the typical drug design target, a small contained enzyme-binding pocket. Computational methods to predict modes of protein-protein interaction, as well as protein interface hot spots, have garnered significant interest, in order to facilitate the development of drugs to successfully disrupt and inhibit protein-protein interactions. This review summarizes some current methods available for computational protein-protein docking, as well as tabulating some examples of the successful design of antagonists and small molecule inhibitors for protein-protein interactions. Several of these drugs are now beginning to appear in the clinic. PMID:22316151

  14. Advanced protein formulations

    PubMed Central

    Wang, Wei

    2015-01-01

    It is well recognized that protein product development is far more challenging than that for small-molecule drugs. The major challenges include inherent sensitivity to different types of stresses during the drug product manufacturing process, high rate of physical and chemical degradation during long-term storage, and enhanced aggregation and/or viscosity at high protein concentrations. In the past decade, many novel formulation concepts and technologies have been or are being developed to address these product development challenges for proteins. These concepts and technologies include use of uncommon/combination of formulation stabilizers, conjugation or fusion with potential stabilizers, site-specific mutagenesis, and preparation of nontraditional types of dosage forms—semiaqueous solutions, nonfreeze-dried solid formulations, suspensions, and other emerging concepts. No one technology appears to be mature, ideal, and/or adequate to address all the challenges. These gaps will likely remain in the foreseeable future and need significant efforts for ultimate resolution. PMID:25858529

  15. Thermodynamics of Protein Aggregation

    NASA Astrophysics Data System (ADS)

    Osborne, Kenneth L.; Barz, Bogdan; Bachmann, Michael; Strodel, Birgit

    Amyloid protein aggregation characterizes many neurodegenerative disorders, including Alzheimer's, Parkinson's, and Creutz- feldt-Jakob disease. Evidence suggests that amyloid aggregates may share similar aggregation pathways, implying simulation of full-length amyloid proteins is not necessary for understanding amyloid formation. In this study we simulate GNNQQNY, the N-terminal prion-determining domain of the yeast protein Sup35 to investigate the thermodynamics of structural transitions during aggregation. We use a coarse-grained model with replica-exchange molecular dynamics to investigate the association of 3-, 6-, and 12-chain GNNQQNY systems and we determine the aggregation pathway by studying aggregation states of GN- NQQNY. We find that the aggregation of the hydrophilic GNNQQNY sequence is mainly driven by H-bond formation, leading to the formation of /3-sheets from the very beginning of the assembly process. Condensation (aggregation) and ordering take place simultaneously, which is underpinned by the occurrence of a single heat capacity peak only.

  16. Collapse transition in proteins.

    PubMed

    Ziv, Guy; Thirumalai, D; Haran, Gilad

    2009-01-01

    The coil-globule transition, a tenet of the physics of polymers, has been identified in recent years as an important unresolved aspect of the initial stages of the folding of proteins. We describe the basics of the collapse transition, starting with homopolymers and continuing with proteins. Studies of denatured-state collapse under equilibrium are then presented. An emphasis is placed on single-molecule fluorescence experiments, which are particularly useful for measuring properties of the denatured state even under conditions of coexistence with the folded state. Attempts to understand the dynamics of collapse, both theoretically and experimentally, are then described. Only an upper limit for the rate of collapse has been obtained so far. Improvements in experimental and theoretical methodology are likely to continue to push our understanding of the importance of the denatured-state thermodynamics and dynamics for protein folding in the coming years. PMID:19081910

  17. Polarizable protein packing.

    PubMed

    Ng, Albert H; Snow, Christopher D

    2011-05-01

    To incorporate protein polarization effects within a protein combinatorial optimization framework, we decompose the polarizable force field AMOEBA into low order terms. Including terms up to the third-order provides a fair approximation to the full energy while maintaining tractability. We represent the polarizable packing problem for protein G as a hypergraph and solve for optimal rotamers with the FASTER combinatorial optimization algorithm. These approximate energy models can be improved to high accuracy [root mean square deviation (rmsd) < 1 kJ mol(-1)] via ridge regression. The resulting trained approximations are used to efficiently identify new, low-energy solutions. The approach is general and should allow combinatorial optimization of other many-body problems. PMID:21264879

  18. Matricellular proteins and biomaterials

    PubMed Central

    Morris, Aaron H.; Kyriakides, Themis R.

    2014-01-01

    Biomaterials are essential to modern medicine as components of reconstructive implants, implantable sensors, and vehicles for localized drug delivery. Advances in biomaterials have led to progression from simply making implants that are nontoxic to making implants that are specifically designed to elicit particular functions within the host. The interaction of implants and the extracellular matrix during the foreign body response is a growing area of concern for the field of biomaterials, because it can lead to implant failure. Expression of matricellular proteins is modulated during the foreign body response and these proteins interact with biomaterials. The design of biomaterials to specifically alter the levels of matricellular proteins surrounding implants provides a new avenue for the design and fabrication of biomimetic biomaterials. PMID:24657843

  19. Electron transfer in proteins.

    PubMed

    Gray, H B; Winkler, J R

    1996-01-01

    Electron-transfer (ET) reactions are key steps in a diverse array of biological transformations ranging from photosynthesis to aerobic respiration. A powerful theoretical formalism has been developed that describes ET rates in terms of two parameters: the nuclear reorganization energy (lambda) and the electronic-coupling strength (HAB). Studies of ET reactions in ruthenium-modified proteins have probed lambda and HAB in several metalloproteins (cytochrome c, myoglobin, azurin). This work has shown that protein reorganization energies are sensitive to the medium surrounding the redox sites and that an aqueous environment, in particular, leads to large reorganization energies. Analyses of electronic-coupling strengths suggest that the efficiency of long-range ET depends on the protein secondary structure: beta sheets appear to mediate coupling more efficiently than alpha-helical structures, and hydrogen bonds play a critical role in both. PMID:8811189

  20. Protein Crystal Serum Albumin

    NASA Technical Reports Server (NTRS)

    1998-01-01

    As the most abundant protein in the circulatory system albumin contributes 80% to colloid osmotic blood pressure. Albumin is also chiefly responsible for the maintenance of blood pH. It is located in every tissue and bodily secretion, with extracellular protein comprising 60% of total albumin. Perhaps the most outstanding property of albumin is its ability to bind reversibly to an incredible variety of ligands. It is widely accepted in the pharmaceutical industry that the overall distribution, metabolism, and efficiency of many drugs are rendered ineffective because of their unusually high affinity for this abundant protein. An understanding of the chemistry of the various classes of pharmaceutical interactions with albumin can suggest new approaches to drug therapy and design. Principal Investigator: Dan Carter/New Century Pharmaceuticals

  1. Protein crystallization studies

    NASA Technical Reports Server (NTRS)

    Lyne, James Evans

    1996-01-01

    The Structural Biology laboratory at NASA Marshall Spaceflight Center uses x-ray crystallographic techniques to conduct research into the three-dimensional structure of a wide variety of proteins. A major effort in the laboratory involves an ongoing study of human serum albumin (the principal protein in human plasma) and its interaction with various endogenous substances and pharmaceutical agents. Another focus is on antigenic and functional proteins from several pathogenic organisms including the human immunodeficiency virus (HIV) and the widespread parasitic genus, Schistosoma. My efforts this summer have been twofold: first, to identify clinically significant drug interactions involving albumin binding displacement and to initiate studies of the three-dimensional structure of albumin complexed with these agents, and secondly, to establish collaborative efforts to extend the lab's work on human pathogens.

  2. New MAPS for misfolded proteins.

    PubMed

    Volkmar, Norbert; Fenech, Emma; Christianson, John C

    2016-06-28

    Clearing misfolded proteins from the cytoplasm is essential to maintain cellular homeostasis. Now, a parallel clearance system is described that uses the deubiquitylase USP19 to enable secretion of misfolded cytoplasmic proteins when conventional proteasomal degradation is compromised. Misfolding-associated protein secretion (MAPS) has important implications for protein quality control and prion-like transmission. PMID:27350445

  3. SOY PROTEIN NANOPARTICLES AND NANOCOMPOSITES

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Soy protein isolate (SPI) is obtained from soybean by removing soybean oil and soy carbohydrates. SPI contains more than 90% protein. Structurally, SPI is a globular protein and its aggregates in water consist of sphere-like protein particles. The number average aggregate size of SPI at pH=5.2 is...

  4. FLOW BEHAVIOR OF PROTEIN BLENDS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Blending proteins can increase textural strength or enhance taste or mouth feel, such as blending soy with whey to improve taste. In this study, we measured the viscosity of various combinations of six proteins (whey protein isolates, calcium caseinate, soy protein isolates, wheat gluten, egg album...

  5. Bioinformatics and Moonlighting Proteins

    PubMed Central

    Hernández, Sergio; Franco, Luís; Calvo, Alejandra; Ferragut, Gabriela; Hermoso, Antoni; Amela, Isaac; Gómez, Antonio; Querol, Enrique; Cedano, Juan

    2015-01-01

    Multitasking or moonlighting is the capability of some proteins to execute two or more biochemical functions. Usually, moonlighting proteins are experimentally revealed by serendipity. For this reason, it would be helpful that Bioinformatics could predict this multifunctionality, especially because of the large amounts of sequences from genome projects. In the present work, we analyze and describe several approaches that use sequences, structures, interactomics, and current bioinformatics algorithms and programs to try to overcome this problem. Among these approaches are (a) remote homology searches using Psi-Blast, (b) detection of functional motifs and domains, (c) analysis of data from protein–protein interaction databases (PPIs), (d) match the query protein sequence to 3D databases (i.e., algorithms as PISITE), and (e) mutation correlation analysis between amino acids by algorithms as MISTIC. Programs designed to identify functional motif/domains detect mainly the canonical function but usually fail in the detection of the moonlighting one, Pfam and ProDom being the best methods. Remote homology search by Psi-Blast combined with data from interactomics databases (PPIs) has the best performance. Structural information and mutation correlation analysis can help us to map the functional sites. Mutation correlation analysis can only be used in very specific situations – it requires the existence of multialigned family protein sequences – but can suggest how the evolutionary process of second function acquisition took place. The multitasking protein database MultitaskProtDB (http://wallace.uab.es/multitask/), previously published by our group, has been used as a benchmark for the all of the analyses. PMID:26157797

  6. Modeling Mercury in Proteins.

    PubMed

    Parks, J M; Smith, J C

    2016-01-01

    Mercury (Hg) is a naturally occurring element that is released into the biosphere both by natural processes and anthropogenic activities. Although its reduced, elemental form Hg(0) is relatively nontoxic, other forms such as Hg(2+) and, in particular, its methylated form, methylmercury, are toxic, with deleterious effects on both ecosystems and humans. Microorganisms play important roles in the transformation of mercury in the environment. Inorganic Hg(2+) can be methylated by certain bacteria and archaea to form methylmercury. Conversely, bacteria also demethylate methylmercury and reduce Hg(2+) to relatively inert Hg(0). Transformations and toxicity occur as a result of mercury interacting with various proteins. Clearly, then, understanding the toxic effects of mercury and its cycling in the environment requires characterization of these interactions. Computational approaches are ideally suited to studies of mercury in proteins because they can provide a detailed molecular picture and circumvent issues associated with toxicity. Here, we describe computational methods for investigating and characterizing how mercury binds to proteins, how inter- and intraprotein transfer of mercury is orchestrated in biological systems, and how chemical reactions in proteins transform the metal. We describe quantum chemical analyses of aqueous Hg(II), which reveal critical factors that determine ligand-binding propensities. We then provide a perspective on how we used chemical reasoning to discover how microorganisms methylate mercury. We also highlight our combined computational and experimental studies of the proteins and enzymes of the mer operon, a suite of genes that confer mercury resistance in many bacteria. Lastly, we place work on mercury in proteins in the context of what is needed for a comprehensive multiscale model of environmental mercury cycling. PMID:27497164

  7. Epistasis in protein evolution.

    PubMed

    Starr, Tyler N; Thornton, Joseph W

    2016-07-01

    The structure, function, and evolution of proteins depend on physical and genetic interactions among amino acids. Recent studies have used new strategies to explore the prevalence, biochemical mechanisms, and evolutionary implications of these interactions-called epistasis-within proteins. Here we describe an emerging picture of pervasive epistasis in which the physical and biological effects of mutations change over the course of evolution in a lineage-specific fashion. Epistasis can restrict the trajectories available to an evolving protein or open new paths to sequences and functions that would otherwise have been inaccessible. We describe two broad classes of epistatic interactions, which arise from different physical mechanisms and have different effects on evolutionary processes. Specific epistasis-in which one mutation influences the phenotypic effect of few other mutations-is caused by direct and indirect physical interactions between mutations, which nonadditively change the protein's physical properties, such as conformation, stability, or affinity for ligands. In contrast, nonspecific epistasis describes mutations that modify the effect of many others; these typically behave additively with respect to the physical properties of a protein but exhibit epistasis because of a nonlinear relationship between the physical properties and their biological effects, such as function or fitness. Both types of interaction are rampant, but specific epistasis has stronger effects on the rate and outcomes of evolution, because it imposes stricter constraints and modulates evolutionary potential more dramatically; it therefore makes evolution more contingent on low-probability historical events and leaves stronger marks on the sequences, structures, and functions of protein families. PMID:26833806

  8. Single-cell proteins

    SciTech Connect

    Litchfield, J.H.

    1983-02-11

    Both photosynthetic and nonphotosynthetic microorganisms, grown on various carbon and energy sources, are used in fermentation processes for the production of single-cell proteins. Commercial-scale production has been limited to two algal processes, one bacterial process, and several yeast and fungal processes. High capital and operating costs and the need for extensive nutritional and toxicological assessments have limited the development and commercialization of new processes. Any increase in commercial-scale production appears to be limited to those regions of the world where low-cost carbon and energy sources are available and conventional animal feedstuff proteins, such as soybean meal or fish meal, are in short supply. (Refs. 59).

  9. Protein-based ferrogels.

    PubMed

    Mody, Puja; Hart, Cassidy; Romano, Siena; El-Magbri, Mariam; Esson, Moira M; Ibeh, Trisha; Knowlton, Elizabeth D; Zhang, Ming; Wagner, Michael J; Hartings, Matthew R

    2016-06-01

    We present a novel synthesis in which hemoglobin and Fe(2+) react, in the presence of KNO3 and KOH, to produce protein microgels that contain magnetic iron oxide nanoparticles. The synthesis results in microgels with polymer properties (denaturing and glass transition temperatures) that are consistent with the dried protein. The iron oxide nanoparticles that exhibit an average diameter of 22nm, are ferrimagnetic, and display properties consistent with Fe3O4. The multiple functional capabilities displayed by these materials: biocompatibility, magnetism, dye uptake and controlled release, and other properties archetypal of hydrogels, will make the magnetic hydrogels attractive for a number of biomedical applications. PMID:26901627

  10. Late embryogenesis abundant proteins

    PubMed Central

    Olvera-Carrillo, Yadira; Reyes, José Luis

    2011-01-01

    Late Embryogenesis Abundant (LEA) proteins accumulate at the onset of seed desiccation and in response to water deficit in vegetative plant tissues. The typical LEA proteins are highly hydrophilic and intrinsically unstructured. They have been classified in different families, each one showing distinctive conserved motifs. In this manuscript we present and discuss some of the recent findings regarding their role in plant adaptation to water deficit, as well as those concerning to their possible function, and how it can be related to their intrinsic structural flexibility. PMID:21447997

  11. Congenital protein hypoglycosylation diseases

    PubMed Central

    Sparks, Susan E

    2012-01-01

    Glycosylation is an essential process by which sugars are attached to proteins and lipids. Complete lack of glycosylation is not compatible with life. Because of the widespread function of glycosylation, inherited disorders of glycosylation are multisystemic. Since the identification of the first defect on N-linked glycosylation in the 1980s, there are over 40 different congenital protein hypoglycosylation diseases. This review will include defects of N-linked glycosylation, O-linked glycosylation and disorders of combined N- and O-linked glycosylation. PMID:23776380

  12. Lipid-transfer proteins.

    PubMed

    Ng, Tzi Bun; Cheung, Randy Chi Fai; Wong, Jack Ho; Ye, Xiujuan

    2012-01-01

    Lipid-transfer proteins (LTPs) are basic proteins found in abundance in higher plants. LTPs play lots of roles in plants such as participation in cutin formation, embryogenesis, defense reactions against phytopathogens, symbiosis, and the adaptation of plants to various environmental conditions. In addition, LTPs from field mustard and Chinese daffodil exhibit antiproliferative activity against human cancer cells. LTPs from chili pepper and coffee manifest inhibitory activity against fungi pathogenic to humans such as Candida species. The intent of this article is to review LTPs in the plant kingdom. PMID:23193591

  13. DELIVERY OF THERAPEUTIC PROTEINS

    PubMed Central

    Pisal, Dipak S.; Kosloski, Matthew P.; Balu-Iyer, Sathy V.

    2009-01-01

    The safety and efficacy of protein therapeutics are limited by three interrelated pharmaceutical issues, in vitro and in vivo instability, immunogenicity and shorter half-lives. Novel drug modifications for overcoming these issues are under investigation and include covalent attachment of poly(ethylene glycol) (PEG), polysialic acid, or glycolic acid, as well as developing new formulations containing nanoparticulate or colloidal systems (e.g. liposomes, polymeric microspheres, polymeric nanoparticles). Such strategies have the potential to develop as next generation protein therapeutics. This review includes a general discussion on these delivery approaches. PMID:20049941

  14. Conformation Distributions in Adsorbed Proteins.

    NASA Astrophysics Data System (ADS)

    Meuse, Curtis W.; Hubbard, Joseph B.; Vrettos, John S.; Smith, Jackson R.; Cicerone, Marcus T.

    2007-03-01

    While the structural basis of protein function is well understood in the biopharmaceutical and biotechnology industries, few methods for the characterization and comparison of protein conformation distributions are available. New methods capable of measuring the stability of protein conformations and the integrity of protein-protein, protein-ligand and protein-surface interactions both in solution and on surfaces are needed to help the development of protein-based products. We are developing infrared spectroscopy methods for the characterization and comparison of molecular conformation distributions in monolayers and in solutions. We have extracted an order parameter describing the orientational and conformational variations of protein functional groups around the average molecular values from a single polarized spectrum. We will discuss the development of these methods and compare them to amide hydrogen/deuterium exchange methods for albumin in solution and on different polymer surfaces to show that our order parameter is related to protein stability.

  15. Extreme multifunctional proteins identified from a human protein interaction network

    PubMed Central

    Chapple, Charles E.; Robisson, Benoit; Spinelli, Lionel; Guien, Céline; Becker, Emmanuelle; Brun, Christine

    2015-01-01

    Moonlighting proteins are a subclass of multifunctional proteins whose functions are unrelated. Although they may play important roles in cells, there has been no large-scale method to identify them, nor any effort to characterize them as a group. Here, we propose the first method for the identification of ‘extreme multifunctional' proteins from an interactome as a first step to characterize moonlighting proteins. By combining network topological information with protein annotations, we identify 430 extreme multifunctional proteins (3% of the human interactome). We show that the candidates form a distinct sub-group of proteins, characterized by specific features, which form a signature of extreme multifunctionality. Overall, extreme multifunctional proteins are enriched in linear motifs and less intrinsically disordered than network hubs. We also provide MoonDB, a database containing information on all the candidates identified in the analysis and a set of manually curated human moonlighting proteins. PMID:26054620

  16. Protein-protein interactions: methods for detection and analysis.

    PubMed Central

    Phizicky, E M; Fields, S

    1995-01-01

    The function and activity of a protein are often modulated by other proteins with which it interacts. This review is intended as a practical guide to the analysis of such protein-protein interactions. We discuss biochemical methods such as protein affinity chromatography, affinity blotting, coimmunoprecipitation, and cross-linking; molecular biological methods such as protein probing, the two-hybrid system, and phage display: and genetic methods such as the isolation of extragenic suppressors, synthetic mutants, and unlinked noncomplementing mutants. We next describe how binding affinities can be evaluated by techniques including protein affinity chromatography, sedimentation, gel filtration, fluorescence methods, solid-phase sampling of equilibrium solutions, and surface plasmon resonance. Finally, three examples of well-characterized domains involved in multiple protein-protein interactions are examined. The emphasis of the discussion is on variations in the approaches, concerns in evaluating the results, and advantages and disadvantages of the techniques. PMID:7708014

  17. Protein Molecular Structures, Protein SubFractions, and Protein Availability Affected by Heat Processing: A Review

    SciTech Connect

    Yu,P.

    2007-01-01

    The utilization and availability of protein depended on the types of protein and their specific susceptibility to enzymatic hydrolysis (inhibitory activities) in the gastrointestine and was highly associated with protein molecular structures. Studying internal protein structure and protein subfraction profiles leaded to an understanding of the components that make up a whole protein. An understanding of the molecular structure of the whole protein was often vital to understanding its digestive behavior and nutritive value in animals. In this review, recently obtained information on protein molecular structural effects of heat processing was reviewed, in relation to protein characteristics affecting digestive behavior and nutrient utilization and availability. The emphasis of this review was on (1) using the newly advanced synchrotron technology (S-FTIR) as a novel approach to reveal protein molecular chemistry affected by heat processing within intact plant tissues; (2) revealing the effects of heat processing on the profile changes of protein subfractions associated with digestive behaviors and kinetics manipulated by heat processing; (3) prediction of the changes of protein availability and supply after heat processing, using the advanced DVE/OEB and NRC-2001 models, and (4) obtaining information on optimal processing conditions of protein as intestinal protein source to achieve target values for potential high net absorbable protein in the small intestine. The information described in this article may give better insight in the mechanisms involved and the intrinsic protein molecular structural changes occurring upon processing.

  18. Divergent signaling pathways cooperatively regulate TGFβ induction of cysteine-rich protein 2 in vascular smooth muscle cells

    PubMed Central

    2014-01-01

    Background Vascular smooth muscle cells (VSMCs) of the arterial wall play a critical role in the development of occlusive vascular diseases. Cysteine-rich protein 2 (CRP2) is a VSMC-expressed LIM-only protein, which functionally limits VSMC migration and protects against pathological vascular remodeling. The multifunctional cytokine TGFβ has been implicated to play a role in the pathogenesis of atherosclerosis through numerous downstream signaling pathways. We showed previously that TGFβ upregulates CRP2 expression; however, the detailed signaling mechanisms remain unclear. Results TGFβ treatment of VSMCs activated both Smad2/3 and ATF2 phosphorylation. Individually knocking down Smad2/3 or ATF2 pathways with siRNA impaired the TGFβ induction of CRP2, indicating that both contribute to CRP2 expression. Inhibiting TβRI kinase activity by SB431542 or TβRI knockdown abolished Smad2/3 phosphorylation but did not alter ATF2 phosphorylation, indicating while Smad2/3 phosphorylation was TβRI-dependent ATF2 phosphorylation was independent of TβRI. Inhibiting Src kinase activity by SU6656 suppressed TGFβ-induced RhoA and ATF2 activation but not Smad2 phosphorylation. Blocking ROCK activity, the major downstream target of RhoA, abolished ATF2 phosphorylation and CRP2 induction but not Smad2 phosphorylation. Furthermore, JNK inhibition with SP600125 reduced TGFβ-induced ATF2 (but not Smad2) phosphorylation and CRP2 protein expression while ROCK inhibition blocked JNK activation. These results indicate that downstream of TβRII, Src family kinase-RhoA-ROCK-JNK signaling pathway mediates TβRI-independent ATF2 activation. Promoter analysis revealed that the TGFβ induction of CRP2 was mediated through the CRE and SBE promoter elements that were located in close proximity. Conclusions Our results demonstrate that two signaling pathways downstream of TGFβ converge on the CRE and SBE sites of the Csrp2 promoter to cooperatively control CRP2 induction in VSMCs, which

  19. Transient protein-protein interactions visualized by solution NMR.

    PubMed

    Liu, Zhu; Gong, Zhou; Dong, Xu; Tang, Chun

    2016-01-01

    Proteins interact with each other to establish their identities in cell. The affinities for the interactions span more than ten orders of magnitude, and KD values in μM-mM regimen are considered transient and are important in cell signaling. Solution NMR including diamagnetic and paramagnetic techniques has enabled atomic-resolution depictions of transient protein-protein interactions. Diamagnetic NMR allows characterization of protein complexes with KD values up to several mM, whereas ultraweak and fleeting complexes can be modeled with the use of paramagnetic NMR especially paramagnetic relaxation enhancement (PRE). When tackling ever-larger protein complexes, PRE can be particularly useful in providing long-range intermolecular distance restraints. As NMR measurements are averaged over the ensemble of complex structures, structural information for dynamic protein-protein interactions besides the stereospecific one can often be extracted. Herein the protein interaction dynamics are exemplified by encounter complexes, alternative binding modes, and coupled binding/folding of intrinsically disordered proteins. Further integration of NMR with other biophysical techniques should allow better visualization of transient protein-protein interactions. In particular, single-molecule data may facilitate the interpretation of ensemble-averaged NMR data. Though same structures of proteins and protein complexes were found in cell as in diluted solution, we anticipate that the dynamics of transient protein protein-protein interactions be different, which awaits awaits exploration by NMR. This article is part of a Special Issue entitled: Physiological Enzymology and Protein Functions. This article is part of a Special Issue entitled: Physiological Enzymology and Protein Functions. PMID:25896389

  20. Preparing Protein Samples

    NASA Technical Reports Server (NTRS)

    2000-01-01

    Cindy Barnes of University Space Research Association (USRA) at NASA's Marshall Space Flight Center pipettes a protein solution in preparation to grow crystals as part of NASA's structural biology program. Research on Earth helps scientists define conditions and specimens they will use in space experiments.

  1. Protein Crystal Bovine Insulin

    NASA Technical Reports Server (NTRS)

    1991-01-01

    The comparison of protein crystal, Bovine Insulin space-grown (left) and earth-grown (right). Facilitates the incorporation of glucose into cells. In diabetics, there is either a decrease in or complete lack of insulin, thereby leading to several harmful complications. Principal Investigator is Larry DeLucas.

  2. Cosolvent assisted protein refolding.

    PubMed

    Cleland, J L; Wang, D I

    1990-12-01

    The use of cosolvents in aqueous systems has been shown to enhance protein refolding and decrease aggregation. In this study, we have used polyethylene glycol (PEG) in the molecular weight range of 1000 to 8000 Daltons to effectively increase the rate of refolding and prevent aggregation of the model protein, bovine carbonic anhydrase B (CAB). At concentrations of 3 and 30 g/l, PEG increased the rate of recovery of active protein in the absence of aggregation. Using 3 g/l PEG (3350 MW), the refolding rate was three fold greater than the observed normal refolding rate. The observed rate enhancement was caused by PEG acting on the first intermediate in the CAB refolding pathway to increase the rate of formation of the second intermediate. The interaction of PEG with the first intermediate also prevented its self-association during refolding and at equilibrium. The stabilization of this first intermediate resulted in complete recovery of active protein under normal aggregating conditions. PMID:1367488

  3. The Protein Ensemble Database.

    PubMed

    Varadi, Mihaly; Tompa, Peter

    2015-01-01

    The scientific community's major conceptual notion of structural biology has recently shifted in emphasis from the classical structure-function paradigm due to the emergence of intrinsically disordered proteins (IDPs). As opposed to their folded cousins, these proteins are defined by the lack of a stable 3D fold and a high degree of inherent structural heterogeneity that is closely tied to their function. Due to their flexible nature, solution techniques such as small-angle X-ray scattering (SAXS), nuclear magnetic resonance (NMR) spectroscopy and fluorescence resonance energy transfer (FRET) are particularly well-suited for characterizing their biophysical properties. Computationally derived structural ensembles based on such experimental measurements provide models of the conformational sampling displayed by these proteins, and they may offer valuable insights into the functional consequences of inherent flexibility. The Protein Ensemble Database (http://pedb.vib.be) is the first openly accessible, manually curated online resource storing the ensemble models, protocols used during the calculation procedure, and underlying primary experimental data derived from SAXS and/or NMR measurements. By making this previously inaccessible data freely available to researchers, this novel resource is expected to promote the development of more advanced modelling methodologies, facilitate the design of standardized calculation protocols, and consequently lead to a better understanding of how function arises from the disordered state. PMID:26387108

  4. Protein Requirements during Aging.

    PubMed

    Courtney-Martin, Glenda; Ball, Ronald O; Pencharz, Paul B; Elango, Rajavel

    2016-01-01

    Protein recommendations for elderly, both men and women, are based on nitrogen balance studies. They are set at 0.66 and 0.8 g/kg/day as the estimated average requirement (EAR) and recommended dietary allowance (RDA), respectively, similar to young adults. This recommendation is based on single linear regression of available nitrogen balance data obtained at test protein intakes close to or below zero balance. Using the indicator amino acid oxidation (IAAO) method, we estimated the protein requirement in young adults and in both elderly men and women to be 0.9 and 1.2 g/kg/day as the EAR and RDA, respectively. This suggests that there is no difference in requirement on a gender basis or on a per kg body weight basis between younger and older adults. The requirement estimates however are ~40% higher than the current protein recommendations on a body weight basis. They are also 40% higher than our estimates in young men when calculated on the basis of fat free mass. Thus, current recommendations may need to be re-assessed. Potential rationale for this difference includes a decreased sensitivity to dietary amino acids and increased insulin resistance in the elderly compared with younger individuals. PMID:27529275

  5. Protein crystal growth

    NASA Technical Reports Server (NTRS)

    2001-01-01

    Atomic force microscopy uses laser technology to reveal a defect, a double-screw dislocation, on the surface of this crystal of canavalin, a major source of dietary protein for humans and domestic animals. When a crystal grows, attachment kinetics and transport kinetics are competing for control of the molecules. As a molecule gets close to the crystal surface, it has to attach properly for the crystal to be usable. NASA has funded investigators to look at those attachment kinetics from a theoretical standpoint and an experimental standpoint. Dr. Alex McPherson of the University of California, Irvine, is one of those investigators. He uses X-ray diffraction and atomic force microscopy in his laboratory to answer some of the many questions about how protein crystals grow. Atomic force microscopy provides a means of looking at how individual molecules are added to the surface of growing protein crystals. This helps McPherson understand the kinetics of protein crystal growth. McPherson asks, How fast do crystals grow? What are the forces involved? Investigators funded by NASA have clearly shown that such factors as the level of supersaturation and the rate of growth all affect the habit [characteristic arrangement of facets] of the crystal and the defects that occur in the crystal.

  6. Protein-protein and protein-salt interactions in aqueous protein solutions containing concentrated electrolytes

    SciTech Connect

    Curtis, R.A.; Blanch, H.W.; Prausnitz, J.M.

    1998-01-05

    Protein-protein and protein-salt interactions have been obtained for ovalbumin in solutions of ammonium sulfate and for lysozyme in solutions of ammonium sulfate, sodium chloride, potassium isothiocyanate, and potassium chloride. The two-body interactions between ovalbumin molecules in concentrated ammonium-sulfate solutions can be described by the DLVO potentials plus a potential that accounts for the decrease in free volume available to the protein due to the presence of the salt ions. The interaction between ovalbumin and ammonium sulfate is unfavorable, reflecting the kosmotropic nature of sulfate anions. Lysozyme-lysozyme interactions cannot be described by the above potentials because anion binding to lysozyme alters these interactions. Lysozyme-isothiocyanate complexes are strongly attractive due to electrostatic interactions resulting from bridging by the isothiocyanate ion. Lysozyme-lysozyme interactions in sulfate solutions are more repulsive than expected, possibly resulting from a larger excluded volume of a lysozyme-sulfate bound complex or perhaps, hydration forces between the lysozyme-sulfate complexes.

  7. Cellulose binding domain proteins

    DOEpatents

    Shoseyov, O.; Shpiegl, I.; Goldstein, M.; Doi, R.

    1998-11-17

    A cellulose binding domain (CBD) having a high affinity for crystalline cellulose and chitin is disclosed, along with methods for the molecular cloning and recombinant production. Fusion products comprising the CBD and a second protein are likewise described. A wide range of applications are contemplated for both the CBD and the fusion products, including drug delivery, affinity separations, and diagnostic techniques. 16 figs.

  8. Cellulose binding domain proteins

    DOEpatents

    Shoseyov, Oded; Shpiegl, Itai; Goldstein, Marc; Doi, Roy

    1998-01-01

    A cellulose binding domain (CBD) having a high affinity for crystalline cellulose and chitin is disclosed, along with methods for the molecular cloning and recombinant production thereof. Fusion products comprising the CBD and a second protein are likewise described. A wide range of applications are contemplated for both the CBD and the fusion products, including drug delivery, affinity separations, and diagnostic techniques.

  9. Ribosome-inactivating proteins

    PubMed Central

    Walsh, Matthew J; Dodd, Jennifer E; Hautbergue, Guillaume M

    2013-01-01

    Ribosome-inactivating proteins (RIPs) were first isolated over a century ago and have been shown to be catalytic toxins that irreversibly inactivate protein synthesis. Elucidation of atomic structures and molecular mechanism has revealed these proteins to be a diverse group subdivided into two classes. RIPs have been shown to exhibit RNA N-glycosidase activity and depurinate the 28S rRNA of the eukaryotic 60S ribosomal subunit. In this review, we compare archetypal RIP family members with other potent toxins that abolish protein synthesis: the fungal ribotoxins which directly cleave the 28S rRNA and the newly discovered Burkholderia lethal factor 1 (BLF1). BLF1 presents additional challenges to the current classification system since, like the ribotoxins, it does not possess RNA N-glycosidase activity but does irreversibly inactivate ribosomes. We further discuss whether the RIP classification should be broadened to include toxins achieving irreversible ribosome inactivation with similar turnovers to RIPs, but through different enzymatic mechanisms. PMID:24071927

  10. Protein Requirements during Aging

    PubMed Central

    Courtney-Martin, Glenda; Ball, Ronald O.; Pencharz, Paul B.; Elango, Rajavel

    2016-01-01

    Protein recommendations for elderly, both men and women, are based on nitrogen balance studies. They are set at 0.66 and 0.8 g/kg/day as the estimated average requirement (EAR) and recommended dietary allowance (RDA), respectively, similar to young adults. This recommendation is based on single linear regression of available nitrogen balance data obtained at test protein intakes close to or below zero balance. Using the indicator amino acid oxidation (IAAO) method, we estimated the protein requirement in young adults and in both elderly men and women to be 0.9 and 1.2 g/kg/day as the EAR and RDA, respectively. This suggests that there is no difference in requirement on a gender basis or on a per kg body weight basis between younger and older adults. The requirement estimates however are ~40% higher than the current protein recommendations on a body weight basis. They are also 40% higher than our estimates in young men when calculated on the basis of fat free mass. Thus, current recommendations may need to be re-assessed. Potential rationale for this difference includes a decreased sensitivity to dietary amino acids and increased insulin resistance in the elderly compared with younger individuals. PMID:27529275

  11. Protein states and proteinquakes.

    PubMed Central

    Ansari, A; Berendzen, J; Bowne, S F; Frauenfelder, H; Iben, I E; Sauke, T B; Shyamsunder, E; Young, R D

    1985-01-01

    After photodissociation of carbon monoxide bound to myoglobin, the protein relaxes to the deoxy equilibrium structure in a quake-like motion. Investigation of the proteinquake and of related intramolecular equilibrium motions shows that states and motions have a hierarchical glass-like structure. PMID:3860839

  12. Thermal unfolding of proteins

    NASA Astrophysics Data System (ADS)

    Cieplak, Marek; Sułkowska, Joanna I.

    2005-11-01

    Thermal unfolding of proteins is compared to folding and mechanical stretching in a simple topology-based dynamical model. We define the unfolding time and demonstrate its low-temperature divergence. Below a characteristic temperature, contacts break at separate time scales and unfolding proceeds approximately in a way reverse to folding. Features in these scenarios agree with experiments and atomic simulations on titin.

  13. Dynamics of protein conformations

    NASA Astrophysics Data System (ADS)

    Stepanova, Maria

    2010-10-01

    A novel theoretical methodology is introduced to identify dynamic structural domains and analyze local flexibility in proteins. The methodology employs a multiscale approach combining identification of essential collective coordinates based on the covariance analysis of molecular dynamics trajectories, construction of the Mori projection operator with these essential coordinates, and analysis of the corresponding generalized Langevin equations [M.Stepanova, Phys.Rev.E 76(2007)051918]. Because the approach employs a rigorous theory, the outcomes are physically transparent: the dynamic domains are associated with regions of relative rigidity in the protein, whereas off-domain regions are relatively soft. This also allows scoring the flexibility in the macromolecule with atomic-level resolution [N.Blinov, M.Berjanskii, D.S.Wishart, and M.Stepanova, Biochemistry, 48(2009)1488]. The applications include the domain coarse-graining and characterization of conformational stability in protein G and prion proteins. The results are compared with published NMR experiments. Potential applications for structural biology, bioinformatics, and drug design are discussed.

  14. Protein denaturing on Nanospheres

    NASA Astrophysics Data System (ADS)

    Forrest, James; Teichroeb, Jonathan

    2007-03-01

    We have used localized surface plasmon resonance (LSPR) to monitor the structural changes that accompany thermal denaturing of Bovine Serum Albumin(BSA) adsorbed onto gold nanospheres of size 5nm-60nm. The effect of the protein on the LSPR was monitored by visible extinction spectroscopy. The position of the resonance is affected by the conformation of the adsorbed protein layer, and as such can be used as a very sensitive probe of thermal denaturing that is specific to the adsorbed protein. The results are compared to detailed calculations and show that full calculations can lead to significant increases in knowledge where gold nanospheres are used as biosensors. Thermal denaturing on spheres with diameter > 20 nm show strong similarity to bulk calorimetric studies of BSA in solution. BSA adsorbed on nanospheres with d<= 15 nm shows a qualitative difference in behavior, suggesting a sensitivity of denaturing characteristics on local surface curvature. Studies of isothermal denaturing kinetics were used to obtain an activatiuon barrier for thermal denaturing. This activation barrier also exhibited a strong dependence on nanoparticle size. These results may have important implications for other protein-nanoparticle interactions.

  15. [ALR, the multifunctional protein].

    PubMed

    Balogh, Tibor; Szarka, András

    2015-03-29

    ALR is a mystic protein. It has a so called "long" 22 kDa and a "short" 15 kDa forms. It has been described after partial hepatectomy and it has just been considered as a key protein of liver regeneration. At the beginning of the 21st century it has been revealed that the "long" form is localized in the mitochondrial intermembrane space and it is an element of the mitochondrial protein import and disulphide relay system. Several proteins of the substrates of the mitochondrial disulphide relay system are necessary for the proper function of the mitochondria, thus any mutation of the ALR gene leads to mitochondrial diseases. The "short" form of ALR functions as a secreted extracellular growth factor and it promotes the protection, regeneration and proliferation of hepatocytes. The results gained on the recently generated conditional ALR mutant mice suggest that ALR can play an important role in the pathogenesis of alcoholic and non-alcoholic steatosis. Since the serum level of ALR is modified in several liver diseases it can be a promising marker molecule in laboratory diagnostics. PMID:25796277

  16. The Protein Data Bank

    PubMed Central

    Berman, Helen M.; Westbrook, John; Feng, Zukang; Gilliland, Gary; Bhat, T. N.; Weissig, Helge; Shindyalov, Ilya N.; Bourne, Philip E.

    2000-01-01

    The Protein Data Bank (PDB; http://www.rcsb.org/pdb/ ) is the single worldwide archive of structural data of biological macromolecules. This paper describes the goals of the PDB, the systems in place for data deposition and access, how to obtain further information, and near-term plans for the future development of the resource. PMID:10592235

  17. Tuber Storage Proteins

    PubMed Central

    SHEWRY, PETER R.

    2003-01-01

    A wide range of plants are grown for their edible tubers, but five species together account for almost 90 % of the total world production. These are potato (Solanum tuberosum), cassava (Manihot esculenta), sweet potato (Ipomoea batatus), yams (Dioscorea spp.) and taro (Colocasia, Cyrtosperma and Xanthosoma spp.). All of these, except cassava, contain groups of storage proteins, but these differ in the biological properties and evolutionary relationships. Thus, patatin from potato exhibits activity as an acylhydrolase and esterase, sporamin from sweet potato is an inhibitor of trypsin, and dioscorin from yam is a carbonic anhydrase. Both sporamin and dioscorin also exhibit antioxidant and radical scavenging activity. Taro differs from the other three crops in that it contains two major types of storage protein: a trypsin inhibitor related to sporamin and a mannose‐binding lectin. These characteristics indicate that tuber storage proteins have evolved independently in different species, which contrasts with the highly conserved families of storage proteins present in seeds. Furthermore, all exhibit biological activities which could contribute to resistance to pests, pathogens or abiotic stresses, indicating that they may have dual roles in the tubers. PMID:12730067

  18. Protein specific polymeric immunomicrospheres

    NASA Technical Reports Server (NTRS)

    Rembaum, Alan (Inventor); Yen, Shiao-Ping S. (Inventor); Dreyer, William J. (Inventor)

    1980-01-01

    Small, round, bio-compatible microspheres capable of covalently bonding proteins and having a uniform diameter below about 3500 A are prepared by substantially instantaneously initiating polymerization of an aqueous emulsion containing no more than 35% total monomer including an acrylic monomer substituted with a covalently bondable group such as hydroxyl, amino or carboxyl and a minor amount of a cross-linking agent.

  19. Perspectives on protein crystallisation

    SciTech Connect

    Ochi, T.; Stojanoff, V.; Bolanos-Garcia, V.M.; Moreno, A.

    2009-12-11

    This final part on 'perspectives' is focused on new strategies that can be used to crystallise proteins and improve the crystal quality of macromolecular complexes using any of the methods reviewed in this focused issue. Some advantages and disadvantages, limitations, and plausible applications to high-resolution X-ray crystallography are discussed.

  20. Bayesian Estimator of Protein-Protein Association Probabilities

    2008-05-28

    The Bayesian Estimator of Protein-Protein Association Probabilities (BEPro3) is a software tool for estimating probabilities of protein-protein association between bait and prey protein pairs using data from multiple-bait, multiple-replicate, protein LC-MS/MS affinity isolation experiments. BEPro3 is public domain software, has been tested on Windows XP and version 10.4 or newer of the Mac OS 10.4, and is freely available. A user guide, example dataset with analysis and additional documentation are included with the BEPro3 download.

  1. Phosphorylation of human link proteins

    SciTech Connect

    Oester, D.A.; Caterson, B.; Schwartz, E.R.

    1986-06-13

    Three link proteins of 48, 44 and 40 kDa were purified from human articular cartilage and identified with monoclonal anti-link protein antibody 8-A-4. Two sets of lower molecular weight proteins of 30-31 kDa and 24-26 kDa also contained link protein epitopes recognized by the monoclonal antibody and were most likely degradative products of the intact link proteins. The link proteins of 48 and 40 kDa were identified as phosphoproteins while the 44 kDa link protein did not contain /sup 32/P. The phosphorylated 48 and 40 kDa link proteins contained approximately 2 moles PO/sub 4//mole link protein.

  2. Identifying the hub proteins from complicated membrane protein network systems.

    PubMed

    Shen, Yi-Zhen; Ding, Yong-Sheng; Gu, Quan; Chou, Kuo-Chen

    2010-05-01

    The so-called "hub proteins" are those proteins in a protein-protein interaction network system that have remarkably higher interaction relations (or degrees) than the others. Therefore, the information of hub proteins can provide very useful insights for selecting or prioritizing targets during drug development. In this paper, by combining the multi-agent-based method with the graphical spectrum analysis and immune-genetic algorithm, a novel simulator for identifying the hub proteins from membrane protein interaction networks is proposed. As a demonstration of using the simulator, two hub membrane proteins, YPL227C and YIL147C, were identified from a complicated network system consisting of 1500 membrane proteins. Meanwhile, along with the two identified hub proteins, their molecular functions, biological processes, and cellular components were also revealed. It is anticipated that the hub-protein-simulator may become a very useful tool for system biology and drug development, particularly in deciphering unknown protein functions, determining protein complexes, and in identifying the key targets from a complicated disease system. PMID:20507268

  3. Direct Probing of Protein-Protein Interactions

    SciTech Connect

    Noy, A; Sulchek, T A; Friddle, R W

    2005-03-10

    This project aimed to establish feasibility of using experimental techniques based on direct measurements of interaction forces on the single molecule scale to characterize equilibrium interaction potentials between individual biological molecules. Such capability will impact several research areas, ranging from rapid interaction screening capabilities to providing verifiable inputs for computational models. It should be one of the enabling technologies for modern proteomics research. This study used a combination of Monte-Carlo simulations, theoretical considerations, and direct experimental measurements to investigate two model systems that represented typical experimental situations: force-induced melting of DNA rigidly attached to the tip, and force-induced unbinding of a protein-antibody pair connected to flexible tethers. Our results establish that for both systems researchers can use force spectroscopy measurements to extract reliable information about equilibrium interaction potentials. However, the approaches necessary to extract these potentials in each case--Jarzynski reconstruction and Dynamic Force Spectroscopy--are very different. We also show how the thermodynamics and kinetics of unbinding process dictates the choice between in each case.

  4. Hydrogels Constructed from Engineered Proteins.

    PubMed

    Li, Hongbin; Kong, Na; Laver, Bryce; Liu, Junqiu

    2016-02-24

    Due to their various potential biomedical applications, hydrogels based on engineered proteins have attracted considerable interest. Benefitting from significant progress in recombinant DNA technology and protein engineering/design techniques, the field of protein hydrogels has made amazing progress. The latest progress of hydrogels constructed from engineered recombinant proteins are presented, mainly focused on biorecognition-driven physical hydrogels as well as chemically crosslinked hydrogels. The various bio-recognition based physical crosslinking strategies are discussed, as well as chemical crosslinking chemistries used to engineer protein hydrogels, and protein hydrogels' various biomedical applications. The future perspectives of this fast evolving field of biomaterials are also discussed. PMID:26707834

  5. Redox control of protein degradation

    PubMed Central

    Pajares, Marta; Jiménez-Moreno, Natalia; Dias, Irundika H.K.; Debelec, Bilge; Vucetic, Milica; Fladmark, Kari E.; Basaga, Huveyda; Ribaric, Samo; Milisav, Irina; Cuadrado, Antonio

    2015-01-01

    Intracellular proteolysis is critical to maintain timely degradation of altered proteins including oxidized proteins. This review attempts to summarize the most relevant findings about oxidant protein modification, as well as the impact of reactive oxygen species on the proteolytic systems that regulate cell response to an oxidant environment: the ubiquitin-proteasome system (UPS), autophagy and the unfolded protein response (UPR). In the presence of an oxidant environment, these systems are critical to ensure proteostasis and cell survival. An example of altered degradation of oxidized proteins in pathology is provided for neurodegenerative diseases. Future work will determine if protein oxidation is a valid target to combat proteinopathies. PMID:26381917

  6. Protein-protein interactions in DNA mismatch repair.

    PubMed

    Friedhoff, Peter; Li, Pingping; Gotthardt, Julia

    2016-02-01

    The principal DNA mismatch repair proteins MutS and MutL are versatile enzymes that couple DNA mismatch or damage recognition to other cellular processes. Besides interaction with their DNA substrates this involves transient interactions with other proteins which is triggered by the DNA mismatch or damage and controlled by conformational changes. Both MutS and MutL proteins have ATPase activity, which adds another level to control their activity and interactions with DNA substrates and other proteins. Here we focus on the protein-protein interactions, protein interaction sites and the different levels of structural knowledge about the protein complexes formed with MutS and MutL during the mismatch repair reaction. PMID:26725162

  7. Functionalizing Microporous Membranes for Protein Purification and Protein Digestion

    NASA Astrophysics Data System (ADS)

    Dong, Jinlan; Bruening, Merlin L.

    2015-07-01

    This review examines advances in the functionalization of microporous membranes for protein purification and the development of protease-containing membranes for controlled protein digestion prior to mass spectrometry analysis. Recent studies confirm that membranes are superior to bead-based columns for rapid protein capture, presumably because convective mass transport in membrane pores rapidly brings proteins to binding sites. Modification of porous membranes with functional polymeric films or TiO2 nanoparticles yields materials that selectively capture species ranging from phosphopeptides to His-tagged proteins, and protein-binding capacities often exceed those of commercial beads. Thin membranes also provide a convenient framework for creating enzyme-containing reactors that afford control over residence times. With millisecond residence times, reactors with immobilized proteases limit protein digestion to increase sequence coverage in mass spectrometry analysis and facilitate elucidation of protein structures. This review emphasizes the advantages of membrane-based techniques and concludes with some challenges for their practical application.

  8. Affinity purification of proteins binding to GST fusion proteins.

    PubMed

    Swaffield, J C; Johnston, S A

    2001-05-01

    This unit describes the use of proteins fused to glutathione-S-transferase (GST fusion proteins) to affinity purify other proteins, a technique also known as GST pulldown purification. The describes a strategy in which a GST fusion protein is bound to agarose affinity beads and the complex is then used to assay the binding of a specific test protein that has been labeled with [35S]methionine by in vitro translation. However, this method can be adapted for use with other types of fusion proteins; for example, His6, biotin tags, or maltose-binding protein fusions (MBP), and these may offer particular advantages. A describes preparation of an E. coli extract that is added to the reaction mixture with purified test protein to reduce nonspecific binding. PMID:18265191

  9. How do oncoprotein mutations rewire protein-protein interaction networks?

    PubMed

    Bowler, Emily H; Wang, Zhenghe; Ewing, Rob M

    2015-01-01

    The acquisition of mutations that activate oncogenes or inactivate tumor suppressors is a primary feature of most cancers. Mutations that directly alter protein sequence and structure drive the development of tumors through aberrant expression and modification of proteins, in many cases directly impacting components of signal transduction pathways and cellular architecture. Cancer-associated mutations may have direct or indirect effects on proteins and their interactions and while the effects of mutations on signaling pathways have been widely studied, how mutations alter underlying protein-protein interaction networks is much less well understood. Systematic mapping of oncoprotein protein interactions using proteomics techniques as well as computational network analyses is revealing how oncoprotein mutations perturb protein-protein interaction networks and drive the cancer phenotype. PMID:26325016

  10. Purine inhibitors of protein kinases, G proteins and polymerases

    DOEpatents

    Gray, Nathanael S.; Schultz, Peter; Kim, Sung-Hou; Meijer, Laurent

    2004-10-12

    The present invention relates to 2-N-substituted 6-(4-methoxybenzylamino)-9-isopropylpurines that inhibit, inter alia, protein kinases, G-proteins and polymerases. In addition, the present invention relates to methods of using such 2-N-substituted 6-(4-methoxybenzylamino)-9-isopropylpurines to inhibit protein kinases, G-proteins, polymerases and other cellular processes and to treat cellular proliferative diseases.

  11. Protein identification and Peptide expression resolver: harmonizing protein identification with protein expression data.

    PubMed

    Kearney, Paul; Butler, Heather; Eng, Kevin; Hugo, Patrice

    2008-01-01

    Proteomic discovery platforms generate both peptide expression information and protein identification information. Peptide expression data are used to determine which peptides are differentially expressed between study cohorts, and then these peptides are targeted for protein identification. In this paper, we demonstrate that peptide expression information is also a powerful tool for enhancing confidence in protein identification results. Specifically, we evaluate the following hypothesis: tryptic peptides originating from the same protein have similar expression profiles across samples in the discovery study. Evidence supporting this hypothesis is provided. This hypothesis is integrated into a protein identification tool, PIPER (Protein Identification and Peptide Expression Resolver), that reduces erroneous protein identifications below 5%. PIPER's utility is illustrated by application to a 72-sample biomarker discovery study where it is demonstrated that false positive protein identifications can be reduced below 5%. Consequently, it is recommended that PIPER methodology be incorporated into proteomic studies where both protein expression and identification data are collected. PMID:18062667

  12. Predicting Disease-Related Proteins Based on Clique Backbone in Protein-Protein Interaction Network

    PubMed Central

    Yang, Lei; Zhao, Xudong; Tang, Xianglong

    2014-01-01

    Network biology integrates different kinds of data, including physical or functional networks and disease gene sets, to interpret human disease. A clique (maximal complete subgraph) in a protein-protein interaction network is a topological module and possesses inherently biological significance. A disease-related clique possibly associates with complex diseases. Fully identifying disease components in a clique is conductive to uncovering disease mechanisms. This paper proposes an approach of predicting disease proteins based on cliques in a protein-protein interaction network. To tolerate false positive and negative interactions in protein networks, extending cliques and scoring predicted disease proteins with gene ontology terms are introduced to the clique-based method. Precisions of predicted disease proteins are verified by disease phenotypes and steadily keep to more than 95%. The predicted disease proteins associated with cliques can partly complement mapping between genotype and phenotype, and provide clues for understanding the pathogenesis of serious diseases. PMID:25013377

  13. Understanding Protein Non-Folding

    PubMed Central

    Uversky, Vladimir N.; Dunker, A. Keith

    2010-01-01

    This review describes the family of intrinsically disordered proteins, members of which fail to form rigid 3-D structures under physiological conditions, either along their entire lengths or only in localized regions. Instead, these intriguing proteins/regions exist as dynamic ensembles within which atom positions and backbone Ramachandran angles exhibit extreme temporal fluctuations without specific equilibrium values. Many of these intrinsically disordered proteins are known to carry out important biological functions which, in fact, depend on the absence of specific 3-D structure. The existence of such proteins does not fit the prevailing structure-function paradigm, which states that unique 3-D structure is a prerequisite to function. Thus, the protein structure-function paradigm has to be expanded to include intrinsically disordered proteins and alternative relationships among protein sequence, structure, and function. This shift in the paradigm represents a major breakthrough for biochemistry, biophysics and molecular biology, as it opens new levels of understanding with regard to the complex life of proteins. This review will try to answer the following questions: How were intrinsically disordered proteins discovered? Why don't these proteins fold? What is so special about intrinsic disorder? What are the functional advantages of disordered proteins/regions? What is the functional repertoire of these proteins? What are the relationships between intrinsically disordered proteins and human diseases? PMID:20117254

  14. Protein Structure Databases.

    PubMed

    Laskowski, Roman A

    2016-01-01

    Web-based protein structure databases come in a wide variety of types and levels of information content. Those having the most general interest are the various atlases that describe each experimentally determined protein structure and provide useful links, analyses, and schematic diagrams relating to its 3D structure and biological function. Also of great interest are the databases that classify 3D structures by their folds as these can reveal evolutionary relationships which may be hard to detect from sequence comparison alone. Related to these are the numerous servers that compare folds-particularly useful for newly solved structures, and especially those of unknown function. Beyond these are a vast number of databases for the more specialized user, dealing with specific families, diseases, structural features, and so on. PMID:27115626

  15. A magnetic protein biocompass.

    PubMed

    Qin, Siying; Yin, Hang; Yang, Celi; Dou, Yunfeng; Liu, Zhongmin; Zhang, Peng; Yu, He; Huang, Yulong; Feng, Jing; Hao, Junfeng; Hao, Jia; Deng, Lizong; Yan, Xiyun; Dong, Xiaoli; Zhao, Zhongxian; Jiang, Taijiao; Wang, Hong-Wei; Luo, Shu-Jin; Xie, Can

    2016-02-01

    The notion that animals can detect the Earth's magnetic field was once ridiculed, but is now well established. Yet the biological nature of such magnetosensing phenomenon remains unknown. Here, we report a putative magnetic receptor (Drosophila CG8198, here named MagR) and a multimeric magnetosensing rod-like protein complex, identified by theoretical postulation and genome-wide screening, and validated with cellular, biochemical, structural and biophysical methods. The magnetosensing complex consists of the identified putative magnetoreceptor and known magnetoreception-related photoreceptor cryptochromes (Cry), has the attributes of both Cry- and iron-based systems, and exhibits spontaneous alignment in magnetic fields, including that of the Earth. Such a protein complex may form the basis of magnetoreception in animals, and may lead to applications across multiple fields. PMID:26569474

  16. Bone Morphogenetic Proteins.

    PubMed

    Katagiri, Takenobu; Watabe, Tetsuro

    2016-01-01

    Bone morphogenetic proteins (BMPs), originally identified as osteoinductive components in extracts derived from bone, are now known to play important roles in a wide array of processes during formation and maintenance of various organs including bone, cartilage, muscle, kidney, and blood vessels. BMPs and the related "growth and differentiation factors" (GDFs) are members of the transforming growth factor β (TGF-β) family, and transduce their signals through type I and type II serine-threonine kinase receptors and their intracellular downstream effectors, including Smad proteins. Furthermore, BMP signals are finely tuned by various agonists and antagonists. Because deregulation of the BMP activity at multiple steps in signal transduction is linked to a wide variety of human diseases, therapeutic use of activators and inhibitors of BMP signaling will provide potential avenues for the treatment of the human disorders that are caused by hypo- and hyperactivation of BMP signals, respectively. PMID:27252362

  17. Protein mediated membrane adhesion

    NASA Astrophysics Data System (ADS)

    Carlson, Andreas; Mahadevan, L.

    2015-05-01

    Adhesion in the context of mechanical attachment, signaling, and movement in cellular dynamics is mediated by the kinetic interactions between membrane-embedded proteins in an aqueous environment. Here, we present a minimal theoretical framework for the dynamics of membrane adhesion that accounts for the kinetics of protein binding, the elastic deformation of the membrane, and the hydrodynamics of squeeze flow in the membrane gap. We analyze the resulting equations using scaling estimates to characterize the spatiotemporal features of the adhesive patterning and corroborate them using numerical simulations. In addition to characterizing aspects of cellular dynamics, our results might also be applicable to a range of phenomena in physical chemistry and materials science where flow, deformation, and kinetics are coupled to each other in slender geometries.

  18. Electron Flow through Proteins

    PubMed Central

    Gray, Harry B.; Winkler, Jay R.

    2009-01-01

    Electron transfers in photosynthesis and respiration commonly occur between metal-containing cofactors that are separated by large molecular distances. Employing laser flash-quench triggering methods, we have shown that 20-Å, coupling-limited FeII to RuIII and CuI to RuIII electron tunneling in Ru-modified cytochromes and blue copper proteins can occur on the microsecond timescale both in solutions and crystals. Redox equivalents can be transferred even longer distances by multistep tunneling, often called hopping, through intervening amino acid side chains. Our work has established that 20-Å hole hopping through an intervening tryptophan is two orders of magnitude faster than single-step electron tunneling in a Re-modified blue copper protein. PMID:20161522

  19. A magnetic protein biocompass

    NASA Astrophysics Data System (ADS)

    Qin, Siying; Yin, Hang; Yang, Celi; Dou, Yunfeng; Liu, Zhongmin; Zhang, Peng; Yu, He; Huang, Yulong; Feng, Jing; Hao, Junfeng; Hao, Jia; Deng, Lizong; Yan, Xiyun; Dong, Xiaoli; Zhao, Zhongxian; Jiang, Taijiao; Wang, Hong-Wei; Luo, Shu-Jin; Xie, Can

    2016-02-01

    The notion that animals can detect the Earth’s magnetic field was once ridiculed, but is now well established. Yet the biological nature of such magnetosensing phenomenon remains unknown. Here, we report a putative magnetic receptor (Drosophila CG8198, here named MagR) and a multimeric magnetosensing rod-like protein complex, identified by theoretical postulation and genome-wide screening, and validated with cellular, biochemical, structural and biophysical methods. The magnetosensing complex consists of the identified putative magnetoreceptor and known magnetoreception-related photoreceptor cryptochromes (Cry), has the attributes of both Cry- and iron-based systems, and exhibits spontaneous alignment in magnetic fields, including that of the Earth. Such a protein complex may form the basis of magnetoreception in animals, and may lead to applications across multiple fields.

  20. The interface of protein structure, protein biophysics, and molecular evolution

    PubMed Central

    Liberles, David A; Teichmann, Sarah A; Bahar, Ivet; Bastolla, Ugo; Bloom, Jesse; Bornberg-Bauer, Erich; Colwell, Lucy J; de Koning, A P Jason; Dokholyan, Nikolay V; Echave, Julian; Elofsson, Arne; Gerloff, Dietlind L; Goldstein, Richard A; Grahnen, Johan A; Holder, Mark T; Lakner, Clemens; Lartillot, Nicholas; Lovell, Simon C; Naylor, Gavin; Perica, Tina; Pollock, David D; Pupko, Tal; Regan, Lynne; Roger, Andrew; Rubinstein, Nimrod; Shakhnovich, Eugene; Sjölander, Kimmen; Sunyaev, Shamil; Teufel, Ashley I; Thorne, Jeffrey L; Thornton, Joseph W; Weinreich, Daniel M; Whelan, Simon

    2012-01-01

    Abstract The interface of protein structural biology, protein biophysics, molecular evolution, and molecular population genetics forms the foundations for a mechanistic understanding of many aspects of protein biochemistry. Current efforts in interdisciplinary protein modeling are in their infancy and the state-of-the art of such models is described. Beyond the relationship between amino acid substitution and static protein structure, protein function, and corresponding organismal fitness, other considerations are also discussed. More complex mutational processes such as insertion and deletion and domain rearrangements and even circular permutations should be evaluated. The role of intrinsically disordered proteins is still controversial, but may be increasingly important to consider. Protein geometry and protein dynamics as a deviation from static considerations of protein structure are also important. Protein expression level is known to be a major determinant of evolutionary rate and several considerations including selection at the mRNA level and the role of interaction specificity are discussed. Lastly, the relationship between modeling and needed high-throughput experimental data as well as experimental examination of protein evolution using ancestral sequence resurrection and in vitro biochemistry are presented, towards an aim of ultimately generating better models for biological inference and prediction. PMID:22528593