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Sample records for damaged d-ala-d-ala binding

  1. A Redesigned Vancomycin Engineered for Dual D-Ala-D-Ala and D-Ala-D-Lac Binding Exhibits Potent Antimicrobial Activity Against Vancomycin-Resistant Bacteria

    PubMed Central

    Xie, Jian; Pierce, Joshua G.; James, Robert C.; Okano, Akinori; Boger, Dale L.

    2011-01-01

    The emergence of bacteria resistant to vancomycin, often the antibiotic of last resort, poses a major health problem. Vancomycin-resistant bacteria sense a glycopeptide antibiotic challenge and remodel their cell wall precursor peptidoglycan terminus from D-Ala-d-Ala to D-Ala-D-Lac, reducing the binding of vancomycin to its target 1000-fold and accounting for the loss in antimicrobial activity. Here, we report [Φ[C(=NH)NH]Tpg4]-vancomycin aglycon designed to exhibit the dual binding to D-Ala-D-Ala and D-Ala-D-Lac needed to reinstate activity against vancomycin-resistant bacteria. Its binding to a model D-Ala-D-Ala ligand was found to be only two-fold less than vancomycin aglycon and this affinity was maintained with a model D-Ala-D-Lac ligand, representing a 600-fold increase relative to vancomycin aglycon. Accurately reflecting these binding characteristics, it exhibits potent antimicrobial activity against vancomycin-resistant bacteria (MIC = 0.31 g/mL, VanA VRE). Thus, a complementary single atom exchange in the vancomycin core structure (O NH) to counter the single atom exchange in the cell wall precursors of resistant bacteria (NH O) reinstates potent antimicrobial activity and charts a rational path forward for the development of antibiotics for the treatment of vancomycin-resistant bacterial infections. PMID:21823662

  2. Total Synthesis of [Ψ[C(=NH)NH]Tpg4]Vancomycin and its (4-Chlorobiphenyl)methyl Derivative: Impact of Peripheral Modifications on Vancomycin Analogues Redesigned for Dual d-Ala-d-Ala and d-Ala-d-Lac Binding

    PubMed Central

    2015-01-01

    The total synthesis of two key analogues of vancomycin containing single-atom exchanges in the binding pocket (residue 4 amidine and thioamide) are disclosed as well as their peripherally modified (4-chlorobiphenyl)methyl (CBP) derivatives. Their assessment indicates that combined pocket amidine and CBP peripherally modified analogues exhibit a remarkable spectrum of antimicrobial activity (VSSA, MRSA, VanA and VanB VRE) and impressive potencies (MIC = 0.06–0.005 μg/mL) against both vancomycin-sensitive and -resistant bacteria and likely benefit from two independent and synergistic mechanisms of action. Like vancomycin, such analogues are likely to display especially durable antibiotic activity not prone to rapidly acquired clinical resistance. PMID:25211770

  3. Structure of vancomycin and a vancomycin/D-Ala-D-Ala complex in solution

    SciTech Connect

    Molinari, H.; Pastore, A. ); Lian, Luyun ); Hawkes, G.E.; Sales, K. )

    1990-03-06

    Restrained molecular dynamics simulations were used to study the interactions between the glycopeptide antibiotic vancomycin and the dipeptide Ac-D-Ala-D-Ala. Restraints were obtained from a combination of homonuclear and heteronuclear two-dimensional NMR experiments (NOESY, ROESY, {sup 1}H-{sup 15}N inverse correlation). The comparison between the structures obtained for vancomycin alone and for the complex suggests a new hypothesis on the binding mode of this system. The numerical simulations were not straightforward because vancomycin is made of building blocks for which standard force-fields are not available. The representation of unusual chemical environments is also mandatory. The authors believe that the extension of the force-field parameters to their system could be of more general interest. Furthermore, they consider vancomycin and its complex a good example for exploring the more general problem of molecular recognition, a challenge that has been widely approached in the past few years but for which no unique and general methodology has, so far, been recognized.

  4. Total Synthesis of [Ψ[C(=S)NH]Tpg4]Vancomycin Aglycon, [Ψ[C(=NH)NH]Tpg4]Vancomycin Aglycon, and Related Key Compounds: Reengineering Vancomycin for Dual d-Ala-d-Ala and d-Ala-d-Lac Binding

    PubMed Central

    Xie, Jian; Okano, Akinori; Pierce, Joshua G.; James, Robert C.; Stamm, Simon; Crane, Christine M.; Boger, Dale L.

    2011-01-01

    The total synthesis of [Ψ[C(=S)NH]Tpg4]vancomycin aglycon (8) and its unique AgOAc-promoted single-step conversion to [Ψ[C(=NH)NH]Tpg4]vancomycin aglycon (7), conducted on a fully deprotected substrate, are disclosed. The synthetic approach not only permits access to 7, but it also allows late stage access to related residue 4 derivatives, alternative access to [Ψ[CH2NH]Tpg4]vancomycin aglycon (6) from a common late stage intermediate, and provides authentic residue 4 thioamide and amidine derivatives of the vancomycin aglycon that will facilitate ongoing efforts on their semisynthetic preparation. In addition to early stage residue 4 thioamide introduction, allowing differentiation of one of seven amide bonds central to the vancomycin core structure, the approach relied on two aromatic nucleophilic substitution reactions for formation of the 16-membered diaryl ethers in the CD/DE ring systems, an effective macrolactamization for closure of the 12-membered biaryl AB ring system, and the defined order of CD, AB, and DE ring closures. This order of ring closures follows their increasing ease of thermal atropisomer equilibration, permitting the recycling of any newly generated unnatural atropisomer under progressively milder thermal conditions where the atropoisomer stereochemistry already set is not impacted. Full details of the evaluation of 7 and 8 along with several related key synthetic compounds containing the core residue 4 amidine and thioamide modifications are reported. The binding affinity of compounds containing the residue 4 amidine with the model d-Ala-d-Ala ligand 2 was found to be only 2–3 times less than the vancomycin aglycon (5) and this binding affinity is maintained with the model d-Ala-d-Lac ligand 4, representing a nearly 600-fold increase in affinity relative to the vancomycin aglycon. Importantly, the amidines display effective dual, balanced binding affinity for both ligands (Ka 2/4 = 0.9–1.05) and they exhibit potent antimicrobial

  5. Total Syntheses and Initial Evaluation of [Ψ[C(=S)NH]Tpg4]vancomycin, [Ψ[C(=NH)NH]Tpg4]vancomycin, [Ψ[CH2NH]Tpg4]vancomycin and their (4-Chlorobiphenyl)methyl Derivatives: Synergistic Binding Pocket and Peripheral Modifications for the Glycopeptide Antibiotics

    PubMed Central

    Okano, Akinori; Nakayama, Atsushi; Wu, Kejia; Lindsey, Erick A.; Schammel, Alex W.; Feng, Yiqing; Collins, Karen C.

    2015-01-01

    Full details of studies are disclosed on the total synthesis of binding pocket analogues of vancomycin, bearing the peripheral L-vancosaminyl-1,2-D-glucosyl disaccharide, that contain changes to a key single atom in the residue 4 amide (residue 4 carbonyl O → S, NH, H2) designed to directly address the underlying molecular basis of resistance to vancomycin. Also disclosed are studies piloting the late stage transformations conducted on the synthetically more accessible C-terminus hydroxymethyl aglycon derivatives and full details of the peripheral chlorobiphenyl functionalization of all the binding pocket modified vancomycin analogues designed for dual D-Ala-D-Ala/D-Ala-D-Lac binding are reported. Their collective assessment indicate that combined binding pocket and chlorobiphenyl peripherally modified analogues exhibit a remarkable spectrum of antimicrobial activity (VSSA, MRSA, VanA and VanB VRE) and impressive potencies against both vancomycin-sensitive and vancomycin-resistant bacteria (MICs = 0.06–0.005 μg/mL and 0.5–0.06 μg/mL for the amidine and methylene analogues, respectively) and likely benefit from two independent and synergistic mechanisms of action, only one of which is dependent on D-Ala-D-Ala/D-Ala-D-Lac binding. Such analogues are likely to display especially durable antibiotic activity not prone to rapidly acquired clinical resistance. PMID:25750995

  6. Altering enzymatic activity: recruitment of carboxypeptidase activity into an RTEM beta-lactamase/penicillin-binding protein 5 chimera.

    PubMed

    Chang, Y H; Labgold, M R; Richards, J H

    1990-04-01

    The D-Ala-D-Ala carboxypeptidases/transpeptidases (penicillin-binding proteins, PBPs) share considerable structural homology with class A beta-lactamases (EC 3.5.2.6), although these beta-lactamases have no observable D-Ala-D-Ala carboxypeptidase activity. With the objective of recruiting such activity into a beta-lactamase background, we have prepared a chimeric protein by inserting a 28-amino acid segment of PBP-5 of Escherichia coli in place of the corresponding region of the RTEM-1 beta-lactamase. The segment thus inserted encompasses two residues conserved in both families: Ser-70, which forms the acyl-enzyme intermediate during beta-lactam hydrolysis, and Lys-73, whose presence has been shown to be necessary for catalysis. This chimera involves changes of 18 residues and gives a protein that differs at 7% of the residues from the parent. Whereas RTEM beta-lactamase has no D-Ala-D-Ala carboxypeptidase activity, that of the chimera is significant and is, in fact, about 1% the activity of PBP-5 on diacetyl-L-Lys-D-Ala-D-Ala; in terms of free energy of activation, the chimera stabilizes the transition state for the reaction to within about 2.7 kcal/mol of the stabilization achieved by PBP-5. Furthermore, the chimera catalyzes hydrolysis exclusively at the carboxyl-terminal amide bond which is the site of cleavage by D-Ala-D-Ala carboxypeptidase. Though containing all those residues that are conserved throughout class A beta-lactamases and are thought to be essential for beta-lactamase activity, the chimera has considerably reduced activity (approximately 10(-5) on penams such as penicillins and ampicillins as substrates. As a catalyst, the chimera shows an induction period of approximately 30 min, reflecting a slow conformational rearrangement from an inactive precursor to the active enzyme.

  7. Implementation of a genetically tuned neural platform in optimizing fluorescence from receptor-ligand binding interactions on microchips.

    PubMed

    Alvarado, Judith; Hanrahan, Grady; Nguyen, Huong T H; Gomez, Frank A

    2012-09-01

    This paper describes the use of a genetically tuned neural network platform to optimize the fluorescence realized upon binding 5-carboxyfluorescein-D-Ala-D-Ala-D-Ala (5-FAM-(D-Ala)(3) ) (1) to the antibiotic teicoplanin from Actinoplanes teichomyceticus electrostatically attached to a microfluidic channel originally modified with 3-aminopropyltriethoxysilane. Here, three parameters: (i) the length of time teicoplanin was in the microchannel; (ii) the length of time 1 was in the microchannel, thereby, in equilibrium with teicoplanin, and; (iii) the amount of time buffer was flushed through the microchannel to wash out any unbound 1 remaining in the channel, are examined at a constant concentration of 1, with neural network methodology applied to optimize fluorescence. Optimal neural structure provided a best fit model, both for the training set (r(2) = 0.985) and testing set (r(2) = 0.967) data. Simulated results were experimentally validated demonstrating efficiency of the neural network approach and proved superior to the use of multiple linear regression and neural networks using standard back propagation.

  8. Characterization of Structural Variations in the Peptidoglycan of Vancomycin-Susceptible Enterococcus faecium: Understanding Glycopeptide-Antibiotic Binding Sites using Mass Spectrometry

    PubMed Central

    Patti, Gary J.; Chen, Jiawei; Schaefer, Jacob; Gross, Michael L.

    2008-01-01

    Enterococcus faecium, an opportunistic pathogen that causes a significant number of hospital-acquired infections each year, presents a serious clinical challenge because an increasing number of infections are resistant to the so-called antibiotic of last resort, vancomycin. Vancomycin and other new glycopeptide derivatives target the bacterial cell wall, thereby perturbing its biosynthesis. To help determine the modes of action of glycopeptide antibiotics, we have developed a bottom-up mass spectrometry approach complemented by solid-state NMR to elucidate important structural characteristics of vancomycin-susceptible E. faecium peptidoglycan. Using accurate-mass measurements and integrating ion-current chromatographic peaks of digested peptidoglycan, we identified individual muropeptide species and approximated the relative amount of each. Even though the organism investigated is susceptible to vancomycin, only 3% of the digested peptidoglycan has the well-known D-Ala-D-Ala vancomycin-binding site. The data are consistent with a previously proposed template model of cell-wall biosynthesis where D-Ala-D-Ala stems that are not cross-linked are cleaved in mature peptidoglycan. Additionally, our mass-spectrometry approach allowed differentiation and quantification of muropeptide species seen as unresolved chromatographic peaks. Our method provides an estimate of the extent of muropeptides containing O-acetylation, amidation and hydroxylation, and the number of species forming cyclic imides. The varieties of muropeptides on which the modifications are detected suggest that significant processing occurs in mature peptidoglycan where several enzymes are active in editing cell-wall structure. PMID:18692403

  9. Molecular basis for the role of Staphylococcus aureus penicillin binding protein 4 in antimicrobial resistance.

    PubMed

    Navratna, Vikas; Nadig, Savitha; Sood, Varun; Prasad, K; Arakere, Gayathri; Gopal, B

    2010-01-01

    Penicillin binding proteins (PBPs) are membrane-associated proteins that catalyze the final step of murein biosynthesis. These proteins function as either transpeptidases or carboxypeptidases and in a few cases demonstrate transglycosylase activity. Both transpeptidase and carboxypeptidase activities of PBPs occur at the D-Ala-D-Ala terminus of a murein precursor containing a disaccharide pentapeptide comprising N-acetylglucosamine and N-acetyl-muramic acid-L-Ala-D-Glu-L-Lys-D-Ala-D-Ala. Beta-lactam antibiotics inhibit these enzymes by competing with the pentapeptide precursor for binding to the active site of the enzyme. Here we describe the crystal structure, biochemical characteristics, and expression profile of PBP4, a low-molecular-mass PBP from Staphylococcus aureus strain COL. The crystal structures of PBP4-antibiotic complexes reported here were determined by molecular replacement, using the atomic coordinates deposited by the New York Structural Genomics Consortium. While the pbp4 gene is not essential for the viability of S. aureus, the knockout phenotype of this gene is characterized by a marked reduction in cross-linked muropeptide and increased vancomycin resistance. Unlike other PBPs, we note that expression of PBP4 was not substantially altered under different experimental conditions, nor did it change across representative hospital- or community-associated strains of S. aureus that were examined. In vitro data on purified recombinant S. aureus PBP4 suggest that it is a beta-lactamase and is not trapped as an acyl intermediate with beta-lactam antibiotics. Put together, the expression analysis and biochemical features of PBP4 provide a framework for understanding the function of this protein in S. aureus and its role in antimicrobial resistance.

  10. Determination of Noncovalent Binding Using a Continuous Stirred Tank Reactor as a Flow Injection Device Coupled to Electrospray Ionization Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Santos, Inês C.; Waybright, Veronica B.; Fan, Hui; Ramirez, Sabra; Mesquita, Raquel B. R.; Rangel, António O. S. S.; Fryčák, Petr; Schug, Kevin A.

    2015-07-01

    Described is a new method based on the concept of controlled band dispersion, achieved by hyphenating flow injection analysis with ESI-MS for noncovalent binding determinations. A continuous stirred tank reactor (CSTR) was used as a FIA device for exponential dilution of an equimolar host-guest solution over time. The data obtained was treated for the noncovalent binding determination using an equimolar binding model. Dissociation constants between vancomycin and Ac-Lys(Ac)-Ala-Ala-OH peptide stereoisomers were determined using both the positive and negative ionization modes. The results obtained for Ac- L-Lys(Ac)- D-Ala- D-Ala (a model for a Gram-positive bacterial cell wall) binding were in reasonable agreement with literature values made by other mass spectrometry binding determination techniques. Also, the developed method allowed the determination of dissociation constants for vancomycin with Ac- L-Lys(Ac)- D-Ala- L-Ala, Ac- L-Lys(Ac)- L-Ala- D-Ala, and Ac- L-Lys(Ac)- L-Ala- L-Ala. Although some differences in measured binding affinities were noted using different ionization modes, the results of each determination were generally consistent. Differences are likely attributable to the influence of a pseudo-physiological ammonium acetate buffer solution on the formation of positively- and negatively-charged ionic complexes.

  11. Determination of Noncovalent Binding Using a Continuous Stirred Tank Reactor as a Flow Injection Device Coupled to Electrospray Ionization Mass Spectrometry.

    PubMed

    Santos, Inês C; Waybright, Veronica B; Fan, Hui; Ramirez, Sabra; Mesquita, Raquel B R; Rangel, António O S S; Fryčák, Petr; Schug, Kevin A

    2015-07-01

    Described is a new method based on the concept of controlled band dispersion, achieved by hyphenating flow injection analysis with ESI-MS for noncovalent binding determinations. A continuous stirred tank reactor (CSTR) was used as a FIA device for exponential dilution of an equimolar host-guest solution over time. The data obtained was treated for the noncovalent binding determination using an equimolar binding model. Dissociation constants between vancomycin and Ac-Lys(Ac)-Ala-Ala-OH peptide stereoisomers were determined using both the positive and negative ionization modes. The results obtained for Ac-L-Lys(Ac)-D-Ala-D-Ala (a model for a Gram-positive bacterial cell wall) binding were in reasonable agreement with literature values made by other mass spectrometry binding determination techniques. Also, the developed method allowed the determination of dissociation constants for vancomycin with Ac-L-Lys(Ac)-D-Ala-L-Ala, Ac-L-Lys(Ac)-L-Ala-D-Ala, and Ac-L-Lys(Ac)-L-Ala-L-Ala. Although some differences in measured binding affinities were noted using different ionization modes, the results of each determination were generally consistent. Differences are likely attributable to the influence of a pseudo-physiological ammonium acetate buffer solution on the formation of positively- and negatively-charged ionic complexes.

  12. Experimental Strategies for Functional Annotation and Metabolism Discovery: Targeted Screening of Solute Binding Proteins and Unbiased Panning of Metabolomes

    PubMed Central

    2015-01-01

    The rate at which genome sequencing data is accruing demands enhanced methods for functional annotation and metabolism discovery. Solute binding proteins (SBPs) facilitate the transport of the first reactant in a metabolic pathway, thereby constraining the regions of chemical space and the chemistries that must be considered for pathway reconstruction. We describe high-throughput protein production and differential scanning fluorimetry platforms, which enabled the screening of 158 SBPs against a 189 component library specifically tailored for this class of proteins. Like all screening efforts, this approach is limited by the practical constraints imposed by construction of the library, i.e., we can study only those metabolites that are known to exist and which can be made in sufficient quantities for experimentation. To move beyond these inherent limitations, we illustrate the promise of crystallographic- and mass spectrometric-based approaches for the unbiased use of entire metabolomes as screening libraries. Together, our approaches identified 40 new SBP ligands, generated experiment-based annotations for 2084 SBPs in 71 isofunctional clusters, and defined numerous metabolic pathways, including novel catabolic pathways for the utilization of ethanolamine as sole nitrogen source and the use of d-Ala-d-Ala as sole carbon source. These efforts begin to define an integrated strategy for realizing the full value of amassing genome sequence data. PMID:25540822

  13. A microplate assay for the coupled transglycosylase-transpeptidase activity of the penicillin binding proteins; a vancomycin-neutralizing tripeptide combination prevents penicillin inhibition of peptidoglycan synthesis.

    PubMed

    Kumar, Vidya P; Basavannacharya, Chandrakala; de Sousa, Sunita M

    2014-07-18

    A microplate, scintillation proximity assay to measure the coupled transglycosylase-transpeptidase activity of the penicillin binding proteins in Escherichia coli membranes was developed. Membranes were incubated with the two peptidoglycan sugar precursors UDP-N-acetyl muramylpentapeptide (UDP-MurNAc(pp)) and UDP-[(3)H]N-acetylglucosamine in the presence of 40 μM vancomycin to allow in situ accumulation of lipid II. In a second step, vancomycin inhibition was relieved by addition of a tripeptide (Lys-D-ala-D-ala) or UDP-MurNAc(pp), resulting in conversion of lipid II to cross-linked peptidoglycan. Inhibitors of the transglycosylase or transpeptidase were added at step 2. Moenomycin, a transglycosylase inhibitor, had an IC50 of 8 nM. Vancomycin and nisin also inhibited the assay. Surprisingly, the transpeptidase inhibitors penicillin and ampicillin showed no inhibition. In a pathway assay of peptidoglycan synthesis, starting from the UDP linked sugar precursors, inhibition by penicillin was reversed by a 'neutral' combination of vancomycin plus tripeptide, suggesting an interaction thus far unreported.

  14. New insights into glycopeptide antibiotic binding to cell wall precursors using SPR and NMR spectroscopy.

    PubMed

    Treviño, Juan; Bayón, Carlos; Ardá, Ana; Marinelli, Flavia; Gandolfi, Raffaella; Molinari, Francesco; Jimenez-Barbero, Jesús; Hernáiz, María J

    2014-06-10

    Glycopeptide antibiotics, such as vancomycin and teicoplanin, are used to treat life-threatening infections caused by multidrug-resistant Gram-positive pathogens. They inhibit bacterial cell wall biosynthesis by binding to the D-Ala-D-Ala C-terminus of peptidoglycan precursors. Vancomycin-resistant bacteria replace the dipeptide with the D-Ala-D-Lac depsipeptide, thus reducing the binding affinity of the antibiotics with their molecular targets. Herein, studies of the interaction of teicoplanin, teicoplanin-like A40926, and of their semisynthetic derivatives (mideplanin, MDL63,246, dalbavancin) with peptide analogues of cell-wall precursors by NMR spectroscopy and surface plasmon resonance (SPR) are reported. NMR spectroscopy revealed the existence of two different complexes in solution, when the different glycopeptides interact with Ac2KdAlaDAlaOH. Despite the NMR experimental conditions, which are different from those employed for the SPR measurements, the NMR spectroscopy results parallel those deduced in the chip with respect to the drastic binding difference existing between the D-Ala and the D-Lac terminating analogues, confirming that all these antibiotics share the same primary molecular mechanism of action and resistance. Kinetic analysis of the interaction between the glycopeptide antibiotics and immobilized AcKdAlaDAlaOH by SPR suggest a dimerization process that was not observed by NMR spectroscopy in DMSO solution. Moreover, in SPR, all glycopeptides with a hydrophobic acyl chain present stronger binding with a hydrophobic surface than vancomycin, indicating that additional interactions through the employed surface are involved. In conclusion, SPR provides a tool to differentiate between vancomycin and other glycopeptides, and the calculated binding affinities at the surface seem to be more relevant to in vitro antimicrobial activity than the estimations from NMR spectroscopy analysis.

  15. Structural variations of the cell wall precursor lipid II and their influence on binding and activity of the lipoglycopeptide antibiotic oritavancin.

    PubMed

    Münch, Daniela; Engels, Ina; Müller, Anna; Reder-Christ, Katrin; Falkenstein-Paul, Hildegard; Bierbaum, Gabriele; Grein, Fabian; Bendas, Gerd; Sahl, Hans-Georg; Schneider, Tanja

    2015-02-01

    Oritavancin is a semisynthetic derivative of the glycopeptide antibiotic chloroeremomycin with activity against Gram-positive pathogens, including vancomycin-resistant staphylococci and enterococci. Compared to vancomycin, oritavancin is characterized by the presence of two additional residues, a hydrophobic 4'-chlorobiphenyl methyl moiety and a 4-epi-vancosamine substituent, which is also present in chloroeremomycin. Here, we show that oritavancin and its des-N-methylleucyl variant (des-oritavancin) effectively inhibit lipid I- and lipid II-consuming peptidoglycan biosynthesis reactions in vitro. In contrast to that for vancomycin, the binding affinity of oritavancin to the cell wall precursor lipid II appears to involve, in addition to the D-Ala-D-Ala terminus, other species-specific binding sites of the lipid II molecule, i.e., the crossbridge and D-isoglutamine in position 2 of the lipid II stem peptide, both characteristic for a number of Gram-positive pathogens, including staphylococci and enterococci. Using purified lipid II and modified lipid II variants, we studied the impact of these modifications on the binding of oritavancin and compared it to those of vancomycin, chloroeremomycin, and des-oritavancin. Analysis of the binding parameters revealed that additional intramolecular interactions of oritavancin with the peptidoglycan precursor appear to compensate for the loss of a crucial hydrogen bond in vancomycin-resistant strains, resulting in enhanced binding affinity. Augmenting previous findings, we show that amidation of the lipid II stem peptide predominantly accounts for the increased binding of oritavancin to the modified intermediates ending in D-Ala-D-Lac. Corroborating our conclusions, we further provide biochemical evidence for the phenomenon of the antagonistic effects of mecA and vanA resistance determinants in Staphylococcus aureus, thus partially explaining the low frequency of methicillin-resistant S. aureus (MRSA) acquiring high

  16. Damage-specific DNA-binding proteins from human cells

    SciTech Connect

    Kanjilal, S.

    1992-01-01

    The primary objective of the study was to detect and characterize factors from human cells that bind DNA damaged by ultraviolet radiation. An application of the gel-shift assay was devised in which a DNA probe was UV-irradiated and compared with non-irradiated probe DNA for the ability to bind to such factors in cell extracts. UV-dose dependent binding proteins were identified. Formation of the DNA-protein complexes was independent of the specific sequence, form or source of the DNA. There was a marked preference for lesions on double stranded DNA over those on single stranded DNA. DNA irradiated with gamma rays did not compete with UV-irradiated DNA for the binding activities. Cell lines from patients with genetic diseases associated with disorders of the DNA repair system were screened for the presence of damaged-DNA-binding activities. Simultaneous occurrence of the clinical symptoms of some of these diseases had been previously documented and possible links between the syndromes proposed. However, supporting biochemical or molecular evidence for such associations were lacking. The data from the present investigations indicate that some cases of Xeroderma Pigmentosum group A, Cockayne's Syndrome, Bloom's Syndrome and Ataxia Telangiectasia, all of which exhibit sensitivity to UV or gamma radiation, share an aberrant damaged-DNA-binding factor. These findings support the hypothesis that some of the repair disorder diseases are closely related and may have arisen from a common defect. Partial purification of the binding activities from HeLa cells was achieved. Size-exclusion chromatography resolved the activities into various peaks, one of which was less damage-specific than the others as determined by competition studies using native or UV-irradiated DNA. Some of the activities were further separated by ion-exchange chromatography. On using affinity chromatography methods, the major damage-binding factor could be eluted in the presence of 2 M KCl and 1% NP-40.

  17. Experimental and Theoretical Studies of the Structures and Interactions of Vancomycin Antibiotics with Cell Wall Analogues

    SciTech Connect

    Yang, Zhibo; Vorpagel, Erich R.; Laskin, Julia

    2008-10-01

    Surface-induced dissociation (SID) of the singly protonated complex of vancomycin antibiotic with cell wall peptide analogue (Nα,Nε-diacetyl-L-Lys-D-Ala-D-Ala) was studied using a 6 T Fourier Transform Ion Cyclotron Resonance Mass Spectrometer (FT-ICR MS) specially configured for SID experiments. The binding energy between the vancomycin and the peptide was obtained from the RRKM modeling of the time- and energy resolved fragmentation efficiency curves (TFECs) of the precursor ion and its fragments. Electronic structure calculations of the geometries, proton affinities and binding energies were performed for several model systems including vancomycin (V), vancomycin aglycon (VA), Nα,Nε-diacetyl-L-Lys-D-Ala-D-Ala, and non-covalent complexes of VA with N-acetyl-D-Ala-D-Ala and Nα,Nε-diacetyl-L-Lys-D-Ala-D-Ala at the B3LYP/6-31G(d) level of theory. Comparison between the experimental and computational results suggests that the most probable structure of the complex observed in our experiments corresponds to the neutral peptide bound to the vancomycin protonated at the secondary amino group of the N-methyl-leucine residue. The experimental binding energy of 30.9 ± 1.8 kcal/mol is in good agreement with the binding energy of 29.3 ± 2.5 kcal/mol calculated for the model system representing the preferred structure of the complex.

  18. The DNA damage-recognition problem in human and other eukaryotic cells: the XPA damage binding protein.

    PubMed Central

    Cleaver, J E; States, J C

    1997-01-01

    The capacity of human and other eukaryotic cells to recognize a disparate variety of damaged sites in DNA, and selectively excise and repair them, resides in a deceptively small simple protein, a 38-42 kDa zinc-finger binding protein, XPA (xeroderma pigmentosum group A), that has no inherent catalytic properties. One key to its damage-recognition ability resides in a DNA-binding domain which combines a zinc finger and a single-strand binding region which may infiltrate small single-stranded regions caused by helix-destabilizing lesions. Another is the augmentation of its binding capacity by interactions with other single-stranded binding proteins and helicases which co-operate in the binding and are unloaded at the binding site to facilitate further unwinding of the DNA and subsequent catalysis. The properties of these reactions suggest there must be considerable conformational changes in XPA and associated proteins to provide a flexible fit to a wide variety of damaged structures in the DNA. PMID:9359827

  19. Hydrogen Bonding Motifs in MutSaphla and their response to binding damaged DNA

    NASA Astrophysics Data System (ADS)

    Negureanu, Lacra; Salsbury, Freddie

    2013-03-01

    Over the past decade, there has been a growing interest in studying the binding of damaged DNA to the MutSalpha protein complex. This protein complex, the Msh2/Msh6 complex in humans, is the initial complex that binds mismatched DNA and other DNA defects that occur during replication. This complex has also been shown to bind at least some types of damaged DNA. As a result of this interest, multiple studies have contrasted the interactions of MutSalpha with its normal mismatched substrate and with the interactions of MutsSalpha to DNA damaged by the chemotherapeutic cisplatin. To complement these studies, we examined the interaction between MutSalpha and DNA damaged by carboplatin via all-atom molecular dynamics simulations. These simulations provide evidence for different hydrogen bonding interactions at the protein/DNA and protein/protein interface. The hydrogen bonding motifs found are broadly similar to those found in binding to the adduct from cis-platin, but have distinct differences. These subtle differences may play a role in the way the different damages are signaled by MutS.

  20. UV damage-specific DNA-binding protein in xeroderma pigmentosum complementation group E

    SciTech Connect

    Kataoka, H.; Fujiwara, Y. )

    1991-03-29

    The gel mobility shift assay method revealed a specifically ultraviolet (UV) damage recognizing, DNA-binding protein in nuclear extracts of normal human cells. The resulted DNA/protein complexes caused the two retarded mobility shifts. Four xeroderma pigmentosum complementation group E (XPE) fibroblast strains derived from unrelated Japanese families were not deficient in such a DNA damage recognition/binding protein because of the normal complex formation and gel mobility shifts, although we confirmed the reported lack of the protein in the European XPE (XP2RO and XP3RO) cells. Thus, the absence of this binding protein is not always commonly observed in all the XPE strains, and the partially repair-deficient and intermediately UV-hypersensitive phenotype of XPE cells are much similar whether or not they lack the protein.

  1. Phospho-Ser/Thr-binding domains: navigating the cell cycle and DNA damage response.

    PubMed

    Reinhardt, H Christian; Yaffe, Michael B

    2013-09-01

    Coordinated progression through the cell cycle is a complex challenge for eukaryotic cells. Following genotoxic stress, diverse molecular signals must be integrated to establish checkpoints specific for each cell cycle stage, allowing time for various types of DNA repair. Phospho-Ser/Thr-binding domains have emerged as crucial regulators of cell cycle progression and DNA damage signalling. Such domains include 14-3-3 proteins, WW domains, Polo-box domains (in PLK1), WD40 repeats (including those in the E3 ligase SCF(βTrCP)), BRCT domains (including those in BRCA1) and FHA domains (such as in CHK2 and MDC1). Progress has been made in our understanding of the motif (or motifs) that these phospho-Ser/Thr-binding domains connect with on their targets and how these interactions influence the cell cycle and DNA damage response.

  2. Eukaryotic damaged DNA-binding proteins: DNA repair proteins or transcription factors?

    SciTech Connect

    Protic, M.

    1994-12-31

    Recognition and removal of structural defects in the genome, caused by diverse physical and chemical agents, are among the most important cell functions. Proteins that recognize and bind to modified DNA, and thereby initiate damage-induced recovery processes, have been identified in prokaryotic and eukaryotic cells. Damaged DNA-binding (DDB) proteins from prokaryotes are either DNA repair enzymes or noncatalytic subunits of larger DNA repair complexes that participate in excision repair, or in recombinational repair and SOS-mutagenesis. Although the methods employed may not have allowed detection of all eukaryotic DDB proteins and identification of their functions, it appears that during evolution cells have developed a wide array of DDB proteins that can discriminate among the diversity of DNA conformations found in the eukaryotic nucleus, as well as a gene-sharing feature found in DDB proteins that also act as transcription factors.

  3. Ubiquitin-binding domains and their role in the DNA damage response.

    PubMed

    Hofmann, Kay

    2009-04-05

    The modification of eukaryotic proteins by covalent attachment of ubiquitin is a versatile signaling event with a wide range of possible consequences. Canonical poly-ubiquitination by Lys-48 linked chains usually destines a protein for degradation by the proteasome. By contrast, attachment of a single ubiquitin or ubiquitin chains linked through Lys-63 or Lys-6 serves a non-proteolytic role. Over the last years, evidence has accumulated that several nuclear proteins become ubiquitinated in response to DNA damage. Typically, these proteins carry mono-ubiquitin or non-classical ubiquitin chains and are localized close to the site of DNA damage. Of particular interest are PCNA and the variant histone H2AX, two key proteins whose ubiquitination serves to recruit factors needed by the cell to cope with the damage. A prerequisite for docking effector proteins to the site of the lesion is the detection of a specific ubiquitin modification, a process that can be mediated by a range of dedicated ubiquitin-binding domains (UBDs). As the same types of ubiquitin modification are involved in entirely different processes, the recognition of the ubiquitin mark has to go along with the recognition of the modified protein. Thus, ubiquitin-binding domains gain their specificity through combination with other recognition domains and motifs. This review discusses ubiquitin-binding domains relevant to the DNA damage response, including their binding mode, their specificity, and their interdependence with other factors. For several repair pathways, current knowledge of the events downstream of the ubiquitin mark is sketchy. A closer look at orphan UBD proteins might lead to the identification of missing pieces in the DNA response puzzle.

  4. A constitutive damage specific DNA-binding protein is synthesized at higher levels in UV-irradiated primate cells

    SciTech Connect

    Hirschfeld, S.; Levine, A.S.; Ozato, K.; Protic, M. )

    1990-05-01

    Using a DNA band shift assay, we have identified a DNA-binding protein complex in primate cells which is present constitutively and has a high affinity for UV-irradiated, double-stranded DNA. Cells pretreated with UV light, mitomycin C, or aphidicolin have higher levels of this damage-specific DNA-binding protein complex, suggesting that the signal for induction can either be damage to the DNA or interference with cellular DNA replication. Physiochemical modifications of the DNA and competition analysis with defined substrates suggest that the most probable target site for the damage-specific DNA-binding protein complex is a 6-4'-(pyrimidine-2'-one)-pyrimidine dimer: specific binding could not be detected with probes which contain -TT- cyclobutane dimers, and damage-specific DNA binding did not decrease after photoreactivation of UV-irradiated DNA. This damage-specific DNA-binding protein complex is the first such inducible protein complex identified in primate cells. Cells from patients with the sun-sensitive cancer-prone disease, xeroderma pigmentosum (group E), are lacking both the constitutive and the induced damage-specific DNA-binding activities. These findings suggest a possible role for this DNA-binding protein complex in lesion recognition and DNA repair of UV-light-induced photoproducts.

  5. Albumin Binding Function: The Potential Earliest Indicator for Liver Function Damage

    PubMed Central

    Ge, Penglei; Yang, Huayu; Lu, Jingfen; Liao, Wenjun; Du, Shunda; Xu, Yingli; Xu, Haifeng; Lu, Xin; Sang, Xinting; Zhong, Shouxian; Huang, Jiefu

    2016-01-01

    Background. Currently there is no indicator that can evaluate actual liver lesion for early stages of viral hepatitis, nonalcoholic fatty liver disease (NAFLD), and cirrhosis. Aim of this study was to investigate if albumin binding function could better reflect liver function in these liver diseases. Methods. An observational study was performed on 193 patients with early NAFLD, viral hepatitis, and cirrhosis. Cirrhosis patients were separated according to Child-Pugh score into A, B, and C subgroup. Albumin metal ion binding capacity (Ischemia-modified albumin transformed, IMAT) and fatty acid binding capacity (total binding sites, TBS) were detected. Results. Both IMAT and TBS were significantly decreased in patients with NAFLD and early hepatitis. In hepatitis group, they declined prior to changes of liver enzymes. IMAT was significantly higher in cirrhosis Child-Pugh class A group than hepatitis patients and decreased in Child-Pugh class B and class C patients. Both IMAT/albumin and TBS/albumin decreased significantly in hepatitis and NAFLD group patients. Conclusions. This is the first study to discover changes of albumin metal ion and fatty acid binding capacities prior to conventional biomarkers for liver damage in early stage of liver diseases. They may become potential earliest sensitive indicators for liver function evaluation. PMID:28101103

  6. Distinct binding of BRCA2 BRC repeats to RAD51 generates differential DNA damage sensitivity

    PubMed Central

    Chatterjee, Gouri; Jimenez-Sainz, Judit; Presti, Thomas; Nguyen, Tiffany; Jensen, Ryan B.

    2016-01-01

    BRCA2 is a multi-faceted protein critical for the proper regulation of homology-directed repair of DNA double-strand breaks. Elucidating the mechanistic features of BRCA2 is crucial for understanding homologous recombination and how patient-derived mutations impact future cancer risk. Eight centrally located BRC repeats in BRCA2 mediate binding and regulation of RAD51 on resected DNA substrates. Herein, we dissect the biochemical and cellular features of the BRC repeats tethered to the DNA binding domain of BRCA2. To understand how the BRC repeats and isolated domains of BRCA2 contribute to RAD51 binding, we analyzed both the biochemical and cellular properties of these proteins. In contrast to the individual BRC repeat units, we find that the BRC5–8 region potentiates RAD51-mediated DNA strand pairing and provides complementation functions exceeding those of BRC repeats 1–4. Furthermore, BRC5–8 can efficiently repair nuclease-induced DNA double-strand breaks and accelerate the assembly of RAD51 repair complexes upon DNA damage. These findings highlight the importance of the BRC5–8 domain in stabilizing the RAD51 filament and promoting homology-directed repair under conditions of cellular DNA damage. PMID:27084934

  7. DNA-Damage Response RNA-Binding Proteins (DDRBPs): Perspectives from a New Class of Proteins and Their RNA Targets.

    PubMed

    Dutertre, Martin; Vagner, Stéphan

    2016-09-29

    Upon DNA damage, cells trigger an early DNA-damage response (DDR) involving DNA repair and cell cycle checkpoints, and late responses involving gene expression regulation that determine cell fate. Screens for genes involved in the DDR have found many RNA-binding proteins (RBPs), while screens for novel RBPs have identified DDR proteins. An increasing number of RBPs are involved in early and/or late DDR. We propose to call this new class of actors of the DDR, which contain an RNA-binding activity, DNA-damage response RNA-binding proteins (DDRBPs). We then discuss how DDRBPs contribute not only to gene expression regulation in the late DDR but also to early DDR signaling, DNA repair, and chromatin modifications at DNA-damage sites through interactions with both long and short noncoding RNAs.

  8. Urinary Excretion of Liver Type Fatty Acid Binding Protein Accurately Reflects the Degree of Tubulointerstitial Damage

    PubMed Central

    Yokoyama, Takeshi; Kamijo-Ikemori, Atsuko; Sugaya, Takeshi; Hoshino, Seiko; Yasuda, Takashi; Kimura, Kenjiro

    2009-01-01

    To investigate the relationship between liver-type fatty acid-binding protein (L-FABP), a biomarker of chronic kidney disease, in the kidney and the degree of tubulointerstitial damage, folic acid (FA)-induced nephropathy was studied in a mouse model system. As renal L-FABP is not expressed in wild-type mice, human L-FABP (hL-FABP) transgenic mice were used in this study. hL-FABP is expressed in the renal proximal tubules of the transgenic mice that were injected intraperitoneally with FA in NaHCO3 (the FA group) or only NaHCO3 (the control group) and oral saline solution daily during the experimental period. The FA group developed severe tubulointerstitial damage with the infiltration of macrophages and the deposition of type I collagen on days 3 and 7 and recovered to the control level on day 14. The gene and protein expression levels of hL-FABP in the kidney were significantly enhanced on days 3 and 7. Urinary hL-FABP in the FA group was elevated on days 3 and 7 and decreased to the control level on day 14. The protein expression levels of hL-FABP in both the kidney and urine significantly correlated with the degree of tubulointerstitial damage, the infiltration of macrophages, and the deposition of type I collagen. In conclusion, renal expression and urinary excretion of hL-FABP significantly reflected the severity of tubulointerstitial damage in FA-induced nephropathy. PMID:19435794

  9. Mannose-binding lectin genotypes: potential role in tubal damage and adverse IVF outcome.

    PubMed

    Laisk, Triin; Peters, Maire; Salumets, Andres

    2011-12-01

    The innate immune system provides the first-line defence against genital tract pathogens and is also involved in establishing and maintaining a successful pregnancy. Genetic variation of factors regulating immune response can be associated with complications after genital tract infections and may lead to unfavourable pregnancy outcomes. This study focused on four polymorphisms in the mannose binding lectin gene (MBL2) and assessed their significance in tubal damage and female fertility by comparing genotype frequencies among 388 controls and women with tubal factor infertility (n=155) or previous ectopic pregnancy (n=178). The high-producing MBL2 genotype HYA/LYA was found to have a protective effect, while the hyper-producing MBL2 genotype HYA/HYA and low-producing MBL2 genotypes were associated with susceptibility to tubal factor infertility. Also, the low-producing genotypes showed association with early pregnancy loss in IVF treatment. In conclusion, these data suggest that certain MBL2 genotypes can be associated with tubal damage in patients with evidence of Chlamydia trachomatis infection and additionally may contribute to the pathogenesis of early pregnancy loss.

  10. Investigation into the functional impact of the vancomycin C-ring aryl chloride

    PubMed Central

    Pinchman, Joseph R.; Boger, Dale L.

    2013-01-01

    A vancomycin aglycon analogue that possesses a reduced C-ring and an intact E-ring chloride was prepared and its antimicrobial activity towards S. aureus and binding affinity to model cell wall ligands were established. Comparison of the derivative with a series of vancomycin aglycon analogues that possess and lack the chloro substituents on the aryl C- and E-rings defines the impact and further refines the role the C-ring chloride plays in promoting both target binding affinity and binding selectivity for d-Ala-d-Ala and its impact on antimicrobial activity. PMID:23880541

  11. Bio-inspired synthetic receptor molecules towards mimicry of vancomycin.

    PubMed

    Monnee, M C; Brouwer, A J; Verbeek, L M; van Wageningen, A M; Liskamp, R M

    2001-06-18

    A 512-member library of bio-inspired synthetic receptor molecules was prepared featuring a triazacyclophane scaffold. The purpose of this scaffold was to orient three (identical) peptide 'binding arms' in order to mimic an antibiotic binding cavity as is present in the vancomycin antibiotics. The library was screened with D-Ala-D-Ala and D-Ala-D-Lac containing ligands, which are present in the cell wall precursors of pathogenic bacteria. Screening and validation led to identification of a synthetic receptor capable of binding these ligands.

  12. A damage-responsive DNA binding protein regulates transcription of the yeast DNA repair gene PHR1.

    PubMed Central

    Sebastian, J; Sancar, G B

    1991-01-01

    The PHR1 gene of Saccharomyces cerevisiae encodes the DNA repair enzyme photolyase. Transcription of PHR1 increases in response to treatment of cells with 254-nm radiation and chemical agents that damage DNA. We report here the identification of a damage-responsive DNA binding protein, termed photolyase regulatory protein (PRP), and its cognate binding site, termed the PHR1 upstream repression sequence, that together regulate induction of PHR1 transcription after DNA damage. PRP activity, monitored by electrophoretic-mobility-shift assay, was detected in cells during normal growth but disappeared within 30 min after irradiation. Copper-phenanthroline footprinting of PRP-DNA complexes revealed that PRP protects a 39-base-pair region of PHR1 5' flanking sequence beginning 40 base pairs upstream from the coding sequence. A prominent feature of the foot-printed region is a 22-base-pair palindrome. Deletion of the PHR1 upstream repression sequence increased the basal level expression of PHR1 in vivo and decreased induction after exposure of cells to UV radiation or methyl methanesulfonate, whereas insertion of the PRP binding site between the CYC1 upstream activation sequence and "TATA" sequence reduced basal level expression and conferred damage responsiveness upon a reporter gene. Thus these observations establish that PRP is a damage-responsive repressor of PHR1 transcription. Images PMID:1763039

  13. Coumarin-chalcone hybrid instigates DNA damage by minor groove binding and stabilizes p53 through post translational modifications

    PubMed Central

    Ashraf, Raghib; Hamidullah; Hasanain, Mohammad; Pandey, Praveen; Maheshwari, Mayank; Singh, L. Ravithej; Siddiqui, M. Quadir; Konwar, Rituraj; Sashidhara, Koneni V.; Sarkar, Jayanta

    2017-01-01

    S009-131, a coumarin-chalcone hybrid, had been shown to possess anti-proliferative and anti-tumour effect by triggering apoptosis. In this report, we investigated role of DNA damage signalling pathway in S009-131 induced cancer cell death. Here we show that S009-131 causes DNA damage by potential binding to the minor groove which led to the phosphorylation and activation of ATM and DNA-PK, but not ATR, at earlier time points in order to initiate repair process. S009-131 induced DNA damage response triggered activation of p53 through phosphorylation at its key residues. Pharmacological inhibition of PIKKs abrogated S009-131 induced phosphorylation of p53 at Ser 15. DNA damage induced phosphorylation resulted in reduced proteasomal degradation of p53 by disrupting p53-MDM2 interaction. Additionally, our docking studies revealed that S009-131 might also contribute to increased cellular p53 level by occupying p53 binding pocket of MDM2. Posttranslational modifications of p53 upon S009-131 treatment led to enhanced affinity of p53 towards responsive elements (p53-RE) in the promoter regions of target genes and increased transcriptional efficiency. Together, the results suggest that S009-131 cleaves DNA through minor groove binding and eventually activates PIKKs associated DNA damage response signalling to promote stabilization and enhanced transcriptional activity of p53 through posttranslational modifications at key residues. PMID:28349922

  14. Phosphorylation of OPTN by TBK1 enhances its binding to Ub chains and promotes selective autophagy of damaged mitochondria

    PubMed Central

    Richter, Benjamin; Sliter, Danielle A.; Herhaus, Lina; Stolz, Alexandra; Wang, Chunxin; Beli, Petra; Zaffagnini, Gabriele; Wild, Philipp; Martens, Sascha; Wagner, Sebastian A.; Youle, Richard J.; Dikic, Ivan

    2016-01-01

    Selective autophagy of damaged mitochondria requires autophagy receptors optineurin (OPTN), NDP52 (CALCOCO2), TAX1BP1, and p62 (SQSTM1) linking ubiquitinated cargo to autophagic membranes. By using quantitative proteomics, we show that Tank-binding kinase 1 (TBK1) phosphorylates all four receptors on several autophagy-relevant sites, including the ubiquitin- and LC3-binding domains of OPTN and p62/SQSTM1 as well as the SKICH domains of NDP52 and TAX1BP1. Constitutive interaction of TBK1 with OPTN and the ability of OPTN to bind to ubiquitin chains are essential for TBK1 recruitment and kinase activation on mitochondria. TBK1 in turn phosphorylates OPTN’s UBAN domain at S473, thereby expanding the binding capacity of OPTN to diverse Ub chains. In combination with phosphorylation of S177 and S513, this posttranslational modification promotes recruitment and retention of OPTN/TBK1 on ubiquitinated, damaged mitochondria. Moreover, phosphorylation of OPTN on S473 enables binding to pS65 Ub chains and is also implicated in PINK1-driven and Parkin-independent mitophagy. Thus, TBK1-mediated phosphorylation of autophagy receptors creates a signal amplification loop operating in selective autophagy of damaged mitochondria. PMID:27035970

  15. Drosophila damaged DNA-binding protein 1 is an essential factor for development.

    PubMed

    Takata, Kei-ichi; Yoshida, Hideki; Yamaguchi, Masamitsu; Sakaguchi, Kengo

    2004-10-01

    The damaged DNA-binding protein (DDB) complex, thought to recognize (6-4) photoproducts and other lesions in DNA, has been implicated to have a role in global genomic nucleotide excision repair (NER) and E2F-1-mediated transcription. The complex consists of a heterodimer of p127 (DDB1) and p48 (DDB2), the latter also being known as XPE. We reported previously that in Drosophila expression of the DDB1 (D-DDB1) gene is controlled by the DRE/DREF system, and external injury to DNA is not essential for D-DDB1 function. In the present study of the function of D-DDB1 in a multicellular system, we prepared transgenic flies, which were knocked down for the D-DDB1 gene due to RNA interference (RNAi), and performed immunocytochemistry to ascertain the distribution of D-DDB1 in the eye imaginal disc. It was found to be abundant in the anterior of the morphogenetic furrow (MF). Whole-body overexpression of dsRNA of D-DDB1 in Drosophila using a GAL4-UAS targeted expression system induced melanotic tumors and caused complete lethality. When limited to the eye imaginal disc, a severe rough eye phenotype resulted. Correspondingly, all of the D-DDB1 gene knocked-out flies also died. D-DDB1 therefore appears to be an essential development-associated factor in a multicellular organism.

  16. Relative binding affinity of carboxylate-, phosphonate-, and bisphosphonate-functionalized gold nanoparticles targeted to damaged bone tissue

    NASA Astrophysics Data System (ADS)

    Ross, Ryan D.; Cole, Lisa E.; Roeder, Ryan K.

    2012-10-01

    Functionalized Au NPs have received considerable recent interest for targeting and labeling cells and tissues. Damaged bone tissue can be targeted by functionalizing Au NPs with molecules exhibiting affinity for calcium. Therefore, the relative binding affinity of Au NPs surface functionalized with either carboxylate ( l-glutamic acid), phosphonate (2-aminoethylphosphonic acid), or bisphosphonate (alendronate) was investigated for targeted labeling of damaged bone tissue in vitro. Targeted labeling of damaged bone tissue was qualitatively verified by visual observation and backscattered electron microscopy, and quantitatively measured by the surface density of Au NPs using field-emission scanning electron microscopy. The surface density of functionalized Au NPs was significantly greater within damaged tissue compared to undamaged tissue for each functional group. Bisphosphonate-functionalized Au NPs exhibited a greater surface density labeling damaged tissue compared to glutamic acid- and phosphonic acid-functionalized Au NPs, which was consistent with the results of previous work comparing the binding affinity of the same functionalized Au NPs to synthetic hydroxyapatite crystals. Targeted labeling was enabled not only by the functional groups but also by the colloidal stability in solution. Functionalized Au NPs were stabilized by the presence of the functional groups, and were shown to remain well dispersed in ionic (phosphate buffered saline) and serum (fetal bovine serum) solutions for up to 1 week. Therefore, the results of this study suggest that bisphosphonate-functionalized Au NPs have potential for targeted delivery to damaged bone tissue in vitro and provide motivation for in vivo investigation.

  17. Interaction of 6 Mercaptopurine with Calf Thymus DNA – Deciphering the Binding Mode and Photoinduced DNA Damage

    PubMed Central

    Rehman, Sayeed Ur; Yaseen, Zahid; Husain, Mohammed Amir; Sarwar, Tarique; Ishqi, Hassan Mubarak; Tabish, Mohammad

    2014-01-01

    DNA is one of the major intracellular targets for a wide range of anticancer and antibiotic drugs. Elucidating the binding between small molecules and DNA provides great help in understanding drug-DNA interactions and in designing of new and promising drugs for clinical use. The ability of small molecules to bind and interfere with DNA replication and transcription provides further insight into how the drugs control the expression of genes. Interaction of an antimetabolite anticancer drug 6mercaptopurine (6MP) with calf thymus DNA was studied using various approaches like UV-visible spectroscopy, fluorescence spectroscopy, CD, viscosity and molecular docking. UV-visible spectroscopy confirmed 6MP-DNA interaction. Steady state fluorescence experiments revealed a moderate binding constant of 7.48×103 M−1 which was consistent with an external binding mode. Competitive displacement assays further confirmed a non-intercalative binding mode of 6MP which was further confirmed by CD and viscosity experiments. Molecular docking further revealed the minimum energy conformation (−119.67 kJ/mole) of the complex formed between DNA and 6MP. Hence, the biophysical techniques and in-silico molecular docking approaches confirmed the groove binding/electrostatic mode of interaction between 6MP and DNA. Further, photo induced generation of ROS by 6MP was studied spectrophotometrically and DNA damage was assessed by plasmid nicking and comet assay. There was a significant increase in ROS generation and consequent DNA damage in the presence of light. PMID:24718609

  18. PARP activation regulates the RNA-binding protein NONO in the DNA damage response to DNA double-strand breaks.

    PubMed

    Krietsch, Jana; Caron, Marie-Christine; Gagné, Jean-Philippe; Ethier, Chantal; Vignard, Julien; Vincent, Michel; Rouleau, Michèle; Hendzel, Michael J; Poirier, Guy G; Masson, Jean-Yves

    2012-11-01

    After the generation of DNA double-strand breaks (DSBs), poly(ADP-ribose) polymerase-1 (PARP-1) is one of the first proteins to be recruited and activated through its binding to the free DNA ends. Upon activation, PARP-1 uses NAD+ to generate large amounts of poly(ADP-ribose) (PAR), which facilitates the recruitment of DNA repair factors. Here, we identify the RNA-binding protein NONO, a partner protein of SFPQ, as a novel PAR-binding protein. The protein motif being primarily responsible for PAR-binding is the RNA recognition motif 1 (RRM1), which is also crucial for RNA-binding, highlighting a competition between RNA and PAR as they share the same binding site. Strikingly, the in vivo recruitment of NONO to DNA damage sites completely depends on PAR, generated by activated PARP-1. Furthermore, we show that upon PAR-dependent recruitment, NONO stimulates nonhomologous end joining (NHEJ) and represses homologous recombination (HR) in vivo. Our results therefore place NONO after PARP activation in the context of DNA DSB repair pathway decision. Understanding the mechanism of action of proteins that act in the same pathway as PARP-1 is crucial to shed more light onto the effect of interference on PAR-mediated pathways with PARP inhibitors, which have already reached phase III clinical trials but are until date poorly understood.

  19. Induction of a novel damage-specific DNA binding protein correlates with enhanced DNA repair in primate cells

    SciTech Connect

    Protic, M.; Hirschfeld, S.; Tsang, A.P.; Wagner, M.; Dixon, K.; Levine, A.S. )

    1989-10-01

    Pretreatment of mammalian cell with DNA-damaging agents, such as UV light or mitomycin C, but not the tumor promoter 12-O-tetradecanoyl-phorbol-13-acetate (TPA), results in the enhanced repair of subsequently transfected UV-damaged expression vectors. To determine the cellular factors that are responsible for this enhancement, the authors have used a modified gel retardation assay to detect the proteins that interact with damaged DNA. They have identified a constitutive DNA binding protein in extracts from primate cells that has a high affinity for UV-irradiated double-stranded DNA. Cells pretreated with UV light, mitomycin C, or aphidicolin, but not TPA or serum starvation, have higher levels of this damage-specific DNA binding (DDB) protein. These results suggest that the signal for induction of DDB protein can either be damage to the DNA or interference with cellular DNA replication. The induction of DDB protein varies among primate cells with different phenotypes: (1) virus-transformed repair-proficient cells have partially or fully lost the ability to induce DDB protein above constitutive levels; (2) primary cells from repair-deficient xeroderma pigmentosum (XP) group C, and transformed XP groups A and D, show constitutive DDB protein, but do not show induced levels of this protein 48 h after UV; and (3) primary and transformed repair-deficient cells from one XP E patient are lacking both the constitutive and the induced DDB activity. The correlation between the induction of the DDB protein and the enhanced repair of UV-damaged expression vectors implies the involvement of the DDB protein in this inducible cellular response.

  20. Identification of the DNA damage-responsive element of RNR2 and evidence that four distinct cellular factors bind it.

    PubMed Central

    Elledge, S J; Davis, R W

    1989-01-01

    The RNR2 gene encodes the small subunit of ribonucleotide reductase, the enzyme that catalyzes the first step in the pathway for the production of the deoxyribonucleotides needed for DNA synthesis. Transcription of this gene is induced approximately 20-fold in response to environmental stimuli that damage DNA or block DNA replication. Deletion and subcloning analysis identified two, and possibly three, upstream activating sequences (UAS) and one repressing (URS) element in the RNR2 regulatory region. A 42-base-pair (bp) fragment from this region was found to be necessary for proper regulation of RNR2 and to be capable of conferring DNA damage inducibility upon a heterologous promoter. This fragment contained both positively and negatively acting sequences. Four DNA-binding factors interacted with the RNR2 regulatory region. One factor was identified as the GRF1 protein, the product of the RAP1 gene. GRF1 bound to the UAS2 element of RNR2, which was found to be directly adjacent to the 42-bp fragment. UAS2 activity was repressed by the 42-bp fragment. Three other factors bound to the 42-bp fragment; one of these factors, RRF3, had a second binding site in the RNR2 promoter. These factors are likely to mediate the response of RNR2 to DNA damage. Images PMID:2685561

  1. A carrier protein strategy yields the structure of dalbavancin

    PubMed Central

    Economou, Nicoleta J.; Nahoum, Virginie; Weeks, Stephen D.; Grasty, Kimberly C.; Zentner, Isaac J.; Townsend, Tracy M.; Bhuiya, Mohammad W.; Cocklin, Simon; Loll, Patrick J.

    2012-01-01

    Many large natural product antibiotics act by specifically binding and sequestering target molecules found on bacterial cells. We have developed a new strategy to expedite the structural analysis of such antibiotic-target complexes, in which we covalently link the target molecules to carrier proteins, and then crystallize the entire carrier/target/antibiotic complex. Using native chemical ligation, we have linked the Lys-d-Ala-d-Ala binding epitope for glycopeptide antibiotics to three different carrier proteins. We show that recognition of this peptide by multiple antibiotics is not compromised by the presence of the carrier protein partner, and use this approach to determine the first-ever crystal structure for the new therapeutic dalbavancin. We also report the first crystal structure of an asymmetric ristocetin antibiotic dimer, as well as the structure of vancomycin bound to a carrier-target fusion. The dalbavancin structure reveals an antibiotic molecule that has closed around its binding partner; it also suggests mechanisms by which the drug can enhance its half-life by binding to serum proteins, and be targeted to bacterial membranes. Notably, the carrier protein approach is not limited to peptide ligands such as Lys-d-Ala-d-Ala, but is applicable to a diverse range of targets. This strategy is likely to yield structural insights that accelerate new therapeutic development. PMID:22352468

  2. Thermodynamics of Damaged DNA Binding and Catalysis by Human AP Endonuclease 1.

    PubMed

    Miroshnikova, A D; Kuznetsova, A A; Kuznetsov, N A; Fedorova, O S

    2016-01-01

    Apurinic/apyrimidinic (AP) endonucleases play an important role in DNA repair and initiation of AP site elimination. One of the most topical problems in the field of DNA repair is to understand the mechanism of the enzymatic process involving the human enzyme APE1 that provides recognition of AP sites and efficient cleavage of the 5'-phosphodiester bond. In this study, a thermodynamic analysis of the interaction between APE1 and a DNA substrate containing a stable AP site analog lacking the C1' hydroxyl group (F site) was performed. Based on stopped-flow kinetic data at different temperatures, the steps of DNA binding, catalysis, and DNA product release were characterized. The changes in the standard Gibbs energy, enthalpy, and entropy of sequential specific steps of the repair process were determined. The thermodynamic analysis of the data suggests that the initial step of the DNA substrate binding includes formation of non-specific contacts between the enzyme binding surface and DNA, as well as insertion of the amino acid residues Arg177 and Met270 into the duplex, which results in the removal of "crystalline" water molecules from DNA grooves. The second binding step involves the F site flipping-out process and formation of specific contacts between the enzyme active site and the everted 5'-phosphate-2'-deoxyribose residue. It was shown that non-specific interactions between the binding surfaces of the enzyme and DNA provide the main contribution into the thermodynamic parameters of the DNA product release step.

  3. Repair of Nerve Cell Membrane Damage by Calcium-Dependent, Membrane-Binding Proteins (Revised)

    DTIC Science & Technology

    2012-09-01

    family exhibit a “ bivalent ” activity resulting in the aggregation of membranes coincident with the binding of the annexin to the membrane [7,8]. Such...10. [12] D.J. Selkoe, Alzheimer’s disease: genes , proteins, and therapy, Physiol Rev 81 (2001) 741-766. [13] A. Demuro, I. Parker, G.E. Stutzmann

  4. Thermodynamics of Damaged DNA Binding and Catalysis by Human AP Endonuclease 1

    PubMed Central

    Miroshnikova, A. D.; Kuznetsova, A. A.; Kuznetsov, N. A.; Fedorova, O. S.

    2016-01-01

    Apurinic/apyrimidinic (AP) endonucleases play an important role in DNA repair and initiation of AP site elimination. One of the most topical problems in the field of DNA repair is to understand the mechanism of the enzymatic process involving the human enzyme APE1 that provides recognition of AP sites and efficient cleavage of the 5’-phosphodiester bond. In this study, a thermodynamic analysis of the interaction between APE1 and a DNA substrate containing a stable AP site analog lacking the C1’ hydroxyl group (F site) was performed. Based on stopped-flow kinetic data at different temperatures, the steps of DNA binding, catalysis, and DNA product release were characterized. The changes in the standard Gibbs energy, enthalpy, and entropy of sequential specific steps of the repair process were determined. The thermodynamic analysis of the data suggests that the initial step of the DNA substrate binding includes formation of non-specific contacts between the enzyme binding surface and DNA, as well as insertion of the amino acid residues Arg177 and Met270 into the duplex, which results in the removal of “crystalline” water molecules from DNA grooves. The second binding step involves the F site flipping-out process and formation of specific contacts between the enzyme active site and the everted 5’-phosphate-2’-deoxyribose residue. It was shown that non-specific interactions between the binding surfaces of the enzyme and DNA provide the main contribution into the thermodynamic parameters of the DNA product release step. PMID:27099790

  5. Correction of the DNA repair defect in xeroderma pigmentosum group E by injection of a DNA damage-binding protein

    SciTech Connect

    Keeney, S.; Brody, T.; Linn, S.; Eker, A.P.M.; Vermeulen, W.; Bootsma, D.; Hoeijmakers, J.H.J.

    1994-04-26

    Cells from a subset of patients with the DNA-repair-defective disease xeroderma pigmentosum complementation group E (XP-E) are known to lack a DNA damage-binding (DDB) activity. Purified human DDB protein was injected into XP-E cells to test whether the DNA-repair defect in these cells is caused by a defect in DDB activity. Injected DDB protein stimulated DNA repair to normal levels in those strains that lack the DDB activity but did not stimulate repair in cells from other xeroderma pigmentosum groups or in XP-E cells that contain the activity. These results provide direct evidence that defective DDB activity causes the repair defect in a subset of XP-E patients, which in turn establishes a role for this activity in nucleotide-excision repair in vivo.

  6. Plant flavone apigenin binds to nucleic acid bases and reduces oxidative DNA damage in prostate epithelial cells.

    PubMed

    Sharma, Haripaul; Kanwal, Rajnee; Bhaskaran, Natarajan; Gupta, Sanjay

    2014-01-01

    Oxidative stress has been linked to prostate carcinogenesis as human prostate tissue is vulnerable to oxidative DNA damage. Apigenin, a dietary plant flavone, possesses anti-proliferative and anticancer effects; however, its antioxidant properties have not been fully elucidated. We investigated sub-cellular distribution of apigenin, it's binding to DNA and protective effects against H2O2-induced DNA damage using transformed human prostate epithelial RWPE-1 cells and prostate cancer LNCaP, PC-3 and DU145 cells. Exposure of cells to apigenin exhibited higher accumulation in RWPE-1 and LNCaP cells, compared to PC-3 and DU145 cells. The kinetics of apigenin uptake in LNCaP cells was estimated with a Km value of 5 µmole/L and Vmax of 190 pmoles/million cells/h. Sub-cellular fractionation demonstrated that nuclear matrix retains the highest concentration of apigenin (45.3%), followed by cytosol (23.9%), nuclear membranes (17.9%) and microsomes (12.9%), respectively. Spectroscopic analysis of apigenin with calf-thymus DNA exhibited intercalation as the dominant binding mode to DNA duplex. Apigenin exposure resulted in significant genoprotective effects in H2O2-stressed RWPE-1 cells by reduction in reactive oxygen species levels. In addition, apigenin exposure suppressed the formation of 8-hydroxy-2' deoxyguanosine and protected exposed cells from apoptosis. Our studies demonstrate that apigenin is readily taken up by normal prostatic epithelial cells and prostate cancer cells, and is incorporated into their nuclei, where its intercalation with nucleic acid bases may account for its antioxidant and chemopreventive activities.

  7. Damage-specific DNA binding protein 1 (DDB1): a protein with a wide range of functions.

    PubMed

    Iovine, Barbara; Iannella, Maria Luigia; Bevilacqua, Maria Assunta

    2011-12-01

    Damage-specific DNA binding protein 1 (DDB1) is a multifunctional protein that was first isolated as a subunit of a heterodimeric complex that recognises the UV-induced DNA lesions in the nucleotide excision repair pathway. DDB1 and DDB2 form a complex that promotes the global genome repair (GG-NER), whereas DDB1 and Cockayne syndrome group A protein (CSA) form a complex that contributes to the transcription-coupled repair (TC-NER) pathway. DDB1 is also a component of an ubiquitin-E3 ligase complex and functions as substrate or adapter protein between Cullin 4A (Cul4A) and CUL4-associated factors (DCAFs) to target substrates for ubiquitination. CUL4-DDB1 E3-ligase complex regulates the selective proteolysis of key proteins in DNA repair, replication and transcription. In addition, DDB1 plays a role in transcriptional regulation of UV-induced genes. It is conceivable that DDB1 acts as a sensor of damage to maintain the balance between genome integrity and cell cycle progression. However, the temporal order between these two events remains to be established.

  8. Acrylamide binding to the DNA and protamine of spermiogenic stages in the mouse and its relationship to genetic damage.

    PubMed

    Sega, G A; Alcota, R P; Tancongco, C P; Brimer, P A

    1989-08-01

    Mice received an intraperitoneal injection of 14C-labeled acrylamide (AA) at an exposure of 125 mg/kg to equal that used in genetic studies carried out by Shelby et al. (1986). Subsequently, spermatozoa were recovered from the reproductive tracts of the animals over a 3-week period and assayed for the amount of bound AA. A strong increase in the level of binding occurred in late-spermatid to early-spermatozoa stages; these same stages are also genetically most sensitive to the action of AA. At all time points, alkylation of DNA within the sperm accounted for a very small fraction (generally less than 0.5%) of the total sperm-head alkylation. However, alkylation of protamine, a protein unique to sperm cells, was found to be correlated with total sperm-head alkylation and accounted for essentially all of the AA binding. Two radioactive adducts were found in hydrolysed protamine samples, one of which co-eluted with a standard of S-carboxyethylcysteine. Protamine alkylation appears to be a significant cause of acrylamide-induced genetic damage in spermiogenic cells of the mouse.

  9. Overexpression of UV-DAMAGED DNA BINDING PROTEIN 1 links plant development and phytonutrient accumulation in high pigment-1 tomato.

    PubMed

    Azari, Raviv; Reuveni, Moshe; Evenor, Dalia; Nahon, Sahadia; Shlomo, Haviva; Chen, Lea; Levin, Ilan

    2010-08-01

    Fruits of tomato plants carrying the high pigment-1 mutations hp-1 and hp-1(w) are characterized by an increased number of plastids coupled with enhanced levels of functional metabolites. Unfortunately, hp-1 mutant plants are also typified by light-dependent retardation in seedling and whole-plant growth and development, which limits their cultivation. These mutations were mapped to the gene encoding UV-DAMAGED DNA BINDING PROTEIN 1 (DDB1) and, recently, fruit-specific RNA interference studies have demonstrated an increased number of plastids and enhanced carotenoid accumulation in the transgenic tomato fruits. However, whole-plant overexpression of DDB1, required to substantiate its effects on seedling and plant development and to couple them with fruit phenotypes, has heretofore been unsuccessful. In this study, five transgenic lines constitutively overexpressing normal DDB1 in hp-1 mutant plants were analysed. Eleven-day-old seedlings, representing these lines, displayed up to approximately 73- and approximately 221-fold overexpression of the gene in hypocotyls and cotyledons, respectively. This overexpression resulted in statistically significant reversion to the non-mutant developmental phenotypes, including more than a full quantitative reversion. This reversion of phenotypes was generally accompanied by correlated responses in chlorophyll accumulation and altered expression of selected light signalling genes: PHYTOCHROME A, CRYPTOCHROME 1, ELONGATED HYPOCOTYL 5, and the gene encoding CHLOROPHYLL A/B-BINDING PROTEIN 4. Cumulatively, these results provide the missing link between DDB1 and its effects on tomato plant development.

  10. The single-strand DNA binding activity of human PC4 preventsmutagenesis and killing by oxidative DNA damage

    SciTech Connect

    Wang, Jen-Yeu; Sarker, Altaf Hossain; Cooper, Priscilla K.; Volkert, Michael R.

    2004-02-01

    Human positive cofactor 4 (PC4) is a transcriptional coactivator with a highly conserved single-strand DNA (ssDNA) binding domain of unknown function. We identified PC4 as a suppressor of the oxidative mutator phenotype of the Escherichia coli fpg mutY mutant and demonstrate that this suppression requires its ssDNA binding activity. Yeast mutants lacking their PC4 ortholog Sub1 are sensitive to hydrogen peroxide and exhibit spontaneous and peroxide induced hypermutability. PC4 expression suppresses the peroxide sensitivity of the yeast sub l{Delta} mutant, suggesting that the human protein has a similar function. A role for yeast and human proteins in DNA repair is suggested by the demonstration that Sub1 acts in a peroxide-resistance pathway involving Rad2 and by the physical interaction of PC4 with the human Rad2 homolog XPG. We show XPG recruits PC4 to a bubble-containing DNA substrate with resulting displacement of XPG and formation of a PC4-DNA complex. We discuss the possible requirement for PC4 in either global or transcription-coupled repair of oxidative DNA damage to mediate the release of XPG bound to its substrate.

  11. Differences of calcium binding proteins immunoreactivities in the young hippocampal CA1 region from the adult following transient ischemic damage.

    PubMed

    Lee, Young Joo; Yan, Bing Chun; Park, Joon Ha; Ahn, Ji Hyeon; Kim, In Hye; Lee, Jae-Chul; Lee, Hui Young; Kim, Young-Myeong; Won, Moo-Ho; Cho, Jun Hwi

    2013-03-15

    It has been reported that the young were much more resistant to transient cerebral ischemia than in the adult. In the present study, we examined that about 90% of CA1 pyramidal cells in the adult gerbil hippocampus died at 4days after ischemia-reperfusion; however, in the young hippocampus, about 56% of them died at 7days after ischemia-reperfusion. We compared immunoreactivities and levels of calcium binding proteins (CBPs), such as calbindin 28k (CB-D28k), calretinin (CR) and parvalbumin (PV). The immunoreactivities and protein levels of all the CBPs in the young sham were higher than those in the adult sham. In the adult, the immunoreactivities and protein levels of all the CBPs were markedly decreased at 4days after ischemia-reperfusion, however, in the young, they were apparently maintained. At 7days after ischemia-reperfusion, they were decreased in the young, however, they were much higher than those in the adult. In brief, the immunoreactivities and levels of CBPs were not decreased in the ischemic CA1 region of the young 4days after transient cerebral ischemia. This finding indicates that the longer maintenance of CBPs may contribute to a less and more delayed neuronal death/damage in the young.

  12. Protection from Inflammatory Organ Damage in a Murine Model of Hemophagocytic Lymphohistiocytosis Using Treatment with IL-18 Binding Protein

    PubMed Central

    Chiossone, Laura; Audonnet, Sandra; Chetaille, Bruno; Chasson, Lionel; Farnarier, Catherine; Berda-Haddad, Yael; Jordan, Stefan; Koszinowski, Ulrich H.; Dalod, Marc; Mazodier, Karin; Novick, Daniela; Dinarello, Charles A.; Vivier, Eric; Kaplanski, Gilles

    2012-01-01

    Hemophagocytic lymphohistiocytosis (HLH) is a life-threatening condition due to the association of an infectious agent with lymphocyte cytotoxicity defects, either of congenital genetic origin in children or presumably acquired in adults. In HLH patients, an excess of lymphocyte or macrophage cytokines, such as IFN-γ and TNFα is present in serum. In animal models of the disease, IFN-γ and TNF-α have been shown to play a central pathogenic role. In humans, unusually high concentrations of IL-18, an inducer of IFN-γ, and TNF-α have been reported, and are associated with an imbalance between IL-18 and its natural inhibitor IL-18 binding protein (IL-18BP) resulting in an excess of free IL-18. Here we studied whether IL-18BP could reduce disease severity in an animal model of HLH. Mouse cytomegalovirus infection in perforin-1 knock-out mice induced a lethal condition similar to human HLH characterized by cytopenia with marked inflammatory lesions in the liver and spleen as well as the presence of hemophagocytosis in bone marrow. IL-18BP treatment decreased hemophagocytosis and reversed liver as well as spleen damage. IL-18BP treatment also reduced both IFN-γ and TNF-α production by CD8+ T and NK cells, as well as Fas ligand expression on NK cell surface. These data suggest that IL-18BP is beneficial in an animal model of HLH and in combination with anti-infectious therapy may be a promising strategy to treat HLH patients. PMID:22891066

  13. The fluorescence properties and binding mechanism of SYTOX green, a bright, low photo-damage DNA intercalating agent.

    PubMed

    Thakur, Shreyasi; Cattoni, Diego I; Nöllmann, Marcelo

    2015-07-01

    DNA intercalators are widely used in cancer therapeutics, to probe protein-DNA interactions and to investigate the statistical-mechanical properties of DNA. Here, we employ single-molecule fluorescence microscopy, magnetic tweezers, and ensemble-binding assays to investigate the fluorescence properties and binding mechanism of SYTOX green, a DNA labeling dye previously used for staining dead cells and becoming of common use for single-molecule methodologies. Specifically, we show that SYTOX green presents several advantages with respect to other dyes: (1) binds DNA rapidly and with high affinity; (2) has a good signal-to-noise ratio even at low concentrations; (3) exhibits a low photobleaching rate; and (4) induces lower light-induced DNA degradation. Finally, we show that SYTOX green is a DNA intercalator that binds DNA cooperatively with a binding site of 3.5 bp, increasing the DNA length upon binding by 43%, while not affecting its mechanical properties.

  14. Enzymatic characterization and crystal structure analysis of the D-alanine-D-alanine ligase from Helicobacter pylori.

    PubMed

    Wu, Dalei; Zhang, Liang; Kong, Yunhua; Du, Jiamu; Chen, Shuai; Chen, Jing; Ding, Jianping; Jiang, Hualiang; Shen, Xu

    2008-09-01

    D-Alanine-D-alanine ligase is the second enzyme in the D-Ala branch of bacterial cell wall peptidoglycan assembly, and recognized as an attractive antimicrobial target. In this work, the D-Ala-D-Ala ligase of Helicobacter pylori strain SS1 (HpDdl) was kinetically and structurally characterized. The determined apparent K(m) of ATP (0.87 microM), the K(m1) (1.89 mM) and K(m2) of D-Ala (627 mM), and the k(cat) (115 min(-1)) at pH 8.0 indicated its relatively weak binding affinity and poor catalytic activity against the substrate D-Ala in vitro. However, by complementary assay of expressing HpDdl in Escherichia coli Delta ddl mutant, HpDdl was confirmed to be capable of D-Ala-D-Ala ligating in vivo. Through sequence alignment with other members of the D-Ala-D-X ligase superfamily, HpDdl keeps two conservatively substituted residues (Ile16 and Leu241) and two nonconserved residues (Leu308 and Tyr311) broadly located in the active region of the enzyme. Kinetic analyses against the corresponding HpDdl mutants (I16V, L241Y, L241F, L308T, and Y311S) suggested that these residues, especially Leu308 and Tyr311, might partly contribute to the unique catalytic properties of the enzyme. This was fairly proved by the crystal structure of HpDdl, which revealed that there is a 3(10)-helix (including residues from Gly306 to Leu312) near the D-Ala binding region in the C-terminal domain, where HpDdl has two sequence deletions compared with other homologs. Such 3(10)-helix may participate in D-Ala binding and conformational change of the enzyme. Our present work hopefully provides useful information for understanding the D-Ala-D-Ala ligase of Helicobacter pylori.

  15. Small RNA-mediated repair of UV-induced DNA lesions by the DNA DAMAGE-BINDING PROTEIN 2 and ARGONAUTE 1.

    PubMed

    Schalk, Catherine; Cognat, Valérie; Graindorge, Stéfanie; Vincent, Timothée; Voinnet, Olivier; Molinier, Jean

    2017-04-04

    As photosynthetic organisms, plants need to prevent irreversible UV-induced DNA lesions. Through an unbiased, genome-wide approach, we have uncovered a previously unrecognized interplay between Global Genome Repair and small interfering RNAs (siRNAs) in the recognition of DNA photoproducts, prevalently in intergenic regions. Genetic and biochemical approaches indicate that, upon UV irradiation, the DNA DAMAGE-BINDING PROTEIN 2 (DDB2) and ARGONAUTE 1 (AGO1) of Arabidopsis thaliana form a chromatin-bound complex together with 21-nt siRNAs, which likely facilitates recognition of DNA damages in an RNA/DNA complementary strand-specific manner. The biogenesis of photoproduct-associated siRNAs involves the noncanonical, concerted action of RNA POLYMERASE IV, RNA-DEPENDENT RNA POLYMERASE-2, and DICER-LIKE-4. Furthermore, the chromatin association/dissociation of the DDB2-AGO1 complex is under the control of siRNA abundance and DNA damage signaling. These findings reveal unexpected nuclear functions for DCL4 and AGO1, and shed light on the interplay between small RNAs and DNA repair recognition factors at damaged sites.

  16. Cdk5 activator-binding protein C53 regulates apoptosis induced by genotoxic stress via modulating the G2/M DNA damage checkpoint.

    PubMed

    Jiang, Hai; Luo, Shouqing; Li, Honglin

    2005-05-27

    In response to DNA damage, the cellular decision of life versus death involves an intricate network of multiple factors that play critical roles in regulation of DNA repair, cell cycle, and cell death. DNA damage checkpoint proteins are crucial for maintaining DNA integrity and normal cellular functions, but they may also reduce the effectiveness of cancer treatment. Here we report the involvement of Cdk5 activator p35-binding protein C53 in regulation of apoptosis induced by genotoxic stress through modulating Cdk1-cyclin B1 function. C53 was originally identified as a Cdk5 activator p35-binding protein and a caspase substrate. Importantly, our results demonstrated that C53 deficiency conferred partial resistance to genotoxic agents such as etoposide and x-ray irradiation, whereas ectopic expression of C53 rendered cells susceptible to multiple genotoxins that usually trigger G(2)/M arrest. Furthermore, we found that Cdk1 activity was required for etoposide-induced apoptosis of HeLa cells. Overexpression of C53 promoted Cdk1 activity and nuclear accumulation of cyclin B1, whereas C53 deficiency led to more cytoplasmic retention of cyclin B1, suggesting that C53 acts as a pivotal player in modulating the G(2)/M DNA damage checkpoint. Finally, C53 and cyclin B1 co-localize and associate in vivo, indicating a direct role of C53 in regulating the Cdk1-cyclin B1 complex. Taken together, our results strongly indicate that in response to genotoxic stress, C53 serves as an important regulatory component of the G(2)/M DNA damage checkpoint. By overriding the G(2)/M checkpoint-mediated inhibition of Cdk1-cyclin B1 function, ectopic expression of C53 may represent a novel approach for chemo- and radio-sensitization of cancer cells.

  17. Fatty Acid Saturation of Albumin Used in Resuscitation Fluids Modulates Cell Damage in Shock: In Vitro Results Using a Novel Technique to Measure Fatty Acid Binding Capacity.

    PubMed

    Penn, Alexander H; Dubick, Michael A; Torres Filho, Ivo P

    2017-03-21

    The use of albumin for resuscitation has not proven as beneficial in human trials as expected from numerous animal studies. One explanation could be the practice of adding fatty acid (FA) during manufacture of pharmaceutical albumin. During ischemia, unbound free FAs (FFA) in the circulation could potentially induce cellular damage. We hypothesized that albumins with higher available binding capacities (ABC) for FFAs may prevent that damage. Therefore, we developed a technique to measure ABC, determined if pharmaceutical human serum albumin (HSA) has decreased ABC compared to FA-free bovine serum albumin (BSA), and if binding capacity would affect hemolysis when blood is mixed with exogenous FFA at levels similar to those observed in shock. The new assay used exogenous oleic acid (OA), glass fiber filtration, and a FFA assay kit. RBC hemolysis was determined by mixing 0-5 mM OA with PBS, HSA, FA-free BSA, or FA-saturated BSA and measuring plasma hemoglobin after incubation with human blood. 5% HSA contained 4.7±0.2 mM FFA, leaving an ABC of 5.0 ± 0.6 mM, compared to FA-free BSA's ABC of 7.0 ± 1.3 mM (P < 0.024). Hemolysis after OA was reduced with FA-free BSA but increased with FA-saturated BSA. HSA provided intermediate results. 25% solutions of FA-free BSA and HSA were more protective, while 25% FA-saturated BSA was more damaging than 5% solutions. These findings suggest that increased FA saturation may reverse albumin's potential benefit to lessen cellular damage and may explain, at least in part, its failure in human trauma studies.

  18. DNA-damage-inducible 1 protein (Ddi1) contains an uncharacteristic ubiquitin-like domain that binds ubiquitin

    PubMed Central

    Nowicka, Urszula; Zhang, Daoning; Walker, Olivier; Krutauz, Daria; Castañeda, Carlos A.; Chaturvedi, Apurva; Chen, Tony Y.; Reis, Noa; Glickman, Michael H.; Fushman, David

    2015-01-01

    SUMMARY Ddi1 belongs to a family of shuttle proteins targeting polyubiquitinated substrates for proteasomal degradation. Unlike the other proteasomal shuttles, Rad23 and Dsk2, Ddi1 remains an enigma: its function is not fully understood and structural properties are poorly characterized. We determined the structure and binding properties of the ubiquitin-like (UBL) and ubiquitin-associated (UBA) domains of Ddi1 from Saccharomyces cerevisiae. We found that, while Ddi1UBA forms a characteristic UBA:ubiquitin complex, Ddi1UBL has entirely uncharacteristic binding preferences. Despite having a ubiquitin-like fold, Ddi1UBL does not interact with typical UBL-receptors but, unexpectedly, binds ubiquitin, forming a unique interface mediated by hydrophobic contacts and by salt-bridges between oppositely-charged residues of Ddi1UBL and ubiquitin. In stark contrast with ubiquitin and other UBLs, the β-sheet surface of Ddi1UBL is negatively charged and, therefore, is recognized in a completely different way. The dual functionality of Ddi1UBL, capable of binding both ubiquitin and proteasome, suggests a novel mechanism for Ddi1 as a proteasomal shuttle. PMID:25703377

  19. Site-specific copper-catalyzed oxidation of α-synuclein: tightening the link between metal binding and protein oxidative damage in Parkinson's disease.

    PubMed

    Miotto, Marco C; Rodriguez, Esaú E; Valiente-Gabioud, Ariel A; Torres-Monserrat, Valentina; Binolfi, Andrés; Quintanar, Liliana; Zweckstetter, Markus; Griesinger, Christian; Fernández, Claudio O

    2014-05-05

    Amyloid aggregation of α-synuclein (AS) has been linked to the pathological effects associated with Parkinson's disease (PD). Cu(II) binds specifically at the N-terminus of AS and triggers its aggregation. Site-specific Cu(I)-catalyzed oxidation of AS has been proposed as a plausible mechanism for metal-enhanced AS amyloid formation. In this study, Cu(I) binding to AS was probed by NMR spectroscopy, in combination with synthetic peptide models, site-directed mutagenesis, and C-terminal-truncated protein variants. Our results demonstrate that both Met residues in the motif (1)MDVFM(5) constitute key structural determinants for the high-affinity binding of Cu(I) to the N-terminal region of AS. The replacement of one Met residue by Ile causes a dramatic decrease in the binding affinity for Cu(I), whereas the removal of both Met residues results in a complete lack of binding. Moreover, these Met residues can be oxidized rapidly after air exposure of the AS-Cu(I) complex, whereas Met-116 and Met-127 in the C-terminal region remain unaffected. Met-1 displays higher susceptibility to oxidative damage compared to Met-5 because it is directly involved in both Cu(II) and Cu(I) coordination, resulting in closer exposure to the reactive oxygen species that may be generated by the redox cycling of copper. Our findings support a mechanism where the interaction of AS with copper ions leads to site-specific metal-catalyzed oxidation in the protein under physiologically relevant conditions. In light of recent biological findings, these results support a role for AS-copper interactions in neurodegeneration in PD.

  20. The DNA damage-binding protein XPC is a frequent target for inactivation in squamous cell carcinomas.

    PubMed

    de Feraudy, Sebastien; Ridd, Katie; Richards, Lauren M; Kwok, Pui-Yan; Revet, Ingrid; Oh, Dennis; Feeney, Luzviminda; Cleaver, James E

    2010-08-01

    XPC, the main damage-recognition protein responsible for nucleotide excision repair of UVB damage to DNA, is lost or mutated in xeroderma pigmentosum group C (XP-C), a rare inherited disease characterized by high incidence and early onset of non-melanoma and melanoma skin cancers. The high incidence of skin cancers in XP-C patients suggests that loss of expression of XPC protein might also provide a selective advantage for initiation and progression of similar cancers in non XP-C patients in the general population. To test whether XPC is selectively lost in squamous cell carcinomas from non XP-C patients, we examined XPC expression by immunohistochemistry on a tissue microarray with 244 tissue cores, including in situ and invasive squamous-cell carcinomas (SCCs), keratoacanthoma (KA), and normal skin samples from both immunocompetent and immunosuppressed patients. We found that XPC expression was lost in 49% of invasive squamous cell carcinomas from immunocompetent patients and 59% from immunosuppressed patients. Loss of expression was correlated with deletions of chromosomal 3p and mutations in the XPC gene. The XPC gene is consequently inactivated or lost in almost half of squamous cell carcinomas from non XP-C patients. Loss or mutation of XPC may be an early event during skin carcinogenesis that provides a selective advantage for initiation and progression of squamous cell carcinomas in non XP-C patients.

  1. Damage-specific DNA binding protein 1 (DDB1) is involved in ubiquitin-mediated proteolysis of p27Kip1 in response to UV irradiation.

    PubMed

    Iovine, Barbara; Iannella, Maria Luigia; Bevilacqua, Maria Assunta

    2011-05-01

    Damage-specific DNA binding protein 1 (DDB1) is a conserved protein component of the damaged DNA binding protein complex (DDB) that recognizes UV-induced DNA lesions and initiates the nucleotide excision repair process. DDB1 is also part of an E3 ubiquitin-ligase complex that targets a variety of substrates for proteolysis including the cyclin-dependent kinase inhibitor p27(Kip1). The mechanism regulating the trafficking of DDB1 and its relationship with UV irradiation is not known, although cell cycle progression is implicated in the molecular machinery driving DDB1 into the nucleus. We evaluated the involvement of DDB1 in ubiquitination of the cdk inhibitor p27(Kip1) in response to UV irradiation. First, we observed that low and high doses of UV irradiation exert different effects on p27(Kip1) protein levels. Indeed, low but not high UV doses induced p27(Kip1) protein proteolysis in several human cell lines and UV-dependent degradation is dominant over other genotoxic agents such as cisplatin. We also demonstrate that p27(Kip1) reduction is not due to transcriptional regulation and that the proteasome inhibitor MG132 affects p27(Kip1) degradation. We observed that at low UV doses the decrease in p27(Kip1) nuclear protein related with DDB1 translocation into the nucleus; conversely, high doses of UV-induced p27(Kip1) accumulation and unchanged level of DDB1. The knockdown of DDB1 or Skp2 prevents UV-induced degradation of p27(Kip1) suggesting that DDB1 is essential to regulation of p27(kip1) turnover after a mild DNA damage. Our findings support the concept that DDB1 contributes to the activation of DNA repair mechanisms and could be a key factor in regulating the cell cycle in response to UV-induced DNA damage. Although the temporal order with which DDB1 contributes to ubiquitination of p27(Kip1) or initiates the nucleotide excision repair process remains to be established, our results represent a major step towards clarifying these issues.

  2. Escherichia coli phage-shock protein A (PspA) binds to membrane phospholipids and repairs proton leakage of the damaged membranes.

    PubMed

    Kobayashi, Ryuji; Suzuki, Toshiharu; Yoshida, Masasuke

    2007-10-01

    Escherichia coli phage-shock protein A (PspA), a 25.3 kDa peripheral membrane protein, is induced under the membrane stress conditions and is assumed to help maintain membrane potential. Here, we report that purified PspA, existing as a large oligomer, is really able to suppress proton leakage of the membranes. This was demonstrated for membrane vesicles prepared from the PspA-lacking E. coli mutants, and for membrane vesicles damaged by ethanol and Triton X-100 prepared from the mutant and the wild-type cells. PspA also suppressed proton leakage of damaged liposomes made from E. coli total lipids. Furthermore, we found that PspA bound preferentially to liposomes containing phosphatidylserine and phosphatidylglycerol. All these effects were not observed for monomer PspA that was prepared by refolding of urea-denatured PspA. These results indicate that oligomers of PspA bind to membrane phospholipids and suppress proton leakage.

  3. New orally active DNA minor groove binding small molecule CT-1 acts against breast cancer by targeting tumor DNA damage leading to p53-dependent apoptosis.

    PubMed

    Saini, Karan Singh; Hamidullah; Ashraf, Raghib; Mandalapu, Dhanaraju; Das, Sharmistha; Siddiqui, Mohd Quadir; Dwivedi, Sonam; Sarkar, Jayanta; Sharma, Vishnu Lal; Konwar, Rituraj

    2017-04-01

    Targeting tumor DNA damage and p53 pathway is a clinically established strategy in the development of cancer chemotherapeutics. Majority of anti-cancer drugs are delivered through parenteral route for reasons like severe toxicity, lack of stability, and poor enteral absorption. Current DNA targeting drugs in clinical like anthracycline suffers from major drawbacks like cardiotoxicity. Here, we report identification of a new orally active small molecule curcumin-triazole conjugate (CT-1) with significant anti-breast cancer activity in vitro and in vivo. CT-1 selectively and significantly inhibits viability of breast cancer cell lines; retards cells cycle progression at S phase and induce mitochondrial-mediated cell apoptosis. CT-1 selectively binds to minor groove of DNA and induces DNA damage leading to increase in p53 along with decrease in its ubiquitination. Inhibition of p53 with pharmacological inhibitor as well as siRNA revealed the necessity of p53 in CT-1-mediated anti-cancer effects in breast cancer cells. Studies using several other intact p53 and deficient p53 cancer cell lines further confirmed necessity of p53 in CT-1-mediated anti-cancer response. Pharmacological inhibition of pan-caspase showed CT-1 induces caspase-dependent cell death in breast cancer cells. Most interestingly, oral administration of CT-1 induces significant inhibition of tumor growth in LA-7 syngeneic orthotropic rat mammary tumor model. CT-1 treated mammary tumor shows enhancement in DNA damage, p53 upregulation, and apoptosis. Collectively, CT-1 exhibits potent anti-cancer effect both in vitro and in vivo and could serve as a safe orally active lead for anti-cancer drug development. © 2016 Wiley Periodicals, Inc.

  4. THAP5 is a DNA-binding transcriptional repressor that is regulated in melanoma cells during DNA damage-induced cell death

    SciTech Connect

    Balakrishnan, Meenakshi P.; Cilenti, Lucia; Ambivero, Camilla; Goto, Yamafumi; Takata, Minoru; Turkson, James; Li, Xiaoman Shawn; Zervos, Antonis S.

    2011-01-07

    Research highlights: {yields} THAP5 is a DNA-binding protein and a transcriptional repressor. {yields} THAP5 is induced in melanoma cells upon exposure to UV or treatment with cisplatin. {yields} THAP5 induction correlates with the degree of apoptosis in melanoma cell population. {yields} THAP5 is a pro-apoptotic protein involved in melanoma cell death. -- Abstract: THAP5 was originally isolated as a specific interactor and substrate of the mitochondrial pro-apoptotic Omi/HtrA2 protease. It is a human zinc finger protein characterized by a restricted pattern of expression and the lack of orthologs in mouse and rat. The biological function of THAP5 is unknown but our previous studies suggest it could regulate G2/M transition in kidney cells and could be involved in human cardiomyocyte cell death associated with coronary artery disease (CAD). In this report, we expanded our studies on the properties and function of THAP5 in human melanoma cells. THAP5 was expressed in primary human melanocytes as well as in all melanoma cell lines that were tested. THAP5 protein level was significantly induced by UV irradiation or cisplatin treatment, conditions known to cause DNA damage. The induction of THAP5 correlated with a significant increase in apoptotic cell death. In addition, we show that THAP5 is a nuclear protein that could recognize and bind a specific DNA motif. THAP5 could also repress the transcription of a reporter gene in a heterologous system. Our work suggests that THAP5 is a DNA-binding protein and a transcriptional repressor. Furthermore, THAP5 has a pro-apoptotic function and it was induced in melanoma cells under conditions that promoted cell death.

  5. Roles of the DNA binding proteins H-NS and StpA in homologous recombination and repair of bleomycin-induced damage in Escherichia coli.

    PubMed

    Shiraishi, Kouya; Ogata, Yasuyuki; Hanada, Katsuhiro; Kano, Yasunobu; Ikeda, Hideo

    2007-10-01

    The DNA binding protein H-NS promotes homologous recombination in Escherichia coli, but the role of its paralog StpA in this process remains unclear. Here we show that an hns mutant, but not an stpA mutant, are marginally defective in conjugational recombination and is sensitive to the double-strand-break-inducing agent bleomycin. Interestingly, the hns stpA double mutant is severely defective in homologous recombination and more bleomycin-sensitive than is the hns or stpA single mutant, indicating that the stpA mutation synergistically enhances the defects of homologous recombination and the increased bleomycin-sensitivity in the hns mutant. In addition, the transduction analysis in the hns stpA double mutant indicated that the stpA mutation also enhances the defect of recombination in the hns mutant. These results suggest that H-NS plays an important role in both homologous recombination and repair of bleomycin-induced damage, while StpA can substitute the H-NS function. The recombination analysis of hns single, stpA single, and hns stpA double mutants in the recBC sbcA and recBC sbcBC backgrounds suggested that the reduction of the hns single or hns stpA double mutants may not be due to the defect in a particular recombination pathway, but may be due to the defect in a common process of the pathways. The model for the functions of H-NS and StpA in homologous recombination and double-strand break repair is discussed.

  6. The tomato homolog of the gene encoding UV-damaged DNA binding protein 1 (DDB1) underlined as the gene that causes the high pigment-1 mutant phenotype.

    PubMed

    Lieberman, Michal; Segev, Orit; Gilboa, Nehama; Lalazar, Avraham; Levin, Ilan

    2004-05-01

    A tomato EST sequence, highly homologous to the human and Arabidopsis thaliana UV-damaged DNA binding protein 1 (DDB1), was mapped to the centromeric region of the tomato chromosome 2. This region was previously shown to harbor the HP-1 gene, encoding the high pigment-1 ( hp-1) and the high pigment-1(w) ( hp-1(w)) mutant phenotypes. Recent results also show that the A. thaliana DDB1 protein interacts both genetically and biochemically with the protein encoded by DEETIOLATED1, a gene carrying three tomato mutations that are in many respects isophenotypic to hp-1: high pigment-2 ( hp-2), high pigment-2(j) ( hp-2(j)) and dark green ( dg). The entire coding region of the DDB1 gene was sequenced in an hp-1 mutant and its near-isogenic normal plant in the cv. Ailsa Craig background, and also in an hp-1(w) mutant and its isogenic normal plant in the GT breeding line background. Sequence analysis revealed a single A(931)-to-T(931) base transversion in the coding sequence of the DDB1 gene in the hp-1 mutant plants. This transversion results in the substitution of the conserved asparagine at position 311 to a tyrosine residue. In the hp-1(w) mutant, on the other hand, a single G(2392)-to-A(2392) transition was observed, resulting in the substitution of the conserved glutamic acid at position 798 to a lysine residue. The single nucleotide polymorphism that differentiates hp-1 mutant and normal plants in the cv. Ailsa Craig background was used to design a pyrosequencing genotyping system. Analysis of a resource F(2) population segregating for the hp-1 mutation revealed a very strong linkage association between the DDB1 locus and the photomorphogenic response of the seedlings, measured as hypocotyl length (25

  7. Binding sequences for RdgB, a DNA damage-responsive transcriptional activator, and temperature-dependent expression of bacteriocin and pectin lyase genes in Pectobacterium carotovorum subsp. carotovorum.

    PubMed

    Yamada, Kazuteru; Kaneko, Jun; Kamio, Yoshiyuki; Itoh, Yoshifumi

    2008-10-01

    Pectobacterium carotovorum subsp. carotovorum strain Er simultaneously produces the phage tail-like bacteriocin carotovoricin (Ctv) and pectin lyase (Pnl) in response to DNA-damaging agents. The regulatory protein RdgB of the Mor/C family of proteins activates transcription of pnl through binding to the promoter. However, the optimal temperature for the synthesis of Ctv (23 degrees C) differs from that for synthesis of Pnl (30 degrees C), raising the question of whether RdgB directly activates ctv transcription. Here we report that RdgB directly regulates Ctv synthesis. Gel mobility shift assays demonstrated RdgB binding to the P(0), P(1), and P(2) promoters of the ctv operons, and DNase I footprinting determined RdgB-binding sequences (RdgB boxes) on these and on the pnl promoters. The RdgB box of the pnl promoter included a perfect 7-bp inverted repeat with high binding affinity to the regulator (K(d) [dissociation constant] = 150 nM). In contrast, RdgB boxes of the ctv promoters contained an imperfect inverted repeat with two or three mismatches that consequently reduced binding affinity (K(d) = 250 to 350 nM). Transcription of the rdgB and ctv genes was about doubled at 23 degrees C compared with that at 30 degrees C. In contrast, the amount of pnl transcription tripled at 30 degrees C. Thus, the inverse synthesis of Ctv and Pnl as a function of temperature is apparently controlled at the transcriptional level, and reduced rdgB expression at 30 degrees C obviously affected transcription from the ctv promoters with low-affinity RdgB boxes. Pathogenicity toward potato tubers was reduced in an rdgB knockout mutant, suggesting that the RdgAB system contributes to the pathogenicity of this bacterium, probably by activating pnl expression.

  8. Chromosomal localization and cDNA cloning of the genes (DDB1 and DDB2) for the p127 and p48 subunits of a human damage-specific DNA binding protein

    SciTech Connect

    Dualan, R.; Brody, T.; Keeney, S.

    1995-09-01

    DDB is a damage-specific DNA binding protein whose binding activity is absent from a minority of cell strains from individuals with xeroderma pigmentosum Group E, a human hereditary disease characterized by defective nucleotide excision DNA repair and an increased incidence of skin cancer. The binding activity from HeLa cells is associated with polypeptides of M{sub r} 124,000 and 41,000 as determined by SDS-polyacrylamide gels. This report describes the isolation of full-length human cDNAs encoding each polypeptide of DDB. The predicted peptide molecular masses based on open reading frames are 127,000 and 48,000. When expressed in an in vitro rabbit reticulocyte system, the p48 subunit migrates with an M{sub r} of 41 kDa on SDS-polyacrylamide gels, similarly to the peptide purified from HeLa cells. There is no significant homology between the derived p48 peptide sequence and any proteins in current databases, and the derived peptide sequence of p127 has homology only with the monkey DDB p127 (98% nucleotide identity and only one conserved amino acid substitution). Using a fluorescence in situ hybridization technique, the DDB p127 locus (DDB1) was assigned to the chromosomal location 11q12-q13, and the DDB p48 locus (DDB2) to 11p11-p12. 34 refs., 3 figs., 1 tab.

  9. CRN13 candidate effectors from plant and animal eukaryotic pathogens are DNA-binding proteins which trigger host DNA damage response.

    PubMed

    Ramirez-Garcés, Diana; Camborde, Laurent; Pel, Michiel J C; Jauneau, Alain; Martinez, Yves; Néant, Isabelle; Leclerc, Catherine; Moreau, Marc; Dumas, Bernard; Gaulin, Elodie

    2016-04-01

    To successfully colonize their host, pathogens produce effectors that can interfere with host cellular processes. Here we investigated the function of CRN13 candidate effectors produced by plant pathogenic oomycetes and detected in the genome of the amphibian pathogenic chytrid fungus Batrachochytrium dendrobatidis (BdCRN13). When expressed in Nicotiana, AeCRN13, from the legume root pathogen Aphanomyces euteiches, increases the susceptibility of the leaves to the oomycete Phytophthora capsici. When transiently expressed in amphibians or plant cells, AeCRN13 and BdCRN13 localize to the cell nuclei, triggering aberrant cell development and eventually causing cell death. Using Förster resonance energy transfer experiments in plant cells, we showed that both CRN13s interact with nuclear DNA and trigger plant DNA damage response (DDR). Mutating key amino acid residues in a predicted HNH-like endonuclease motif abolished the interaction of AeCRN13 with DNA, the induction of DDR and the enhancement of Nicotiana susceptibility to P. capsici. Finally, H2AX phosphorylation, a marker of DNA damage, and enhanced expression of genes involved in the DDR were observed in A. euteiches-infected Medicago truncatula roots. These results show that CRN13 from plant and animal eukaryotic pathogens promotes host susceptibility by targeting nuclear DNA and inducing DDR.

  10. Light scattered by model phantom bacteria reveals molecular interactions at their surface

    NASA Astrophysics Data System (ADS)

    Ghetta, A.; Prosperi, D.; Mantegazza, F.; Panza, L.; Riva, S.; Bellini, T.

    2005-11-01

    Testing molecular interactions is an ubiquitous need in modern biology and molecular medicine. Here, we present a qualitative and quantitative method rooted in the basic properties of the scattering of light, enabling detailed measurement of ligand-receptor interactions occurring on the surface of colloids. The key factor is the use of receptor-coated nanospheres matched in refractive index with water and therefore optically undetectable ("phantom") when not involved in adhesion processes. At the occurrence of ligand binding at the receptor sites, optically unmatched material adsorbs on the nanoparticle surface, giving rise to an increment in their scattering cross section up to a maximum corresponding to saturated binding sites. The analysis of the scattering growth pattern enables extracting the binding affinity. This label-free method has been assessed through the determination of the binding constant of the antibiotic vancomycin with the tripeptide L-Lys-D-Ala-D-Ala and of the vancomycin dimerization constant. We shed light on the role of chelate effect and molecular hindrance in the activity of this glycopeptide. binding affinity | nanoparticles | vancomycin | ligand-receptor recognition

  11. A novel cis-acting element from the 3'UTR of DNA damage-binding protein 2 mRNA links transcriptional and post-transcriptional regulation of gene expression.

    PubMed

    Melanson, Brian D; Cabrita, Miguel A; Bose, Reetesh; Hamill, Jeffrey D; Pan, Elysia; Brochu, Christian; Marcellus, Kristen A; Zhao, Tong T; Holcik, Martin; McKay, Bruce C

    2013-06-01

    The DNA damage-binding protein 2 (DDB2) is an adapter protein that can direct a modular Cul4-DDB1-RING E3 Ligase complex to sites of ultraviolet light-induced DNA damage to ubiquitinate substrates during nucleotide excision repair. The DDB2 transcript is ultraviolet-inducible; therefore, its regulation is likely important for its function. Curiously, the DDB2 mRNA is reportedly short-lived, but the transcript does not contain any previously characterized cis-acting determinants of mRNA stability in its 3' untranslated region (3'UTR). Here, we used a tetracycline regulated d2EGFP reporter construct containing specific 3'UTR sequences from DDB2 to identify novel cis-acting elements that regulate mRNA stability. Synthetic 3'UTRs corresponding to sequences as short as 25 nucleotides from the central region of the 3'UTR of DDB2 were sufficient to accelerate decay of the heterologous reporter mRNA. Conversely, these same 3'UTRs led to more rapid induction of the reporter mRNA, export of the message to the cytoplasm and the subsequent accumulation of the encoded reporter protein, indicating that this newly identified cis-acting element affects transcriptional and post-transciptional processes. These results provide clear evidence that nuclear and cytoplasmic processing of the DDB2 mRNA is inextricably linked.

  12. The p127 subunit (DDB1) of the UV-DNA damage repair binding protein is essential for the targeted degradation of STAT1 by the V protein of the paramyxovirus simian virus 5.

    PubMed

    Andrejeva, J; Poole, E; Young, D F; Goodbourn, S; Randall, R E

    2002-11-01

    The V protein of simian virus 5 (SV5) blocks interferon signaling by targeting STAT1 for proteasome-mediated degradation. Here we present three main pieces of evidence which demonstrate that the p127 subunit (DDB1) of the UV damage-specific DNA binding protein (DDB) plays a central role in this degradation process. First, the V protein of an SV5 mutant which fails to target STAT1 for degradation does not bind DDB1. Second, mutations in the N and C termini of V which abolish the binding of V to DDB1 also prevent V from blocking interferon (IFN) signaling. Third, treatment of HeLa/SV5-V cells, which constitutively express the V protein of SV5 and thus lack STAT1, with short interfering RNAs specific for DDB1 resulted in a reduction in DDB1 levels with a concomitant increase in STAT1 levels and a restoration of IFN signaling. Furthermore, STAT1 is degraded in GM02415 (2RO) cells, which have a mutation in DDB2 (the p48 subunit of DDB) which abolishes its ability to interact with DDB1, thereby demonstrating that the role of DDB1 in STAT1 degradation is independent of its association with DDB2. Evidence is also presented which demonstrates that STAT2 is required for the degradation of STAT1 by SV5. These results suggest that DDB1, STAT1, STAT2, and V may form part of a large multiprotein complex which leads to the targeted degradation of STAT1 by the proteasome.

  13. A possible role of histone-like DNA-binding protein of Streptococcus intermedius in the pathogenesis of bile duct damage in primary biliary cirrhosis.

    PubMed

    Haruta, Ikuko; Kikuchi, Ken; Hashimoto, Etsuko; Kato, Hidehito; Hirota, Katsuhiko; Kobayashi, Makio; Miyake, Yoichiro; Uchiyama, Takehiko; Yagi, Junji; Shiratori, Keiko

    2008-05-01

    Bacterial infection has become a focus of attention in the pathogenesis of primary biliary cirrhosis (PBC). It was reported that anti-histone autoantibody was detected in PBC, suggesting that bacterial histone-like DNA-binding protein (HLP) may be involved in the pathogenesis of PBC. To identify bacterial species in PBC to confirm this possibility, serum reactivity to bacterial cells was studied by ELISA. The IgM class Streptococcus intermedius titers were significantly higher in PBC than chronic hepatitis due to hepatitis C virus (CH-C) and healthy subjects. Among the streptococci, S. intermedius was selected for further study. The antigenic peptide of S. intermedius of HLP was synthesized to examine the serum reactivity to Si-HLP. IgM class anti-Si-HLP peptide titers were significantly higher in PBC. Immunoreactivity to anti-Si-HLP was detected in the cytoplasm of biliary epithelial cells and inflammatory cells in the portal area in PBC patients' livers. Streptococci, especially S. intermedius, might play a key role in the pathogenesis of PBC, possibly involving HLP.

  14. Central Role of the Copper-Binding Motif in the Complex Mechanism of Action of Ixosin: Enhancing Oxidative Damage and Promoting Synergy with Ixosin B.

    PubMed

    Libardo, M Daben J; Gorbatyuk, Vitaliy Y; Angeles-Boza, Alfredo M

    2016-01-08

    Ticks transmit multiple pathogens to different hosts without compromising their health. Their ability to evade microbial infections is largely a result of their effective innate immune response including various antimicrobial peptides. Therefore, a deep understanding of how ticks (and other arthropod vectors) control microbial loads could lead to the design of broad-spectrum antimicrobial agents. In this paper we study the role of the amino-terminal copper and nickel (ATCUN)-binding sequence in the peptide ixosin, isolated from the salivary glands of the hard tick Ixodes sinensis. Our results indicate that the ATCUN motif is not essential to the potency of ixosin, but is indispensable to its oxidative mechanism of action. Specifically, the ATCUN motif promotes dioxygen- and copper-dependent lipid (per)oxidation of bacterial membranes in a temporal fashion coinciding with the onset of bacterial death. Microscopy and studies on model membranes indicate that the oxidized phospholipids are utilized as potential targets of ixosin B (another tick salivary gland peptide) involving its delocalization to the bacterial membrane, thus resulting in a synergistic effect. Our proposed mechanism of action highlights the centrality of the ATCUN motif to ixosin's mechanism of action and demonstrates a novel way in which (tick) antimicrobial peptides (AMPs) utilize metal ions in its activity. This study suggests that ticks employ a variety of effectors to generate an amplified immune response, possibly justifying its vector competence.

  15. Self-Resistance and Cell Wall Composition in the Glycopeptide Producer Amycolatopsis balhimycina▿

    PubMed Central

    Schäberle, Till F.; Vollmer, Waldemar; Frasch, Hans-Jörg; Hüttel, Stephan; Kulik, Andreas; Röttgen, Marlene; von Thaler, Anna-Katharina; Wohlleben, Wolfgang; Stegmann, Evi

    2011-01-01

    The prevailing resistance mechanism against glycopeptides in Gram-positive pathogens involves reprogramming the biosynthesis of peptidoglycan precursors, resulting in d-alanyl-d-lactate depsipeptide termini. Amycolatopsis balhimycina produces the vancomycin-like glycopeptide balhimycin and therefore has to protect itself from the action of the glycopeptide. We studied the roles of the accessory resistance gene orthologs vanYb, vnlRb, and vnlSb, which are part of the balhimycin biosynthetic gene cluster (represented by the subscript “b”). The VanYb carboxypeptidase cleaved the terminal d-Ala from peptidoglycan precursors, and its heterologous expression enhanced glycopeptide resistance in Streptomyces coelicolor. The VanRS-like two component system VnlRSb was not involved in glycopeptide resistance or in the expression of the vanHAX glycopeptide resistance genes. Mature A. balhimycina peptidoglycan contained mainly tri- and tetrapeptides, with only traces of the d-Ala-d-Ala-ending pentapeptides that are binding sites for the antibiotic produced. The structure of the peptidoglycan precursor is consistent with the presence of vanHAX genes, which were identified outside the balhimycin synthesis cluster. Both wild-type and non-antibiotic-producing mutant strains synthesized peptidoglycan precursors ending mainly with d-Lac, indicating constitutive synthesis of a resistant cell wall. A. balhimycina could provide a model for an ancestral glycopeptide producer with constitutively expressed resistance genes. PMID:21690280

  16. Probing the Role of the Vancomycin E-Ring Aryl Chloride: Selective Divergent Synthesis and Evaluation of Alternatively Substituted E-Ring Analogues

    PubMed Central

    Pinchman, Joseph R.; Boger, Dale L.

    2013-01-01

    The selective functionalization of vancomycin aglycon derivatives through conversion of the E-ring aryl chloride to a reactive boronic acid, and its use in the synthesis of a systematic series of vancomycin E-ring analogues are described. The series was used to examine the E-ring chloride impact in binding d-Ala-d-Ala and on antimicrobial activity. In contrast to the reduced activity of the unsubstituted E-ring derivatives, hydrophobic and relatively non-polar substituents approach or match the chloro substituted vancomycin and was insensitive to the electronic character of the substituent (e.g. Cl vs CN/OMe), whereas highly polar substituents fail to provide the enhancements. Moreover, the active permethylated vancomycin aglycon derivatives exhibit VanB VRE antimicrobial activity at levels that approach (typically within 2-fold) their activity against sensitive bacteria. The robust borylation reaction also enabled the functionalization of a minimally protected vancomycin aglycon (N-Boc-vancomycin aglycon), and provides a direct method for the preparation of previously inaccessible analogues. PMID:23617725

  17. Duplex real-time PCR assay for rapid detection of ampicillin-resistant Enterococcus faecium.

    PubMed

    Mohn, Stein Christian; Ulvik, Arve; Jureen, Roland; Willems, Rob J L; Top, Janetta; Leavis, Helen; Harthug, Stig; Langeland, Nina

    2004-02-01

    Rapid and accurate identification of carriers of resistant microorganisms is an important aspect of efficient infection control in hospitals. Traditional identification methods of antibiotic-resistant bacteria usually take at least 3 to 4 days after sampling. A duplex real-time PCR assay was developed for rapid detection of ampicillin-resistant Enterococcus faecium (ARE). Primers and probes that are used in this assay specifically detected the D-Ala-D-Ala ligase gene of E. faecium and the modified penicillin-binding protein 5 gene (pbp5) carrying the Glu-to-Val substitution at position 629 (Val-629) in a set of 129 tested E. faecium strains with known pbp5 sequence. Presence of the Val-629 in the strain set from 11 different countries was highly correlated with ampicillin resistance. In a screening of hospitalized patients, the real-time PCR assay yielded a sensitivity and a specificity for the detection of ARE colonization of 95% and 100%, respectively. The results were obtained 4 h after samples were harvested from overnight broth of rectal swab samples, identifying both species and the resistance marker mutation in pbp5. This novel assay reliably identifies ARE 2 to 3 days more quickly than traditional culture methods, thereby increasing laboratory throughput, making it useful for rectal screening of ARE. The assay demonstrates the advantages of real-time PCR for detection of nosocomial pathogens.

  18. CCAAT/enhancer binding protein β (C/EBPβ) regulates the transcription of growth arrest and DNA damage-inducible protein 45 β (GADD45β) in articular chondrocytes.

    PubMed

    Shimada, Hirofumi; Otero, Miguel; Tsuchimochi, Kaneyuki; Yamasaki, Satoshi; Sakakima, Harutoshi; Matsuda, Fumiyo; Sakasegawa, Megumi; Setoguchi, Takao; Xu, Lin; Goldring, Mary B; Tanimoto, Akihide; Komiya, Setsuro; Ijiri, Kosei

    2016-04-01

    Osteoarthritis (OA) is a whole joint disease characterized by cartilage degradation, which causes pain and disability in older adults. Our previous work showed that growth arrest and DNA damage-inducible protein 45 β (GADD45β) is upregulated in chondrocyte clusters in OA cartilage, especially in the early stage of this disease. CCAAT/enhancer binding protein β (C/EBPβ) is expressed in the hypertrophic growth plate chondrocytes and functions in synergy with GADD45β. Here, the presence and localization of these proteins was assessed by immunohistochemistry using articular cartilage from OA patients, revealing colocalization of C/EBPβ and GADD45β in OA chondrocytes. GADD45β promoter analysis was performed to determine whether C/EBPβ directly regulates GADD45β transcription. Furthermore, we analyzed the effect of C/EBPβ on Gadd45β gene regulation in articular chondrocytes in vivo and in vitro. Immunohistochemical analysis of C/ebpβ-haploinsufficient mice (C/ebpβ(+/-)) cartilage showed that C/ebpβ haploinsufficiency led to reduced Gadd45β gene expression in these cells. In vitro, we evaluated the effects of conditional C/EBPβ overexpression driven by the cartilage oligomeric matrix protein (Comp) promoter in mComp-tTA;pTRE-Tight-BI-DsRed-mC/ebpβ transgenic mice. C/EBPβ overexpression significantly stimulated Gadd45β gene expression in articular chondrocytes. Taken together, our data demonstrate that C/EBPβ plays a central role in controlling Gadd45β gene expression in these cells.

  19. Right Hemisphere Brain Damage

    MedlinePlus

    ... Language and Swallowing / Disorders and Diseases Right Hemisphere Brain Damage [ en Español ] What is right hemisphere brain ... right hemisphere brain damage ? What is right hemisphere brain damage? Right hemisphere brain damage (RHD) is damage ...

  20. Binding Procurement

    NASA Technical Reports Server (NTRS)

    Rao, Gopalakrishna M.; Vaidyanathan, Hari

    2007-01-01

    This viewgraph presentation reviews the use of the binding procurement process in purchasing Aerospace Flight Battery Systems. NASA Engineering and Safety Center (NESC) requested NASA Aerospace Flight Battery Systems Working Group to develop a set of guideline requirements document for Binding Procurement Contracts.

  1. Structure of the Bacteriophage [phi]KZ Lytic Transglycosylase gp144

    SciTech Connect

    Fokine, Andrei; Miroshnikov, Konstantin A.; Shneider, Mikhail M.; Mesyanzhinov, Vadim V.; Rossmann, Michael G.

    2008-04-02

    Lytic transglycosylases are enzymes that act on the peptidoglycan of bacterial cell walls. They cleave the glycosidic linkage between N-acetylmuramoyl and N-acetylglucosaminyl residues with the concomitant formation of a 1,6-anhydromuramoyl product. The x-ray structure of the lytic transglycosylase gp144 from the Pseudomonas bacteriophage {phi}KZ has been determined to 2.5-{angstrom} resolution. This protein is probably employed by the bacteriophage in the late stage of the virus reproduction cycle to destroy the bacterial cell wall to release the phage progeny. {phi}KZ gp144 is a 260-residue {alpha}-helical protein composed of a 70-residue N-terminal cell wall-binding domain and a C-terminal catalytic domain. The fold of the N-terminal domain is similar to the peptidoglycan-binding domain from Streptomyces albus G d-Ala-d-Ala carboxypeptidase and to the N-terminal prodomain of human metalloproteinases that act on extracellular matrices. The C-terminal catalytic domain of gp144 has a structural similarity to the catalytic domain of the transglycosylase Slt70 from Escherichia coli and to lysozymes. The gp144 catalytic domain has an elongated groove that can bind at least five sugar residues at sites A-E. As in other lysozymes, the peptidoglycan cleavage (catalyzed by Glu{sup 115} in gp144) occurs between sugar-binding subsites D and E. The x-ray structure of the {phi}KZ transglycosylase complexed with the chitotetraose (N-acetylglucosamine){sub 4} has been determined to 2.6-{angstrom} resolution. The N-acetylglucosamine residues of the chitotetraose bind in sites A-D.

  2. Damaged Skylab

    NASA Technical Reports Server (NTRS)

    1973-01-01

    The Saturn V vehicle, carrying the unmarned orbital workshop for the Skylab-1 mission, lifted off successfully and all systems performed normally. Sixty-three seconds into the flight, engineers in the operation support and control center saw an unexpected telemetry indication that signalled that damages occurred on one solar array and the micrometeoroid shield during the launch. The micrometeoroid shield, a thin protective cylinder surrounding the workshop protecting it from tiny space particles and the sun's scorching heat, ripped loose from its position around the workshop. This caused the loss of one solar wing and jammed the other. Still unoccupied, the Skylab was stricken with the loss of the heat shield and sunlight beat mercilessly on the lab's sensitive skin. Internal temperatures soared, rendering the station uninhabitable, threatening foods, medicines, films, and experiments. This image, taken during a fly-around inspection by the Skylab-2 crew, shows a crippled Skylab in orbit. The crew found their home in space to be in serious shape; the heat shield gone, one solar wing gone, and the other jammed. The Marshall Space Flight Center (MSFC) developed, tested, rehearsed, and approved three repair options. These options included a parasol sunshade and a twin-pole sunshade to restore the temperature inside the workshop, and a set of metal cutting tools to free the jammed solar panel.

  3. Penicillin Binding Protein 1 Is Important in the Compensatory Response of Staphylococcus aureus to Daptomycin-Induced Membrane Damage and Is a Potential Target for β-Lactam–Daptomycin Synergy

    PubMed Central

    Berti, Andrew D.; Theisen, Erin; Sauer, John-Demian; Nonejuie, Poochit; Olson, Joshua; Pogliano, Joseph; Sakoulas, George; Nizet, Victor; Proctor, Richard A.

    2015-01-01

    The activity of daptomycin (DAP) against methicillin-resistant Staphylococcus aureus (MRSA) is enhanced in the presence of β-lactam antibiotics. This effect is more pronounced with β-lactam antibiotics that exhibit avid binding to penicillin binding protein 1 (PBP1). Here, we present evidence that PBP1 has a significant role in responding to DAP-induced stress on the cell. Expression of the pbpA transcript, encoding PBP1, was specifically induced by DAP exposure whereas expression of pbpB, pbpC, and pbpD, encoding PBP2, PBP3, and PBP4, respectively, remained unchanged. Using a MRSA COL strain with pbpA under an inducible promoter, increased pbpA transcription was accompanied by reduced susceptibility to, and killing by, DAP in vitro. Exposure to β-lactams that preferentially inactivate PBP1 was not associated with increased DAP binding, suggesting that synergy in the setting of anti-PBP1 pharmacotherapy results from increased DAP potency on a per-molecule basis. Combination exposure in an in vitro pharmacokinetic/pharmacodynamic model system with β-lactams that preferentially inactivate PBP1 (DAP-meropenem [MEM] or DAP-imipenem [IPM]) resulted in more-rapid killing than did combination exposure with DAP-nafcillin (NAF) (nonselective), DAP-ceftriaxone (CRO) or DAP-cefotaxime (CTX) (PBP2 selective), DAP-cefaclor (CEC) (PBP3 selective), or DAP-cefoxitin (FOX) (PBP4 selective). Compared to β-lactams with poor PBP1 binding specificity, exposure of S. aureus to DAP plus PBP1-selective β-lactams resulted in an increased frequency of septation and cell wall abnormalities. These data suggest that PBP1 activity may contribute to survival during DAP-induced metabolic stress. Therefore, targeted inactivation of PBP1 may enhance the antimicrobial efficiency of DAP, supporting the use of DAP–β-lactam combination therapy for serious MRSA infections, particularly when the β-lactam undermines the PBP1-mediated compensatory response. PMID:26525797

  4. Influence of the Charge State on the Structures and Interactions of Vancomycin Antibiotics with Cell-Wall Analogue Peptides: Experimental and Theoretical Studies

    SciTech Connect

    Yang, Zhibo; Vorpagel, Erich R.; Laskin, Julia

    2009-02-16

    In this study we examined the effect of the charge state on the energetics and dynamics of dissociation of the non-covalent complex between the vancomycin and the cell wall peptide analogue Nα,Nε-diacetyl-L-Lys-D-Ala-D-Ala (V-Ac2KDADA). The binding energies between the vancomycin and the peptide were obtained from the RRKM modeling of the time- and energy resolved surface-induced dissociation (SID) experiments. Our results demonstrate that the stability of the complex toward fragmentation increases in the order: [V+Ac2KDADA+H]+2 < [V+Ac2KDADA+H]+ < [V+Ac2KDADA-H]-. Dissociation of the singly protonated and singly deprotonated complex is characterized by very large entropy effects indicating substantial increase in the conformational flexibility of the resulting products. The experimental threshold energies of 1.75 eV and 1.34 eV obtained for the [V+Ac2KDADA-H]- and [V+Ac2KDADA+H]+ , respectively, are in excellent agreement with the results of density functional theory (DFT) calculations. The increased stability of the deprotonated complex observed experimentally is attributed to the presence of three charged sites in the deprotonated complex as compared to only one charged site in the singly protonated complex. The low binding energy of 0.93 eV obtained for the doubly protonated complex suggests that this ion is destabilized by Coulomb repulsion between the singly protonated vancomycin and the singly protonated peptide comprising the complex.

  5. Characterization of serine hydroxymethyltransferase GlyA as a potential source of D-alanine in Chlamydia pneumoniae.

    PubMed

    De Benedetti, Stefania; Bühl, Henrike; Gaballah, Ahmed; Klöckner, Anna; Otten, Christian; Schneider, Tanja; Sahl, Hans-Georg; Henrichfreise, Beate

    2014-01-01

    For intracellular Chlamydiaceae, there is no need to withstand osmotic challenges, and a functional cell wall has not been detected in these pathogens so far. Nevertheless, penicillin inhibits cell division in Chlamydiaceae resulting in enlarged aberrant bodies, a phenomenon known as chlamydial anomaly. D-alanine is a unique and essential component in the biosynthesis of bacterial cell walls. In free-living bacteria like Escherichia coli, penicillin-binding proteins such as monofunctional transpeptidases PBP2 and PBP3, the putative targets of penicillin in Chlamydiaceae, cross-link adjacent peptidoglycan strands via meso-diaminopimelic acid and D-Ala-D-Ala moieties of pentapeptide side chains. In the absence of genes coding for alanine racemase Alr and DadX homologs, the source of D-Ala and thus the presence of substrates for PBP2 and PBP3 activity in Chlamydiaceae has puzzled researchers for years. Interestingly, Chlamydiaceae genomes encode GlyA, a serine hydroxymethyltransferase that has been shown to exhibit slow racemization of D- and L-alanine as a side reaction in E. coli. We show that GlyA from Chlamydia pneumoniae can serve as a source of D-Ala. GlyA partially reversed the D-Ala auxotrophic phenotype of an E. coli racemase double mutant. Moreover, purified chlamydial GlyA had racemase activity on L-Ala in vitro and was inhibited by D-cycloserine, identifying GlyA, besides D-Ala ligase MurC/Ddl, as an additional target of this competitive inhibitor in Chlamydiaceae. Proof of D-Ala biosynthesis in Chlamydiaceae helps to clarify the structure of cell wall precursor lipid II and the role of chlamydial penicillin-binding proteins in the development of non-dividing aberrant chlamydial bodies and persistence in the presence of penicillin.

  6. Self-resistance mechanisms of actinomycetes producing lipid II-targeting antibiotics.

    PubMed

    Stegmann, Evi; Frasch, Hans-Joerg; Kilian, Regina; Pozzi, Roberta

    2015-02-01

    Glycopeptides and several lantibiotics are lipid II-targeting antibiotics produced by actinomycetes. To protect themselves from their own product, antibiotic producers developed self-resistance mechanisms. Inspection of different producer strains revealed that their resistance is not only based on a single determinant but on the synergistic action of different factors. Glycopeptide producers possess different ways to synthesize a modified peptidoglycan to prevent the binding of the glycopeptide antibiotic. One possible modification is the synthesis of peptidoglycan precursors terminating with a D-alanyl-D-lactate (D-Ala-D-Lac) rather than with a D-alanyl-D-alanine (D-Ala-D-Ala) resulting in a 1000-fold decreased binding affinity of the glycopeptide to its target. The reprogramming of the peptidoglycan precursor biosynthesis is based on the action of VanHAX or paralogous enzymes as it was shown for Amycolatopsis balhimycina. A second peptidoglycan modification resulting in glycopeptide resistance was investigated in the glycopeptide A40926 producer Nonomuraea ATCC 39727. Nonomuraea eliminates the glycopeptide target by synthesizing a peptidoglycan with 3-3 cross-linked peptide stems. The carboxypeptidase VanYn provides tetrapeptides which serve as substrates for the L,D-transpeptidase catalyzing the formation of 3-3 cross-links. The occurrence of 3-3 cross-linked dimers is also an important feature of the lantibiotic NAI-107 producer Microbispora ATCC PTA-5024. Moreover, the D-Ala in the fourth position in the acceptor peptide of muropeptides is exchanged to glycine or serine in Microbispora, a side reaction of the L,D-transpeptidase. Together with the lipoprotein MlbQ, the ABC transporter MlbYZ and the transmembrane protein MlbJ it might contribute to the self-resistance in Microbispora ATCC PTA-5024.

  7. AmpH, a Bifunctional dd-Endopeptidase and dd-Carboxypeptidase of Escherichia coli▿

    PubMed Central

    González-Leiza, Silvia M.; de Pedro, Miguel A.; Ayala, Juan A.

    2011-01-01

    In Escherichia coli, low-molecular-mass penicillin-binding proteins (LMM PBPs) are important for correct cell morphogenesis. These enzymes display dd-carboxypeptidase and/or dd-endopeptidase activities associated with maturation and remodeling of peptidoglycan (PG). AmpH has been classified as an AmpH-type class C LMM PBP, a group closely related to AmpC β-lactamases. AmpH has been associated with PG recycling, although its enzymatic activity remained uncharacterized until now. Construction and purification of His-tagged AmpH from E. coli permitted a detailed study of its enzymatic properties. The N-terminal export signal of AmpH is processed, but the protein remains membrane associated. The PBP nature of AmpH was demonstrated by its ability to bind the β-lactams Bocillin FL (a fluorescent penicillin) and cefmetazole. In vitro assays with AmpH and specific muropeptides demonstrated that AmpH is a bifunctional dd–endopeptidase and dd-carboxypeptidase. Indeed, the enzyme cleaved the cross-linked dimers tetrapentapeptide (D45) and tetratetrapeptide (D44) with efficiencies (kcat/Km) of 1,200 M−1 s−1 and 670 M−1 s−1, respectively, and removed the terminal d-alanine from muropeptides with a C-terminal d-Ala-d-Ala dipeptide. Both dd-peptidase activities were inhibited by 40 μM cefmetazole. AmpH also displayed a weak β-lactamase activity for nitrocefin of 1.4 × 10−3 nmol/μg protein/min, 1/1,000 the rate obtained for AmpC under the same conditions. AmpH was also active on purified sacculi, exhibiting the bifunctional character that was seen with pure muropeptides. The wide substrate spectrum of the dd-peptidase activities associated with AmpH supports a role for this protein in PG remodeling or recycling. PMID:22001512

  8. Rotational resonance determination of the structure of an enzyme-inhibitor complex: Phosphorylation of an (aminoalkyl)phosphinate inhibitor of D-alanyl-D-alanine ligase by ATP

    SciTech Connect

    McDermott, A.E.; Creuzet, F.; Griffin, R.G. ); Zawadzke, L.E.; Ye, Qizhuang; Walsh, C.T. )

    1990-06-19

    The authors have used a newly developed solid-state NMR method, rotational resonance, to establish the structure of an inhibited complex formed upon reaction of D-alanyl-D-alanine ligase, ATP, and the aminoalkyl dipeptide analogue (1(S)-aminoethyl)(2-carboxy-2(R)-methyl-1-ethyl)phosphinic acid (Ib). Analogue Ib was determined to be an ATP-dependent, slow-binding inhibitor of the D-Ala-D-Ala ligase from Salmonella typhimurium, with an enzyme-inhibitor half-life of 17 days at 37{degree}C. The inhibited complex shows a {sup 31}P NMR spectrum which is very different from that which would arise from a mixture of the free inhibitor and ATP. Four well-resolved lines were observed. A rotational resonance the spectrum shows evidence for strong dipolar couplings between the phosphinate phosphorus and a phosphate ester species. The dipolar coupling between the phosphorus signals at 53 and {minus}3 ppm was measured at rotational resonance by use of numerical simulations of both the line shape of the signal and the profile of magnetization transfer between the two sites. The measured coupling, 1.0 {plus minus} 0.2 kHz, indicates that the two species are bridged in a P-O-P linkage, with a P-P through-space distance of 2.7 {plus minus} 0.2 {angstrom}. This proves that the mechanism of inactivation involves phosphorylation of the enzyme-bound inhibitor by ATP to form a phosphoryl-phosphinate adduct.

  9. Quantification of the d-Ala-d-Lac-Terminated Peptidoglycan Structure in Vancomycin-Resistant Enterococcus faecalis Using a Combined Solid-State Nuclear Magnetic Resonance and Mass Spectrometry Analysis.

    PubMed

    Chang, James D; Foster, Erin E; Yang, Hao; Kim, Sung Joon

    2017-01-31

    Induction of vancomycin resistance in vancomycin-resistant enterococci (VRE) involves replacement of the d-Ala-d-Ala terminus of peptidoglycan (PG) stems with d-Ala-d-Lac, dramatically reducing the binding affinity of vancomycin for lipid II. Effects from vancomycin resistance induction in Enterococcus faecalis (ATCC 51299) were characterized using a combined solid-state nuclear magnetic resonance (NMR) and liquid chromatography-mass spectrometry (LC-MS) analysis. Solid-state NMR directly measured the total amounts of d-Lac and l,d-Ala metabolized from [2-(13)C]pyruvate, accumulated Park's nucleotide, and changes to the PG bridge-linking density during the early exponential growth phase (OD660 = 0.4) in intact whole cells of VRE. A high level of accumulation of depsipeptide-substituted Park's nucleotide consistent with the inhibition of the transglycosylation step of PG biosynthesis during the initial phase of vancomycin resistance was observed, while no changes to the PG bridge-linking density following the induction of vancomycin resistance were detected. This indicated that the attachment of the PG bridge to lipid II by the peptidyl transferases was not inhibited by the d-Ala-d-Lac-substituted PG stem structure in VRE. Compositions of mutanolysin-digested isolated cell walls of VRE grown with and without vancomycin resistance induction were determined by LC-MS. Muropeptides with PG stems terminating in d-Ala-d-Lac were found only in VRE grown in the presence of vancomycin. Percentages of muropeptides with a pentapeptide stem terminating in d-Ala-d-Lac for VRE grown in the presence of vancomycin were 26% for the midexponential phase (OD660 = 0.6) and 57% for the stationary growth phase (OD660 = 1.0). These high percentages indicate that d-Ala-d-Lac-substituted lipid II was efficiently utilized for PG biosynthesis in VRE.

  10. Characterization of VanYn, a novel D,D-peptidase/D,D-carboxypeptidase involved in glycopeptide antibiotic resistance in Nonomuraea sp. ATCC 39727.

    PubMed

    Binda, Elisa; Marcone, Giorgia L; Pollegioni, Loredano; Marinelli, Flavia

    2012-09-01

    VanY(n) is a novel protein involved in the mechanism of self-resistance in Nonomuraea sp. ATCC 39727, which produces the glycopeptide antibiotic A40926, the precursor of the second-generation dalbavancin, which is in phase III of clinical development. VanY(n) (196 residues) is encoded by the dbv7 gene within the dbv biosynthetic cluster devoted to A40926 production. C-terminal His6-tagged VanY(n) was successfully expressed as a soluble and active protein in Escherichia coli. The analysis of the sequence suggests the presence of a hydrophobic transmembrane portion and two conserved sequences (SxHxxGxAxD and ExxH) in the extracytoplasmic domain that are potentially involved in coordination of Zn(2+) and catalytic activity. The presence of these conserved sequences indicates a similar mechanism of action and substrate binding in VanY(n) as in VanY, VanX and VanXY Zn(2+)-dependent D,D-carboxypeptidases and D-Ala-D-Ala dipeptidases acting on peptidoglycan maturation and involved in glycopeptide resistance in pathogens. On substrates mimicking peptidoglycan precursors, VanY(n) shows D,D-carboxypeptidase and D,D-dipeptidase activity, but lacks D,D-carboxyesterase ability on D-Ala-D-Lac-terminating peptides. VanY(n) belongs to the metallo-D,D-carboxypeptidase family, but it is inhibited by β-lactams. Its characterization provides new insights into the evolution and transfer of resistance determinants from environmental glycopeptide-producing actinomycetes (such as Nonomuraea sp.) to glycopeptide-resistant pathogens (enterococci and staphylococci). It may also contribute to an early warning system for emerging resistance mechanisms following the introduction into clinics of a second-generation glycopeptide such as dalbavancin.

  11. DNA Damage and Pulmonary Hypertension

    PubMed Central

    Ranchoux, Benoît; Meloche, Jolyane; Paulin, Roxane; Boucherat, Olivier; Provencher, Steeve; Bonnet, Sébastien

    2016-01-01

    Pulmonary hypertension (PH) is defined by a mean pulmonary arterial pressure over 25 mmHg at rest and is diagnosed by right heart catheterization. Among the different groups of PH, pulmonary arterial hypertension (PAH) is characterized by a progressive obstruction of distal pulmonary arteries, related to endothelial cell dysfunction and vascular cell proliferation, which leads to an increased pulmonary vascular resistance, right ventricular hypertrophy, and right heart failure. Although the primary trigger of PAH remains unknown, oxidative stress and inflammation have been shown to play a key role in the development and progression of vascular remodeling. These factors are known to increase DNA damage that might favor the emergence of the proliferative and apoptosis-resistant phenotype observed in PAH vascular cells. High levels of DNA damage were reported to occur in PAH lungs and remodeled arteries as well as in animal models of PH. Moreover, recent studies have demonstrated that impaired DNA-response mechanisms may lead to an increased mutagen sensitivity in PAH patients. Finally, PAH was linked with decreased breast cancer 1 protein (BRCA1) and DNA topoisomerase 2-binding protein 1 (TopBP1) expression, both involved in maintaining genome integrity. This review aims to provide an overview of recent evidence of DNA damage and DNA repair deficiency and their implication in PAH pathogenesis. PMID:27338373

  12. DNA damage may drive nucleosomal reorganization to facilitate damage detection

    NASA Astrophysics Data System (ADS)

    LeGresley, Sarah E.; Wilt, Jamie; Antonik, Matthew

    2014-03-01

    One issue in genome maintenance is how DNA repair proteins find lesions at rates that seem to exceed diffusion-limited search rates. We propose a phenomenon where DNA damage induces nucleosomal rearrangements which move lesions to potential rendezvous points in the chromatin structure. These rendezvous points are the dyad and the linker DNA between histones, positions in the chromatin which are more likely to be accessible by repair proteins engaged in a random search. The feasibility of this mechanism is tested by considering the statistical mechanics of DNA containing a single lesion wrapped onto the nucleosome. We consider lesions which make the DNA either more flexible or more rigid by modeling the lesion as either a decrease or an increase in the bending energy. We include this energy in a partition function model of nucleosome breathing. Our results indicate that the steady state for a breathing nucleosome will most likely position the lesion at the dyad or in the linker, depending on the energy of the lesion. A role for DNA binding proteins and chromatin remodelers is suggested based on their ability to alter the mechanical properties of the DNA and DNA-histone binding, respectively. We speculate that these positions around the nucleosome potentially serve as rendezvous points where DNA lesions may be encountered by repair proteins which may be sterically hindered from searching the rest of the nucleosomal DNA. The strength of the repositioning is strongly dependent on the structural details of the DNA lesion and the wrapping and breathing of the nucleosome. A more sophisticated evaluation of this proposed mechanism will require detailed information about breathing dynamics, the structure of partially wrapped nucleosomes, and the structural properties of damaged DNA.

  13. Corrosion Damage Functions

    SciTech Connect

    Jones, Russell H.

    2002-11-30

    Corrosion damage can lead to reduced operational lifetimes. Often this damage is not as obvious as general corrosion but takes the form of pits, intergranular corrosion, crevice corrosion and hydrogen absorption. These types of corrosion damage lead to stress corrosion cracking, hydrogen induced cracking and corrosion fatigue. A critical step in defining a corrosion damage function is determining the relationship between the corrosion damage, the resulting crack propagation mechanism and component lifetimes. The sequence of events is often some localized corrosion event such as pitting, transition of the pit to a planar crack, propagation of this short crack, transition of the short crack to long crack conditions and continued propagation through Stage I, II, and III of the long crack SCC regimes. A description of critical corrosion damage processes and examples of the transition to long crack SCC conditions will be discussed.

  14. Chemically Induced Damage to the Hippocampal Formation,

    DTIC Science & Technology

    1986-05-01

    31-42. Siesj0, B K (1981): Cell damage in the brain: A speculative synthe i, J Cereb Blood Flow Metabol 1, 155-85. Stefanis, C (1964): Hippocampal... serotonin effects on central neurons, Advanc Pharmacol 6A, 414-18. Storm-Mathisen, J (1972): Glutamate decarboxylase in the rat hippo- campal region...and chro- nic toluene inhalation on behavior and [ H]- serotonin binding in rat, Life Sciences 30, 1997-2002. Zimmer, L, Woolley, D and Chang, L (1985

  15. Impact damage of composites

    NASA Technical Reports Server (NTRS)

    Wu, Hsi-Young T.; Springer, George S.

    1986-01-01

    A model is described for estimating the impact damage of fiber reinforced composite plates. The displacements and stresses are calculated by a three dimensional transient, finite element method of solution of the governing equations applicable to a linearly elastic body. The region in which damage occurs is estimated using the Tsai-Wu failure criterion. A computer code was developed which can be used to calculate the impact force, displacements and velocities of the plate and the impact body, stresses and strains in the plate, and the damage area. Sample numerical results are presented illustrating the type of information provided by the code. Comparisons between measured and calculated damage areas are also given.

  16. Damage Tolerance of Composites

    NASA Technical Reports Server (NTRS)

    Hodge, Andy

    2007-01-01

    Fracture control requirements have been developed to address damage tolerance of composites for manned space flight hardware. The requirements provide the framework for critical and noncritical hardware assessment and testing. The need for damage threat assessments, impact damage protection plans, and nondestructive evaluation are also addressed. Hardware intended to be damage tolerant have extensive coupon, sub-element, and full-scale testing requirements in-line with the Building Block Approach concept from the MIL-HDBK-17, Department of Defense Composite Materials Handbook.

  17. Comparative Analysis of Interaction of Human and Yeast DNA Damage Recognition Complexes with Damaged DNA in Nucleotide Excision Repair*

    PubMed Central

    Krasikova, Yuliya S.; Rechkunova, Nadejda I.; Maltseva, Ekaterina A.; Pestryakov, Pavel E.; Petruseva, Irina O.; Sugasawa, Kaoru; Chen, Xuejing; Min, Jung-Hyun; Lavrik, Olga I.

    2013-01-01

    The human XPC-RAD23B complex and its yeast ortholog, Rad4-Rad23, are the primary initiators of global genome nucleotide excision repair. The interaction of these proteins with damaged DNA was analyzed using model DNA duplexes containing a single fluorescein-substituted dUMP analog as a lesion. An electrophoretic mobility shift assay revealed similarity between human and yeast proteins in DNA binding. Quantitative analyses of XPC/Rad4 binding to the model DNA structures were performed by fluorescent depolarization measurements. XPC-RAD23B and Rad4-Rad23 proteins demonstrate approximately equal binding affinity to the damaged DNA duplex (KD ∼ (0.5 ± 0.1) and (0.6 ± 0.3) nm, respectively). Using photoreactive DNA containing 5-iodo-dUMP in defined positions, XPC/Rad4 location on damaged DNA was shown. Under conditions of equimolar binding to DNA both proteins exhibited the highest level of cross-links to 5I-dUMP located exactly opposite the damaged nucleotide. The positioning of the XPC and Rad4 proteins on damaged DNA by photocross-linking footprinting is consistent with x-ray analysis of the Rad4-DNA crystal complex. The identity of the XPC and Rad4 location illustrates the common principles of structure organization of DNA damage-scanning proteins from different Eukarya organisms. PMID:23443653

  18. The RNA Splicing Response to DNA Damage.

    PubMed

    Shkreta, Lulzim; Chabot, Benoit

    2015-10-29

    The number of factors known to participate in the DNA damage response (DDR) has expanded considerably in recent years to include splicing and alternative splicing factors. While the binding of splicing proteins and ribonucleoprotein complexes to nascent transcripts prevents genomic instability by deterring the formation of RNA/DNA duplexes, splicing factors are also recruited to, or removed from, sites of DNA damage. The first steps of the DDR promote the post-translational modification of splicing factors to affect their localization and activity, while more downstream DDR events alter their expression. Although descriptions of molecular mechanisms remain limited, an emerging trend is that DNA damage disrupts the coupling of constitutive and alternative splicing with the transcription of genes involved in DNA repair, cell-cycle control and apoptosis. A better understanding of how changes in splice site selection are integrated into the DDR may provide new avenues to combat cancer and delay aging.

  19. Damage and intensity survey

    USGS Publications Warehouse

    Reagor, G.; Brewer, L.R.

    1992-01-01

    A field team (the tuhors) from the National Earthquake Information Center (USGS) conducted a damage survey of the epicentral area in the week following the earthquakes. Detailed information about damage and where and how strongly the earthquakes were felt was obtained through interviews with local residents and personal observations. 

  20. An inducible long noncoding RNA amplifies DNA damage signaling

    PubMed Central

    Schmitt, Adam M.; Garcia, Julia T.; Hung, Tiffany; Flynn, Ryan A.; Shen, Ying; Qu, Kun; Payumo, Alexander Y.; Peres-da-Silva, Ashwin; Broz, Daniela Kenzelmann; Baum, Rachel; Guo, Shuling; Chen, James K.; Attardi, Laura D.; Chang, Howard Y.

    2016-01-01

    Long noncoding RNAs (lncRNAs) are prevalent genes with frequently exquisite regulation but mostly unknown functions. Here we demonstrate a role of lncRNAs in guiding organismal DNA damage response. DNA damage activates transcription of DINO (Damage Induced NOncoding) via p53. DINO is required for p53-dependent gene expression, cell cycle arrest, and apoptosis in response to DNA damage, and DINO expression suffice to activate damage signaling and cell cycle arrest in the absence of DNA damage. DINO binds to and promotes p53 protein stabilization, mediating a p53 auto-amplification loop. Dino knockout or promoter inactivation in mice dampens p53 signaling and ameliorates acute radiation syndrome in vivo. Thus, inducible lncRNA can create a feedback loop with its cognate transcription factor to amplify cellular signaling networks. PMID:27668660

  1. RNA protects a nucleoprotein complex against radiation damage

    SciTech Connect

    Bury, Charles S.; McGeehan, John E.; Antson, Alfred A.; Carmichael, Ian; Gerstel, Markus; Shevtsov, Mikhail B.; Garman, Elspeth F.

    2016-04-26

    Radiation damage during macromolecular X-ray crystallographic data collection is still the main impediment for many macromolecular structure determinations. Even when an eventual model results from the crystallographic pipeline, the manifestations of radiation-induced structural and conformation changes, the so-called specific damage, within crystalline macromolecules can lead to false interpretations of biological mechanisms. Although this has been well characterized within protein crystals, far less is known about specific damage effects within the larger class of nucleoprotein complexes. We developed a methodology whereby per-atom density changes could be quantified with increasing dose over a wide (1.3–25.0 MGy) range and at higher resolution (1.98 Å) than the previous systematic specific damage study on a protein–DNA complex. Specific damage manifestations were determined within the largetrpRNA-binding attenuation protein (TRAP) bound to a single-stranded RNA that forms a belt around the protein. Over a large dose range, the RNA was found to be far less susceptible to radiation-induced chemical changes than the protein. The availability of two TRAP molecules in the asymmetric unit, of which only one contained bound RNA, allowed a controlled investigation into the exact role of RNA binding in protein specific damage susceptibility. The 11-fold symmetry within each TRAP ring permitted statistically significant analysis of the Glu and Asp damage patterns, with RNA binding unexpectedly being observed to protect these otherwise highly sensitive residues within the 11 RNA-binding pockets distributed around the outside of the protein molecule. In addition, the method enabled a quantification of the reduction in radiation-induced Lys and Phe disordering upon RNA binding directly from the electron density.

  2. RNA protects a nucleoprotein complex against radiation damage

    DOE PAGES

    Bury, Charles S.; McGeehan, John E.; Antson, Alfred A.; ...

    2016-04-26

    Radiation damage during macromolecular X-ray crystallographic data collection is still the main impediment for many macromolecular structure determinations. Even when an eventual model results from the crystallographic pipeline, the manifestations of radiation-induced structural and conformation changes, the so-called specific damage, within crystalline macromolecules can lead to false interpretations of biological mechanisms. Although this has been well characterized within protein crystals, far less is known about specific damage effects within the larger class of nucleoprotein complexes. We developed a methodology whereby per-atom density changes could be quantified with increasing dose over a wide (1.3–25.0 MGy) range and at higher resolution (1.98more » Å) than the previous systematic specific damage study on a protein–DNA complex. Specific damage manifestations were determined within the largetrpRNA-binding attenuation protein (TRAP) bound to a single-stranded RNA that forms a belt around the protein. Over a large dose range, the RNA was found to be far less susceptible to radiation-induced chemical changes than the protein. The availability of two TRAP molecules in the asymmetric unit, of which only one contained bound RNA, allowed a controlled investigation into the exact role of RNA binding in protein specific damage susceptibility. The 11-fold symmetry within each TRAP ring permitted statistically significant analysis of the Glu and Asp damage patterns, with RNA binding unexpectedly being observed to protect these otherwise highly sensitive residues within the 11 RNA-binding pockets distributed around the outside of the protein molecule. In addition, the method enabled a quantification of the reduction in radiation-induced Lys and Phe disordering upon RNA binding directly from the electron density.« less

  3. War Damage Assessment

    NASA Technical Reports Server (NTRS)

    1994-01-01

    During and after the Persian Gulf war, hundreds of "oil lakes" were created in Kuwait by oil released from damaged wells. The lakes are a hazard to the Kuwait atmosphere, soil and ground water and must be carefully monitored. Boston University Center for Remote Sensing, assisted by other organizations, has accurately mapped the lakes using Landsat and Spot imagery. The war damage included the formation of over 300 oil lakes, oil pollution and sand dune movement. Total damage area is over 5,400 square kilometers - 30 percent of Kuwait's total surface area.

  4. Investigation of Friction-induced Damage to the Pig Cornea.

    PubMed

    Barros, Raquel C; Van Kooten, Theo G; Veeregowda, Deepak Halenahally

    2015-10-01

    Mechanical friction causes damage to the cornea. A friction measurement device with minimal intervention with the pig cornea tear film revealed a low friction coefficient of 0.011 in glycerine solution. Glycerine molecules presumably bind to water, mucins, and epithelial cells and therewith improve both squeeze film and boundary lubrication. Using confocal microscopy, we determined that glycerine solution reduced damage to epithelial cells by 50% compared with the phosphate buffer saline.

  5. Court Disallows Damage Claims

    ERIC Educational Resources Information Center

    Tomson, Bernard; Coplan, Norman

    1976-01-01

    In rejecting claims for damages, the Court finds that contract's "increase or decrease of cost" language is not applicable to added overhead costs and loss of labor efficiency resulting from delays over which the contractor has no control. (Author)

  6. Composites Damage Tolerance Workshop

    NASA Technical Reports Server (NTRS)

    Gregg, Wayne

    2006-01-01

    The Composite Damage Tolerance Workshop included participants from NASA, academia, and private industry. The objectives of the workshop were to begin dialogue in order to establish a working group within the Agency, create awareness of damage tolerance requirements for Constellation, and discuss potential composite hardware for the Crew Launch Vehicle (CLV) Upper Stage (US) and Crew Module. It was proposed that a composites damage tolerance working group be created that acts within the framework of the existing NASA Fracture Control Methodology Panel. The working group charter would be to identify damage tolerance gaps and obstacles for implementation of composite structures into manned space flight systems and to develop strategies and recommendations to overcome these obstacles.

  7. LSD and Genetic Damage

    ERIC Educational Resources Information Center

    Dishotsky, Norman I.; And Others

    1971-01-01

    Reviews studies of the effects of lysergic acid diethylamide (LSD) on man and other organisms. Concludes that pure LSD injected in moderate doses does not cause chromosome or detectable genetic damage and is not a teratogen or carcinogen. (JM)

  8. Nerve Damage (Diabetic Neuropathies)

    MedlinePlus

    ... may include numbness or insensitivity to pain or temperature a tingling, burning, or prickling sensation sharp pains ... from working properly, the body cannot regulate its temperature as it should. Nerve damage can also cause ...

  9. Diabetes and nerve damage

    MedlinePlus

    Diabetic neuropathy; Diabetes - neuropathy; Diabetes - peripheral neuropathy ... In people with diabetes, the body's nerves can be damaged by decreased blood flow and a high blood sugar level. This condition is ...

  10. Flexure with damage

    NASA Astrophysics Data System (ADS)

    Manaker, David M.; Turcotte, Donald L.; Kellogg, Louise H.

    2006-09-01

    Ductile behaviour in rocks is often associated with plasticity due to dislocation motion or diffusion under high pressures and temperatures. However, ductile behaviour can also occur in brittle materials. An example would be cataclastic flow associated with folding at shallow crustal levels. Engineers utilize damage mechanics to model the continuum deformation of brittle materials. In this paper we utilize a modified form of damage mechanics that includes a yield stress. Here, damage represents a reduction in frictional strength. We use this empirical approach to simulate bending of the lithosphere through the problem of plate flexure. We use numerical simulations to obtain quasi-static solutions to the Navier equations of elasticity. We use the program GeoFEST v. 4.5 (Geophysical Finite Element Simulation Tool), developed by NASA Jet Propulsion Laboratory, to generate solutions for each time step. When the von Mises stress exceeds the critical stress on an element we apply damage to reduce the shear modulus of the element. Damage is calculated at each time step by a power-law relationship of the ratio of the critical stress to the von Mises stress and the critical strain to the von Mises strain. This results in the relaxation of the material due to increasing damage. To test our method, we apply our damage rheology to a semi-infinite plate deforming under its own weight. Where the von Mises stress exceeds the critical stress, we simulate the formation of damage and observe the time-dependent relaxation of the stress and strain to near the yield strength. We simulate a wide range of behaviours from slow relaxation to instantaneous failure, over timescales that span six orders of magnitude. Using this method, stress relaxation produces perfectly plastic behaviour in cases where failure does not occur. For cases of failure, we observe a rapid increase in damage, analogous to the acceleration of microcrack formation and acoustic emissions prior to failure. Thus

  11. Assessing Tropical Cyclone Damage

    NASA Astrophysics Data System (ADS)

    Done, J.; Czajkowski, J.

    2012-12-01

    Landfalling tropical cyclones impact large coastal and inland areas causing direct damage due to winds, storm-surge flooding, tornadoes, and precipitation; as well as causing substantial indirect damage such as electrical outages and business interruption. The likely climate change impact of increased tropical cyclone intensity, combined with increases in exposure, bring the possibility of increased damage in the future. A considerable amount of research has focused on modeling economic damage due to tropical cyclones, and a series of indices have been developed to assess damages under climate change. We highlight a number of ways this research can be improved through a series of case study analyses. First, historical loss estimates are revisited to properly account for; time, impacted regions, the source of damage by type, and whether the damage was direct/indirect and insured/uninsured. Second, the drivers of loss from both the socio-economic and physical side are examined. A case is made to move beyond the use of maximum wind speed to more stable metrics and the use of other characteristics of the wind field such as direction, degree of gustiness, and duration is explored. A novel approach presented here is the potential to model losses directly as a function of climate variables such as sea surface temperature, greenhouse gases, and aerosols. This work is the first stage in the development of a tropical cyclone loss model to enable projections of losses under scenarios of both socio-economic change (such as population migration or altered policy) and physical change (such as shifts in tropical cyclone activity one from basin to another or within the same basin).

  12. Reducing Radiation Damage

    SciTech Connect

    Blankenbecler, Richard

    2006-06-05

    This talk describes the use of a modified treatment sequence, i.e., radiation dose, geometry, dwell time, etc., to mitigate some of the deleterious effects of cancer radiotherapy by utilizing natural cell repair processes. If bad side effects can be reduced, a more aggressive therapy can be put into place. Cells contain many mechanisms that repair damage of various types. If the damage can not be repaired, cells will undergo apoptosis (cell death). Data will be reviewed that support the fact that a small dose of radiation will activate damage repair genes within a cell. Once the mechanisms are fully active, they will efficiently repair the severe damage from a much larger radiation dose. The data ranges from experiments on specific cell cultures using microarray (gene chip) techniques to experiments on complete organisms. The suggested effect and treatment is consistent with the assumption that all radiation is harmful, no matter how small the dose. Nevertheless, the harm can be reduced. These mechanisms need to be further studied and characterized. In particular, their time dependence needs to be understood before the proposed treatment can be optimized. Under certain situations it is also possible that the deleterious effects of chemotherapy can be mitigated and the damage to radiation workers can be reduced.

  13. Crumpling Damaged Graphene

    PubMed Central

    Giordanelli, I.; Mendoza, M.; Andrade Jr., J. S.; Gomes, M. A. F.; Herrmann, H. J.

    2016-01-01

    Through molecular mechanics we find that non-covalent interactions modify the fractality of crumpled damaged graphene. Pristine graphene membranes are damaged by adding random vacancies and carbon-hydrogen bonds. Crumpled membranes exhibit a fractal dimension of 2.71 ± 0.02 when all interactions between carbon atoms are considered, and 2.30 ± 0.05 when non-covalent interactions are suppressed. The transition between these two values, obtained by switching on/off the non-covalent interactions of equilibrium configurations, is shown to be reversible and independent on thermalisation. In order to explain this transition, we propose a theoretical model that is compatible with our numerical findings. Finally, we also compare damaged graphene membranes with other crumpled structures, as for instance polymerised membranes and paper sheets, that share similar scaling properties. PMID:27173442

  14. Damage Tolerance Assessment Branch

    NASA Technical Reports Server (NTRS)

    Walker, James L.

    2013-01-01

    The Damage Tolerance Assessment Branch evaluates the ability of a structure to perform reliably throughout its service life in the presence of a defect, crack, or other form of damage. Such assessment is fundamental to the use of structural materials and requires an integral blend of materials engineering, fracture testing and analysis, and nondestructive evaluation. The vision of the Branch is to increase the safety of manned space flight by improving the fracture control and the associated nondestructive evaluation processes through development and application of standards, guidelines, advanced test and analytical methods. The Branch also strives to assist and solve non-aerospace related NDE and damage tolerance problems, providing consultation, prototyping and inspection services.

  15. Ureaplasma urealyticum binds mannose-binding lectin.

    PubMed

    Benstein, Barbara D; Ourth, Donald D; Crouse, Dennis T; Shanklin, D Radford

    2004-10-01

    Mannose-binding C-type lectin (MBL) is an important component of innate immunity in mammals. Mannose-binding lectin (MBL), an acute phase protein, acts as an opsonin for phagocytosis and also activates the mannan-binding lectin complement pathway. It may play a particularly significant role during infancy before adequate specific protection can be provided by the adaptive immune system. Ureaplasma urealyticum has been linked to several diseases including pneumonia and chronic lung disease (CLD) in premature infants. We therefore investigated the ability of U. urealyticum to bind MBL. A guinea pig IgG anti-rabbit-MBL antiserum was produced. An immunoblot (dot-blot) assay done on nitrocellulose membrane determined that the anti-MBL antibody had specificity against both rabbit and human MBL. Pure cultures of U. urealyticum, serotype 3, were used to make slide preparations. The slides containing the organisms were then incubated with nonimmune rabbit serum containing MBL. Ureaplasma was shown to bind rabbit MBL with an immunocytochemical assay using the guinea pig IgG anti-rabbit MBL antiserum. Horseradish peroxidase (HRP)-labeled anti-guinea pig IgG was used to localize the reaction. The anti-MBL antiserum was also used in an immunocytochemical assay to localize U. urealyticum in histological sections of lungs from mice specifically infected with this organism. The same method also indicated binding of MBL by ureaplasma in human lung tissue obtained at autopsy from culture positive infants. Our results demonstrate that ureaplasma has the capacity to bind MBL. The absence of MBL may play a role in the predisposition of diseases related to this organism.

  16. Courtside: A Damaging Lesson

    ERIC Educational Resources Information Center

    Zirkel, Perry A.

    2004-01-01

    This case presents a costly lesson for teachers and for districts that include a liquidated, or stipulated, damages clause in their teacher employment contracts. Although the court enforced the clause in this case, in this well-reasoned recent decision and in most of the much older, canvassed case law from other jurisdictions, the answer to the…

  17. Loss and damage

    NASA Astrophysics Data System (ADS)

    Huq, Saleemul; Roberts, Erin; Fenton, Adrian

    2013-11-01

    Loss and damage is a relative newcomer to the climate change agenda. It has the potential to reinvigorate existing mitigation and adaptation efforts, but this will ultimately require leadership from developed countries and enhanced understanding of several key issues, such as limits to adaptation.

  18. Coping with brain damage

    NASA Technical Reports Server (NTRS)

    Waring, W.

    1974-01-01

    Two neurological disorders, cerebral palsy, and traumatic brain damage as from an accident, are considered. The discussion covers the incidence of disabilities, their characteristics, and what is now being done to deal with them, particularly in reference to areas in which the capabilities of the engineer can be effectively applied.

  19. Modifying Radiation Damage

    PubMed Central

    Kim, Kwanghee; McBride, William H.

    2011-01-01

    Radiation leaves a fairly characteristic footprint in biological materials, but this is rapidly all but obliterated by the canonical biological responses to the radiation damage. The innate immune recognition systems that sense “danger” through direct radiation damage and through associated collateral damage set in motion a chain of events that, in a tissue compromised by radiation, often unwittingly result in oscillating waves of molecular and cellular responses as tissues attempt to heal. Understanding “nature’s whispers” that inform on these processes will lead to novel forms of intervention targeted more precisely towards modifying them in an appropriate and timely fashion so as to improve the healing process and prevent or mitigate the development of acute and late effects of normal tissue radiation damage, whether it be accidental, as a result of a terrorist incident, or of therapeutic treatment of cancer. Here we attempt to discuss some of the non-free radical scavenging mechanisms that modify radiation responses and comment on where we see them within a conceptual framework of an evolving radiation-induced lesion. PMID:20583981

  20. Evolving nucleotide binding surfaces

    NASA Technical Reports Server (NTRS)

    Kieber-Emmons, T.; Rein, R.

    1981-01-01

    An analysis is presented of the stability and nature of binding of a nucleotide to several known dehydrogenases. The employed approach includes calculation of hydrophobic stabilization of the binding motif and its intermolecular interaction with the ligand. The evolutionary changes of the binding motif are studied by calculating the Euclidean deviation of the respective dehydrogenases. Attention is given to the possible structural elements involved in the origin of nucleotide recognition by non-coded primordial polypeptides.

  1. Fluorescent reports for detection and measurement of DNA damage

    SciTech Connect

    Uziel, M.; Houck, K. )

    1993-01-01

    Epidemiological studies of real populations are complicated by the inevitable coexistence of exposure to multiple agents within the target population. An alternative method for characterizing these types of exposures is to use the reactive chemical functional group as the toxic agent identify the corresponding classes (families) of damage as markers of effects. We have begun studies to develop spectrometric reporters of DNA damage that can be measured on intact DNA. The direct measurement of adducts on microgram levels of DNA from tissue biopsy may succeed because of the high sensitivity and selectivity of different reporter compounds. While one cannot readily distinguish between recent or persistent exposures, baseline values for individuals may be constructed. For example, normal oxidative metabolism and environmental radiation create oxidation processes that continually damage DNA. These reactions create lesions that can be measured with the reporter compound FABA [N- (5- fluoresceinyl), N[prime]-(3-boronatophenyl)thioureal]. We report preliminary observations with binding FABA (selective for cis, vicdiol structures) to damage sites present on intact nonirradiated and irradiated DNA from C3H10T[sub 1/2] cells. We have observed binding of 42,000 FABA per mouse tetraploid genome (9 billion base pairs) to the putative thymidylic glycol resulting from normal oxidative processes in nonirradiated DNA. Additional binding of FABA to DNA from cells exposed to 100, 300, and 500 rad shows an exponential increase in binding sites of up to 140,000 with 500 rad exposure. This damage reporter may prove useful in characterizing levels of nonovert and overt oxidative damage to DNA.

  2. Signaling domain of Sonic Hedgehog as cannibalistic calcium-regulated zinc-peptidase.

    PubMed

    Rebollido-Rios, Rocio; Bandari, Shyam; Wilms, Christoph; Jakuschev, Stanislav; Vortkamp, Andrea; Grobe, Kay; Hoffmann, Daniel

    2014-07-01

    Sonic Hedgehog (Shh) is a representative of the evolutionary closely related class of Hedgehog proteins that have essential signaling functions in animal development. The N-terminal domain (ShhN) is also assigned to the group of LAS proteins (LAS = Lysostaphin type enzymes, D-Ala-D-Ala metalloproteases, Sonic Hedgehog), of which all members harbor a structurally well-defined Zn2+ center; however, it is remarkable that ShhN so far is the only LAS member without proven peptidase activity. Another unique feature of ShhN in the LAS group is a double-Ca2+ center close to the zinc. We have studied the effect of these calcium ions on ShhN structure, dynamics, and interactions. We find that the presence of calcium has a marked impact on ShhN properties, with the two calcium ions having different effects. The more strongly bound calcium ion significantly stabilizes the overall structure. Surprisingly, the binding of the second calcium ion switches the putative catalytic center from a state similar to LAS enzymes to a state that probably is catalytically inactive. We describe in detail the mechanics of the switch, including the effect on substrate co-ordinating residues and on the putative catalytic water molecule. The properties of the putative substrate binding site suggest that ShhN could degrade other ShhN molecules, e.g. by cleavage at highly conserved glycines in ShhN. To test experimentally the stability of ShhN against autodegradation, we compare two ShhN mutants in vitro: (1) a ShhN mutant unable to bind calcium but with putative catalytic center intact, and thus, according to our hypothesis, a constitutively active peptidase, and (2) a mutant carrying additionally mutation E177A, i.e., with the putative catalytically active residue knocked out. The in vitro results are consistent with ShhN being a cannibalistic zinc-peptidase. These experiments also reveal that the peptidase activity depends on pH.

  3. DNA binding studies of tartrazine food additive.

    PubMed

    Kashanian, Soheila; Zeidali, Sahar Heidary

    2011-07-01

    The interaction of native calf thymus DNA with tartrazine in 10 mM Tris-HCl aqueous solution at neutral pH 7.4 was investigated. Tartrazine is a nitrous derivative and may cause allergic reactions, with a potential of toxicological risk. Also, tartrazine induces oxidative stress and DNA damage. Its DNA binding properties were studied by UV-vis and circular dichroism spectra, competitive binding with Hoechst 33258, and viscosity measurements. Tartrazine molecules bind to DNA via groove mode as illustrated by hyperchromism in the UV absorption band of tartrazine, decrease in Hoechst-DNA solution fluorescence, unchanged viscosity of DNA, and conformational changes such as conversion from B-like to C-like in the circular dichroism spectra of DNA. The binding constants (K(b)) of DNA with tartrazine were calculated at different temperatures. Enthalpy and entropy changes were calculated to be +37 and +213 kJ mol(-1), respectively, according to the Van't Hoff equation, which indicated that the reaction is predominantly entropically driven. Also, tartrazine does not cleave plasmid DNA. Tartrazine interacts with calf thymus DNA via a groove interaction mode with an intrinsic binding constant of 3.75 × 10(4) M(-1).

  4. Melanin-binding radiopharmaceuticals

    SciTech Connect

    Packer, S; Fairchild, R G; Watts, K P; Greenberg, D; Hannon, S J

    1980-01-01

    The scope of this paper is limited to an analysis of the factors that are important to the relationship of radiopharmaceuticals to melanin. While the authors do not attempt to deal with differences between melanin-binding vs. melanoma-binding, a notable variance is assumed. (PSB)

  5. Tornado damage risk assessment

    SciTech Connect

    Reinhold, T.A.; Ellingwood, B.

    1982-09-01

    Several proposed models were evaluated for predicting tornado wind speed probabilities at nuclear plant sites as part of a program to develop statistical data on tornadoes needed for probability-based load combination analysis. A unified model was developed which synthesized the desired aspects of tornado occurrence and damage potential. The sensitivity of wind speed probability estimates to various tornado modeling assumptions are examined, and the probability distributions of tornado wind speed that are needed for load combination studies are presented.

  6. Battle Damage Modeling

    DTIC Science & Technology

    2010-05-01

    The compressive matrix damage zone was found along the whole central thickness for CFRP models only in the case of 875 g blast load, while is...high specific properties, also fiber- reinforced polymers are being considered for energy absorption applications in military armors. A deep insight...equilibrium. Due to their high specific properties, also fiber-reinforced polymers are being considered for energy absorption applications in

  7. Earthquake damage to schools

    USGS Publications Warehouse

    McCullough, Heather

    1994-01-01

    These unusual slides show earthquake damage to school and university buildings around the world. They graphically illustrate the potential danger to our schools, and to the welfare of our children, that results from major earthquakes. The slides range from Algeria, where a collapsed school roof is held up only by students' desks; to Anchorage, Alaska, where an elementary school structure has split in half; to California and other areas, where school buildings have sustained damage to walls, roofs, and chimneys. Interestingly, all the United States earthquakes depicted in this set of slides occurred either on a holiday or before or after school hours, except the 1935 tremor in Helena, Montana, which occurred at 11:35 am. It undoubtedly would have caused casualties had the schools not been closed days earlier by Helena city officials because of a damaging foreshock. Students in Algeria, the People's Republic of China, Armenia, and other stricken countries were not so fortunate. This set of slides represents 17 destructive earthquakes that occurred in 9 countries, and covers more than a century--from 1886 to 1988. Two of the tremors, both of which occurred in the United States, were magnitude 8+ on the Richter Scale, and four were magnitude 7-7.9. The events represented by the slides (see table below) claimed more than a quarter of a million lives.

  8. DAMCAL; Damage Reach Stage-Damage Calculation: Users Manual

    DTIC Science & Technology

    1979-02-01

    M AD-A273 611 US Army Corps of Engineers Hydrologic Engineering Center GENERALIZED COMPUTER PROGRAM DTIC "k ELECTE DEC 13 1993 DAMCAL A Damage Reach...Stage- Damage Calculation User’s Manual February 1979 93-30134 ApprOved for Public Release. Distribution Unlimited. CPD-35 93 12 100 3 8 DAMCAL Damage ...Reach Stage- Damage Calculation Accesion For User’s Manual TIS ’C&I DTIC TA.3 Juhfificatoc.n By ............. .......................... February 1979

  9. Chimeric Proteins to Detect DNA Damage and Mismatches

    SciTech Connect

    McCutchen-Maloney, S; Malfatti, M; Robbins, K M

    2002-01-14

    The goal of this project was to develop chimeric proteins composed of a DNA mismatch or damage binding protein and a nuclease, as well as methods to detect DNA mismatches and damage. We accomplished this through protein engineering based on using polymerase chain reactions (PCRs) to create chimeras with novel functions for damage and mismatch detection. This project addressed fundamental questions relating to disease susceptibility and radiation-induced damage in cells. It also supported and enhanced LLNL's competency in the emerging field of proteomics. In nature, DNA is constantly being subjected to damaging agents such as exposure to ultraviolet (UV) radiation and various environmental and dietary carcinogens. If DNA damage is not repaired however, mutations in DNA result that can eventually manifest in cancer and other diseases. In addition to damage-induced DNA mutations, single nucleotide polymorphisms (SNPs), which are variations in the genetic sequence between individuals, may predispose some to disease. As a result of the Human Genome Project, the integrity of a person's DNA can now be monitored. Therefore, methods to detect DNA damage, mutations, and SNPs are useful not only in basic research but also in the health and biotechnology industries. Current methods of detection often use radioactive labeling and rely on expensive instrumentation that is not readily available in many research settings. Our methods to detect DNA damage and mismatches employ simple gel electrophoresis and flow cytometry, thereby alleviating the need for radioactive labeling and expensive equipment. In FY2001, we explored SNP detection by developing methods based on the ability of the chimeric proteins to detect mismatches. Using multiplex assays with flow cytometry and fluorescent beads to which the DNA substrates where attached, we showed that several of the chimeras possess greater affinity for damaged and mismatched DNA than for native DNA. This affinity was demonstrated in

  10. How cells recognize damaged DNA: Clues from xeroderma pigmentosum and yeast

    SciTech Connect

    Chu, G.; Chang, E.; Patterson, M. )

    1990-01-01

    Xeroderma pigmentosum (XP) is characterized by the defective excision repair of DNA damaged by many agents, including ultraviolet radiation (UV) and cisplatin. We have identified a factor in human cells that recognizes multiple forms of DNA damage and is absent in XP complementation group E. Denoted XPE binding factor, it is expressed at five-fold higher levels in tumor cell lines resistant to the antitumor drug cisplatin. Finally, although it does not have photoreactivating activity, XPE binding factor shares multiple binding characteristics with yeast photolyase, suggesting that it is the human homolog of photolyase.

  11. Low Level Laser Retinal Damage

    DTIC Science & Technology

    1990-03-01

    18 Related Projects ........................ . . ....... 20 References . . . . .......................... 22 2 INTRODUCTION The objectives of...fluorescein is a potent phototoxic agent in the retina.26 The damage threshold for blue light retinal damage is lowered by a factor of ten after an... Related to the Probiem of Retinal Light Damage 1. Corneal Holography 2. Hematoporphyrin Studies 3. Fluorescein Fluorescence Measurements 7 EQUIPMENT

  12. Urinary intestinal fatty acid binding protein predicts necrotizing enterocolitis.

    PubMed

    Gregory, Katherine E; Winston, Abigail B; Yamamoto, Hidemi S; Dawood, Hassan Y; Fashemi, Titilayo; Fichorova, Raina N; Van Marter, Linda J

    2014-06-01

    Necrotizing enterocolitis, characterized by sudden onset and rapid progression, remains the most significant gastrointestinal disorder among premature infants. In seeking a predictive biomarker, we found intestinal fatty acid binding protein, an indicator of enterocyte damage, was substantially increased within three and seven days before the diagnosis of necrotizing enterocolitis.

  13. Tokamak ARC damage

    SciTech Connect

    Murray, J.G.; Gorker, G.E.

    1985-01-01

    Tokamak fusion reactors will have large plasma currents of approximately 10 MA with hundreds of megajoules stored in the magnetic fields. When a major plasma instability occurs, the disruption of the plasma current induces voltage in the adjacent conducting structures, giving rise to large transient currents. The induced voltages may be sufficiently high to cause arcing across sector gaps or from one protruding component to another. This report reviews a tokamak arcing scenario and provides guidelines for designing tokamaks to minimize the possibility of arc damage.

  14. Does orthodontics damage faces?

    PubMed

    DiBiase, A T; Sandler, P J

    2001-03-01

    With the increasing provision of orthodontic care in this country, certain practitioners have raised concerns regarding the use of elective extractions and retraction mechanics, especially the effects on the facial profile and the TMJ. The non-extraction versus extraction debate spans the history of orthodontics, and the concepts of facial attractiveness are subject to change as fashions change. Within the realms of evidence-based practice, there is little or no evidence to suggest that the philosophies and mechanics of contemporary orthodontics, in the vast majority of cases, cause damage to the profile or are directly linked to the development of TMJ dysfunction.

  15. Memory binding and white matter integrity in familial Alzheimer's disease.

    PubMed

    Parra, Mario A; Saarimäki, Heini; Bastin, Mark E; Londoño, Ana C; Pettit, Lewis; Lopera, Francisco; Della Sala, Sergio; Abrahams, Sharon

    2015-05-01

    Binding information in short-term and long-term memory are functions sensitive to Alzheimer's disease. They have been found to be affected in patients who meet criteria for familial Alzheimer's disease due to the mutation E280A of the PSEN1 gene. However, only short-term memory binding has been found to be affected in asymptomatic carriers of this mutation. The neural correlates of this dissociation are poorly understood. The present study used diffusion tensor magnetic resonance imaging to investigate whether the integrity of white matter structures could offer an account. A sample of 19 patients with familial Alzheimer's disease, 18 asymptomatic carriers and 21 non-carrier controls underwent diffusion tensor magnetic resonance imaging, neuropsychological and memory binding assessment. The short-term memory binding task required participants to detect changes across two consecutive screens displaying arrays of shapes, colours, or shape-colour bindings. The long-term memory binding task was a Paired Associates Learning Test. Performance on these tasks were entered into regression models. Relative to controls, patients with familial Alzheimer's disease performed poorly on both memory binding tasks. Asymptomatic carriers differed from controls only in the short-term memory binding task. White matter integrity explained poor memory binding performance only in patients with familial Alzheimer's disease. White matter water diffusion metrics from the frontal lobe accounted for poor performance on both memory binding tasks. Dissociations were found in the genu of corpus callosum which accounted for short-term memory binding impairments and in the hippocampal part of cingulum bundle which accounted for long-term memory binding deficits. The results indicate that white matter structures in the frontal and temporal lobes are vulnerable to the early stages of familial Alzheimer's disease and their damage is associated with impairments in two memory binding functions known to

  16. Oxidative base damage in RNA detected by reverse transcriptase.

    PubMed Central

    Rhee, Y; Valentine, M R; Termini, J

    1995-01-01

    Oxidative base damage in DNA and metabolic defects in the recognition and removal of such damage play important roles in mutagenesis and human disease. The extent to which cellular RNA is a substrate for oxidative damage and the possible biological consequences of RNA base oxidation, however, remain largely unexplored. Since oxidatively modified RNA may contribute to the high mutability of retroviral genomic DNA, we have been interested in developing methods for the sequence specific detection of such damage. We show here that a primer extension assay using AMV reverse transcriptase (RT) can be used to reveal oxidatively damaged sites in RNA. This finding extends the currently known range of RNA modifications detectable with AMV reverse transcriptase. Analogous assays using DNA polymerases to detect base damage in DNA substrates appear to be restricted to lesions at thymine. Oxidative base damage in the absence of any detectable chain breaks was produced by dye photosensitization of RNA. Six out of 20 dyes examined were capable of producing RT detectable lesions. RT stops were seen predominantly at purines, although many pyrimidine sites were also detected. Dye specific photofootprints revealed by RT analysis suggests differential dye binding to the RNA substrate. Some of the photoreactive dyes described here may have potential utility in RNA structural analysis, particularly in the identification of stem-loop regions in complex RNAs. Images PMID:7545285

  17. Metallochaperones: bind and deliver

    SciTech Connect

    Rosenzweig, A.C.

    2010-03-08

    Metallochaperones deliver metal ions directly to target proteins via specific protein-protein interactions. Recent research has led to a molecular picture of how some metallochaperones bind metal ions, recognize their partner proteins, and accomplish metal ion transfer.

  18. SHBG (Sex Hormone Binding Globulin)

    MedlinePlus

    ... as: Testosterone-estrogen Binding Globulin; TeBG Formal name: Sex Hormone Binding Globulin Related tests: Testosterone , Free Testosterone, ... I should know? How is it used? The sex hormone binding globulin (SHBG) test may be used ...

  19. Thoracic damage control surgery.

    PubMed

    Gonçalves, Roberto; Saad, Roberto

    2016-01-01

    The damage control surgery came up with the philosophy of applying essential maneuvers to control bleeding and abdominal contamination in trauma patients who are within the limits of their physiological reserves. This concept was extended to thoracic injuries, where relatively simple maneuvers can shorten operative time of in extremis patients. This article aims to revise the various damage control techniques in thoracic organs that must be known to the surgeon engaged in emergency care. RESUMO A cirurgia de controle de danos surgiu com a filosofia de se aplicar manobras essenciais para controle de sangramento e contaminação abdominal, em doentes traumatizados, nos limites de suas reservas fisiológicas. Este conceito se estendeu para as lesões torácicas, onde manobras relativamente simples, podem abreviar o tempo operatório de doentes in extremis. Este artigo tem como objetivo, revisar as diversas técnicas de controle de dano em órgãos torácicos, que devem ser de conhecimento do cirurgião que atua na emergência.

  20. New DNA-binding radioprotectors

    NASA Astrophysics Data System (ADS)

    Martin, Roger

    The normal tissue damage associated with cancer radiotherapy has motivated the development at Peter Mac of a new class of DNA-binding radioprotecting drugs that could be applied top-ically to normal tissues at risk. Methylproamine (MP), the lead compound, reduces radiation induced cell kill at low concentrations. For example, experiments comparing the clonogenic survival of transformed human keratinocytes treated with 30 micromolar MP before and dur-ing various doses of ionising radiation, with the radiation dose response for untreated cells, indicate a dose reduction factor (DRF) of 2. Similar survival curve experiments using various concentrations of MP, with parallel measurements of uptake of MP into cell nuclei, have en-abled the relationship between drug uptake and extent of radioprotection to be established. Radioprotection has also been demonstrated after systemic administration to mice, for three different endpoints, namely lung, jejunum and bone marrow (survival at 30 days post-TBI). The results of pulse radiolysis studies indicated that the drugs act by reduction of transient radiation-induced oxidative species on DNA. This hypothesis was substantiated by the results of experiments in which MP radioprotection of radiation-induced DNA double-strand breaks, assessed as -H2AX foci, in the human keratinocyte cell line. For both endpoints, the extent of radioprotection increased with MP concentration up to a maximal value. These results are consistent with the hypothesis that radioprotection by MP is mediated by attenuation of the extent of initial DNA damage. However, although MP is a potent radioprotector, it becomes cytotoxic at higher concentrations. This limitation has been addressed in an extensive program of lead optimisation and some promising analogues have emerged from which the next lead will be selected. Given the clinical potential of topical radioprotection, the new analogues are being assessed in terms of delivery to mouse oral mucosa. This is

  1. The Sunscreen Octyl Methoxycinnamate Binds to DNA

    NASA Astrophysics Data System (ADS)

    Norrell, Johannes; Vohra, Shikhar; Nordlund, T. M.

    2000-03-01

    Sunscreens are designed to prevent skin cancer by absorbing ultraviolet radiation from the sun before it gets to the DNA in skin cells. The purpose of this work is to determine whether or not octyl methoxycinnamate, an active ingredient in many sunscreens, will bind to DNA. If so, the sunscreen could transfer the energy it absorbed from the sun to the DNA and cause damage. To determine this, we prepared samples with varying concentrations of cinnamate added to herring sperm DNA, sonicating the mixture to disperse the hydrophobic sunscreen into solution. Absorption and fluorescence spectra of the mixtures showed (i) much more sunscreen was dispersed into solution when DNA was present, and (ii) the spectra of both DNA and sunscreen differed from those of the separate solutions. We conclude that the octyl methoxycinnamate can indeed bind to DNA in aqueous solution. Energy transfer experiments from DNA to sunscreen and from sunscreen to 2-aminopurine- (a fluorescent DNA base) labeled DNA will be presented.

  2. Multivariate pluvial flood damage models

    SciTech Connect

    Van Ootegem, Luc; Verhofstadt, Elsy; Van Herck, Kristine; Creten, Tom

    2015-09-15

    Depth–damage-functions, relating the monetary flood damage to the depth of the inundation, are commonly used in the case of fluvial floods (floods caused by a river overflowing). We construct four multivariate damage models for pluvial floods (caused by extreme rainfall) by differentiating on the one hand between ground floor floods and basement floods and on the other hand between damage to residential buildings and damage to housing contents. We do not only take into account the effect of flood-depth on damage, but also incorporate the effects of non-hazard indicators (building characteristics, behavioural indicators and socio-economic variables). By using a Tobit-estimation technique on identified victims of pluvial floods in Flanders (Belgium), we take into account the effect of cases of reported zero damage. Our results show that the flood depth is an important predictor of damage, but with a diverging impact between ground floor floods and basement floods. Also non-hazard indicators are important. For example being aware of the risk just before the water enters the building reduces content damage considerably, underlining the importance of warning systems and policy in this case of pluvial floods. - Highlights: • Prediction of damage of pluvial floods using also non-hazard information • We include ‘no damage cases’ using a Tobit model. • The damage of flood depth is stronger for ground floor than for basement floods. • Non-hazard indicators are especially important for content damage. • Potential gain of policies that increase awareness of flood risks.

  3. NBS1 and multiple regulations of DNA damage response

    PubMed Central

    Komatsu, Kenshi

    2016-01-01

    DNA damage response is finely tuned, with several pathways including those for DNA repair, chromatin remodeling and cell cycle checkpoint, although most studies to date have focused on single pathways. Genetic diseases characterized by genome instability have provided novel insights into the underlying mechanisms of DNA damage response. NBS1, a protein responsible for the radiation-sensitive autosomal recessive disorder Nijmegen breakage syndrome, is one of the first factors to accumulate at sites of DNA double-strand breaks (DSBs). NBS1 binds to at least five key proteins, including ATM, RPA, MRE11, RAD18 and RNF20, in the conserved regions within a limited span of the C terminus, functioning in the regulation of chromatin remodeling, cell cycle checkpoint and DNA repair in response to DSBs. In this article, we reviewed the functions of these binding proteins and their comprehensive association with NBS1. PMID:27068998

  4. Sigma Receptor Binding Assays.

    PubMed

    Chu, Uyen B; Ruoho, Arnold E

    2015-12-08

    Sigma receptors, both Sigma-1(S1R) and Sigma-2 (S2R), are small molecule-regulated, primarily endoplasmic reticulum (ER) membrane-associated sites. A number of drugs bind to sigma receptors, including the antipsychotic haloperidol and (+)-pentazocine, an opioid analgesic. Sigma receptors are implicated in many central nervous system disorders, in particular Alzheimer's disease and conditions associated with motor control, such as Amyotrophic Lateral Sclerosis (ALS). Described in this unit are radioligand binding assays used for the pharmacological characterization of S1R and S2R. Methods detailed include a radioligand saturation binding assay for defining receptor densities and a competitive inhibition binding assay employing [³H]-(+)-pentazocine for identifying and characterizing novel ligands that interact with S1R. Procedures using [³H]-1,3-di(2-tolyl)guanidine ([³H]-DTG), a nonselective sigma receptor ligand, are described for conducting a saturation binding and competitive inhibition assays for the S2R site. These protocols are of value in drug discovery in identifying new sigma ligands and in the characterization of these receptors.

  5. High-affinity RNA binding by a hyperthermophilic single-stranded DNA-binding protein.

    PubMed

    Morten, Michael J; Gamsjaeger, Roland; Cubeddu, Liza; Kariawasam, Ruvini; Peregrina, Jose; Penedo, J Carlos; White, Malcolm F

    2017-03-01

    Single-stranded DNA-binding proteins (SSBs), including replication protein A (RPA) in eukaryotes, play a central role in DNA replication, recombination, and repair. SSBs utilise an oligonucleotide/oligosaccharide-binding (OB) fold domain to bind DNA, and typically oligomerise in solution to bring multiple OB fold domains together in the functional SSB. SSBs from hyperthermophilic crenarchaea, such as Sulfolobus solfataricus, have an unusual structure with a single OB fold coupled to a flexible C-terminal tail. The OB fold resembles those in RPA, whilst the tail is reminiscent of bacterial SSBs and mediates interaction with other proteins. One paradigm in the field is that SSBs bind specifically to ssDNA and much less strongly to RNA, ensuring that their functions are restricted to DNA metabolism. Here, we use a combination of biochemical and biophysical approaches to demonstrate that the binding properties of S. solfataricus SSB are essentially identical for ssDNA and ssRNA. These features may represent an adaptation to a hyperthermophilic lifestyle, where DNA and RNA damage is a more frequent event.

  6. Radiation damage in macromolecular cryocrystallography.

    PubMed

    Ravelli, Raimond B G; Garman, Elspeth F

    2006-10-01

    X-ray radiation damage to cryocooled ( approximately 100 K) macromolecular crystals has emerged as a general problem, especially since the advent of third generation synchrotron undulator sources. Interest in understanding the physical and chemical phenomena behind the observed effects is growing rapidly. The specific structural damage seen in electron density maps has to be accounted for when studying intermediates, and can sometimes be related to biological function. Radiation damage induces non-isomorphism, thus hampering traditional phasing methods. However, specific damage can also be used to obtain phases. With an increased knowledge of expected crystal lifetime, beamline characteristics and types of damage, macromolecular crystallographers might soon be able to account for radiation damage in data collection, processing and phasing.

  7. Aluminum binding by humus

    SciTech Connect

    Benedetti, M.F.; Hiemstra, T.; Riemsdijk, W. van; Kinniburgh, D.

    1996-10-01

    The need for qualitative and quantitative description of the chemical speciation of Al, in particular and other metal ions in general, is stressed by the increased mobilization of metal ions in water and soils due to acid rain deposition. In this paper we present new data of Al binding to two humic acids. These new data sets and the some previously published data will be analyzed with the NICA-Donnan model using one set of parameters to describe the Al binding to the different humic substances. Once the experimental data is described with the NICA-Donnan approach, we will show the effect of Ca on Al binding and surface speciation as well as the effect of Al on the charge of the humic particles. The parameters derived from the laboratory experiments will be used to describe the variation of the field based Al partition coefficient.

  8. The nucleosome: orchestrating DNA damage signaling and repair within chromatin.

    PubMed

    Agarwal, Poonam; Miller, Kyle M

    2016-10-01

    DNA damage occurs within the chromatin environment, which ultimately participates in regulating DNA damage response (DDR) pathways and repair of the lesion. DNA damage activates a cascade of signaling events that extensively modulates chromatin structure and organization to coordinate DDR factor recruitment to the break and repair, whilst also promoting the maintenance of normal chromatin functions within the damaged region. For example, DDR pathways must avoid conflicts between other DNA-based processes that function within the context of chromatin, including transcription and replication. The molecular mechanisms governing the recognition, target specificity, and recruitment of DDR factors and enzymes to the fundamental repeating unit of chromatin, i.e., the nucleosome, are poorly understood. Here we present our current view of how chromatin recognition by DDR factors is achieved at the level of the nucleosome. Emerging evidence suggests that the nucleosome surface, including the nucleosome acidic patch, promotes the binding and activity of several DNA damage factors on chromatin. Thus, in addition to interactions with damaged DNA and histone modifications, nucleosome recognition by DDR factors plays a key role in orchestrating the requisite chromatin response to maintain both genome and epigenome integrity.

  9. Prediction of tissue thermal damage.

    PubMed

    Li, Xin; Zhong, Yongmin; Subic, Aleksandar; Jazar, Reza; Smith, Julian; Gu, Chengfan

    2016-04-29

    This paper presents a method to characterize tissue thermal damage by taking into account the thermal-mechanical effect of soft tissues for thermal ablation. This method integrates the bio-heating conduction and non-rigid motion dynamics to describe thermal-mechanical behaviors of soft tissues and further extends the traditional tissue damage model to characterize thermal-mechanical damage of soft tissues. Simulations and comparison analysis demonstrate that the proposed method can effectively predict tissue thermal damage and it also provides reliable guidelines for control of the thermal ablation procedure.

  10. Inhibition of selectin binding

    DOEpatents

    Nagy, Jon O.; Spevak, Wayne R.; Dasgupta, Falguni; Bertozzi, Carolyn

    1999-10-05

    This invention provides a system for inhibiting the binding between two cells, one expressing P- or L-selectin on the surface and the other expressing the corresponding ligand. A covalently crosslinked lipid composition is prepared having saccharides and acidic group on separate lipids. The composition is then interposed between the cells so as to inhibit binding. Inhibition can be achieved at an effective oligosaccharide concentration as low as 10.sup.6 fold below that of the free saccharide. Since selectins are involved in recruiting cells to sites of injury, this system can be used to palliate certain inflammatory and immunological conditions.

  11. Inhibition of selectin binding

    DOEpatents

    Nagy, Jon O.; Spevak, Wayne R.; Dasgupta, Falguni; Bertozzi, Caroline

    2001-10-09

    This invention provides compositions for inhibiting the binding between two cells, one expressing P- or L-selectin on the surface and the other expressing the corresponding ligand. A covalently crosslinked lipid composition is prepared having saccharides and acidic group on separate lipids. The composition is then interposed between the cells so as to inhibit binding. Inhibition can be achieved at an effective oligosaccharide concentration as low as 10.sup.6 fold below that of the free saccharide. Since selectins are involved in recruiting cells to sites of injury, these composition scan be used to palliate certain inflammatory and immunological conditions.

  12. Inhibition of selectin binding

    DOEpatents

    Nagy, Jon O.; Spevak, Wayne R.; Dasgupta, Falguni; Bertozzi, Caroline

    1999-01-01

    This invention provides compositions for inhibiting the binding between two cells, one expressing P- or L-selectin on the surface and the other expressing the corresponding ligand. A covalently crosslinked lipid composition is prepared having saccharides and acidic group on separate lipids. The composition is then interposed between the cells so as to inhibit binding. Inhibition can be achieved at an effective oligosaccharide concentration as low as 10.sup.6 fold below that of the free saccharide. Since selectins are involved in recruiting cells to sites of injury, these composition scan be used to palliate certain inflammatory and immunological conditions.

  13. Glycosylases utilize ``stop and go'' motion to locate DNA damage

    NASA Astrophysics Data System (ADS)

    Nelson, Shane

    2015-03-01

    Oxidative damage to DNA results in alterations that are mutagenic or even cytotoxic. Base excision repair is a mechanism that functions to identify and correct these lesions, and is present in organisms ranging from bacteria to humans. DNA glycosylases are the first enzymes in this pathway and function to locate and remove oxidatively damaged bases, and do so utilizing only thermal energy. However, the question remains of how these enzymes locate and recognize a damaged base among millions of undamaged bases. Utilizing fluorescence video microscopy with high spatial and temporal resolution, we have observed a number of different fluorescently labeled glycosylases (including bacterial FPG, NEI, and NTH as well as mammalian MutyH and OGG). These enzymes diffuse along DNA tightropes at approximately 0.01 +/- 0.005 μm2/s with binding lifetimes ranging from one second to several minutes. Chemically induced damage to the DNA substrate causes a ~ 50% reduction in diffusion coefficients and a ~ 400% increase in binding lifetimes, while mutation of the key ``wedge residue'' - which has been shown to be responsible for damage detection - results in a 200% increase in the diffusion coefficient. Utilizing a sliding window approach to measure diffusion coefficients within individual trajectories, we observe that distributions of diffusion coefficients are bimodal, consistent with periods of diffusive motion interspersed with immobile periods. Utilizing a unique chemo-mechanical simulation approach, we demonstrate that the motion of these glycosylases can be explained as free diffusion along the helical pitch of the DNA, punctuated with two different types of pauses: 1) rapid, short-lived pauses as the enzyme rapidly probes DNA bases to interrogate for damage and, 2) less frequent, longer lived pauses that reflect the enzyme bound to and catalytically removing a damaged base. These simulations also indicate that the wedge residue is critical for interrogation and recognition of

  14. Road Damage Following Earthquake

    NASA Technical Reports Server (NTRS)

    1989-01-01

    Ground shaking triggered liquefaction in a subsurface layer of water-saturated sand, producing differential lateral and vertical movement in a overlying carapace of unliquified sand and slit, which moved from right to left towards the Pajaro River. This mode of ground failure, termed lateral spreading, is a principal cause of liquefaction-related earthquake damage caused by the Oct. 17, 1989, Loma Prieta earthquake. Sand and soil grains have faces that can cause friction as they roll and slide against each other, or even cause sticking and form small voids between grains. This complex behavior can cause soil to behave like a liquid under certain conditions such as earthquakes or when powders are handled in industrial processes. Mechanics of Granular Materials (MGM) experiments aboard the Space Shuttle use the microgravity of space to simulate this behavior under conditons that carnot be achieved in laboratory tests on Earth. MGM is shedding light on the behavior of fine-grain materials under low effective stresses. Applications include earthquake engineering, granular flow technologies (such as powder feed systems for pharmaceuticals and fertilizers), and terrestrial and planetary geology. Nine MGM specimens have flown on two Space Shuttle flights. Another three are scheduled to fly on STS-107. The principal investigator is Stein Sture of the University of Colorado at Boulder. Credit: S.D. Ellen, U.S. Geological Survey

  15. Shock Initiation of Damaged Explosives

    SciTech Connect

    Chidester, S K; Vandersall, K S; Tarver, C M

    2009-10-22

    Explosive and propellant charges are subjected to various mechanical and thermal insults that can increase their sensitivity over the course of their lifetimes. To quantify this effect, shock initiation experiments were performed on mechanically and thermally damaged LX-04 (85% HMX, 15% Viton by weight) and PBX 9502 (95% TATB, 5% Kel-F by weight) to obtain in-situ manganin pressure gauge data and run distances to detonation at various shock pressures. We report the behavior of the HMX-based explosive LX-04 that was damaged mechanically by applying a compressive load of 600 psi for 20,000 cycles, thus creating many small narrow cracks, or by cutting wedge shaped parts that were then loosely reassembled, thus creating a few large cracks. The thermally damaged LX-04 charges were heated to 190 C for long enough for the beta to delta solid - solid phase transition to occur, and then cooled to ambient temperature. Mechanically damaged LX-04 exhibited only slightly increased shock sensitivity, while thermally damaged LX-04 was much more shock sensitive. Similarly, the insensitive explosive PBX 9502 was mechanically damaged using the same two techniques. Since PBX 9502 does not undergo a solid - solid phase transition but does undergo irreversible or 'rachet' growth when thermally cycled, thermal damage to PBX 9502 was induced by this procedure. As for LX-04, the thermally damaged PBX 9502 demonstrated a greater shock sensitivity than mechanically damaged PBX 9502. The Ignition and Growth reactive flow model calculated the increased sensitivities by igniting more damaged LX-04 and PBX 9502 near the shock front based on the measured densities (porosities) of the damaged charges.

  16. MD-2 binds cholesterol.

    PubMed

    Choi, Soo-Ho; Kim, Jungsu; Gonen, Ayelet; Viriyakosol, Suganya; Miller, Yury I

    2016-02-19

    Cholesterol is a structural component of cellular membranes, which is transported from liver to peripheral cells in the form of cholesterol esters (CE), residing in the hydrophobic core of low-density lipoprotein. Oxidized CE (OxCE) is often found in plasma and in atherosclerotic lesions of subjects with cardiovascular disease. Our earlier studies have demonstrated that OxCE activates inflammatory responses in macrophages via toll-like receptor-4 (TLR4). Here we demonstrate that cholesterol binds to myeloid differentiation-2 (MD-2), a TLR4 ancillary molecule, which is a binding receptor for bacterial lipopolysaccharide (LPS) and is indispensable for LPS-induced TLR4 dimerization and signaling. Cholesterol binding to MD-2 was competed by LPS and by OxCE-modified BSA. Furthermore, soluble MD-2 in human plasma and MD-2 in mouse atherosclerotic lesions carried cholesterol, the finding supporting the biological significance of MD-2 cholesterol binding. These results help understand the molecular basis of TLR4 activation by OxCE and mechanisms of chronic inflammation in atherosclerosis.

  17. MD-2 binds cholesterol

    PubMed Central

    Choi, Soo-Ho; Kim, Jungsu; Gonen, Ayelet; Viriyakosol, Suganya; Miller, Yury I.

    2016-01-01

    Cholesterol is a structural component of cellular membranes, which is transported from liver to peripheral cells in the form of cholesterol esters (CE), residing in the hydrophobic core of low-density lipoprotein. Oxidized CE (OxCE) is often found in plasma and in atherosclerotic lesions of subjects with cardiovascular disease. Our earlier studies have demonstrated that OxCE activates inflammatory responses in macrophages via toll-like receptor-4 (TLR4). Here we demonstrate that cholesterol binds to myeloid differentiation-2 (MD-2), a TLR4 ancillary molecule, which is a binding receptor for bacterial lipopolysaccharide (LPS) and is indispensable for LPS-induced TLR4 dimerization and signaling. Cholesterol binding to MD-2 was competed by LPS and by OxCE-modified BSA. Furthermore, soluble MD-2 in human plasma and MD-2 in mouse atherosclerotic lesions carried cholesterol, the finding supporting the biological significance of MD-2 cholesterol binding. These results help understand the molecular basis of TLR4 activation by OxCE and mechanisms of chronic inflammation in atherosclerosis. PMID:26806306

  18. Cellulose binding domain proteins

    SciTech Connect

    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.

  19. 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.

  20. Sequential memory: Binding dynamics

    NASA Astrophysics Data System (ADS)

    Afraimovich, Valentin; Gong, Xue; Rabinovich, Mikhail

    2015-10-01

    Temporal order memories are critical for everyday animal and human functioning. Experiments and our own experience show that the binding or association of various features of an event together and the maintaining of multimodality events in sequential order are the key components of any sequential memories—episodic, semantic, working, etc. We study a robustness of binding sequential dynamics based on our previously introduced model in the form of generalized Lotka-Volterra equations. In the phase space of the model, there exists a multi-dimensional binding heteroclinic network consisting of saddle equilibrium points and heteroclinic trajectories joining them. We prove here the robustness of the binding sequential dynamics, i.e., the feasibility phenomenon for coupled heteroclinic networks: for each collection of successive heteroclinic trajectories inside the unified networks, there is an open set of initial points such that the trajectory going through each of them follows the prescribed collection staying in a small neighborhood of it. We show also that the symbolic complexity function of the system restricted to this neighborhood is a polynomial of degree L - 1, where L is the number of modalities.

  1. SIGMA RECEPTOR BINDING ASSAYS

    PubMed Central

    CHU, UYEN B.; RUOHO, ARNOLD E.

    2016-01-01

    Sigma receptors belong to a class of small molecule-regulated, primarily endoplasmic reticulum (ER) membrane-associated receptors, of which there are two subtypes: the Sigma-1 receptor (S1R) and the Sigma-2 receptor (S2R). Both S1R and S2R bind to a number of drugs including antipsychotic, haloperidol, and the opioid analgesic, (+)-pentazocine. Sigma receptors are implicated in multiple disease pathologies associated with the nervous system including diseases affecting motor control such as Amyotrophic Lateral Sclerosis (ALS) and Alzeimher's disease. This unit describes methods for the pharmacological characterization of S1R and S2R using radioligand-binding assays. In the first section, radioligand saturation binding assay to determine receptor densities and competitive inhibition assays to characterize affinities of novel compounds are presented for S1R using the selective S1R ligand, [3H]-(+)-pentazocine. The second section describes radioligand saturation binding assay and competitive inhibition assays for the S2R using a non-selective S1R and S2R ligand, [3H]-1,3-di(2-tolyl)guanidine ([3H]-DTG). PMID:26646191

  2. Targeted DNA damage at individual telomeres disrupts their integrity and triggers cell death

    PubMed Central

    Sun, Luxi; Tan, Rong; Xu, Jianquan; LaFace, Justin; Gao, Ying; Xiao, Yanchun; Attar, Myriam; Neumann, Carola; Li, Guo-Min; Su, Bing; Liu, Yang; Nakajima, Satoshi; Levine, Arthur S.; Lan, Li

    2015-01-01

    Cellular DNA is organized into chromosomes and capped by a unique nucleoprotein structure, the telomere. Both oxidative stress and telomere shortening/dysfunction cause aging-related degenerative pathologies and increase cancer risk. However, a direct connection between oxidative damage to telomeric DNA, comprising <1% of the genome, and telomere dysfunction has not been established. By fusing the KillerRed chromophore with the telomere repeat binding factor 1, TRF1, we developed a novel approach to generate localized damage to telomere DNA and to monitor the real time damage response at the single telomere level. We found that DNA damage at long telomeres in U2OS cells is not repaired efficiently compared to DNA damage in non-telomeric regions of the same length in heterochromatin. Telomeric DNA damage shortens the average length of telomeres and leads to cell senescence in HeLa cells and cell death in HeLa, U2OS and IMR90 cells, when DNA damage at non-telomeric regions is undetectable. Telomere-specific damage induces chromosomal aberrations, including chromatid telomere loss and telomere associations, distinct from the damage induced by ionizing irradiation. Taken together, our results demonstrate that oxidative damage induces telomere dysfunction and underline the importance of maintaining telomere integrity upon oxidative damage. PMID:26082495

  3. The oxidative damage initiation hypothesis for meiosis.

    PubMed

    Hörandl, Elvira; Hadacek, Franz

    2013-12-01

    The maintenance of sexual reproduction in eukaryotes is still a major enigma in evolutionary biology. Meiosis represents the only common feature of sex in all eukaryotic kingdoms, and thus, we regard it a key issue for discussing its function. Almost all asexuality modes maintain meiosis either in a modified form or as an alternative pathway, and facultatively apomictic plants increase frequencies of sexuality relative to apomixis after abiotic stress. On the physiological level, abiotic stress causes oxidative stress. We hypothesize that repair of oxidative damage on nuclear DNA could be a major driving force in the evolution of meiosis. We present a hypothetical model for the possible redox chemistry that underlies the binding of the meiosis-specific protein Spo11 to DNA. During prophase of meiosis I, oxidized sites at the DNA molecule are being targeted by the catalytic tyrosine moieties of Spo11 protein, which acts like an antioxidant reducing the oxidized target. The oxidized tyrosine residues, tyrosyl radicals, attack the phosphodiester bonds of the DNA backbone causing DNA double strand breaks that can be repaired by various mechanisms. Polyploidy in apomictic plants could mitigate oxidative DNA damage and decrease Spo11 activation. Our hypothesis may contribute to explaining various enigmatic phenomena: first, DSB formation outnumbers crossovers and, thus, effective recombination events by far because the target of meiosis may be the removal of oxidative lesions; second, it offers an argument for why expression of sexuality is responsive to stress in many eukaryotes; and third, repair of oxidative DNA damage turns meiosis into an essential characteristic of eukaryotic reproduction.

  4. The ATM Kinase Induces MicroRNA Biogenesis in the DNA Damage Response

    PubMed Central

    Zhang, Xinna; Wan, Guohui; Berger, Franklin G.; He, Xiaoming; Lu, Xiongbin

    2011-01-01

    SUMMARY The DNA damage response involves a complex network of processes that detect and repair DNA damage. Here we show that miRNA biogenesis is globally induced upon DNA damage in an ATM-dependent manner. About one fourth of miRNAs are significantly up-regulated after DNA damage, while loss of ATM abolishes their induction. KSRP (KH-type splicing regulatory protein) is a key player that translates DNA damage signaling to miRNA biogenesis. The ATM kinase directly binds to and phosphorylates KSRP, leading to enhanced interaction between KSRP and pri-miRNAs and increased KSRP activity in miRNA processing. Mutations of the ATM phosphorylation sites of KSRP impaired its activity in regulating miRNAs. These findings reveal a mechanism by which DNA damage signaling is linked to miRNA biogenesis. PMID:21329876

  5. Real-Time Tracking of Parental Histones Reveals Their Contribution to Chromatin Integrity Following DNA Damage.

    PubMed

    Adam, Salomé; Dabin, Juliette; Chevallier, Odile; Leroy, Olivier; Baldeyron, Céline; Corpet, Armelle; Lomonte, Patrick; Renaud, Olivier; Almouzni, Geneviève; Polo, Sophie E

    2016-10-06

    Chromatin integrity is critical for cell function and identity but is challenged by DNA damage. To understand how chromatin architecture and the information that it conveys are preserved or altered following genotoxic stress, we established a system for real-time tracking of parental histones, which characterize the pre-damage chromatin state. Focusing on histone H3 dynamics after local UVC irradiation in human cells, we demonstrate that parental histones rapidly redistribute around damaged regions by a dual mechanism combining chromatin opening and histone mobilization on chromatin. Importantly, parental histones almost entirely recover and mix with new histones in repairing chromatin. Our data further define a close coordination of parental histone dynamics with DNA repair progression through the damage sensor DDB2 (DNA damage-binding protein 2). We speculate that this mechanism may contribute to maintaining a memory of the original chromatin landscape and may help preserve epigenome stability in response to DNA damage.

  6. Molecular level detection and localization of mechanical damage in collagen enabled by collagen hybridizing peptides

    NASA Astrophysics Data System (ADS)

    Zitnay, Jared L.; Li, Yang; Qin, Zhao; San, Boi Hoa; Depalle, Baptiste; Reese, Shawn P.; Buehler, Markus J.; Yu, S. Michael; Weiss, Jeffrey A.

    2017-03-01

    Mechanical injury to connective tissue causes changes in collagen structure and material behaviour, but the role and mechanisms of molecular damage have not been established. In the case of mechanical subfailure damage, no apparent macroscale damage can be detected, yet this damage initiates and potentiates in pathological processes. Here, we utilize collagen hybridizing peptide (CHP), which binds unfolded collagen by triple helix formation, to detect molecular level subfailure damage to collagen in mechanically stretched rat tail tendon fascicle. Our results directly reveal that collagen triple helix unfolding occurs during tensile loading of collagenous tissues and thus is an important damage mechanism. Steered molecular dynamics simulations suggest that a likely mechanism for triple helix unfolding is intermolecular shearing of collagen α-chains. Our results elucidate a probable molecular failure mechanism associated with subfailure injuries, and demonstrate the potential of CHP targeting for diagnosis, treatment and monitoring of tissue disease and injury.

  7. Molecular level detection and localization of mechanical damage in collagen enabled by collagen hybridizing peptides

    PubMed Central

    Zitnay, Jared L.; Li, Yang; Qin, Zhao; San, Boi Hoa; Depalle, Baptiste; Reese, Shawn P.; Buehler, Markus J.; Yu, S. Michael; Weiss, Jeffrey A.

    2017-01-01

    Mechanical injury to connective tissue causes changes in collagen structure and material behaviour, but the role and mechanisms of molecular damage have not been established. In the case of mechanical subfailure damage, no apparent macroscale damage can be detected, yet this damage initiates and potentiates in pathological processes. Here, we utilize collagen hybridizing peptide (CHP), which binds unfolded collagen by triple helix formation, to detect molecular level subfailure damage to collagen in mechanically stretched rat tail tendon fascicle. Our results directly reveal that collagen triple helix unfolding occurs during tensile loading of collagenous tissues and thus is an important damage mechanism. Steered molecular dynamics simulations suggest that a likely mechanism for triple helix unfolding is intermolecular shearing of collagen α-chains. Our results elucidate a probable molecular failure mechanism associated with subfailure injuries, and demonstrate the potential of CHP targeting for diagnosis, treatment and monitoring of tissue disease and injury. PMID:28327610

  8. Damage Progression in Bolted Composites

    NASA Technical Reports Server (NTRS)

    Minnetyan, Levon; Chamis, Christos; Gotsis, Pascal K.

    1998-01-01

    Structural durability,damage tolerance,and progressive fracture characteristics of bolted graphite/epoxy composite laminates are evaluated via computational simulation. Constituent material properties and stress and strain limits are scaled up to the structure level to evaluate the overall damage and fracture propagation for bolted composites. Single and double bolted composite specimens with various widths and bolt spacings are evaluated. The effect of bolt spacing is investigated with regard to the structural durability of a bolted joint. Damage initiation, growth, accumulation, and propagation to fracture are included in the simulations. Results show the damage progression sequence and structural fracture resistance during different degradation stages. A procedure is outlined for the use of computational simulation data in the assessment of damage tolerance, determination of sensitive parameters affecting fracture, and interpretation of experimental results with insight for design decisions.

  9. Damage Progression in Bolted Composites

    NASA Technical Reports Server (NTRS)

    Minnetyan, Levon; Chamis, Christos C.; Gotsis, Pascal K.

    1998-01-01

    Structural durability, damage tolerance, and progressive fracture characteristics of bolted graphite/epoxy composite laminates are evaluated via computational simulation. Constituent material properties and stress and strain limits are scaled up to the structure level to evaluate the overall damage and fracture propagation for bolted composites. Single and double bolted composite specimens with various widths and bolt spacings are evaluated. The effect of bolt spacing is investigated with regard to the structural durability of a bolted joint. Damage initiation, growth, accumulation, and propagation to fracture are included in the simulations. Results show the damage progression sequence and structural fracture resistance during different degradation stages. A procedure is outlined for the use of computational simulation data in the assessment of damage tolerance, determination of sensitive parameters affecting fracture, and interpretation of experimental results with insight for design decisions.

  10. Cellular Concentrations of DDB2 Regulate Dynamic Binding of DDB1 at UV-Induced DNA Damage▿

    PubMed Central

    Alekseev, Sergey; Luijsterburg, Martijn S.; Pines, Alex; Geverts, Bart; Mari, Pierre-Olivier; Giglia-Mari, Giuseppina; Lans, Hannes; Houtsmuller, Adriaan B.; Mullenders, Leon H. F.; Hoeijmakers, Jan H. J.; Vermeulen, Wim

    2008-01-01

    Nucleotide excision repair (NER) is the principal pathway for counteracting cytotoxic and mutagenic effects of UV irradiation. To provide insight into the in vivo regulation of the DNA damage recognition step of global genome NER (GG-NER), we constructed cell lines expressing fluorescently tagged damaged DNA binding protein 1 (DDB1). DDB1 is a core subunit of a number of cullin 4-RING ubiquitin ligase complexes. UV-activated DDB1-DDB2-CUL4A-ROC1 ubiquitin ligase participates in the initiation of GG-NER and triggers the UV-dependent degradation of its subunit DDB2. We found that DDB1 rapidly accumulates on DNA damage sites. However, its binding to damaged DNA is not static, since DDB1 constantly dissociates from and binds to DNA lesions. DDB2, but not CUL4A, was indispensable for binding of DDB1 to DNA damage sites. The residence time of DDB1 on the damage site is independent of the main damage-recognizing protein of GG-NER, XPC, as well as of UV-induced proteolysis of DDB2. The amount of DDB1 that is temporally immobilized on damaged DNA critically depends on DDB2 levels in the cell. We propose a model in which UV-dependent degradation of DDB2 is important for the release of DDB1 from continuous association to unrepaired DNA and makes DDB1 available for its other DNA damage response functions. PMID:18936169

  11. Binding energy effects in cascade evolution and sputtering

    SciTech Connect

    Robinson, M.T.

    1995-06-01

    The MARLOWE model was extended to include a binding energy dependent on the local crystalline order, so that atoms are bound less strongly to their lattice sites near surfaces or associated damage. Sputtering and cascade evolution were studied on the examples of self-ion irradiations of Cu and Au monocrystals. In cascades, the mean binding energy is reduced {approximately}8% in Cu with little dependence on the initial recoil energy; in Au, it is reduced {approximately}9% at 1 keV and {approximately}15% at 100 keV. In sputtering, the mean binding energy is reduced {approximately}8% in Cu and {approximately}15% in Au with little energy dependence; the yields are increased about half as much. Most sites from which sputtered atoms originate are isolated in both metals. Small clusters of such sites occur in Cu, but there are some large clusters in Au, especially in [111] targets. There are always more large clusters with damage-dependent binding than with a constant binding energy, but only a few clusters are compact enough to be regarded as pits.

  12. Molecular Evolution of the Oxygen-Binding Hemerythrin Domain

    PubMed Central

    Alvarez-Carreño, Claudia; Becerra, Arturo; Lazcano, Antonio

    2016-01-01

    Background The evolution of oxygenic photosynthesis during Precambrian times entailed the diversification of strategies minimizing reactive oxygen species-associated damage. Four families of oxygen-carrier proteins (hemoglobin, hemerythrin and the two non-homologous families of arthropodan and molluscan hemocyanins) are known to have evolved independently the capacity to bind oxygen reversibly, providing cells with strategies to cope with the evolutionary pressure of oxygen accumulation. Oxygen-binding hemerythrin was first studied in marine invertebrates but further research has made it clear that it is present in the three domains of life, strongly suggesting that its origin predated the emergence of eukaryotes. Results Oxygen-binding hemerythrins are a monophyletic sub-group of the hemerythrin/HHE (histidine, histidine, glutamic acid) cation-binding domain. Oxygen-binding hemerythrin homologs were unambiguously identified in 367/2236 bacterial, 21/150 archaeal and 4/135 eukaryotic genomes. Overall, oxygen-binding hemerythrin homologues were found in the same proportion as single-domain and as long protein sequences. The associated functions of protein domains in long hemerythrin sequences can be classified in three major groups: signal transduction, phosphorelay response regulation, and protein binding. This suggests that in many organisms the reversible oxygen-binding capacity was incorporated in signaling pathways. A maximum-likelihood tree of oxygen-binding hemerythrin homologues revealed a complex evolutionary history in which lateral gene transfer, duplications and gene losses appear to have played an important role. Conclusions Hemerythrin is an ancient protein domain with a complex evolutionary history. The distinctive iron-binding coordination site of oxygen-binding hemerythrins evolved first in prokaryotes, very likely prior to the divergence of Firmicutes and Proteobacteria, and spread into many bacterial, archaeal and eukaryotic species. The later

  13. A systematic analysis of factors localized to damaged chromatin reveals PARP-dependent recruitment of transcription factors

    PubMed Central

    Izhar, Lior; Adamson, Britt; Ciccia, Alberto; Lewis, Jedd; Pontano-Vaites, Laura; Leng, Yumei; Liang, Anthony C.; Westbrook, Thomas F.; Harper, J. Wade; Elledge, Stephen J.

    2015-01-01

    Localization to sites of DNA damage is a hallmark of DNA damage response (DDR) proteins. To identify new DDR factors, we screened epitope-tagged proteins for localization to sites of chromatin damaged by UV laser microirradiation and found >120 proteins that localize to damaged chromatin. These include the BAF tumor suppressor complex and the ALS candidate protein TAF15. TAF15 contains multiple domains that bind damaged chromatin in a PARP-dependent manner, suggesting a possible role as glue that tethers multiple PAR chains together. Many positives were transcription factors and >70% of randomly tested transcription factors localized to sites of DNA damage and approximately 90% were PARP-dependent for localization. Mutational analyses showed that localization to damaged chromatin is DNA-binding domain-dependent. By examining Hoechst staining patterns at damage sites, we see evidence of chromatin decompaction that is PARP-dependent. We propose that PARP-regulated chromatin remodeling at sites of damage allows transient accessibility of DNA-binding proteins. PMID:26004182

  14. Equivalent damage: A critical assessment

    NASA Technical Reports Server (NTRS)

    Laflen, J. R.; Cook, T. S.

    1982-01-01

    Concepts in equivalent damage were evaluated to determine their applicability to the life prediction of hot path components of aircraft gas turbine engines. Equivalent damage was defined as being those effects which influence the crack initiation life-time beyond the damage that is measured in uniaxial, fully-reversed sinusoidal and isothermal experiments at low homologous temperatures. Three areas of equivalent damage were examined: mean stress, cumulative damage, and multiaxiality. For each area, a literature survey was conducted to aid in selecting the most appropriate theories. Where possible, data correlations were also used in the evaluation process. A set of criteria was developed for ranking the theories in each equivalent damage regime. These criteria considered aspects of engine utilization as well as the theoretical basis and correlative ability of each theory. In addition, consideration was given to the complex nature of the loading cycle at fatigue critical locations of hot path components; this loading includes non-proportional multiaxial stressing, combined temperature and strain fluctuations, and general creep-fatigue interactions. Through applications of selected equivalent damage theories to some suitable data sets it was found that there is insufficient data to allow specific recommendations of preferred theories for general applications. A series of experiments and areas of further investigations were identified.

  15. Estimating bird damage from damage incidence in wine grape vineyards

    USGS Publications Warehouse

    DeHaven, R.W.; Hothem, R.L.

    1981-01-01

    Bird damage was measured during 1977 and 1978 at 32 wine grape vineyards in the San Joaquin Valley and North Coastal Region of California. Both the percentage bird loss (PBL) and the percentage of bunches damaged (BDI = bird damage incidence) were determined during 55 total-damage assessments, and the resulting data pairs were used to develop a regression of PBL on BDI. The final prediction equation was loge (PBL + 1) = 0.0385 BDI, for which the SE = 9.6297 10-4, and it accounted for 97% of the observed variation. We conclude that by using that equation, reasonably accurate predictions of PBL can be obtained from relatively quick and inexpensive estimates of BDI. Guidelines for the use of the prediction method and the accuracy of some PBL predictions are discussed.

  16. MMOD Impact Damage to ISS

    NASA Technical Reports Server (NTRS)

    Hyde, James L.; Christiansen, Eric; Lear, Dana M.

    2014-01-01

    Paper will describe micrometeoroid and orbital debris (MMOD) damage that has been observed on the International Space Station (ISS). Several hundred documented MMOD damage sites on ISS have been identified through imagery from the windows of ISS modules or docked vehicles. Sites that are observable from ISS or shuttle windows exhibiting distinct features of hypervelocity impact damage are usually greater than 5mm in diameter. Many smaller features are revealed in on-orbit imagery are typically less distinct and difficult to characterize but could be MMOD damage. Additional images of on-orbit damage features have been collected by astronauts during extra vehicular activities. Ground inspection of returned ISS hardware has also contributed to the database of ISS MMOD impact damage. A handful of orbital replacement units (ORU) from the ISS active thermal control and electrical power subsystems were swapped out and returned during the Space Shuttle program. In addition, a reusable logistics module was deployed on ISS for a total 59.4 days on 11 shuttle missions between 2001 and 2011 and then brought back in the shuttle payload bay. All of this returned hardware was subjected to detailed post-flight inspections for MMOD damage, and a database with nearly 1000 impact records has been collected. A description of the largest observed damages will be provided in the paper. In addition, a discussion of significant MMOD impact sites with operational or design aspects will be presented. Some of the ISS modules/subsystems damaged by MMOD to be included in the discussion are (1) Solar Arrays, (2) US and Russian windows, (3) EVA handrails, (4) Radiators, and (5) Russian FGB module.

  17. Earthquake damage to transportation systems

    USGS Publications Warehouse

    McCullough, Heather

    1994-01-01

    Earthquakes represent one of the most destructive natural hazards known to man. A large magnitude earthquake near a populated area can affect residents over thousands of square kilometers and cause billions of dollars in property damage. Such an event can kill or injure thousands of residents and disrupt the socioeconomic environment for months, sometimes years. A serious result of a large-magnitude earthquake is the disruption of transportation systems, which limits post-disaster emergency response. Movement of emergency vehicles, such as police cars, fire trucks and ambulances, is often severely restricted. Damage to transportation systems is categorized below by cause including: ground failure, faulting, vibration damage, and tsunamis.

  18. Structural Damage Assessment under Uncertainty

    NASA Astrophysics Data System (ADS)

    Lopez Martinez, Israel

    Structural damage assessment has applications in the majority of engineering structures and mechanical systems ranging from aerospace vehicles to manufacturing equipment. The primary goals of any structural damage assessment and health monitoring systems are to ascertain the condition of a structure and to provide an evaluation of changes as a function of time as well as providing an early-warning of an unsafe condition. There are many structural heath monitoring and assessment techniques developed for research using numerical simulations and scaled structural experiments. However, the transition from research to real-world structures has been rather slow. One major reason for this slow-progress is the existence of uncertainty in every step of the damage assessment process. This dissertation research involved the experimental and numerical investigation of uncertainty in vibration-based structural health monitoring and development of robust detection and localization methods. The basic premise of vibration-based structural health monitoring is that changes in structural characteristics, such as stiffness, mass and damping, will affect the global vibration response of the structure. The diagnostic performance of vibration-based monitoring system is affected by uncertainty sources such as measurement errors, environmental disturbances and parametric modeling uncertainties. To address diagnostic errors due to irreducible uncertainty, a pattern recognition framework for damage detection has been developed to be used for continuous monitoring of structures. The robust damage detection approach developed is based on the ensemble of dimensional reduction algorithms for improved damage-sensitive feature extraction. For damage localization, the determination of an experimental structural model was performed based on output-only modal analysis. An experimental model correlation technique is developed in which the discrepancies between the undamaged and damaged modal data are

  19. Conciliating binding efficiency and polypharmacology.

    PubMed

    Mestres, Jordi; Gregori-Puigjané, Elisabet

    2009-09-01

    The association between molecular size and risk of failure has promoted the use of binding efficiency as a prioritization metric in lead selection. Even though by extension it is often referred to as "ligand efficiency", the concept was originally conceived to be strictly applicable to comparing the binding efficiencies of ligands for a single target. With current trends in designing drugs to bind efficiently to multiple targets, a revision of the original binding efficiency definition is carried out. To this aim, the dependency of binding efficiency on polypharmacology is highlighted in a retrospective analysis of a set of antipsychotic drugs. Statistical standardization of target binding efficiencies relative to basal values obtained from a large background of medicinal chemistry compounds is proposed as a means to conciliate the concepts of binding efficiency and polypharmacology. Finally, the interplay between binding efficiency and therapeutic efficacy for optimizing natural products, random hits, and fragments is discussed.

  20. Types and Consequences of DNA Damage

    EPA Science Inventory

    This review provides a concise overview of the types of DNA damage and the molecular mechanisms by which a cell senses DNA damage, repairs the damage, converts the damage into a mutation, or dies as a consequence of unrepaired DNA damage. Such information is important in consid...

  1. The Interaction between Polynucleotide Kinase Phosphatase and the DNA Repair Protein XRCC1 Is Critical for Repair of DNA Alkylation Damage and Stable Association at DNA Damage Sites*

    PubMed Central

    Della-Maria, Julie; Hegde, Muralidhar L.; McNeill, Daniel R.; Matsumoto, Yoshihiro; Tsai, Miaw-Sheue; Ellenberger, Tom; Wilson, David M.; Mitra, Sankar; Tomkinson, Alan E.

    2012-01-01

    XRCC1 plays a key role in the repair of DNA base damage and single-strand breaks. Although it has no known enzymatic activity, XRCC1 interacts with multiple DNA repair proteins and is a subunit of distinct DNA repair protein complexes. Here we used the yeast two-hybrid genetic assay to identify mutant versions of XRCC1 that are selectively defective in interacting with a single protein partner. One XRCC1 mutant, A482T, that was defective in binding to polynucleotide kinase phosphatase (PNKP) not only retained the ability to interact with partner proteins that bind to different regions of XRCC1 but also with aprataxin and aprataxin-like factor whose binding sites overlap with that of PNKP. Disruption of the interaction between PNKP and XRCC1 did not impact their initial recruitment to localized DNA damage sites but dramatically reduced their retention there. Furthermore, the interaction between PNKP and the DNA ligase IIIα-XRCC1 complex significantly increased the efficiency of reconstituted repair reactions and was required for complementation of the DNA damage sensitivity to DNA alkylation agents of xrcc1 mutant cells. Together our results reveal novel roles for the interaction between PNKP and XRCC1 in the retention of XRCC1 at DNA damage sites and in DNA alkylation damage repair. PMID:22992732

  2. Nucleotide Accumulation Induced in Staphylococcus aureus by Glycine

    PubMed Central

    Strominger, Jack L.; Birge, Claire H.

    1965-01-01

    Strominger, Jack L. (Washington University School of Medicine, St. Louis, Mo.), and Claire H. Birge. Nucleotide accumulation induced in Staphylococcus aureus by glycine. J. Bacteriol. 89:1124–1127. 1965.—High concentrations of glycine induce accumulation of four uridine nucleotides in Staphylococcus aureus. Investigations of their structure suggest that these compounds are uridine diphosphate (UDP)-acetylmuramic acid, UDP-acetylmuramyl-gly-d-glu-l-lys, UDP-acetylmuramyl-l-ala-d-glu-l-lys and UDP-acetylmuramyl-gly-d-glu-l-lys-d-ala-d-ala. The mechanism by which glycine may induce uridine nucleotide accumulation and protoplast formation is discussed. Images PMID:14276106

  3. Optical detection of DNA damage

    NASA Astrophysics Data System (ADS)

    Rogers, Kim R.; Apostol, A.; Cembrano, J.

    1999-02-01

    A rapid and sensitive fluorescence assay for oxidative damage to calf thymus DNA is reported. A decrease in the transition temperature for strand separation resulted from exposure of the DNA to the reactive decomposition products of 3- morpholinosydnonimine (SIN-1) (i.e., nitric oxide, superoxide, peroxynitrite, hydrogen peroxide, and hydroxyl radicals). A decrease in melting temperature of 12 degrees Celsius was indicative of oxidative damage including single strand chain breaks. Double stranded (ds) and single stranded (ss) forms of DNA were determined using the indicator dyes ethidium bromide and PicoGreen. The change in DNA 'melting' curves was dependant on the concentration of SIN-1 and was most pronounced at 75 degrees Celsius. This chemically induced damage was significantly inhibited by sodium citrate, tris(hydroxymethyl)aminomethane (Tris), and diethylenetriaminepentaacetic acid (DTPA), but was unaffected by superoxide dismutase (SOD), catalase, ethylenediamine tetraacietic acid (EDTA), or deferoxamine. Lowest observable effect level for SIN-1-induced damage was 200 (mu) M.

  4. Loss and damage post Paris

    NASA Astrophysics Data System (ADS)

    Petherick, Anna

    2016-08-01

    The Paris Agreement gave the Warsaw International Mechanism for Loss and Damage a permanent and potentially prominent place in climate negotiations, but beyond that its impact remains wide open for interpretation.

  5. Radiolytic Damage to Genetic Material.

    ERIC Educational Resources Information Center

    Ward, John F.

    1981-01-01

    Describes some basic findings in the radiation chemistry of genetic material derived from studies of model systems. Uses these findings to extrapolate the consequences of radiation damage to DNA within cells. (CS)

  6. Pain Medicines and Kidney Damage

    MedlinePlus

    ... Work Kidney Disease A-Z Pain Medicine and Kidney Damage An analgesic is any medicine intended to ... of chronic kidney disease called analgesic nephropathy. Acute Kidney Failure Some patient case reports have attributed incidents ...

  7. Excitation optimization for damage detection

    SciTech Connect

    Bement, Matthew T; Bewley, Thomas R

    2009-01-01

    A technique is developed to answer the important question: 'Given limited system response measurements and ever-present physical limits on the level of excitation, what excitation should be provided to a system to make damage most detectable?' Specifically, a method is presented for optimizing excitations that maximize the sensitivity of output measurements to perturbations in damage-related parameters estimated with an extended Kalman filter. This optimization is carried out in a computationally efficient manner using adjoint-based optimization and causes the innovations term in the extended Kalman filter to be larger in the presence of estimation errors, which leads to a better estimate of the damage-related parameters in question. The technique is demonstrated numerically on a nonlinear 2 DOF system, where a significant improvement in the damage-related parameter estimation is observed.

  8. Damaging effects of visible light

    NASA Astrophysics Data System (ADS)

    Williams, T. P.; Baker, B. N.

    1982-02-01

    The right eyes of anesthetized, ten week old albino rats are exposed to constant photon fluxes at 6 wavelengths for 6 hours. The left eye of each animal is patched during the exposure and is used as control. Histologic examination of retinal sections disclosed a region in the superior retina which is more damaged than are other areas. Attempting to ascertain an action spectrum by measuring outer nuclear layer (ONL) lost in this sensitive region fails. However, it is shown that when ONL thickness is integrated over the entire retinal sections, a rhodopsin action-spectrum emerges. It is concluded that retinal light damage in the albina rat under these conditions is rhodopsin mediated; and assessment of the extent of damage is best made by some method which integrates over the entire retinal section. The latter methodology is not routinely incorporated into studies of retinal light-damage but probably should be.

  9. BDS thin film damage competition

    SciTech Connect

    Stolz, C J; Thomas, M D; Griffin, A J

    2008-10-24

    A laser damage competition was held at the 2008 Boulder Damage Symposium in order to determine the current status of thin film laser resistance within the private, academic, and government sectors. This damage competition allows a direct comparison of the current state-of-the-art of high laser resistance coatings since they are all tested using the same damage test setup and the same protocol. A normal incidence high reflector multilayer coating was selected at a wavelength of 1064 nm. The substrates were provided by the submitters. A double blind test assured sample and submitter anonymity so only a summary of the results are presented here. In addition to the laser resistance results, details of deposition processes, coating materials, and layer count will also be shared.

  10. Chemical Protection Against Radiation Damage

    ERIC Educational Resources Information Center

    Campaigne, Ernest

    1969-01-01

    Discusses potential war time and medical uses for chemical compounds giving protection against radiation damage. Describes compounds known to protect, research aimed at discovering such compounds, and problems of toxicity. (EB)

  11. Probabilistic Fatigue Damage Program (FATIG)

    NASA Technical Reports Server (NTRS)

    Michalopoulos, Constantine

    2012-01-01

    FATIG computes fatigue damage/fatigue life using the stress rms (root mean square) value, the total number of cycles, and S-N curve parameters. The damage is computed by the following methods: (a) traditional method using Miner s rule with stress cycles determined from a Rayleigh distribution up to 3*sigma; and (b) classical fatigue damage formula involving the Gamma function, which is derived from the integral version of Miner's rule. The integration is carried out over all stress amplitudes. This software solves the problem of probabilistic fatigue damage using the integral form of the Palmgren-Miner rule. The software computes fatigue life using an approach involving all stress amplitudes, up to N*sigma, as specified by the user. It can be used in the design of structural components subjected to random dynamic loading, or by any stress analyst with minimal training for fatigue life estimates of structural components.

  12. Library Binding Manual. Revised Edition.

    ERIC Educational Resources Information Center

    Lakhanpal, S. K.

    This procedural manual is designed to be used in bindery sections in public, university and special libraries. It briefly discusses these general matters: administrative control; selection of a binder; when and what to bind; conventional binding; routines; missing issues; schedule for shipments; temporary binding; rare books, maps and newspapers;…

  13. The cardiotoxicity and myocyte damage caused by small molecule anticancer tyrosine kinase inhibitors is correlated with lack of target specificity

    SciTech Connect

    Hasinoff, Brian B.

    2010-04-15

    The use of the new anticancer tyrosine kinase inhibitors (TKI) has revolutionized the treatment of certain cancers. However, the use of some of these results in cardiotoxicity. Large-scale profiling data recently made available for the binding of 7 of the 9 FDA-approved tyrosine kinase inhibitors to a panel of 317 kinases has allowed us to correlate kinase inhibitor binding selectivity scores with TKI-induced damage to neonatal rat cardiac myocytes. The tyrosine kinase selectivity scores, but not the serine-threonine kinase scores, were highly correlated with the myocyte damaging effects of the TKIs. Additionally, we showed that damage to myocytes gave a good rank order correlation with clinical cardiotoxicity. Finally, strength of TKI binding to colony-stimulating factor 1 receptor (CSF1R) was highly correlated with myocyte damage, thus possibly implicating this kinase in contributing to TKI-induced cardiotoxicity.

  14. The contribution of collision cascades to sputtering and radiation damage.

    PubMed

    Thompson, M W

    2004-01-15

    Measurement of the velocity distribution of sputtered atoms has tested theories of sputtering and radiation damage. Experiments measuring the time of flight of sputtered atoms in precisely defined directions from the emitting surface, from which the related velocity and energy distributions may be deduced, are described. The energy distribution from polycrystalline targets often fits the form E/(E + Eb)3 derived from a theoretical model in which the bombarding ions initiate collision cascades that eject atoms through the surface. It is assumed that atoms are bound to the surface by a binding force normal to the surface, represented as a binding energy, in the formula. It is shown how this theory relates to that of radiation damage. Departures from the formula seem to correlate with high-energy density in cascades and/or low values of Eb/kT0, with T0 the target temperature. A second component then appears in the energy spectrum approximated by [formula: see text], where deltaT is an effective local temperature rise induced by cascades. The velocity distributions from single crystals are strongly affected by both the direction of ion incidence, indicative of ion channelling, and by emission in directions close to simple crystal axes, indicative of momentum focusing within cascades. Models of the cascade region, and the local heating it causes, have been deduced from sputtering experiments and have advanced our understanding of defect structures caused by radiation damage. Momentum focusing processes are active in creating interstitial-vacancy pairs in both radiation damage and sputtering and their properties have been deduced from these experiments. It is shown how the study of sputtering has enhanced the understanding of radiation damage.

  15. DNA Damage Induced Neuronal Death

    DTIC Science & Technology

    1999-10-01

    Experiments are proposed to examine the molecular mechanism by which mustard chemical warfare agents induce neuronal cell death . DNA damage is the...proposed underlying mechanism of mustard-induced neuronal cell death . We propose a novel research strategy to test this hypothesis by using mice with...perturbed DNA repair to explore the relationship between mustard-induced DNA damage and neuronal cell death . Initial in vitro studies (Years 1, 2 & 3

  16. Heat transfer in damaged material

    NASA Astrophysics Data System (ADS)

    Kruis, J.

    2013-10-01

    Fully coupled thermo-mechanical analysis of civil engineering problems is studied. The mechanical analysis is based on damage mechanics which is useful for modeling of behaviour of quasi-brittle materials, especially in tension. The damage is assumed to be isotropic. The heat transfer is assumed in the form of heat conduction governed by the Fourier law and heat radiation governed by the Stefan-Boltzmann law. Fully coupled thermo-mechanical problem is formulated.

  17. Military Robotics and Collateral Damage

    DTIC Science & Technology

    2004-06-01

    Applications and Operations Military Robotics and Collateral Damage Robert Douglass (Primary POC) SET Associates 3811 N. Fairfax...2004 4. TITLE AND SUBTITLE Military Robotics and Collateral Damage 5a. CONTRACT NUMBER 5b. GRANT NUMBER 5c. PROGRAM ELEMENT NUMBER 6. AUTHOR(S...Standard Form 298 (Rev. 8-98) Prescribed by ANSI Std Z39-18 Abstract We explore a concept of a combined force of air and ground combat robots

  18. Replicating damaged DNA in eukaryotes.

    PubMed

    Chatterjee, Nimrat; Siede, Wolfram

    2013-12-01

    DNA damage is one of many possible perturbations that challenge the mechanisms that preserve genetic stability during the copying of the eukaryotic genome in S phase. This short review provides, in the first part, a general introduction to the topic and an overview of checkpoint responses. In the second part, the mechanisms of error-free tolerance in response to fork-arresting DNA damage will be discussed in some detail.

  19. Damage assessment in systemic vasculitis.

    PubMed

    Silveira, Luis H

    2008-12-01

    Systemic vasculitides were initially reported as acute, progressive, severe, and life-threatening diseases. The introduction of glucocorticoids and cyclophosphamide for the treatment of vasculitis improved survival dramatically, but morbidity has remained high. Damage develops as a consequence of recurrent or persistent active vasculitis or its treatment. It is defined as the accumulation of nonhealing scars that are unlikely to respond to immunosuppressive therapy. Damage assessment is essential in systemic vasculitis because it may facilitate patient stratification in clinical trials and possibly in clinical practice. Moreover, it may avoid unnecessary use of immunosuppressive therapy. The Vasculitis Damage Index, developed and validated in 1997, has been very useful in solving many matters in systemic vasculitis and is currently the only validated damage-assessment tool available. However, the vasculitis community has recognized that there is a growing need to improve the evaluation of damage in vasculitis. The development of a Combined Damage Assessment index, which would permit a more appropriate and standardized approach to disease assessment applicable to systemic vasculitis, has been proposed.

  20. Americium binding to humic acid.

    PubMed

    Peters, A J; Hamilton-Taylor, J; Tipping, E

    2001-09-01

    The binding of americium (Am) by peat humic acid (PHA) has been investigated at Am concentrations between 10(-1) and 10(-7) M at pH approximately 2.6 in the presence and absence of Cu as a competing ion. Cu-PHA binding was also investigated in order to derive independent binding constants for use in modeling the competitive binding studies. Humic ion-binding model VI was used to compare the acquired data with previously published binding data and to investigate the importance of high-affinity binding sites in metal-PHA binding. Am was not observed to bind to high-affinity, low-concentration binding sites. The model VI parameter deltaLK2 takes into accountthe small number of strong sites in PHA and was found to be important for Cu-PHA binding but not for Am-PHA binding, regardless of whether Cu was present. Analysis of the PHA sample revealed that it contained a considerable quantity of Fe not removed by the extraction procedure, much of which is believed to be present as Fe(III). Model VI was then used to investigate the possible importance of the presence of Fe(III) in the Am-PHA binding experiments. When Fe(III) was assumed to be present, improved descriptions of the data by model VI were obtained by assuming that all of the metals [Am, Cu, and Fe(III)] undergo strong binding. This highlights the importance of Fe(III) competition in metal-PHA binding studies and possible shortcomings in the extraction procedure used to extract PHA.

  1. Carboplatin binding to histidine

    SciTech Connect

    Tanley, Simon W. M.; Diederichs, Kay; Kroon-Batenburg, Loes M. J.; Levy, Colin; Schreurs, Antoine M. M.; Helliwell, John R.

    2014-08-29

    An X-ray crystal structure showing the binding of purely carboplatin to histidine in a model protein has finally been obtained. This required extensive crystallization trials and various novel crystal structure analyses. Carboplatin is a second-generation platinum anticancer agent used for the treatment of a variety of cancers. Previous X-ray crystallographic studies of carboplatin binding to histidine (in hen egg-white lysozyme; HEWL) showed the partial conversion of carboplatin to cisplatin owing to the high NaCl concentration used in the crystallization conditions. HEWL co-crystallizations with carboplatin in NaBr conditions have now been carried out to confirm whether carboplatin converts to the bromine form and whether this takes place in a similar way to the partial conversion of carboplatin to cisplatin observed previously in NaCl conditions. Here, it is reported that a partial chemical transformation takes place but to a transplatin form. Thus, to attempt to resolve purely carboplatin binding at histidine, this study utilized co-crystallization of HEWL with carboplatin without NaCl to eliminate the partial chemical conversion of carboplatin. Tetragonal HEWL crystals co-crystallized with carboplatin were successfully obtained in four different conditions, each at a different pH value. The structural results obtained show carboplatin bound to either one or both of the N atoms of His15 of HEWL, and this particular variation was dependent on the concentration of anions in the crystallization mixture and the elapsed time, as well as the pH used. The structural details of the bound carboplatin molecule also differed between them. Overall, the most detailed crystal structure showed the majority of the carboplatin atoms bound to the platinum centre; however, the four-carbon ring structure of the cyclobutanedicarboxylate moiety (CBDC) remained elusive. The potential impact of the results for the administration of carboplatin as an anticancer agent are described.

  2. The lack of target specificity of small molecule anticancer kinase inhibitors is correlated with their ability to damage myocytes in vitro

    SciTech Connect

    Hasinoff, Brian B. Patel, Daywin

    2010-12-01

    Many new targeted small molecule anticancer kinase inhibitors are actively being developed. However, the clinical use of some kinase inhibitors has been shown to result in cardiotoxicity. In most cases the mechanisms by which they exert their cardiotoxicity are not well understood. We have used large scale profiling data on 8 FDA-approved tyrosine kinase inhibitors and 10 other kinase inhibitors to a panel of 317 kinases in order to correlate binding constants and kinase inhibitor binding selectivity scores with kinase inhibitor-induced damage to neonatal rat cardiac myocytes. The 18 kinase inhibitors that were the subject of this study were: canertinib, dasatinib, dovitinib, erlotinib, flavopiridol, gefitinib, imatinib, lapatinib, midostaurin, motesanib, pazopanib, sorafenib, staurosporine, sunitinib, tandutinib, tozasertib, vandetanib and vatalanib. The combined tyrosine kinase and serine-threonine kinase selectivity scores were highly correlated with the myocyte-damaging effects of the kinase inhibitors. This result suggests that myocyte damage was due to a lack of target selectivity to binding of both tyrosine kinases and serine-threonine kinases, and was not due to binding to either group specifically. Finally, the strength of kinase inhibitor binding for 290 kinases was examined for correlations with myocyte damage. Kinase inhibitor binding was significantly correlated with myocyte damage for 12 kinases. Thus, myocyte damage may be multifactorial in nature with the inhibition of a number of kinases involved in producing kinase inhibitor-induced myocyte damage.

  3. Boulder Damage Symposium annual thin-film laser damage competition

    NASA Astrophysics Data System (ADS)

    Stolz, Christopher J.

    2012-12-01

    Optical instruments and laser systems are often fluence-limited by multilayer thin films deposited on the optical surfaces. When comparing publications within the laser damage literature, there can be confusing and conflicting laser damage results. This is due to differences in testing protocols between research groups studying very different applications. In this series of competitions, samples from multiple vendors are compared under identical testing parameters and a single testing service. Unlike a typical study where a hypothesis is tested within a well-controlled experiment with isolated variables, this competition isolates the laser damage testing variables so that trends can be observed between different deposition processes, coating materials, cleaning techniques, and multiple coating suppliers. This series of damage competitions has also been designed to observe general trends of damage morphologies and mechanisms over a wide range of coating types (high reflector and antireflector), wavelengths (193 to 1064 nm), and pulse lengths (180 fs to 13 ns). For each of the competitions, a double blind test assured sample and submitter anonymity so only a summary of the deposition process, coating materials, layer count and spectral results are presented. In summary, laser resistance was strongly affected by substrate cleaning, coating deposition method, and coating material selection whereas layer count and spectral properties had minimal impact.

  4. Boulder damage symposium annual thin film laser damage competition

    SciTech Connect

    Stolz, Christopher J.

    2012-11-28

    Optical instruments and laser systems are often fluence-limited by multilayer thin films deposited on the optical surfaces. When comparing publications within the laser damage literature, there can be confusing and conflicting laser damage results. This is due to differences in testing protocols between research groups studying very different applications. In this series of competitions, samples from multiple vendors are compared under identical testing parameters and a single testing service. Unlike a typical study where a hypothesis is tested within a well-controlled experiment with isolated variables, this competition isolates the laser damage testing variables so that trends can be observed between different deposition processes, coating materials, cleaning techniques, and multiple coating suppliers. The resulting series of damage competitions has also been designed to observe general trends of damage morphologies and mechanisms over a wide range of coating types (high reflector and antireflector), wavelengths (193 to 1064 nm), and pulse lengths (180 fs to 13 ns). A double blind test assured sample and submitter anonymity were used in each of the competitions so only a summary of the deposition process, coating materials, layer count and spectral results are presented. Laser resistance was strongly affected by substrate cleaning, coating deposition method, and coating material selection whereas layer count and spectral properties had minimal impact.

  5. Boulder damage symposium annual thin film laser damage competition

    DOE PAGES

    Stolz, Christopher J.

    2012-11-28

    Optical instruments and laser systems are often fluence-limited by multilayer thin films deposited on the optical surfaces. When comparing publications within the laser damage literature, there can be confusing and conflicting laser damage results. This is due to differences in testing protocols between research groups studying very different applications. In this series of competitions, samples from multiple vendors are compared under identical testing parameters and a single testing service. Unlike a typical study where a hypothesis is tested within a well-controlled experiment with isolated variables, this competition isolates the laser damage testing variables so that trends can be observed betweenmore » different deposition processes, coating materials, cleaning techniques, and multiple coating suppliers. The resulting series of damage competitions has also been designed to observe general trends of damage morphologies and mechanisms over a wide range of coating types (high reflector and antireflector), wavelengths (193 to 1064 nm), and pulse lengths (180 fs to 13 ns). A double blind test assured sample and submitter anonymity were used in each of the competitions so only a summary of the deposition process, coating materials, layer count and spectral results are presented. Laser resistance was strongly affected by substrate cleaning, coating deposition method, and coating material selection whereas layer count and spectral properties had minimal impact.« less

  6. A model for damage of microheterogeneous kidney stones

    NASA Astrophysics Data System (ADS)

    Szeri, Andrew J.; Zohdi, Tarek I.; Blake, John R.

    2005-04-01

    In this paper, a theoretical framework is developed for the mechanics of kidney stones with an isotropic, random microstructure-such as those comprised of cystine or struvite. The approach is based on a micromechanical description of kidney stones comprised of crystals in a binding matrix. Stress concentration functions are developed to determine load sharing of the particle phase and the binding matrix phase. As an illustration of the theory, the fatigue of kidney stones subject to shock wave lithotripsy is considered. Stress concentration functions are used to construct fatigue life estimates for each phase, as a function of the volume fraction and of the mechanical properties of the constituents, as well as the loading from SWL. The failure of the binding matrix is determined explicitly in a model for the accumulation of distributed damage. Also considered is the amount of material damaged in a representative non-spherical collapse of a cavitation bubble near the stone surface. The theory can be used to assess the importance of microscale heterogeneity on the comminution of renal calculi and to estimate the number of cycles to failure in terms of measurable material properties.

  7. Neural networks for damage identification

    SciTech Connect

    Paez, T.L.; Klenke, S.E.

    1997-11-01

    Efforts to optimize the design of mechanical systems for preestablished use environments and to extend the durations of use cycles establish a need for in-service health monitoring. Numerous studies have proposed measures of structural response for the identification of structural damage, but few have suggested systematic techniques to guide the decision as to whether or not damage has occurred based on real data. Such techniques are necessary because in field applications the environments in which systems operate and the measurements that characterize system behavior are random. This paper investigates the use of artificial neural networks (ANNs) to identify damage in mechanical systems. Two probabilistic neural networks (PNNs) are developed and used to judge whether or not damage has occurred in a specific mechanical system, based on experimental measurements. The first PNN is a classical type that casts Bayesian decision analysis into an ANN framework; it uses exemplars measured from the undamaged and damaged system to establish whether system response measurements of unknown origin come from the former class (undamaged) or the latter class (damaged). The second PNN establishes the character of the undamaged system in terms of a kernel density estimator of measures of system response; when presented with system response measures of unknown origin, it makes a probabilistic judgment whether or not the data come from the undamaged population. The physical system used to carry out the experiments is an aerospace system component, and the environment used to excite the system is a stationary random vibration. The results of damage identification experiments are presented along with conclusions rating the effectiveness of the approaches.

  8. Collagen binding to Staphylococcus aureus

    SciTech Connect

    Holderbaum, D.; Hall, G.S.; Ehrhart, L.A.

    1986-11-01

    Staphylococcus aureus can bind soluble collagen in a specific, saturable manner. We have previously shown that some variability exists in the degree of collagen binding between different strains of heat-killed, formaldehyde-fixed S. aureus which are commercially available as immunologic reagents. The present study demonstrates that live S. aureus of the Cowan 1 strain binds amounts of collagen per organism equivalent to those demonstrated previously in heat-killed, formaldehyde-fixed bacteria but has an affinity over 100 times greater, with Kd values of 9.7 X 10(-11) M and 4.3 X 10(-8) M for live and heat-killed organisms, respectively. Studies were also carried out with S. aureus killed by ionizing radiation, since this method of killing the organism seemed less likely to alter the binding moieties on the surface than did heat killing. Bacteria killed by exposure to gamma radiation bound collagen in a manner essentially indistinguishable from that of live organisms. Binding of collagen to irradiated cells of the Cowan 1 strain was rapid, with equilibrium reached by 30 min at 22 degrees C, and was fully reversible. The binding was not inhibited by fibronectin, fibrinogen, C1q, or immunoglobulin G, suggesting a binding site for collagen distinct from those for these proteins. Collagen binding was virtually eliminated in trypsin-treated organisms, indicating that the binding site has a protein component. Of four strains examined, Cowan 1 and S. aureus ATCC 25923 showed saturable, specific binding, while strains Woods and S4 showed a complete lack of binding. These results suggest that some strains of S. aureus contain high-affinity binding sites for collagen. While the number of binding sites per bacterium varied sixfold in the two collagen-binding strains, the apparent affinity was similar.

  9. STS-118 Radiator Impact Damage

    NASA Technical Reports Server (NTRS)

    Lear, Dana M.; Hyde, J.; Christiansen, E.; Herrin, J.; Lyons, F.

    2008-01-01

    During the August 2007 STS-118 mission to the International Space Station, a micro-meteoroid or orbital debris (MMOD) particle impacted and completely penetrated one of shuttle Endeavour s radiator panels and the underlying thermal control system (TCS) blanket, leaving deposits on (but no damage to) the payload bay door. While it is not unusual for shuttle orbiters to be impacted by small MMOD particles, the damage from this impact is larger than any previously seen on the shuttle radiator panels. A close-up photograph of the radiator impact entry hole is shown in Figure 1, and the location of the impact on Endeavour s left-side aft-most radiator panel is shown in Figure 2. The aft radiator panel is 0.5-inches thick and consists of 0.011 inch thick aluminum facesheets on the front and back of an aluminum honeycomb core. The front facesheet is additionally covered by a 0.005 inch thick layer of silver-Teflon thermal tape. The entry hole in the silver-Teflon tape measured 8.1 mm by 6.4 mm (0.32 inches by 0.25 inches). The entry hole in the outer facesheet measured 7.4 mm by 5.3 mm (0.29 inches by 0.21 inches) (0.23 inches). The impactor also perforated an existing 0.012 inch doubler that had been bonded over the facesheet to repair previous impact damage (an example that lightning can strike the same place twice, even for MMOD impact). The peeled-back edge around the entry hole, or lip , is a characteristic of many hypervelocity impacts. High velocity impact with the front facesheet fragmented the impacting particle and caused it to spread out into a debris cloud. The debris cloud caused considerable damage to the internal honeycomb core with 23 honeycomb cells over a region of 28 mm by 26 mm (1.1 inches by 1.0 inches) having either been completely destroyed or partially damaged. Figure 3 is a view of the exit hole in the rear facesheet, and partially shows the extent of the honeycomb core damage and clearly shows the jagged petaled exit hole through the backside

  10. Avian pathogenic Escherichia coli bind fibronectin and laminin.

    PubMed

    Ramírez, Rosa María; Almanza, Yolanda; González, Rafael; García, Santos; Heredia, Norma

    2009-04-01

    Avian colisepticemia frequently occurs after respiratory tract damage, the primary site for infection allows bacteria to encounter an exposed basement membrane, where laminin and fibronectin are important components. We investigated the ability of an isolate of avian pathogenic Escherichia coli to bind fibronectin and laminin. Using Far-western dot blot analysis, we demonstrated the ability of this microorganism to bind basement membrane proteins fibronectin and laminin. Results from an ELISA-based approach indicate that the binding to these membrane proteins was bacterial-dose dependent. Furthermore, two specific E. coli polypeptides, of 32 kDa and 130 kDa, reacted with laminin and fibronectin, respectively. Further evaluation of these potential bacterial adhesins may provide insights into the pathogenesis of colibacillosis.

  11. Autophagy in DNA damage response.

    PubMed

    Czarny, Piotr; Pawlowska, Elzbieta; Bialkowska-Warzecha, Jolanta; Kaarniranta, Kai; Blasiak, Janusz

    2015-01-23

    DNA damage response (DDR) involves DNA repair, cell cycle regulation and apoptosis, but autophagy is also suggested to play a role in DDR. Autophagy can be activated in response to DNA-damaging agents, but the exact mechanism underlying this activation is not fully understood, although it is suggested that it involves the inhibition of mammalian target of rapamycin complex 1 (mTORC1). mTORC1 represses autophagy via phosphorylation of the ULK1/2-Atg13-FIP200 complex thus preventing maturation of pre-autophagosomal structures. When DNA damage occurs, it is recognized by some proteins or their complexes, such as poly(ADP)ribose polymerase 1 (PARP-1), Mre11-Rad50-Nbs1 (MRN) complex or FOXO3, which activate repressors of mTORC1. SQSTM1/p62 is one of the proteins whose levels are regulated via autophagic degradation. Inhibition of autophagy by knockout of FIP200 results in upregulation of SQSTM1/p62, enhanced DNA damage and less efficient damage repair. Mitophagy, one form of autophagy involved in the selective degradation of mitochondria, may also play role in DDR. It degrades abnormal mitochondria and can either repress or activate apoptosis, but the exact mechanism remains unknown. There is a need to clarify the role of autophagy in DDR, as this process may possess several important biomedical applications, involving also cancer therapy.

  12. Multipose binding in molecular docking.

    PubMed

    Atkovska, Kalina; Samsonov, Sergey A; Paszkowski-Rogacz, Maciej; Pisabarro, M Teresa

    2014-02-14

    Molecular docking has been extensively applied in virtual screening of small molecule libraries for lead identification and optimization. A necessary prerequisite for successful differentiation between active and non-active ligands is the accurate prediction of their binding affinities in the complex by use of docking scoring functions. However, many studies have shown rather poor correlations between docking scores and experimental binding affinities. Our work aimed to improve this correlation by implementing a multipose binding concept in the docking scoring scheme. Multipose binding, i.e., the property of certain protein-ligand complexes to exhibit different ligand binding modes, has been shown to occur in nature for a variety of molecules. We conducted a high-throughput docking study and implemented multipose binding in the scoring procedure by considering multiple docking solutions in binding affinity prediction. In general, improvement of the agreement between docking scores and experimental data was observed, and this was most pronounced in complexes with large and flexible ligands and high binding affinities. Further developments of the selection criteria for docking solutions for each individual complex are still necessary for a general utilization of the multipose binding concept for accurate binding affinity prediction by molecular docking.

  13. Rapid Recovery of Damaged Ecosystems

    PubMed Central

    Jones, Holly P.; Schmitz, Oswald J.

    2009-01-01

    Background Recent reports on the state of the global environment provide evidence that humankind is inflicting great damage to the very ecosystems that support human livelihoods. The reports further predict that ecosystems will take centuries to recover from damages if they recover at all. Accordingly, there is despair that we are passing on a legacy of irreparable damage to future generations which is entirely inconsistent with principles of sustainability. Methodology/Principal Findings We tested the prediction of irreparable harm using a synthesis of recovery times compiled from 240 independent studies reported in the scientific literature. We provide startling evidence that most ecosystems globally can, given human will, recover from very major perturbations on timescales of decades to half-centuries. Significance/Conclusions Accordingly, we find much hope that humankind can transition to more sustainable use of ecosystems. PMID:19471645

  14. Remote detection of forest damage

    NASA Technical Reports Server (NTRS)

    Rock, B. N.; Vogelmann, J. E.; Vogelmann, A. F.; Hoshizaki, T.; Williams, D. L.

    1986-01-01

    The use of remote sensing to discriminate, measure, and map forest damage is evaluated. TM spectal coverage, a helicopter-mounted radiometer, and ground-based surveys were utilized to examine the responses of the spruces and firs of Camels Hump Mountain, Vermont to stresses, such as pollution and trace metals. The basic spectral properties of vegetation are described. Forest damage at the site was estimated as 11.8-76.0 percent for the spruces and 19-43.8 percent for the balsam firs. Shifts in the spectra of the conifers in particular in the near IR region are analyzed, and variations in the mesophyll cell anatomy and pigment content of the spruces and firs are investigated. The relations between canopy moisture and damage is studied. The TM data are compared to aircraft data and found to be well correlated.

  15. System for estimating fatigue damage

    DOEpatents

    LeMonds, Jeffrey; Guzzo, Judith Ann; Liu, Shaopeng; Dani, Uttara Ashwin

    2017-03-14

    In one aspect, a system for estimating fatigue damage in a riser string is provided. The system includes a plurality of accelerometers which can be deployed along a riser string and a communications link to transmit accelerometer data from the plurality of accelerometers to one or more data processors in real time. With data from a limited number of accelerometers located at sensor locations, the system estimates an optimized current profile along the entire length of the riser including riser locations where no accelerometer is present. The optimized current profile is then used to estimate damage rates to individual riser components and to update a total accumulated damage to individual riser components. The number of sensor locations is small relative to the length of a deepwater riser string, and a riser string several miles long can be reliably monitored along its entire length by fewer than twenty sensor locations.

  16. Chromatin perturbations during the DNA damage response in higher eukaryotes.

    PubMed

    Bakkenist, Christopher J; Kastan, Michael B

    2015-12-01

    The DNA damage response is a widely used term that encompasses all signaling initiated at DNA lesions and damaged replication forks as it extends to orchestrate DNA repair, cell cycle checkpoints, cell death and senescence. ATM, an apical DNA damage signaling kinase, is virtually instantaneously activated following the introduction of DNA double-strand breaks (DSBs). The MRE11-RAD50-NBS1 (MRN) complex, which has a catalytic role in DNA repair, and the KAT5 (Tip60) acetyltransferase are required for maximal ATM kinase activation in cells exposed to low doses of ionizing radiation. The sensing of DNA lesions occurs within a highly complex and heterogeneous chromatin environment. Chromatin decondensation and histone eviction at DSBs may be permissive for KAT5 binding to H3K9me3 and H3K36me3, ATM kinase acetylation and activation. Furthermore, chromatin perturbation may be a prerequisite for most DNA repair. Nucleosome disassembly during DNA repair was first reported in the 1970s by Smerdon and colleagues when nucleosome rearrangement was noted during the process of nucleotide excision repair of UV-induced DNA damage in human cells. Recently, the multi-functional protein nucleolin was identified as the relevant histone chaperone required for partial nucleosome disruption at DBSs, the recruitment of repair enzymes and for DNA repair. Notably, ATM kinase is activated by chromatin perturbations induced by a variety of treatments that do not directly cause DSBs, including treatment with histone deacetylase inhibitors. Central to the mechanisms that activate ATR, the second apical DNA damage signaling kinase, outside of a stalled and collapsed replication fork in S-phase, is chromatin decondensation and histone eviction associated with DNA end resection at DSBs. Thus, a stress that is common to both ATM and ATR kinase activation is chromatin perturbations, and we argue that chromatin perturbations are both sufficient and required for induction of the DNA damage response.

  17. The CATDAT damaging earthquakes database

    NASA Astrophysics Data System (ADS)

    Daniell, J. E.; Khazai, B.; Wenzel, F.; Vervaeck, A.

    2011-08-01

    The global CATDAT damaging earthquakes and secondary effects (tsunami, fire, landslides, liquefaction and fault rupture) database was developed to validate, remove discrepancies, and expand greatly upon existing global databases; and to better understand the trends in vulnerability, exposure, and possible future impacts of such historic earthquakes. Lack of consistency and errors in other earthquake loss databases frequently cited and used in analyses was a major shortcoming in the view of the authors which needed to be improved upon. Over 17 000 sources of information have been utilised, primarily in the last few years, to present data from over 12 200 damaging earthquakes historically, with over 7000 earthquakes since 1900 examined and validated before insertion into the database. Each validated earthquake includes seismological information, building damage, ranges of social losses to account for varying sources (deaths, injuries, homeless, and affected), and economic losses (direct, indirect, aid, and insured). Globally, a slightly increasing trend in economic damage due to earthquakes is not consistent with the greatly increasing exposure. The 1923 Great Kanto (214 billion USD damage; 2011 HNDECI-adjusted dollars) compared to the 2011 Tohoku (>300 billion USD at time of writing), 2008 Sichuan and 1995 Kobe earthquakes show the increasing concern for economic loss in urban areas as the trend should be expected to increase. Many economic and social loss values not reported in existing databases have been collected. Historical GDP (Gross Domestic Product), exchange rate, wage information, population, HDI (Human Development Index), and insurance information have been collected globally to form comparisons. This catalogue is the largest known cross-checked global historic damaging earthquake database and should have far-reaching consequences for earthquake loss estimation, socio-economic analysis, and the global reinsurance field.

  18. Autophagy Induced by Calcium Phosphate Precipitates Targets Damaged Endosomes*

    PubMed Central

    Chen, Xi; Khambu, Bilon; Zhang, Hao; Gao, Wentao; Li, Min; Chen, Xiaoyun; Yoshimori, Tamotsu; Yin, Xiao-Ming

    2014-01-01

    Calcium phosphate precipitates (CPPs) form complexes with DNA, which enter cells via endocytosis. Under this condition CPPs induce autophagy via the canonic autophagy machinery. Here we showed that CPP-induced autophagy was also dependent on endocytosis as the process was significantly inhibited by methyl-β-cyclodextrin and dynasore, which suppress clathrin-dependent endocytosis. Consistently, CPP treatment triggered the formation of filipin-positive intracellular vesicles whose membranes are rich in cholesterol. Unexpectedly, these vesicles were also positive for galectin 3, suggesting that they were damaged and the membrane glycans became accessible to galectins to bind. Endosome damage was caused by endocytosis of CPPs and was reversed by calcium chelators or by endocytosis inhibitors. Notably, CPP-induced LC3-positive autophagosomes were colocalized with galectin 3, ubiquitin, and p62/SQSTM1. Inhibition of galectin 3 reduced p62 puncta and autophagosome formation. Knockdown of p62 additionally inhibited the colocalization of autophagosomes with galectins. Furthermore, most of the galectin 3-positive vesicles were colocalized with Rab7 or LAMP1. Agents that affect endosome/lysosome maturation and function, such as bafilomycin A1, also significantly affected CPP-induced tubulovesicular autophagosome formation. These findings thus indicate that endocytosed CPPs caused endosome damage and recruitment of galectins, particularly at the later endosome stage, which led to the interaction of the autophagosomal membranes with the damaged endosome in the presence of p62. PMID:24619419

  19. NBQX and TCP prevent soman-induced hippocampal damage

    SciTech Connect

    Lallement, G.; Carpentier, P.; Pernot-Marino, I.; Baubichon, D.; Blanchet, G.

    1993-05-13

    In a previous investigation we demonstrated that the measurement of w3 (peripheral-type benzodiazepine) binding site densities could be of widespread applicability in the localization and quantification of soman-induced damage in the central nervous system. We thus used this marker to assess, in mouse hippocampus, the neuroprotective activity against soman-induced brain damage of NBQX and TCP which are respective antagonists of non-NMDA and NMDA glutamatergic receptors. Injection of NBQX at 20 or 40 mg/kg 5 min prior to soman totally prevented the neuronal damage. Comparatively, TCP had neuroprotective efficacy when administered at l mg/kg 5 min prior to soman followed by a reinjection 1 hour after. These results demonstrate that both NBQX and TCP afford a satisfactory neuroprotection against soman-induced brain damage. Since it is known that the neuropathology due to soman is closely seizure-related, it is likely that the neuroprotective activities of NBQX and TCP are related to the respective roles of non-NMDA and NMDA receptors in the onset and maintenance of soman-induced seizures.

  20. 7 CFR 51.3067 - Serious damage.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 7 Agriculture 2 2010-01-01 2010-01-01 false Serious damage. 51.3067 Section 51.3067 Agriculture... Standards for Florida Avocados Definitions § 51.3067 Serious damage. Serious damage means any defect which... serious damage: (a) Anthracnose when any spot exceeds the area of a circle one-fourth inch in diameter,...

  1. 7 CFR 51.2960 - Damage.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 7 Agriculture 2 2010-01-01 2010-01-01 false Damage. 51.2960 Section 51.2960 Agriculture... Standards for Grades of Walnuts in the Shell Definitions § 51.2960 Damage. Damage means any specific defect... considered as damage: (a) Broken shells when the area from which a portion of the shell is missing is...

  2. Global deformation facilitates flipping of damaged 8-oxo-guanine and guanine in DNA

    PubMed Central

    La Rosa, Giuseppe; Zacharias, Martin

    2016-01-01

    Oxidation of guanine (Gua) to form 7,8-dihydro-8-oxoguanine (8oxoG) is a frequent mutagenic DNA lesion. DNA repair glycosylases such as the bacterial MutM can effciently recognize and eliminate the 8oxoG damage by base excision. The base excision requires a 8oxoG looping out (flipping) from an intrahelical base paired to an extrahelical state where the damaged base is in the enzyme active site. It is still unclear how the damage is identified and flipped from an energetically stable stacked and paired state without any external energy source. Free energy simulations have been employed to study the flipping process for globally deformed DNA conformational states. DNA deformations were generated by systematically untwisting the DNA to mimic its conformation in repair enzyme encounter complex. The simulations indicate that global DNA untwisting deformation toward the enzyme bound form alone (without protein) significantly reduces the penalty for damage flipping to about half of the penalty observed in regular DNA. The finding offers a mechanistic explanation how binding free energy that is transformed to binding induced DNA deformation facilitates flipping and helps to rapidly detect a damaged base. It is likely of general relevance since repair enzyme binding frequently results in significant deformation of the target DNA. PMID:27651459

  3. Certification of damage tolerant composite structure

    NASA Technical Reports Server (NTRS)

    Rapoff, Andrew J.; Dill, Harold D.; Sanger, Kenneth B.; Kautz, Edward F.

    1990-01-01

    A reliability based certification testing methodology for impact damage tolerant composite structure was developed. Cocured, adhesively bonded, and impact damaged composite static strength and fatigue life data were statistically analyzed to determine the influence of test parameters on the data scatter. The impact damage resistance and damage tolerance of various structural configurations were characterized through the analysis of an industry wide database of impact test results. Realistic impact damage certification requirements were proposed based on actual fleet aircraft data. The capabilities of available impact damage analysis methods were determined through correlation with experimental data. Probabilistic methods were developed to estimate the reliability of impact damaged composite structures.

  4. Strength and failure of a damaged material

    SciTech Connect

    Cerreta, Ellen K.; Gray III, George T.; Trujillo, Carl P.; Potocki, Mark L.; Vachhani, Shraddha; Martinez, Daniel T.; Lovato, Manual L.

    2015-09-07

    Under complex, dynamic loading conditions, damage can occur within a material. Should this damage not lead to catastrophic failure, the material can continue to sustain further loading. But, little is understood about how to represent the mechanical response of a material that has experienced dynamic loading leading to incipient damage. We examine this effect in copper. Copper is shock loaded to impart an incipient state of damage to the material. Thereafter compression and tensile specimens were sectioned from the dynamically damaged specimen to quantify the subsequent properties of the material in the region of intense incipient damage and in regions far from the damage. Finally, we observed that enhanced yield stresses result from the damaged material even over material, which has simply been shock loaded and not damaged. These results are rationalized in terms of stored plastic work due to the damage process.

  5. Strength and failure of a damaged material

    DOE PAGES

    Cerreta, Ellen K.; Gray III, George T.; Trujillo, Carl P.; ...

    2015-09-07

    Under complex, dynamic loading conditions, damage can occur within a material. Should this damage not lead to catastrophic failure, the material can continue to sustain further loading. But, little is understood about how to represent the mechanical response of a material that has experienced dynamic loading leading to incipient damage. We examine this effect in copper. Copper is shock loaded to impart an incipient state of damage to the material. Thereafter compression and tensile specimens were sectioned from the dynamically damaged specimen to quantify the subsequent properties of the material in the region of intense incipient damage and in regionsmore » far from the damage. Finally, we observed that enhanced yield stresses result from the damaged material even over material, which has simply been shock loaded and not damaged. These results are rationalized in terms of stored plastic work due to the damage process.« less

  6. Antioxidant flavonoids bind human serum albumin

    NASA Astrophysics Data System (ADS)

    Kanakis, C. D.; Tarantilis, P. A.; Polissiou, M. G.; Diamantoglou, S.; Tajmir-Riahi, H. A.

    2006-10-01

    Human serum albumin (HSA) is a principal extracellular protein with a high concentration in blood plasma and carrier for many drugs to different molecular targets. Flavonoids are powerful antioxidants and prevent DNA damage. The antioxidative protections are related to their binding modes to DNA duplex and complexation with free radicals in vivo. However, flavonoids are known to inhibit the activities of several enzymes such as calcium phospholipid-dependent protein kinase, tyrosine protein kinase from rat lung, phosphorylase kinase, phosphatidylinositol 3-kinase and DNA topoisomerases that exhibit the importance of flavonoid-protein interaction. This study was designed to examine the interaction of human serum albumin (HSA) with quercetin (que), kaempferol (kae) and delphinidin (del) in aqueous solution at physiological conditions, using constant protein concentration of 0.25 mM (final) and various drug contents of 1 μM-1 mM. FTIR and UV-vis spectroscopic methods were used to determine the polyphenolic binding mode, the binding constant and the effects of flavonoid complexation on protein secondary structure. The spectroscopic results showed that flavonoids are located along the polypeptide chains through H-bonding interactions with overall affinity constant of Kque = 1.4 × 10 4 M -1, Kkae = 2.6 × 10 5 M -1 and Kdel = 4.71 × 10 5 M -1. The protein secondary structure showed no alterations at low pigment concentration (1 μM), whereas at high flavonoid content (1 mM), major reduction of α-helix from 55% (free HSA) to 42-46% and increase of β-sheet from 15% (free HSA) to 17-19% and β-anti from 7% (free HSA) to 10-20% occurred in the flavonoid-HSA adducts. The major reduction of HSA α-helix is indicative of a partial protein unfolding upon flavonoid interaction.

  7. Viral interference with DNA repair by targeting of the single-stranded DNA binding protein RPA.

    PubMed

    Banerjee, Pubali; DeJesus, Rowena; Gjoerup, Ole; Schaffhausen, Brian S

    2013-10-01

    Correct repair of damaged DNA is critical for genomic integrity. Deficiencies in DNA repair are linked with human cancer. Here we report a novel mechanism by which a virus manipulates DNA damage responses. Infection with murine polyomavirus sensitizes cells to DNA damage by UV and etoposide. Polyomavirus large T antigen (LT) alone is sufficient to sensitize cells 100 fold to UV and other kinds of DNA damage. This results in activated stress responses and apoptosis. Genetic analysis shows that LT sensitizes via the binding of its origin-binding domain (OBD) to the single-stranded DNA binding protein replication protein A (RPA). Overexpression of RPA protects cells expressing OBD from damage, and knockdown of RPA mimics the LT phenotype. LT prevents recruitment of RPA to nuclear foci after DNA damage. This leads to failure to recruit repair proteins such as Rad51 or Rad9, explaining why LT prevents repair of double strand DNA breaks by homologous recombination. A targeted intervention directed at RPA based on this viral mechanism could be useful in circumventing the resistance of cancer cells to therapy.

  8. PARP-2 domain requirements for DNA damage-dependent activation and localization to sites of DNA damage.

    PubMed

    Riccio, Amanda A; Cingolani, Gino; Pascal, John M

    2016-02-29

    Poly(ADP-ribose) polymerase-2 (PARP-2) is one of three human PARP enzymes that are potently activated during the cellular DNA damage response (DDR). DDR-PARPs detect DNA strand breaks, leading to a dramatic increase in their catalytic production of the posttranslational modification poly(ADP-ribose) (PAR) to facilitate repair. There are limited biochemical and structural insights into the functional domains of PARP-2, which has restricted our understanding of how PARP-2 is specialized toward specific repair pathways. PARP-2 has a modular architecture composed of a C-terminal catalytic domain (CAT), a central Trp-Gly-Arg (WGR) domain and an N-terminal region (NTR). Although the NTR is generally considered the key DNA-binding domain of PARP-2, we report here that all three domains of PARP-2 collectively contribute to interaction with DNA damage. Biophysical, structural and biochemical analyses indicate that the NTR is natively disordered, and is only required for activation on specific types of DNA damage. Interestingly, the NTR is not essential for PARP-2 localization to sites of DNA damage. Rather, the WGR and CAT domains function together to recruit PARP-2 to sites of DNA breaks. Our study differentiates the functions of PARP-2 domains from those of PARP-1, the other major DDR-PARP, and highlights the specialization of the multi-domain architectures of DDR-PARPs.

  9. PARP-2 domain requirements for DNA damage-dependent activation and localization to sites of DNA damage

    PubMed Central

    Riccio, Amanda A.; Cingolani, Gino; Pascal, John M.

    2016-01-01

    Poly(ADP-ribose) polymerase-2 (PARP-2) is one of three human PARP enzymes that are potently activated during the cellular DNA damage response (DDR). DDR-PARPs detect DNA strand breaks, leading to a dramatic increase in their catalytic production of the posttranslational modification poly(ADP-ribose) (PAR) to facilitate repair. There are limited biochemical and structural insights into the functional domains of PARP-2, which has restricted our understanding of how PARP-2 is specialized toward specific repair pathways. PARP-2 has a modular architecture composed of a C-terminal catalytic domain (CAT), a central Trp-Gly-Arg (WGR) domain and an N-terminal region (NTR). Although the NTR is generally considered the key DNA-binding domain of PARP-2, we report here that all three domains of PARP-2 collectively contribute to interaction with DNA damage. Biophysical, structural and biochemical analyses indicate that the NTR is natively disordered, and is only required for activation on specific types of DNA damage. Interestingly, the NTR is not essential for PARP-2 localization to sites of DNA damage. Rather, the WGR and CAT domains function together to recruit PARP-2 to sites of DNA breaks. Our study differentiates the functions of PARP-2 domains from those of PARP-1, the other major DDR-PARP, and highlights the specialization of the multi-domain architectures of DDR-PARPs. PMID:26704974

  10. CTCF participates in DNA damage response via poly(ADP-ribosyl)ation

    PubMed Central

    Han, Deqiang; Chen, Qian; Shi, Jiazhong; Zhang, Feng; Yu, Xiaochun

    2017-01-01

    CCCTC-binding factor (CTCF) plays an essential role in regulating the structure of chromatin by binding DNA strands for defining the boundary between active and heterochromatic DNA. However, the role of CTCF in DNA damage response remains elusive. Here, we show that CTCF is quickly recruited to the sites of DNA damage. The fast recruitment is mediated by the zinc finger domain and poly (ADP-ribose) (PAR). Further analyses show that only three zinc finger motifs are essential for PAR recognition. Moreover, CTCF-deficient cells are hypersensitive to genotoxic stress such as ionizing radiation (IR). Taken together, these results suggest that CTCF participate in DNA damage response via poly(ADP-ribosylation). PMID:28262757

  11. Lamin A/C-dependent interaction with 53BP1 promotes cellular responses to DNA damage

    PubMed Central

    Gibbs-Seymour, Ian; Markiewicz, Ewa; Bekker-Jensen, Simon; Mailand, Niels; Hutchison, Christopher J

    2015-01-01

    Lamins A/C have been implicated in DNA damage response pathways. We show that the DNA repair protein 53BP1 is a lamin A/C binding protein. In undamaged human dermal fibroblasts (HDF), 53BP1 is a nucleoskeleton protein. 53BP1 binds to lamins A/C via its Tudor domain, and this is abrogated by DNA damage. Lamins A/C regulate 53BP1 levels and consequently lamin A/C-null HDF display a 53BP1 null-like phenotype. Our data favour a model in which lamins A/C maintain a nucleoplasmic pool of 53BP1 in order to facilitate its rapid recruitment to sites of DNA damage and could explain why an absence of lamin A/C accelerates aging. PMID:25645366

  12. Ultrasonic Evaluation of Fatigue Damage

    NASA Astrophysics Data System (ADS)

    Bayer, P.; Singher, L.; Notea, A.

    2004-02-01

    Despite the fact that most engineers and designers are aware of fatigue, many severe breakdowns of industrial plant and machinery still occur due to fatigue. In effect, it's been estimated that fatigue causes at least 80% of the failures in modern engineering components. From an operational point of view, the detection of fatigue damage, preferably at a very early stage, is a critically important consideration in order to prevent possible catastrophic equipment failure and associated losses. This paper describes the investigation involving the use of ultrasonic waves as a potential tool for early detection of fatigue damage. The parameters investigated were the ultrasonic wave velocities (longitudinal and transverse waves) and attenuation coefficient before fatigue damage and after progressive stages of fatigue. Although comparatively small uncertainties were observed, the feasibility of utilizing the velocity of ultrasonic waves as a fatigue monitor was barely substantiated within actual research conditions. However, careful measurements of the ultrasonic attenuation parameter had demonstrated its potential to provide an early assessment of damage during fatigue.

  13. Sulfur Dioxide and Material Damage

    ERIC Educational Resources Information Center

    Gillette, Donald G.

    1975-01-01

    This study relates sulfur dioxide levels with material damage in heavily populated or polluted areas. Estimates of loss were determined from increased maintenance and replacement costs. The data indicate a decrease in losses during the past five years probably due to decline in pollution levels established by air quality standards. (MR)

  14. Compensation for oil pollution damage

    NASA Astrophysics Data System (ADS)

    Matugina, E. G.; Glyzina, T. S.; Kolbysheva, Yu V.; Klyuchnikov, A. S.; Vusovich, O. V.

    2015-11-01

    The commitment of national industries to traditional energy sources, as well as constantly growing energy demand combined with adverse environmental impact of petroleum production and transportation urge to establish and maintain an appropriate legal and administrative framework for oil pollution damage compensation. The article considers management strategies for petroleum companies that embrace not only production benefits but also environmental issues.

  15. Cisplatin-resistant cells express increased levels of a factor that recognizes damaged DNA

    SciTech Connect

    Chu, G.; Chang, E. )

    1990-05-01

    Cancer treatment with the drug cisplatin is often thwarted by the emergence of drug-resistant cells. To study this phenomenon, the authors identified two independent cellular factors that recognize cisplatin-damaged DNA. One of the two factors, designated XPE binding factor, is deficient in complementation group E of xeroderma pigmentosum, an inherited disease characterized by defective repair of DNA damaged by ultraviolet radiation, cisplatin, and other agents. Human tumor cell lines selected for resistance to cisplatin showed more efficient DNA repair and increased expression of XPE binding factor. These results suggest that XPE binding factor may be responsible, at least in part, for the development of cisplatin resistance in human tumors and that the mechanism may be increased DNA repair.

  16. Plate tectonics, damage and inheritance.

    PubMed

    Bercovici, David; Ricard, Yanick

    2014-04-24

    The initiation of plate tectonics on Earth is a critical event in our planet's history. The time lag between the first proto-subduction (about 4 billion years ago) and global tectonics (approximately 3 billion years ago) suggests that plates and plate boundaries became widespread over a period of 1 billion years. The reason for this time lag is unknown but fundamental to understanding the origin of plate tectonics. Here we suggest that when sufficient lithospheric damage (which promotes shear localization and long-lived weak zones) combines with transient mantle flow and migrating proto-subduction, it leads to the accumulation of weak plate boundaries and eventually to fully formed tectonic plates driven by subduction alone. We simulate this process using a grain evolution and damage mechanism with a composite rheology (which is compatible with field and laboratory observations of polycrystalline rocks), coupled to an idealized model of pressure-driven lithospheric flow in which a low-pressure zone is equivalent to the suction of convective downwellings. In the simplest case, for Earth-like conditions, a few successive rotations of the driving pressure field yield relic damaged weak zones that are inherited by the lithospheric flow to form a nearly perfect plate, with passive spreading and strike-slip margins that persist and localize further, even though flow is driven only by subduction. But for hotter surface conditions, such as those on Venus, accumulation and inheritance of damage is negligible; hence only subduction zones survive and plate tectonics does not spread, which corresponds to observations. After plates have developed, continued changes in driving forces, combined with inherited damage and weak zones, promote increased tectonic complexity, such as oblique subduction, strike-slip boundaries that are subparallel to plate motion, and spalling of minor plates.

  17. Lactoperoxidase binding to streptococci.

    PubMed Central

    Pruitt, K M; Adamson, M; Arnold, R

    1979-01-01

    There have been conflicting reports regarding the binding of lactoperoxidase to bacterial cell surfaces. We describe here the effects of cell-bound lactoperoxidase on acid production by suspensions of Streptococcus mutans (NCTC 10449) in the presence of hydrogen peroxide and thiocyanate. Saline suspensions of log-phase bacteria were treated with 0.1 mg of lactoperoxidase per ml and were then washed thoroughly. The addition of hydrogen peroxide and thiocyanate markedly reduced the acid production of these lactoperoxidase-treated bacteria but had no effect on the acid production of untreated controls. After a 3-h incubation in saline, the lactoperoxidase-treated bacteria produced acid in the presence of hydrogen peroxide and thiocyanate at the same rate as untreated bacteria. These observations suggest that lactoperoxidase is initially bound to the cell surface in an enzymatically active form at a concentration sufficient to inhibit acid production. The lactoperoxidase is slowly degraded or desorbed as the bacteria stand in saline suspension. PMID:39032

  18. Regulation of DNA damage responses and cell cycle progression by hMOB2

    PubMed Central

    Gomez, Valenti; Gundogdu, Ramazan; Gomez, Marta; Hoa, Lily; Panchal, Neelam; O’Driscoll, Mark; Hergovich, Alexander

    2014-01-01

    Mps one binder proteins (MOBs) are conserved regulators of essential signalling pathways. Biochemically, human MOB2 (hMOB2) can inhibit NDR kinases by competing with hMOB1 for binding to NDRs. However, biological roles of hMOB2 have remained enigmatic. Here, we describe novel functions of hMOB2 in the DNA damage response (DDR) and cell cycle regulation. hMOB2 promotes DDR signalling, cell survival and cell cycle arrest after exogenously induced DNA damage. Under normal growth conditions in the absence of exogenously induced DNA damage hMOB2 plays a role in preventing the accumulation of endogenous DNA damage and a subsequent p53/p21-dependent G1/S cell cycle arrest. Unexpectedly, these molecular and cellular phenotypes are not observed upon NDR manipulations, indicating that hMOB2 performs these functions independent of NDR signalling. Thus, to gain mechanistic insight, we screened for novel binding partners of hMOB2, revealing that hMOB2 interacts with RAD50, facilitating the recruitment of the MRE11-RAD50-NBS1 (MRN) DNA damage sensor complex and activated ATM to DNA damaged chromatin. Taken together, we conclude that hMOB2 supports the DDR and cell cycle progression. PMID:25460043

  19. The PIN domain of EXO1 recognizes poly(ADP-ribose) in DNA damage response

    PubMed Central

    Zhang, Feng; Shi, Jiazhong; Chen, Shih-Hsun; Bian, Chunjing; Yu, Xiaochun

    2015-01-01

    Following DNA double-strand breaks, poly(ADP-ribose) (PAR) is quickly and heavily synthesized to mediate fast and early recruitment of a number of DNA damage response factors to the sites of DNA lesions and facilitates DNA damage repair. Here, we found that EXO1, an exonuclease for DNA damage repair, is quickly recruited to the sites of DNA damage via PAR-binding. With further dissection of the functional domains of EXO1, we report that the PIN domain of EXO1 recognizes PAR both in vitro and in vivo and the interaction between the PIN domain and PAR is sufficient for the recruitment. We also found that the R93G variant of EXO1, generated by a single nucleotide polymorphism, abolishes the interaction and the early recruitment. Moreover, our study suggests that the PAR-mediated fast recruitment of EXO1 facilities early DNA end resection, the first step of homologous recombination repair. We observed that other PIN domains could also recognize DNA damage-induced PAR. Taken together, our study demonstrates a novel class of PAR-binding module that plays an important role in DNA damage response. PMID:26400172

  20. Reprogramming cellular events by poly(ADP-ribose)-binding proteins

    PubMed Central

    Pic, Émilie; Ethier, Chantal; Dawson, Ted M.; Dawson, Valina L.; Masson, Jean-Yves; Poirier, Guy G.; Gagné, Jean-Philippe

    2013-01-01

    Poly(ADP-ribosyl)ation is a posttranslational modification catalyzed by the poly(ADP-ribose) polymerases (PARPs). These enzymes covalently modify glutamic, aspartic and lysine amino acid side chains of acceptor proteins by the sequential addition of ADP-ribose (ADPr) units. The poly(ADP-ribose) (pADPr) polymers formed alter the physico-chemical characteristics of the substrate with functional consequences on its biological activities. Recently, non-covalent binding to pADPr has emerged as a key mechanism to modulate and coordinate several intracellular pathways including the DNA damage response, protein stability and cell death. In this review, we describe the basis of non-covalent binding to pADPr that has led to the emerging concept of pADPr-responsive signaling pathways. This review emphasizes the structural elements and the modular strategies developed by pADPr-binding proteins to exert a fine-tuned control of a variety of pathways. Poly(ADP-ribosyl)ation reactions are highly regulated processes, both spatially and temporally, for which at least four specialized pADPr-binding modules accommodate different pADPr structures and reprogram protein functions. In this review, we highlight the role of well-characterized and newly discovered pADPr-binding modules in a diverse set of physiological functions. PMID:23268355

  1. Managing a Library Binding Program.

    ERIC Educational Resources Information Center

    Merrill-Oldham, Jan

    Library binding is one of the activities typically included in newly created preservation departments, but librarians continue to discover that transforming a traditional binding program into one that better meets preservation objectives requires considerable investment of time. This resource guide is intended to help libraries review their…

  2. Binding Energy and Enzymatic Catalysis.

    ERIC Educational Resources Information Center

    Hansen, David E.; Raines, Ronald T.

    1990-01-01

    Discussed is the fundamental role that the favorable free energy of binding of the rate-determining transition state plays in catalysis. The principle that all of the catalytic factors discussed are realized by the use of this binding energy is reviewed. (CW)

  3. Empirically Unbinding the Double Bind.

    ERIC Educational Resources Information Center

    Olson, David H.

    The theoretical concept of the double bind and the possibilities for researching it are discussed. The author has observed that theory and research, which should be reciprocal and mutually beneficial, have been working, as concerns the double bind, at odds with one another. Two approaches to empirically investigating the concept are considered via…

  4. Hybrid molecular dynamics simulation for plasma induced damage analysis

    NASA Astrophysics Data System (ADS)

    Matsukuma, Masaaki

    2016-09-01

    In order to enable further device size reduction (also known as Moore's law) and improved power performance, the semiconductor industry is introducing new materials and device structures into the semiconductor fabrication process. Materials now include III-V compounds, germanium, cobalt, ruthenium, hafnium, and others. The device structure in both memory and logic has been evolving from planar to three dimensional (3D). One such device is the FinFET, where the transistor gate is a vertical fin made either of silicon, silicon-germanium or germanium. These changes have brought renewed interests in the structural damages caused by energetic ion bombardment of the fin sidewalls which are exposed to the ion flux from the plasma during the fin-strip off step. Better control of the physical damage of the 3D devices requires a better understanding of the damage formation mechanisms on such new materials and structures. In this study, the damage formation processes by ion bombardment have been simulated for Si and Ge substrate by Quantum Mechanics/Molecular Mechanics (QM/MM) hybrid simulations and compared to the results from the classical molecular dynamics (MD) simulations. In our QM/MM simulations, the highly reactive region in which the structural damage is created is simulated with the Density Functional based Tight Binding (DFTB) method and the region remote from the primary region is simulated using classical MD with the Stillinger-Weber and Moliere potentials. The learn on the fly method is also used to reduce the computational load. Hence our QM/MM simulation is much faster than the full QC-MD simulations and the original QM/MM simulations. The amorphous layers profile simulated with QM/MM have obvious differences in their thickness for silicon and germanium substrate. The profile of damaged structure in the germanium substrate is characterized by a deeper tail then in silicon. These traits are also observed in the results from the mass selected ion beam

  5. Expression Profile of DNA Damage Signaling Genes in Proton Exposed Mouse Brain

    NASA Astrophysics Data System (ADS)

    Ramesh, Govindarajan; Wu, Honglu

    Exposure of living systems to radiation results in a wide assortment of lesions, the most signif-icant of is damage to genomic DNA which induce several cellular functions such as cell cycle arrest, repair, apoptosis etc. The radiation induced DNA damage investigation is one of the im-portant area in biology, but still the information available regarding the effects of proton is very limited. In this report, we investigated the differential gene expression pattern of DNA damage signaling genes particularly, damaged DNA binding, repair, cell cycle arrest, checkpoints and apoptosis using quantitative real-time RT-PCR array in proton exposed mouse brain tissues. The expression profiles showed significant changes in DNA damage related genes in 2Gy proton exposed mouse brain tissues as compared with control brain tissues. Furthermore, we also show that significantly increased levels of apoptotic related genes, caspase-3 and 8 activities in these cells, suggesting that in addition to differential expression of DNA damage genes, the alteration of apoptosis related genes may also contribute to the radiation induced DNA damage followed by programmed cell death. In summary, our findings suggest that proton exposed brain tissue undergo severe DNA damage which in turn destabilize the chromatin stability.

  6. Damage of hybrid composite laminates

    NASA Astrophysics Data System (ADS)

    Haery, Haleh A.; Kim, Ho Sung

    2013-08-01

    Hybrid laminates consisting of woven glass fabric/epoxy composite plies and woven carbon fabric/epoxy composite plies are studied for fatigue damage and residual strength. A theoretical framework based on the systems approach is proposed as a guide to deal with the complexity involving uncertainties and a large number of variables in the hybrid composite system. A relative damage sensitivity factor expression was developed for quantitative comparisons between non-hybrid and hybrid composites. Hypotheses derived from the theoretical framework were tested and verified. The first hypothesis was that the difference between two different sets of properties produces shear stress in interface between carbon fibre reinforced plastics (CRP) and glass fibre reinforced plastics (GRP), and eventually become a source for CRP/GRP interfacial delamination or longitudinal cracking. The second hypothesis was that inter-fibre bundle delamination occurs more severely to CRP sub-system than GRP sub-system.

  7. Radiation damage evolution in ceramics

    SciTech Connect

    Devanathan, Ramaswami

    2009-09-15

    A review is presented of recent results on radiation damage production, defect accumulation and dynamic annealing in a number of ceramics, such as silicon carbide, zircon and zirconia. Under energetic particle irradiation, ceramics can undergo amorphization by the accumulation of point defects and defect clusters (silicon carbide) or direct impact amorphization (zircon). Ceramics that resist radiation-induced amorphization have mechanisms to dissipate the primary knock-on atom energy, such as replacement collision sequences that leave the lattice undisturbed and low-energy cation site exchange. The presence of engineered mobile defects, such as structural vacancies in stabilized zirconia, can dynamically anneal radiation damage. Thus, defect engineering is a promising strategy to design radiation tolerance for applications such as nuclear waste disposal.

  8. Seismic damage estimation for buried pipeline systems

    SciTech Connect

    Heubach, W.F.

    1995-12-31

    A methodology for estimating seismic damage rates for buried pipeline systems is presented. The methodology is intended for damage estimation of buried pipeline systems in areas where use of more rigorous structural analysis techniques is not practical. Damage is estimated for areas subjected to ground shaking and permanent ground deformation. Although the methodology employs previously developed ground shaking damage algorithms, new damage algorithms are developed for permanent ground deformation. These new algorithms reflect the high levels of damage observed in areas of soil liquefaction.

  9. Method for producing damage resistant optics

    DOEpatents

    Hackel, Lloyd A.; Burnham, Alan K.; Penetrante, Bernardino M.; Brusasco, Raymond M.; Wegner, Paul J.; Hrubesh, Lawrence W.; Kozlowski, Mark R.; Feit, Michael D.

    2003-01-01

    The present invention provides a system that mitigates the growth of surface damage in an optic. Damage to the optic is minimally initiated. In an embodiment of the invention, damage sites in the optic are initiated, located, and then treated to stop the growth of the damage sites. The step of initiating damage sites in the optic includes a scan of the optic using a laser to initiate defects. The exact positions of the initiated sites are identified. A mitigation process is performed that locally or globally removes the cause of subsequent growth of the damaged sites.

  10. Smart accelerometer. [vibration damage detection

    NASA Technical Reports Server (NTRS)

    Bozeman, Richard J., Jr. (Inventor)

    1994-01-01

    The invention discloses methods and apparatus for detecting vibrations from machines which indicate an impending malfunction for the purpose of preventing additional damage and allowing for an orderly shutdown or a change in mode of operation. The method and apparatus is especially suited for reliable operation in providing thruster control data concerning unstable vibration in an electrical environment which is typically noisy and in which unrecognized ground loops may exist.

  11. Fire Damage and Strategic Targeting

    DTIC Science & Technology

    1984-06-01

    both thermally and blast-induced ignitions. The vulnerability is expressed as an incident radiation flux level or as an overpressure, and thus...massive thermal influx. In either case, the probability of fire damage is a function of the incident thermal flux , the level of blast effects, and...considered as variable parameters and were allowed to assume a range of values. The target vulnerability to the thermal flux depends on the type and

  12. Epidemiology of Helicopter Battle Damage

    DTIC Science & Technology

    2010-05-01

    A through Class C accident data , and consider the data to be representative of the spectrum of damage that must be dealt with by deployed...maintenance units. An analysis of the data by year dating back to 1995 was done to look at the effect of OIF and OEF on overall aircraft incident rate. This...per response, including the time for reviewing instructions, searching existing data sources, gathering and maintaining the data needed, and

  13. Blood-Induced Joint Damage

    PubMed Central

    Roosendaal, Goris; Jansen, Nathalie W.D.; Lafeber, Floris P.J.G.; Mastbergen, Simon C.

    2013-01-01

    Objective. Four days of blood exposure leads to irreversible cartilage damage in vitro. In contrast, intermittent intra-articular blood injections twice a week during 4 weeks (mimicking micro-bleeds) in a canine model resulted in transient damage only. In this study, it was evaluated whether acute joint bleeds are more harmful than micro-bleeds in a canine model of knee arthropathy. Design. Seven dogs received 4 sequential daily intra-articular blood injections twice in 2 weeks (mimicking 2 acute 4-day joint bleeds). Seven other dogs received the same blood load but in a total of 8 injections intermittently over the 4-week period with at least 1 day in between (mimicking micro-bleeds over the same timespan). Contralateral knees served as controls. Ten weeks after the last injection cartilage matrix turnover and synovial inflammation were evaluated. Results. Only after the acute joint bleeds the release of newly formed and total (resident) cartilage matrix glycosaminoglycans were increased (P = 0.04 and P = 0.01, respectively). Furthermore, in animals with the acute joint bleeds cartilage glycosaminoglycan content was decreased (P = 0.01) and not in animals with micro-bleeds. Mild synovial inflammation was observed in both groups (both P < 0.0001) but was not different between groups. Conclusions. In contrast to micro-bleeds, 2 acute joint bleeds lead to prolonged cartilage damage independent of the level of synovial inflammation. This model suggests that micro-bleeds are less devastating than acute joint bleeds with respect to joint damage, which might be of relevance to treatment of joint bleeds in clinical practice. PMID:26069675

  14. Ovalbumin with Glycated Carboxyl Groups Shows Membrane-Damaging Activity

    PubMed Central

    Tang, Ching-Chia; Shi, Yi-Jun; Chen, Ying-Jung; Chang, Long-Sen

    2017-01-01

    The aim of the present study was to investigate whether glycated ovalbumin (OVA) showed novel activity at the lipid-water interface. Mannosylated OVA (Man-OVA) was prepared by modification of the carboxyl groups with p-aminophenyl α-dextro (d)-mannopyranoside. An increase in the number of modified carboxyl groups increased the membrane-damaging activity of Man-OVA on cell membrane-mimicking vesicles, whereas OVA did not induce membrane permeability in the tested phospholipid vesicles. The glycation of carboxyl groups caused a notable change in the gross conformation of OVA. Moreover, owing to their spatial positions, the Trp residues in Man-OVA were more exposed, unlike those in OVA. Fluorescence quenching studies suggested that the Trp residues in Man-OVA were located on the interface binds with the lipid vesicles, and their microenvironment was abundant in positively charged residues. Although OVA and Man-OVA showed a similar binding affinity for lipid vesicles, the lipid-interacting feature of Man-OVA was distinct from that of OVA. Chemical modification studies revealed that Lys and Arg residues, but not Trp residues, played a crucial role in the membrane-damaging activity of Man-OVA. Taken together, our data suggest that glycation of carboxyl groups causes changes in the structural properties and membrane-interacting features of OVA, generating OVA with membrane-perturbing activities at the lipid-water interface. PMID:28264493

  15. Economic measurement of environment damages

    SciTech Connect

    Krawiec, F.

    1980-05-01

    The densities, energy consumption, and economic development of the increasing population exacerbate environmental degradation. Air and water pollution is a major environmental problem affecting life and health, outdoor recreation, household soiling, vegetation, materials, and production. The literature review indicated that numerous studies have assessed the physical and monetary damage to populations at risk from excessive concentrations of major air and water pollutants-sulfur dioxide, total suspended particulate matter, oxidants, and carbon monoxide in air; and nutrients, oil, pesticides, and toxic metals and others in water. The measurement of the damages was one of the most controversial issues in pollution abatement. The methods that have been used to estimate the societal value of pollution abatement are: (1) chain of effects, (2) market approaches, and (3) surveys. National gross damages of air pollution of $20.2 billion and of water pollution of $11.1 billion for 1973 are substantial. These best estimates, updated for the economic and demographic conditions, could provide acceptable control totals for estimating and predicting benefits and costs of abating air and water pollution emissions. The major issues to be resolved are: (1) lack of available noneconomic data, (2) theoretical and empirical difficulties of placing a value on human life and health and on benefits such as aesthetics, and (3) lack of available demographic and economic data.

  16. (/sup 3/)tetrahydrotrazodone binding. Association with serotonin binding sites

    SciTech Connect

    Kendall, D.A.; Taylor, D.P.; Enna, S.J.

    1983-05-01

    High (17 nM) and low (603 nM) affinity binding sites for (/sup 3/)tetrahydrotrazodone ((/sup 3/) THT), a biologically active analogue of trazodone, have been identified in rat brain membranes. The substrate specificity, concentration, and subcellular and regional distributions of these sites suggest that they may represent a component of the serotonin transmitter system. Pharmacological analysis of (/sup 3/)THT binding, coupled with brain lesion and drug treatment experiments, revealed that, unlike other antidepressants, (/sup 3/) THT does not attach to either a biogenic amine transporter or serotonin binding sites. Rather, it would appear that (/sup 3/)THT may be an antagonist ligand for the serotonin binding site. This probe may prove of value in defining the mechanism of action of trazodone and in further characterizing serotonin receptors.

  17. An Improved Method for Identifying Specific DNA-Protein-Binding Sites In Vitro.

    PubMed

    Wang, Liangyan; Lu, Huizhi; Wang, Yunguang; Yang, Su; Xu, Hong; Cheng, Kaiying; Zhao, Ye; Tian, Bing; Hua, Yuejin

    2017-03-01

    Binding of proteins to specific DNA sequences is essential for a variety of cellular processes such as DNA replication, transcription and responses to external stimuli. Chromatin immunoprecipitation is widely used for determining intracellular DNA fragments bound by a specific protein. However, the subsequent specific or accurate DNA-protein-binding sequence is usually determined by DNA footprinting. Here, we report an alternative method for identifying specific sites of DNA-protein-binding (designated SSDP) in vitro. This technique is mainly dependent on antibody-antigen immunity, simple and convenient, while radioactive isotope labeling and optimization of partial degradation by deoxyribonuclease (DNase) are avoided. As an example, the specific binding sequence of a target promoter by DdrO (a DNA damage response protein from Deinococcus radiodurans) in vitro was determined by the developed method. The central sequence of the binding site could be easily located using this technique.

  18. PUMA binding induces partial unfolding within BCL-xL to disrupt p53 binding and promote apoptosis.

    PubMed

    Follis, Ariele Viacava; Chipuk, Jerry E; Fisher, John C; Yun, Mi-Kyung; Grace, Christy R; Nourse, Amanda; Baran, Katherine; Ou, Li; Min, Lie; White, Stephen W; Green, Douglas R; Kriwacki, Richard W

    2013-03-01

    Following DNA damage, nuclear p53 induces the expression of PUMA, a BH3-only protein that binds and inhibits the antiapoptotic BCL-2 repertoire, including BCL-xL. PUMA, unique among BH3-only proteins, disrupts the interaction between cytosolic p53 and BCL-xL, allowing p53 to promote apoptosis via direct activation of the BCL-2 effector molecules BAX and BAK. Structural investigations using NMR spectroscopy and X-ray crystallography revealed that PUMA binding induced partial unfolding of two α-helices within BCL-xL. Wild-type PUMA or a PUMA mutant incapable of causing binding-induced unfolding of BCL-xL equivalently inhibited the antiapoptotic BCL-2 repertoire to sensitize for death receptor-activated apoptosis, but only wild-type PUMA promoted p53-dependent, DNA damage-induced apoptosis. Our data suggest that PUMA-induced partial unfolding of BCL-xL disrupts interactions between cytosolic p53 and BCL-xL, releasing the bound p53 to initiate apoptosis. We propose that regulated unfolding of BCL-xL provides a mechanism to promote PUMA-dependent signaling within the apoptotic pathways.

  19. Yeast DNA-repair gene RAD14 encodes a zinc metalloprotein with affinity for ultraviolet-damaged DNA

    SciTech Connect

    Guzder, S.N.; Sung, P.; Prakash, S. ); Prakash, L. )

    1993-06-15

    Xeroderma pigmentosum (XP) patients suffer from a high incidence of skin cancers due to a defect in excision repair of UV light-damaged DNA. Of the seven XP complementation groups, A--G, group A represents a severe and frequent form of the disease. The Saccharomyces cerevisiae RAD14 gene is a homolog of the XP-A correcting (XPAC) gene. Like XP-A cells, rad14-null mutants are defective in the incision step of excision repair of UV-damaged DNA. The authors have purified RAD14 protein to homogeneity from extract of a yeast strain genetically tailored to overexpress RAD14. As determined by atomic emission spectroscopy, RAD14 contains one zinc atom. They also show in vitro that RAD14 binds zinc but does not bind other divalent metal ions. In DNA mobility-shift assays, RAD14 binds specifically to UV-damaged DNA. Removal of cyclobutane pyrimidine dimers from damaged DNA by enzymatic photoreactivation has no effect on binding, strongly suggesting that RAD14 recognizes pyrimidine(6-4)pyrimidone photoproduct sites. These findings indicate that RAD14 functions in damage recognition during excision repair. 37 refs., 4 figs.

  20. Evidence against the nuclear in situ binding of arsenicals-oxidative stress theory of arsenic carcinogenesis

    EPA Science Inventory

    A large amount of evidence suggests that arsenicals act via oxidative stress in causing cancer in humans and experimental animals. It is possible that arsenicals could bind in situ close to nuclear DNA followed by Haber-Weiss type oxidative DNA damage. Therefore, we tested this...

  1. Damage Tolerance: Assessment Handbook. Volume 2: Airframe Damage Tolerance Evaluation

    DTIC Science & Technology

    1993-10-01

    Airframe Damage Tolerance Evaluation NJ 084u5 DTIC FLE 15 1994 Research and Special Programs Administration John A. Volpe National Transportation Systems...permission of John Wiley and Sons, New York, N.Y.] (4-5] CORRODED END Magnes;um Magnesium alloys Zinc Galvanized steel or galvanized wrought iron Aluminum...Reprinted from M M. Ratwani and D.P. Wilhem , iDeelopment andEvaluation of Methods of Plane Strain Fractuire Analysis, Northrop Corporation, AFFDL-TR-73-42

  2. 7 CFR 51.573 - Damage.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ..., CERTIFICATION, AND STANDARDS) United States Standards for Celery Definitions § 51.573 Damage. Damage, unless... the edible or shipping quality of the celery stalk or the general appearance of the stalks in...

  3. 7 CFR 51.573 - Damage.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... Standards for Celery Definitions § 51.573 Damage. Damage, unless otherwise specifically defined in this... the celery stalk or the general appearance of the stalks in the container. Any one of the...

  4. 7 CFR 51.573 - Damage.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... Standards for Celery Definitions § 51.573 Damage. Damage, unless otherwise specifically defined in this... the celery stalk or the general appearance of the stalks in the container. Any one of the...

  5. 7 CFR 51.586 - Serious damage.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... Standards for Celery Definitions § 51.586 Serious damage. Serious damage, unless otherwise specifically... shipping quality of the celery stalk or the general appearance of the stalks in the container. Any one...

  6. 7 CFR 51.586 - Serious damage.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... Standards for Celery Definitions § 51.586 Serious damage. Serious damage, unless otherwise specifically... shipping quality of the celery stalk or the general appearance of the stalks in the container. Any one...

  7. 7 CFR 29.3017 - Damage.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... Damage. The effect of mold, must, rot, black rot, or other fungous or bacterial diseases which attack tobacco in its cured state. Tobacco having the odor of mold, must, or rot is considered damaged. (See...

  8. 7 CFR 29.3017 - Damage.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... Damage. The effect of mold, must, rot, black rot, or other fungous or bacterial diseases which attack tobacco in its cured state. Tobacco having the odor of mold, must, or rot is considered damaged. (See...

  9. 7 CFR 51.908 - Serious damage.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... Definitions § 51.908 Serious damage. Serious damage means any defect or any combination of defects which... berries which are split, crushed, wet, affected by decay or waterberry, or affected by heat or...

  10. 7 CFR 29.3017 - Damage.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... Damage. The effect of mold, must, rot, black rot, or other fungous or bacterial diseases which attack tobacco in its cured state. Tobacco having the odor of mold, must, or rot is considered damaged. (See...

  11. 7 CFR 29.3017 - Damage.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... Damage. The effect of mold, must, rot, black rot, or other fungous or bacterial diseases which attack tobacco in its cured state. Tobacco having the odor of mold, must, or rot is considered damaged. (See...

  12. 7 CFR 29.3017 - Damage.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... Damage. The effect of mold, must, rot, black rot, or other fungous or bacterial diseases which attack tobacco in its cured state. Tobacco having the odor of mold, must, or rot is considered damaged. (See...

  13. Damage Tolerance of Large Shell Structures

    NASA Technical Reports Server (NTRS)

    Minnetyan, L.; Chamis, C. C.

    1999-01-01

    Progressive damage and fracture of large shell structures is investigated. A computer model is used for the assessment of structural response, progressive fracture resistance, and defect/damage tolerance characteristics. Critical locations of a stiffened conical shell segment are identified. Defective and defect-free computer models are simulated to evaluate structural damage/defect tolerance. Safe pressurization levels are assessed for the retention of structural integrity at the presence of damage/ defects. Damage initiation, growth, accumulation, and propagation to fracture are included in the simulations. Damage propagation and burst pressures for defective and defect-free shells are compared to evaluate damage tolerance. Design implications with regard to defect and damage tolerance of a large steel pressure vessel are examined.

  14. 7 CFR 51.1413 - Damage.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... Standards for Grades of Pecans in the Shell 1 Definitions § 51.1413 Damage. Damage means any specific defect... marketing quality of the individual pecan or the general appearance of the pecans in the lot. The...

  15. 7 CFR 51.1413 - Damage.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... Standards for Grades of Pecans in the Shell 1 Definitions § 51.1413 Damage. Damage means any specific defect... marketing quality of the individual pecan or the general appearance of the pecans in the lot. The...

  16. Progressive damage detection using noncontact measurements

    NASA Astrophysics Data System (ADS)

    Venkatappa, Suhas; Petro, Samer H.; EnChen, Shen; GangaRao, Hota V. S.

    1998-03-01

    This paper presents work in progress toward the development of a bridge condition assessment system. The system combines remote laser vibration sensing technology and a strain-energy- based damage detection algorithm. The results from vibration tests conducted on laboratory specimens with different degrees of damage are presented. The vibration signatures are acquired using Scanning Laser Vibrometers (SLV). The extracted mode shapes from these tests are then used in the damage detection algorithm. The preliminary results indicate that the strain energy differences are highly sensitive to damage, and can be used to locate and distinguish progressive damages. The combination of SLV technology and the damage detection algorithm makes remote sensing attractive for the monitoring and inspection of structures. Finite element simulation of a progressive damage at a single location is also presented to illustrate the sensitivity of the algorithm to increasing damages.

  17. DNA Damage and Repair in Vascular Disease.

    PubMed

    Uryga, Anna; Gray, Kelly; Bennett, Martin

    2016-01-01

    DNA damage affecting both genomic and mitochondrial DNA is present in a variety of both inherited and acquired vascular diseases. Multiple cell types show persistent DNA damage and a range of lesions. In turn, DNA damage activates a variety of DNA repair mechanisms, many of which are activated in vascular disease. Such DNA repair mechanisms either stall the cell cycle to allow repair to occur or trigger apoptosis or cell senescence to prevent propagation of damaged DNA. Recent evidence has indicated that DNA damage occurs early, is progressive, and is sufficient to impair function of cells composing the vascular wall. The consequences of persistent genomic and mitochondrial DNA damage, including inflammation, cell senescence, and apoptosis, are present in vascular disease. DNA damage can thus directly cause vascular disease, opening up new possibilities for both prevention and treatment. We review the evidence for and the causes, types, and consequences of DNA damage in vascular disease.

  18. Thermodynamics of interactions of vancomycin and synthetic surrogates of bacterial cell wall.

    PubMed

    Rekharsky, Mikhail; Hesek, Dusan; Lee, Mijoon; Meroueh, Samy O; Inoue, Yoshihisa; Mobashery, Shahriar

    2006-06-21

    Glycopeptide antibiotics, including vancomycin, form complexes via a set of five hydrogen bonds with the acyl-l-Lys-d-Ala-d-Ala portion of the peptidyl stems of the bacterial cell wall peptidoglycan. This complexation deprives the organism from the ability to cross-link peptidyl stems of the peptidoglycan, leading to bacterial cell death. Four synthetic fragments as surrogates of the components of the bacterial cell wall have been prepared in our lab in multistep syntheses. These synthetic samples were used in investigations of the thermodynamics properties (DeltaG degrees , DeltaH degrees , and TDeltaS degrees ) for the complexation with vancomycin by isothermal titration calorimetry (ITC). Complexation with the glycopeptide analogues is largely enthalpy-driven (formation of five hydrogen bonds), and in the analogues with a single peptidyl stem, the complexation is 1:1. The complexation is more complicated with an approximately 2 kDa cell wall surrogate (compound 4), which possesses two peptidyl stems. The data were suggestive of interactions between the two vancomycin molecules, with an entropic penalty attributable to restriction of molecular movements within the complex due to restriction of motion of the highly mobile acyl-d-Ala-d-Ala moiety of the peptidyl stems. These data were reconciled with the recently determined NMR solution structure for the peptidoglycan fragment 4 and its implications for the larger cell wall.

  19. Peptidase activity of beta-lactamases.

    PubMed Central

    Rhazi, N; Galleni, M; Page, M I; Frère, J M

    1999-01-01

    Although beta-lactamases have generally been considered as being devoid of peptidase activity, a low but significant hydrolysis of various N-acylated dipeptides was observed with representatives of each class of beta-lactamases. The kcat/Km values were below 0.1 M(-1). s(-1), but the enzyme rate enhancement factors were in the range 5000-20000 for the best substrates. Not unexpectedly, the best 'peptidase' was the class C beta-lactamase of Enterobacter cloacae P99, but, more surprisingly, the activity was always higher with the phenylacetyl- and benzoyl-d-Ala-d-Ala dipeptides than with the diacetyl- and alpha-acetyl-l-Lys-d-Ala-d-Ala tripeptides, which are the preferred substrates of the low-molecular-mass, soluble dd-peptidases. A comparison between the beta-lactamases and dd-peptidases showed that it might be as difficult for a dd-peptidase to open the beta-lactam ring as it is for the beta-lactamases to hydrolyse the peptides, an observation which can be explained by geometric and stereoelectronic considerations. PMID:10393100

  20. Glycopeptide resistance determinants from the teicoplanin producer Actinoplanes teichomyceticus.

    PubMed

    Serina, Stefania; Radice, Francesca; Maffioli, Sonia; Donadio, Stefano; Sosio, Margherita

    2004-11-01

    In enterococci and other pathogenic bacteria, high-level resistance to vancomycin and other glycopeptide antibiotics requires the action of the van genes, which direct the synthesis of peptidoglycan terminating in the depsipeptide D-alanyl-D-lactate, in place of the usual D-Ala-D-Ala. The Actinoplanes teichomyceticus tcp cluster, devoted to the biosynthesis of the glycopeptide antibiotic teicoplanin, contains van genes associated to a murF-like sequence (murF2). We show that A. teichomyceticus contains also a house-keeping murF1 gene, capable of complementing a temperature sensitive Escherichia coli murF mutant. MurF1, expressed in Streptomyces lividans, can catalyze the addition of either D-Ala-D-Ala or D-Ala-D-Lac to the UDP-N-acetyl-muramyl-L-Ala-D-Glu-d-Lys. However, similarly expressed MurF2 shows a small enzymatic activity only with D-Ala-D-lactate. Introduction of a single copy of the entire set of van genes confers resistance to teicoplanin-type glycopeptides to S. coelicolor.

  1. Moss Chloroplasts Are Surrounded by a Peptidoglycan Wall Containing D-Amino Acids[OPEN

    PubMed Central

    Hirano, Takayuki; Tanidokoro, Koji; Shimizu, Yasuhiro; Kawarabayasi, Yutaka; Ohshima, Toshihisa; Sato, Momo; Tadano, Shinji; Ishikawa, Hayato; Takio, Susumu; Takechi, Katsuaki; Takano, Hiroyoshi

    2016-01-01

    It is believed that the plastids in green plants lost peptidoglycan (i.e., a bacterial cell wall-containing d-amino acids) during their evolution from an endosymbiotic cyanobacterium. Although wall-like structures could not be detected in the plastids of green plants, the moss Physcomitrella patens has the genes required to generate peptidoglycan (Mur genes), and knocking out these genes causes defects in chloroplast division. Here, we generated P. patens knockout lines (∆Pp-ddl) for a homolog of the bacterial peptidoglycan-synthetic gene encoding d-Ala:d-Ala ligase. ∆Pp-ddl had a macrochloroplast phenotype, similar to other Mur knockout lines. The addition of d-Ala-d-Ala (DA-DA) to the medium suppressed the appearance of giant chloroplasts in ∆Pp-ddl, but the addition of l-Ala-l-Ala (LA-LA), DA-LA, LA-DA, or d-Ala did not. Recently, a metabolic method for labeling bacterial peptidoglycan was established using ethynyl-DA-DA (EDA-DA) and click chemistry to attach an azide-modified fluorophore to the ethynyl group. The ∆Pp-ddl line complemented with EDA-DA showed that moss chloroplasts are completely surrounded by peptidoglycan. Our findings strongly suggest that the moss plastids have a peptidoglycan wall containing d-amino acids. By contrast, no plastid phenotypes were observed in the T-DNA tagged ddl mutant lines of Arabidopsis thaliana. PMID:27325639

  2. Avionics Box Cold Plate Damage Prevention

    NASA Technical Reports Server (NTRS)

    Stambolian, Damon; Larcher, Steven; Henderson, Gena; Tran, Donald

    2011-01-01

    Over the years there have been several occurrences of damage to Space Shuttle Orbiter cold plates during removal and replacement of avionics boxes. Thus a process improvement team was put together to determine ways to prevent these kinds of damage. From this effort there were many solutions including, protective covers, training, and improved operations instructions. The focus of this paper is to explain the cold plate damage problem and the corrective actions for preventing future damage to aerospace avionics cold plate designs.

  3. Multilayer Thin Film Sensors for Damage Diagnostics

    NASA Astrophysics Data System (ADS)

    Protasov, A. G.; Gordienko, Y. G.; Zasimchuk, E. E.

    2006-03-01

    The new innovative approach to damage diagnostics within the production and maintenance/servicing procedures in industry is proposed. It is based on the real-time multiscale monitoring of the smart-designed multilayer thin film sensors of fatigue damage with the standard electrical input/output interfaces which can be connected to the embedded and on-board computers. The multilayer thin film sensors supply information about the actual unpredictable deformation damage, actual fatigue life, strain localization places, damage spreading, etc.

  4. Damage experiments in cylindrical geometry update

    SciTech Connect

    Kaul, Anne; Holtkamp, David; Rodriguez, George

    2009-01-01

    Using a cylindrical configuration to study spallation damage allows for a natural recollection of the damaged material under proper driving conditions. Previous experiments provided data about failure initiation in aluminum in a cylindrical geometry and the behavior of material recollected after damage from pressures in the damage initiation regime. The current series of experiments studied the behavior of material recollected after complete failure. Results from the current experiments will be presented.

  5. Sphingolipids in the DNA Damage Response

    PubMed Central

    Carroll, Brittany; Donaldson, Cat; Obeid, Lina

    2014-01-01

    Recently, sphingolipid metabolizing enzymes have emerged as important targets of many chemotherapeutics and DNA damaging agents and therefore play significant roles in mediating the physiological response of the cell to DNA damage. In this review we will highlight points of connection between the DNA damage response (DDR) and sphingolipid metabolism; specifically how certain sphingolipid enzymes are regulated in response to DNA damage and how the bioactive lipids produced by these enzymes affect cell fate. PMID:25434743

  6. Large area damage testing of optics

    SciTech Connect

    Sheehan, L.; Kozlowski, M.; Stolz, C.

    1996-04-26

    The damage threshold specifications for the National Ignition Facility will include a mixture of standard small-area tests and new large-area tests. During our studies of laser damage and conditioning processes of various materials we have found that some damage morphologies are fairly small and this damage does not grow with further illumination. This type of damage might not be detrimental to the laser performance. We should therefore assume that some damage can be allowed on the optics, but decide on a maximum damage allowance of damage. A new specification of damage threshold termed {open_quotes}functional damage threshold{close_quotes} was derived. Further correlation of damage size and type to system performance must be determined in order to use this measurement, but it is clear that it will be a large factor in the optics performance specifications. Large-area tests have verified that small-area testing is not always sufficient when the optic in question has defect-initiated damage. This was evident for example on sputtered polarizer and mirror coatings where the defect density was low enough that the features could be missed by standard small- area testing. For some materials, the scale-length at which damage non-uniformities occur will effect the comparison of small-area and large-area tests. An example of this was the sub-aperture tests on KD*P crystals on the Beamlet test station. The tests verified the large-area damage threshold to be similar to that found when testing a small-area. Implying that for this KD*P material, the dominate damage mechanism is of sufficiently small scale-length that small-area testing is capable of determining the threshold. The Beamlet test station experiments also demonstrated the use of on-line laser conditioning to increase the crystals damage threshold.

  7. Detecting ATM-Dependent Chromatin Modification in DNA Damage Response

    PubMed Central

    Udayakumar, Durga; Horikoshi, Nobuo; Mishra, Lope; Hunt, Clayton; Pandita, Tej K.

    2015-01-01

    Loss of function or mutation of the ataxia–telangiectasia mutated gene product (ATM) results in inherited genetic disorders characterized by neurodegeneration, immunodeficiency, and cancer. Ataxia-telangiectasia mutated (ATM) gene product belongs to the PI3K-like protein kinase (PIKKs) family and is functionally implicated in mitogenic signal transduction, chromosome condensation, meiotic recombination, cell-cycle control, and telomere maintenance. The ATM protein kinase is primarily activated in response to DNA double strand breaks (DSBs), the most deleterious form of DNA damage produced by ionizing radiation (IR) or radiomimetic drugs. It is detected at DNA damage sites, where ATM autophosphorylation causes dissociation of the inactive homodimeric form to the activated monomeric form. Interestingly, heat shock can activate ATM independent of the presence of DNA strand breaks. ATM is an integral part of the sensory machinery that detects DSBs during meiosis, mitosis, or DNA breaks mediated by free radicals. These DNA lesions can trigger higher order chromatin reorganization fuelled by posttranslational modifications of histones and histone binding proteins. Our group, and others, have shown that ATM activation is tightly regulated by chromatin modifications. This review summarizes the multiple approaches used to discern the role of ATM and other associated proteins in chromatin modification in response to DNA damage. PMID:25827888

  8. MECHANISTIC AND BIOLOGICAL ASPECTS OF HELICASE ACTION ON DAMAGED DNA

    PubMed Central

    Suhasini, Avvaru N.; Brosh, Robert M.

    2010-01-01

    Helicases catalytically unwind structured nucleic acids in a nucleoside-triphosphate-dependent and directionally specific manner, and are essential for virtually all aspects of nucleic acid metabolism. ATPase-driven helicases which translocate along nucleic acids play a role in damage recognition or unwinding of a DNA tract containing the lesion. Although classical biochemical experiments provided evidence that bulky covalent adducts inhibit DNA unwinding catalyzed by certain DNA helicases in a strand-specific manner (i.e. , block to DNA unwinding restricted to adduct residence in the strand the helicase translocates), recent studies suggest more complex arrangements that may depend on the helicase under study, its assembly in a protein complex, and the type of structural DNA perturbation. Moreover, base and sugar phosphate backbone modifications exert effects on DNA helicases that suggest specialized tracking mechanisms. As a component of the replication stress response, the single-stranded DNA binding protein Replication Protein A (RPA) may serve to enable eukaryotic DNA helicases to overcome certain base lesions. Helicases play important roles in DNA damage signaling which also involve their partnership with RPA. In this review, we will discuss our current understanding of mechanistic and biological aspects of helicase action on damaged DNA. PMID:20574162

  9. Rate process analysis of thermal damage in cartilage.

    PubMed

    Díaz, Sergio H; Nelson, J Stuart; Wong, Brian J F

    2003-01-07

    Cartilage laser thermoforming (CLT) is a new surgical procedure that allows in situ treatment of deformities in the head and neck with less morbidity than traditional approaches. While some animal and human studies have shown promising results, the clinical feasibility of CLT depends on preservation of chondrocyte viability, which has not been extensively studied. The present paper characterizes cellular damage due to heat in rabbit nasal cartilage. Damage was modelled as a first orderrate process for which two experimentally derived coefficients, A = 1.2 x 10(70) s(-1) and Ea = 4.5 x 10(5) J mole(-1), were determined by quantifying the decrease in concentration of healthy chondrocytes in tissue samples as a function of exposure time to constant-temperature water baths. After immersion, chondrocytes were enzymatically isolated from the matrix and stained with a two-component fluorescent dye. The dye binds nuclear DNA differentially depending upon chondrocyte viability. A flow cytometer was used to detect differential cell fluorescence to determine the percentage of live and dead cells in each sample. As a result, a damage kinetic model was obtained that can be used to predict the onset, extent and severity of cellular injury to thermal exposure.

  10. Rate process analysis of thermal damage in cartilage

    NASA Astrophysics Data System (ADS)

    Díaz, Sergio H.; Nelson, J. Stuart; Wong, Brian J. F.

    2003-01-01

    Cartilage laser thermoforming (CLT) is a new surgical procedure that allows in situ treatment of deformities in the head and neck with less morbidity than traditional approaches. While some animal and human studies have shown promising results, the clinical feasibility of CLT depends on preservation of chondrocyte viability, which has not been extensively studied. The present paper characterizes cellular damage due to heat in rabbit nasal cartilage. Damage was modelled as a first order rate process for which two experimentally derived coefficients, A = 1.2 × 1070 s-1 and Ea = 4.5 × 105 J mole-1, were determined by quantifying the decrease in concentration of healthy chondrocytes in tissue samples as a function of exposure time to constant-temperature water baths. After immersion, chondrocytes were enzymatically isolated from the matrix and stained with a two-component fluorescent dye. The dye binds nuclear DNA differentially depending upon chondrocyte viability. A flow cytometer was used to detect differential cell fluorescence to determine the percentage of live and dead cells in each sample. As a result, a damage kinetic model was obtained that can be used to predict the onset, extent and severity of cellular injury to thermal exposure.

  11. Superresolution microscopy with transient binding.

    PubMed

    Molle, Julia; Raab, Mario; Holzmeister, Susanne; Schmitt-Monreal, Daniel; Grohmann, Dina; He, Zhike; Tinnefeld, Philip

    2016-06-01

    For single-molecule localization based superresolution, the concentration of fluorescent labels has to be thinned out. This is commonly achieved by photophysically or photochemically deactivating subsets of molecules. Alternatively, apparent switching of molecules can be achieved by transient binding of fluorescent labels. Here, a diffusing dye yields bright fluorescent spots when binding to the structure of interest. As the binding interaction is weak, the labeling is reversible and the dye ligand construct diffuses back into solution. This approach of achieving superresolution by transient binding (STB) is reviewed in this manuscript. Different realizations of STB are discussed and compared to other localization-based superresolution modalities. We propose the development of labeling strategies that will make STB a highly versatile tool for superresolution microscopy at highest resolution.

  12. Chiral discrimination in optical binding

    NASA Astrophysics Data System (ADS)

    Forbes, Kayn A.; Andrews, David L.

    2015-05-01

    The laser-induced intermolecular force that exists between two or more particles in the presence of an electromagnetic field is commonly termed "optical binding." Distinct from the single-particle forces that are at play in optical trapping at the molecular level, the phenomenon of optical binding is a manifestation of the coupling between optically induced dipole moments in neutral particles. In other, more widely known areas of optics, there are many examples of chiral discrimination—signifying the different response a chiral material has to the handedness of an optical input. In the present analysis, extending previous work on chiral discrimination in optical binding, a mechanism is identified using a quantum electrodynamical approach. It is shown that the optical binding force between a pair of chiral molecules can be significantly discriminatory in nature, depending upon both the handedness of the interacting particles and the polarization of the incident light, and it is typically several orders of magnitude larger than previously reported.

  13. Microbial starch-binding domain.

    PubMed

    Rodríguez-Sanoja, Romina; Oviedo, Norma; Sánchez, Sergio

    2005-06-01

    Glucosidic bonds from different non-soluble polysaccharides such as starch, cellulose and xylan are hydrolyzed by amylases, cellulases and xylanases, respectively. These enzymes are produced by microorganisms. They have a modular structure that is composed of a catalytic domain and at least one non-catalytic domain that is involved in polysaccharide binding. Starch-binding modules are present in microbial enzymes that are involved in starch metabolism; these are classified into several different families on the basis of their amino acid sequence similarities. Such binding domains promote attachment to the substrate and increase its concentration at the active site of the enzyme, which allows microorganisms to degrade non-soluble starch. Fold similarities are better conserved than sequences; nevertheless, it is possible to notice two evolutionary clusters of microbial starch-binding domains. These domains have enormous potential as tags for protein immobilization, as well as for the tailoring of enzymes that play a part in polysaccharide metabolism.

  14. 7 CFR 51.2933 - Serious damage.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... Standards for Grades of Apricots Definitions § 51.2933 Serious damage. Serious Damage means any specific... or shipping quality of the apricot. The dimensions given for these defects are based on an apricot... larger or smaller apricots. The following specific defects shall be considered as serious damage:...

  15. 7 CFR 51.2933 - Serious damage.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... Standards for Grades of Apricots Definitions § 51.2933 Serious damage. Serious Damage means any specific... or shipping quality of the apricot. The dimensions given for these defects are based on an apricot... larger or smaller apricots. The following specific defects shall be considered as serious damage:...

  16. 7 CFR 51.2933 - Serious damage.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... Standards for Grades of Apricots Definitions § 51.2933 Serious damage. Serious Damage means any specific... or shipping quality of the apricot. The dimensions given for these defects are based on an apricot... larger or smaller apricots. The following specific defects shall be considered as serious damage:...

  17. 7 CFR 51.2128 - Damage.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 7 Agriculture 2 2010-01-01 2010-01-01 false Damage. 51.2128 Section 51.2128 Agriculture... Standards for Grades of Shelled Almonds Definitions § 51.2128 Damage. Damage means any defect which... the affected area on an individual kernel aggregates more than the equivalent of a circle...

  18. 7 CFR 51.2966 - Serious damage.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 7 Agriculture 2 2010-01-01 2010-01-01 false Serious damage. 51.2966 Section 51.2966 Agriculture... Standards for Grades of Walnuts in the Shell Definitions § 51.2966 Serious damage. Serious damage means any... discoloration covering a smaller area if the appearance is equally objectionable; (b) Perforated shells when...

  19. 7 CFR 51.1913 - Serious damage.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 7 Agriculture 2 2010-01-01 2010-01-01 false Serious damage. 51.1913 Section 51.1913 Agriculture... Consumer Standards for Fresh Tomatoes Definitions § 51.1913 Serious damage. Serious damage means any defect... diameter on a tomato 21/2 inches in diameter, or lighter colored, shallow scars covering a greater...

  20. 7 CFR 51.2655 - Damage.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 7 Agriculture 2 2010-01-01 2010-01-01 false Damage. 51.2655 Section 51.2655 Agriculture... Standards for Grades for Sweet Cherries 1 Definitions § 51.2655 Damage. Damage means any specific defect... appearance; (c) Hail injury when deep or not well healed, or when the aggregate area exceeds the area of...

  1. Damage Caused by the Rogue Trustee

    ERIC Educational Resources Information Center

    O'Banion, Terry

    2009-01-01

    Fifty-nine community college presidents and chancellors in 16 states report on the damage caused by rogue trustees. While the damage to presidents, other trustees, and faculty and staff is alarming, the damage these trustees cause the college suggests that the rogue trustee may be the single most destructive force ever to plague an educational…

  2. 7 CFR 51.573 - Damage.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 2 2014-01-01 2014-01-01 false Damage. 51.573 Section 51.573 Agriculture Regulations..., CERTIFICATION, AND STANDARDS) United States Standards for Celery Definitions § 51.573 Damage. Damage, unless... above the point of attachment to the base and the first node, or between a point 11/2 inches below...

  3. 7 CFR 51.2293 - Serious damage.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 2 2014-01-01 2014-01-01 false Serious damage. 51.2293 Section 51.2293 Agriculture... § 51.2293 Serious damage. Serious damage means any defect, other than color, which seriously affects... more than one-fourth of the kernel is severely shriveled, or a greater area is affected by...

  4. 7 CFR 51.3067 - Serious damage.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 2 2014-01-01 2014-01-01 false Serious damage. 51.3067 Section 51.3067 Agriculture..., CERTIFICATION, AND STANDARDS) United States Standards for Florida Avocados Definitions § 51.3067 Serious damage... any one defect, shall be considered as serious damage: (a) Anthracnose when any spot exceeds the...

  5. Damaging effects of visible light. Progress report

    SciTech Connect

    Williams, T.P.

    1980-01-01

    Research progress in studies of photodynamic damage of visual photoreceptors is presented. It was found the retina is not homogeneous in its susceptibility to light damage. Steady state rhodopsin levels have been evaluated in albino rats and in pigmented rats at several light intensities. Studies have continued of the effects of peroxidative photodynamic damage on the properties of rod outer segments. (ACR)

  6. Statistical Analysis of Japanese Structural Damage Data

    DTIC Science & Technology

    1977-01-01

    Calculated Peak Overpressure Data ......... .. 36 Figure 4. Frequency Functions for Assumed Damage Laws ............ .. 38 Figure 5. Conversions to Value of...Buildings .... .......... .. 92 Figure 20. Effect of Damage Law on Confidence Regions ............ .. 97 * Figure 21. Comparison of Confidence Limits on...Value of ad (Cumulative Log Normal Damage Law ) ...... ............ .. 99 * Figure 22. Comparison of Confidence Limits on Value of ad (Cumulative Log

  7. Myeloperoxidase binds to and kills Entamoeba histolytica trophozoites.

    PubMed

    Pacheco-Yépez, J; Rivera-Aguilar, V; Barbosa-Cabrera, E; Rojas Hernández, S; Jarillo-Luna, R A; Campos-Rodríguez, R

    2011-05-01

    During amebic invasion, neutrophils are a key component in either protecting against invading trophozoites or contributing to tissue damage. Upon degranulating or being lysed, neutrophils release toxic substances that can kill amebas as well as damage host tissue. In a previous study we identified a protein from nonspecifically stimulated peritoneal exudates of hamster that has peroxidase and marked amebicidal activity. In the current study we analyzed the in vitro amebicidal effect of purified hamster myeloperoxidase (MPO). The results demonstrate that MPO must bind directly to the surface of Entamoeba histolytica trophozoites in order to carry out amebicidal activity by using the H(2) O(2) produced by the amebas themselves. Myeloperoxidase-incubated amebas showed important morphological and ultrastructural alterations that increased with incubation time. Changes included an increase of vacuoles in the cytoplasm, a decrease of glycogen, alterations of nuclear morphology and disturbances in the plasma membrane culminating in complete ameba destruction.

  8. Boxing and chronic brain damage.

    PubMed

    Stiller, J W; Weinberger, D R

    1985-06-01

    A chronic, and at times, progressive neurologic syndrome associated with boxing has been recognized for some time by boxing fans and people involved with the sport. Since Martland's first description of the syndrome in 1929, there has been only one randomly selected study of ex-boxers, that of Roberts, which showed a 17 per cent prevalence of this syndrome among boxers who fought between 1929 and 1955. This syndrome can be progressive but often is not. Furthermore, the extent of occupational exposure is probably a significant risk factor. Because of this, it would be expected that the prevalance of the syndrome in the modern boxer, as well as the amateur, would be significantly less than during the first half of the century, and, indeed, several studies appear to support this. Recent studies provide evidence that brain damage does exist in modern boxers and suggests that "subclinical" brain damage is likely to be more prevalent than obvious clinical dysfunction. There is clearly a discrepancy between subclinical evidence of neurologic involvement (for example, an abnormal CT scan) and signs of clinical neurologic dysfunction (for example, clinical exam and neuropsychological testing). The latter tend to show less frequent and consistent evidence of brain damage in boxers than does the CT scan. Although it is tempting to assume that an abnormal CT scan presages the development of neurologic dysfunction, it is not clear that this is the case. The prevalence of the syndrome, risk for progression to functional deficit, warning signs, and the natural history cannot be defined at this time. The only way to better define these parameters would be a controlled prospective study, which has yet to be undertaken.

  9. Oxidative damage in dengue fever.

    PubMed

    Seet, Raymond C S; Lee, Chung-Yung J; Lim, Erle C H; Quek, Amy M L; Yeo, Leonard L L; Huang, Shan-Hong; Halliwell, Barry

    2009-08-15

    Oxidative stress may be important in the pathogenesis of dengue infection. Using accurate markers of oxidative damage, we assessed the extent of oxidative damage in dengue patients. The levels of hydroxyeicosatetraenoic acid products (HETEs), F(2)-isoprostanes (F(2)-IsoPs), and cholesterol oxidation products (COPs) were measured in 28 adult dengue patients and 28 age-matched study controls during the febrile, defervescent, and convalescent stages of infection. We compared the absolute and the percentage change in these markers in relation to key clinical parameters and inflammatory markers. The levels of total HETEs and total HETEs/arachidonate, total F(2)-IsoPs/arachidonate, and COPs/cholesterol were higher during the febrile compared to the convalescent level. Total HETEs correlated positively with admission systolic blood pressure (r=0.52, p<0.05), whereas an inverse relationship was found between 7beta-hydroxycholesterol and systolic and diastolic blood pressure (r=-0.61 and -0.59, respectively, p<0.01). The urinary F(2)-IsoP level was higher in urine during the febrile stage compared to the convalescent level. Despite lower total cholesterol levels during the febrile stage compared to convalescent levels, a higher percentage of cholesterol was found as COPs (7beta-, 24-, and 27-hydroxycholesterol). The levels of platelet-activating factor-acetylhydrolase activity, vascular cellular adhesion molecule-1, tumor necrosis factor-alpha, and high-sensitivity C-reactive protein were higher during the febrile stage compared to their convalescent levels (p<0.01). Markers of oxidative damage are altered during the various stages of dengue infection.

  10. Multi-Dimensional Damage Detection

    NASA Technical Reports Server (NTRS)

    Gibson, Tracy L. (Inventor); Williams, Martha K. (Inventor); Lewis, Mark E. (Inventor); Roberson, Luke B. (Inventor); Snyder, Sarah J. (Inventor); Medelius, Pedro J. (Inventor); Parks, Steven L. (Inventor)

    2016-01-01

    Methods and systems may provide for a structure having a plurality of interconnected panels, wherein each panel has a plurality of detection layers separated from one another by one or more non-detection layers. The plurality of detection layers may form a grid of conductive traces. Additionally, a monitor may be coupled to each grid of conductive traces, wherein the monitor is configured to detect damage to the plurality of interconnected panels in response to an electrical property change with respect to one or more of the conductive traces. In one example, the structure is part of an inflatable space platform such as a spacecraft or habitat.

  11. Spall Damage of Concrete Structures

    DTIC Science & Technology

    1988-06-01

    likely also strain-rate sensitive . Fiber-reinforced concrete has been extensively studied. Various types of fiber material, including nylon, glass, and...60/40) 0.95 AMATOL (50/50) 0.97 ANFO (9416 Am Nil Fuel oil) 0.82 1-100 Composition A-3 1.09 1.07 5-50 Composition B 1.11 0.98 5-50 Composition C-4 1.37...defines the boundary between threshold spall and medium spall and deflections less than 4.17 percent of the free span. The heavy damage curve defines

  12. Chemical binding affinity estimation using MSB

    NASA Astrophysics Data System (ADS)

    Weaver, John B.; Rauwerdink, Adam M.

    2011-03-01

    Binding affinity can be estimated in several ways in the laboratory but there is no viable way to estimate binding affinity in vivo without assumptions on the number of binding sites. Magnetic spectroscopy of nanoparticle Brownian motion, MSB, measures the rotational Brownian motion. The MSB signal is affected by nanoparticle binding affinity so it provides a mechanism to measure the chemical binding affinity. We present a possible mechanism to quantify the binding affinity and test that mechanism using viscous solutions.

  13. A Binary-Encounter-Bethe Approach to Simulate DNA Damage by the Direct Effect

    NASA Technical Reports Server (NTRS)

    Plante, Ianik; Cucinotta, Francis A.

    2013-01-01

    The DNA damage is of crucial importance in the understanding of the effects of ionizing radiation. The main mechanisms of DNA damage are by the direct effect of radiation (e.g. direct ionization) and by indirect effect (e.g. damage by.OH radicals created by the radiolysis of water). Despite years of research in this area, many questions on the formation of DNA damage remains. To refine existing DNA damage models, an approach based on the Binary-Encounter-Bethe (BEB) model was developed[1]. This model calculates differential cross sections for ionization of the molecular orbitals of the DNA bases, sugars and phosphates using the electron binding energy, the mean kinetic energy and the occupancy number of the orbital. This cross section has an analytic form which is quite convenient to use and allows the sampling of the energy loss occurring during an ionization event. To simulate the radiation track structure, the code RITRACKS developed at the NASA Johnson Space Center is used[2]. This code calculates all the energy deposition events and the formation of the radiolytic species by the ion and the secondary electrons as well. We have also developed a technique to use the integrated BEB cross section for the bases, sugar and phosphates in the radiation transport code RITRACKS. These techniques should allow the simulation of DNA damage by ionizing radiation, and understanding of the formation of double-strand breaks caused by clustered damage in different conditions.

  14. Pharmacological activity of metal binding agents that alter copper bioavailability

    PubMed Central

    Helsel, Marian E.

    2015-01-01

    Iron, copper and zinc are required nutrients for many organisms but also potent toxins if misappropriated. An overload of any of these metals can be cytotoxic and ultimately lead to organ failure, whereas deficiencies can result in anemia, weakened immune system function, and other medical conditions. Cellular metal imbalances have been implicated in neurodegenerative diseases, cancer and infection. It is therefore critical for living organisms to maintain careful control of both the total levels and subcellular distributions of these metals to maintain healthy function. This perspective explores several strategies envisioned to alter the bioavailability of metal ions by using synthetic metal-binding agents targeted for diseases where misappropriated metal ions are suspected of exacerbating cellular damage. Specifically, we discuss chemical properties that influence the pharmacological outcome of a subset of metal-binding agents known as ionophores, and review several examples that have shown multiple pharmacological activities in metal-related diseases, with a specific focus on copper. PMID:25797044

  15. Non-contact intracellular binding of chloroplasts in vivo

    NASA Astrophysics Data System (ADS)

    Li, Yuchao; Xin, Hongbao; Liu, Xiaoshuai; Li, Baojun

    2015-06-01

    Non-contact intracellular binding and controllable manipulation of chloroplasts in vivo was demonstrated using an optical fiber probe. Launching a 980-nm laser beam into a fiber, which was placed about 3 μm above the surface of a living plant (Hydrilla verticillata) leaf, enabled stable binding of different numbers of chloroplasts, as well as their arrangement into one-dimensional chains and two-dimensional arrays inside the leaf without damaging the chloroplasts. Additionally, the formed chloroplast chains were controllably transported inside the living cells. The optical force exerted on the chloroplasts was calculated to explain the experimental results. This method provides a flexible method for studying intracellular organelle interaction with highly organized organelle-organelle contact in vivo in a non-contact manner.

  16. Structure-based Analysis to Hu-DNA Binding

    SciTech Connect

    Swinger,K.; Rice, P.

    2007-01-01

    HU and IHF are prokaryotic proteins that induce very large bends in DNA. They are present in high concentrations in the bacterial nucleoid and aid in chromosomal compaction. They also function as regulatory cofactors in many processes, such as site-specific recombination and the initiation of replication and transcription. HU and IHF have become paradigms for understanding DNA bending and indirect readout of sequence. While IHF shows significant sequence specificity, HU binds preferentially to certain damaged or distorted DNAs. However, none of the structurally diverse HU substrates previously studied in vitro is identical with the distorted substrates in the recently published Anabaena HU(AHU)-DNA cocrystal structures. Here, we report binding affinities for AHU and the DNA in the cocrystal structures. The binding free energies for formation of these AHU-DNA complexes range from 10-14.5 kcal/mol, representing K{sub d} values in the nanomolar to low picomolar range, and a maximum stabilization of at least 6.3 kcal/mol relative to complexes with undistorted, non-specific DNA. We investigated IHF binding and found that appropriate structural distortions can greatly enhance its affinity. On the basis of the coupling of structural and relevant binding data, we estimate the amount of conformational strain in an IHF-mediated DNA kink that is relieved by a nick (at least 0.76 kcal/mol) and pinpoint the location of the strain. We show that AHU has a sequence preference for an A+T-rich region in the center of its DNA-binding site, correlating with an unusually narrow minor groove. This is similar to sequence preferences shown by the eukaryotic nucleosome.

  17. Structure-based analysis of HU-DNA binding.

    PubMed

    Swinger, Kerren K; Rice, Phoebe A

    2007-01-26

    HU and IHF are prokaryotic proteins that induce very large bends in DNA. They are present in high concentrations in the bacterial nucleoid and aid in chromosomal compaction. They also function as regulatory cofactors in many processes, such as site-specific recombination and the initiation of replication and transcription. HU and IHF have become paradigms for understanding DNA bending and indirect readout of sequence. While IHF shows significant sequence specificity, HU binds preferentially to certain damaged or distorted DNAs. However, none of the structurally diverse HU substrates previously studied in vitro is identical with the distorted substrates in the recently published Anabaena HU(AHU)-DNA cocrystal structures. Here, we report binding affinities for AHU and the DNA in the cocrystal structures. The binding free energies for formation of these AHU-DNA complexes range from approximately 10-14.5 kcal/mol, representing K(d) values in the nanomolar to low picomolar range, and a maximum stabilization of at least approximately 6.3 kcal/mol relative to complexes with undistorted, non-specific DNA. We investigated IHF binding and found that appropriate structural distortions can greatly enhance its affinity. On the basis of the coupling of structural and relevant binding data, we estimate the amount of conformational strain in an IHF-mediated DNA kink that is relieved by a nick (at least 0.76 kcal/mol) and pinpoint the location of the strain. We show that AHU has a sequence preference for an A+T-rich region in the center of its DNA-binding site, correlating with an unusually narrow minor groove. This is similar to sequence preferences shown by the eukaryotic nucleosome.

  18. Towards a canonical elastoplastic damage model

    NASA Astrophysics Data System (ADS)

    Taher, Salah El-Din F.; Baluch, Mohammed H.; Al-Gadhib, Ali H.

    1994-05-01

    Fundamental aspects of elastoplastic damage are outlined. Time-independent isotropic damage is considered in order to study material degradation. By splitting the total strain tensor into its components of elastic damage and plastic damage and using recoverable energy equivalence, three distinct modes of behavior are particularized. For each mode of behavior, a suitable damage variable is culled. An in-depth analysis of this formulation reveals a certain incongruity in the assumptions postulated in some of the previously proposed models. The suggested generalized concepts are supported by experimental evidence.

  19. Binding of cellulose binding modules reveal differences between cellulose substrates

    PubMed Central

    Arola, Suvi; Linder, Markus B.

    2016-01-01

    The interaction between cellulase enzymes and their substrates is of central importance to several technological and scientific challenges. Here we report that the binding of cellulose binding modules (CBM) from Trichoderma reesei cellulases Cel6A and Cel7A show a major difference in how they interact with substrates originating from wood compared to bacterial cellulose. We found that the CBM from TrCel7A recognizes the two substrates differently and as a consequence shows an unexpected way of binding. We show that the substrate has a large impact on the exchange rate of the studied CBM, and moreover, CBM-TrCel7A seems to have an additional mode of binding on wood derived cellulose but not on cellulose originating from bacterial source. This mode is not seen in double CBM (DCBM) constructs comprising both CBM-TrCel7A and CBM-TrCel6A. The linker length of DCBMs affects the binding properties, and slows down the exchange rates of the proteins and thus, can be used to analyze the differences between the single CBM. These results have impact on the cellulase research and offer new understanding on how these industrially relevant enzymes act. PMID:27748440

  20. The binding domain structure of retinoblastoma-binding proteins.

    PubMed Central

    Figge, J.; Breese, K.; Vajda, S.; Zhu, Q. L.; Eisele, L.; Andersen, T. T.; MacColl, R.; Friedrich, T.; Smith, T. F.

    1993-01-01

    The retinoblastoma gene product (Rb), a cellular growth suppressor, complexes with viral and cellular proteins that contain a specific binding domain incorporating three invariant residues: Leu-X-Cys-X-Glu, where X denotes a nonconserved residue. Hydrophobic and electrostatic properties are strongly conserved in this segment even though the nonconserved amino acids vary considerably from one Rb-binding protein to another. In this report, we present a diagnostic computer pattern for a high-affinity Rb-binding domain featuring the three conserved residues as well as the conserved physico-chemical properties. Although the pattern encompasses only 10 residues (with only 4 of these explicitly defined), it exhibits 100% sensitivity and 99.95% specificity in database searches. This implies that a certain pattern of structural and physico-chemical properties encoded by this short sequence is sufficient to govern specific Rb binding. We also present evidence that the secondary structural conformation through this region is important for effective Rb binding. PMID:8382993

  1. Effect of polishing induced subsurface damages on laser induced damage in fused silica optics

    NASA Astrophysics Data System (ADS)

    He, Xiang; Zhao, Heng; Huang, Ying; Cai, Chao; Hu, JiangChuan; Ma, Ping

    2016-10-01

    Conventional used ceria polishing would induce both of Ce contaminants and subsurface damages, which mainly restricts the laser induced damage resistance of fused silica optics. To control the near surface defects, nanometer sized colloidal silica are used to polish fused silica optics after the normal ceria polishing process. Then the contaminant elements and subsurface damages of the polished samples were analyzed by secondary ion mass spectrometry and Nomarski microscopy. It reveals that ceria polishing would introduce lots of subsurface damages whereas colloidal silica polishing induces much fewer subsurface damages especially no fracture induced severe subsurface damages. The laser damage tests reveal that subsequent colloidal silica polishing of the ceria pre-polished samples could gradually eliminate the ceria polishing induced subsurface damages and lower the laser induced damage density accordingly with the increased polishing time. But unlike the damage density, only the severe subsurface damages are totally eliminated could the damage threshold be substantially improved. These results incline to indicate that the subsurface damages have great influence on the laser induced damage density and the fracture related severe subsurface damages will greatly restrict the damage threshold in polished optics.

  2. Screening and Initial Binding Assessment of Fumonisin B1 Aptamers

    PubMed Central

    McKeague, Maureen; Bradley, Charlotte R.; De Girolamo, Annalisa; Visconti, Angelo; Miller, J. David; DeRosa, Maria C.

    2010-01-01

    Fumonisins are mycotoxins produced by Fusarium verticillioides and F. proliferatum, fungi that are ubiquitous in corn (maize). Insect damage and some other environmental conditions result in the accumulation of fumonisins in corn-based products worldwide. Current methods of fumonisin detection rely on the use of immunoaffinity columns and high-performance liquid chromatography (HPLC). The use of aptamers offers a good alternative to the use of antibodies in fumonisin cleanup and detection due to lower costs and improved stability. Aptamers are single-stranded oligonucleotides that are selected using Systematic Evolution of Ligands by EXponential enrichment (SELEX) for their ability to bind to targets with high affinity and specificity. Sequences obtained after 18 rounds of SELEX were screened for their ability to bind to fumonisin B1. Six unique sequences were obtained, each showing improved binding to fumonisin B1 compared to controls. Sequence FB1 39 binds to fumonisin with a dissociation constant of 100 ± 30 nM and shows potential for use in fumonisin biosensors and solid phase extraction columns. PMID:21614178

  3. Putting life on ice: bacteria that bind to frozen water

    PubMed Central

    Bernheim, Reut; Guo, Shuaiqi; Davies, Peter L.; Braslavsky, Ido

    2016-01-01

    Ice-binding proteins (IBPs) are typically small, soluble proteins produced by cold-adapted organisms to help them avoid ice damage by either resisting or tolerating freezing. By contrast, the IBP of the Antarctic bacterium Marinomonas primoryensis is an extremely long, 1.5 MDa protein consisting of five different regions. The fourth region, a 34 kDa domain, is the only part that confers ice binding. Bioinformatic studies suggest that this IBP serves as an adhesin that attaches the bacteria to ice to keep it near the top of the water column, where oxygen and nutrients are available. Using temperature-controlled cells and a microfluidic apparatus, we show that M. primoryensis adheres to ice and is only released when melting occurs. Binding is dependent on the mobility of the bacterium and the functionality of the IBP domain. A polyclonal antibody raised against the IBP region blocks bacterial ice adhesion. This concept may be the basis for blocking biofilm formation in other bacteria, including pathogens. Currently, this IBP is the only known example of an adhesin that has evolved to bind ice. PMID:27534698

  4. Repair of Nerve Cell Membrance Damage by Calcium-Dependent, Membrane-Binding Proteins

    DTIC Science & Technology

    2013-09-01

    properties of the annexin core domain. Once developed for this primordial function , the domain may have been adopted to mediate additional calcium-regulated...described below (Aim 4) to test the functionality of the protein in membrane repair. Figure 2: Application of copine to a supported bilayer with defects...aggregation, the release rate was analyzed as a function of the vesicle concentration, anticipating that the rate would be second order relative to

  5. Repair of Nerve Cell Membrane Damage by Calcium-Dependent, Membrane-Binding Proteins

    DTIC Science & Technology

    2011-09-01

    peroxidation by hydrogen peroxide (see Figures 3 and 4 of the appended Preliminary Patent Application). 9 Similar studies are now being conducted with...described in the statement of work. Aim 1. Express human copine as a recombinant protein in yeast and purify. Human copine I (a close homolog of...copine VI expressed in neural tissue) was expressed from a galactose 10 (GAL10) -regulated yeast expression plasmid (4) and purified by calcium

  6. Chromatin structure and DNA damage

    SciTech Connect

    Gale, J.M.

    1987-01-01

    This dissertation examines the structure and structural transitions of chromatin in relation to DNA damage. The ability of intact and histone H1 depleted chromatin fibers to fold into higher ordered structures in vitro was examined following DNA photodamage introduced by two different agents. (1) 254-nm UV radiation and (2) trimethylpsoralen (plus near-UV radiation). Both agents are highly specific for DNA and form adducts predicted to cause different degrees of distortion in the DNA helix. The salt-induced structural transitions of intact and histone H1 depleted chromatin fibers were monitored by both analytical ultracentrifugation and light scattering. Our results show that even in the presence of extremely large, nonphysiological amounts of photodamage by either agent the ability of chromatin to fold into higher ordered structures is not affected. The compact, 30 nm fiber must therefore be able to accommodate a large amount of DNA damage without any measurable changes in the overall size or degree of compaction of this structure. The distribution of pyrimidine dimers was mapped at the single nucleotide level in nucleosome core DNA from UV-irradiated mononucleosomes, chromatin fibers, and human cells in culture using the 3' ..-->.. 5' exonuclease activity of T4 DNA polymerase.

  7. Air pollution and brain damage.

    PubMed

    Calderón-Garcidueñas, Lilian; Azzarelli, Biagio; Acuna, Hilda; Garcia, Raquel; Gambling, Todd M; Osnaya, Norma; Monroy, Sylvia; DEL Tizapantzi, Maria Rosario; Carson, Johnny L; Villarreal-Calderon, Anna; Rewcastle, Barry

    2002-01-01

    Exposure to complex mixtures of air pollutants produces inflammation in the upper and lower respiratory tract. Because the nasal cavity is a common portal of entry, respiratory and olfactory epithelia are vulnerable targets for toxicological damage. This study has evaluated, by light and electron microscopy and immunohistochemical expression of nuclear factor-kappa beta (NF-kappaB) and inducible nitric oxide synthase (iNOS), the olfactory and respiratory nasal mucosae, olfactory bulb, and cortical and subcortical structures from 32 healthy mongrel canine residents in Southwest Metropolitan Mexico City (SWMMC), a highly polluted urban region. Findings were compared to those in 8 dogs from Tlaxcala, a less polluted, control city. In SWMMC dogs, expression of nuclear neuronal NF-kappaB and iNOS in cortical endothelial cells occurred at ages 2 and 4 weeks; subsequent damage included alterations of the blood-brain barrier (BBB), degenerating cortical neurons, apoptotic glial white matter cells, deposition of apolipoprotein E (apoE)-positive lipid droplets in smooth muscle cells and pericytes, nonneuritic plaques, and neurofibrillary tangles. Persistent pulmonary inflammation and deteriorating olfactory and respiratory barriers may play a role in the neuropathology observed in the brains of these highly exposed canines. Neurodegenerative disorders such as Alzheimer's may begin early in life with air pollutants playing a crucial role.

  8. Malnutrition, liver damage, and cancer.

    PubMed

    Grasso, P

    1981-01-01

    There is no clear indication that malnutrition, per se, is a principal cause of cancer in man, but the prevalence of liver cancer in areas where malnutrition exists supports this hypothesis. Liver damage and liver cancer have been induced in laboratory rats by diets consisting of peanut meal and proteins deficient in some essential amino acids. However, liver damage, but not cancer, was produced when the diets contained no peanut meal but consisted of a mixture of amino acids deficient in methionine and cysteine, so that it is possible that aflatoxin, a contaminant of peanut meal, may have been responsible for the malignancies seen in the earlier experiments. Liver cancer developes in a high proportion of mice allowed to feed ad libitum or given a diet containing a high proportion of fat (groundnut oil) or protein (casein). Dietary restriction reduced the incidences of this cancer. This findings lends some support to current thinking that diet may be a factor in the development of cancer in man.

  9. Genetic Doping and Health Damages

    PubMed Central

    Fallahi, AA; Ravasi, AA; Farhud, DD

    2011-01-01

    Background: Use of genetic doping or gene transfer technology will be the newest and the lethal method of doping in future and have some unpleasant consequences for sports, athletes, and outcomes of competitions. The World Anti-Doping Agency (WADA) defines genetic doping as “the non-therapeutic use of genes, genetic elements, and/or cells that have the capacity to enhance athletic performance ”. The purpose of this review is to consider genetic doping, health damages and risks of new genes if delivered in athletes. Methods: This review, which is carried out by reviewing relevant publications, is primarily based on the journals available in GOOGLE, ELSEVIER, PUBMED in fields of genetic technology, and health using a combination of keywords (e.g., genetic doping, genes, exercise, performance, athletes) until July 2010. Conclusion: There are several genes related to sport performance and if they are used, they will have health risks and sever damages such as cancer, autoimmunization, and heart attack. PMID:23113049

  10. Cell damage seen from Chernobyl

    SciTech Connect

    Not Available

    1992-07-24

    The 30 kilometer radius forbidden zone around the Chernobyl atomic plant serves as a sobering reminder of the world's worst nuclear accident. But for former Soviet biologists, it's also a unique natural laboratory. And one scientist, Nadejda Gulaya of Kiev's Pallaguine Institute of Biochemistry, has been doing studies that she claims offer surprising evidence of Chernobyl's after-effects. Prolonged exposure to radioactive fallout from the 1986 accident, she says, has caused damage to cell membranes in both animals and humans. For the past year, Gulaya has been comparing tissues from animals such as mink, pigs, and rodents inhabiting the Chernobyl area with those from other parts of Ukraine. Her conclusion: Exposure to radiation has, in many cases, caused alterations to membrane phospholipids. These changes, are similar to those that disrupt cellular metabolism following exposure to oxidizing free radicals. Gulaya also has preliminary data from human studies. She claims to have found similar alterations in the neurons of people who have died since being exposed to Chernobyl radiation. That leads her to speculate that the frequent psychiatric disorders may not just be from mental stress or radiophobia, but might reflect actual damage to the central nervous system.

  11. Flood damage modelling: ambition and reality

    NASA Astrophysics Data System (ADS)

    Gerl, Tina; Kreibich, Heidi; Franco, Guillermo; Marechal, David; Schröter, Kai

    2015-04-01

    Flood damage modelling is of increasing importance for reliable risk assessment and management. Research efforts have improved the understanding of damaging processes and more sophisticated flood damage models have been developed. However, research seems to focus on a limited number of sectors and regions and validation of models still receives too little attention. We present a global inventory of flood damage models which is compiled from a review of scientific papers and research reports on flood damage models. The models are catalogued according to model specifications, geographical characteristics, sectors addressed, input variables used, model validation, transferability and model functions. The inventory is evaluated to position the current state of science and technology in flood damage modelling as well as to derive requirements for benchmarking damage models.

  12. Wiring Damage Analyses for STS OV-103

    NASA Technical Reports Server (NTRS)

    Thomas, Walter, III

    2006-01-01

    This study investigated the Shuttle Program s belief that Space Transportation System (STS) wiring damage occurrences are random, that is, a constant occurrence rate. Using Problem Reporting and Corrective Action (PRACA)-derived data for STS Space Shuttle OV-103, wiring damage was observed to increase over the vehicle s life. Causal factors could include wiring physical deterioration, maintenance and inspection induced damage, and inspection process changes resulting in more damage events being reported. Induced damage effects cannot be resolved with existent data. Growth analysis (using Crow-AMSAA, or CA) resolved maintenance/inspection effects (e.g., heightened awareness) on all wire damages and indicated an overall increase since Challenger Return-to-Flight (RTF). An increasing failure or occurrence rate per flight cycle was seen for each wire damage mode; these (individual) rates were not affected by inspection process effects, within statistical error.

  13. Brittle damage models in DYNA2D

    SciTech Connect

    Faux, D.R.

    1997-09-01

    DYNA2D is an explicit Lagrangian finite element code used to model dynamic events where stress wave interactions influence the overall response of the system. DYNA2D is often used to model penetration problems involving ductile-to-ductile impacts; however, with the advent of the use of ceramics in the armor-anti-armor community and the need to model damage to laser optics components, good brittle damage models are now needed in DYNA2D. This report will detail the implementation of four brittle damage models in DYNA2D, three scalar damage models and one tensor damage model. These new brittle damage models are then used to predict experimental results from three distinctly different glass damage problems.

  14. Damage experiments in a cylindrical geometry

    SciTech Connect

    Kaul, Ann M

    2010-09-21

    Studying spallation damage with a cylindrical configuration allows for a natural recollection of the damaged material under proper driving conditions. Additionally, the damaged material can come to a complete rest without the application of further stopping forces. Specific areas of research include the damage initiation regime in convergent geometry, behavior of material recollected after damage, and effects of convergent geometry on the material response. Such experiments produce unique strain and shear stress states, motivating improvements in existing computational material models and increasing the predictive capabilities of codes. A LANL/VNIIEF joint experimental series has produced cylindrical aluminum failure initiation data and studied the behavior of material recollected after damage initiation and after complete failure. In addition to post-shot collection of the damaged target material for subsequent metallographic analysis, dynamic in-situ experimental diagnostics include velocimetry and transverse radial radiography. This paper will discuss the current experimental status.

  15. Damage prediction in incremental forming by using Lemaitre damage model

    NASA Astrophysics Data System (ADS)

    Wu, Shenghua; Reis, Ana; Teixeira, Pedro; da Rocha, A. Barata; Lino, Jorge

    2012-09-01

    Incremental forming is an innovative flexible method used for manufacturing of the sheet metal products and brings a great insight for the small-batch-size or customized sheet products. Some experiments show that incremental sheet metal forming can undergo higher deformations than traditional sheet metal forming. The traditional method to evaluate formability like forming limit curve (FLD) etc can't give the right answer in incremental forming which is subjected to highly non-monotonic serrated strain paths. In this paper, the Lemaitre' damage model is presented and fully coupled with finite element simulation in commercial software ABAQUS to predict the failure in incremental forming. Results show that the prediction makes a great agreement with the relevant experiments.

  16. Damage threshold measurements: Self-focusing or intrinsic damage?

    NASA Astrophysics Data System (ADS)

    Efimov, Oleg

    2013-11-01

    The laser-induced damage (LID) thresholds of pure fused silica (Corning 7980) have been measured with single temporal mode nanosecond pulses at 1064 nm. The laser beam has been focused by spherical and conical lenses into 1.6 μm diameter spots. In the case of pseudo-Bessel beam (conical lens) which inherently was not subjected to self-focusing the threshold has been close to the intrinsic threshold in fused silica. However, the measurement with pseudo-Gaussian beam (spherical lens) has shown about 30% lower value of threshold. Complete identity in the cross-section distributions of beam intensities and considerable difference in measured thresholds indicate that self-focusing influence on the LID of dielectrics even for tight focused laser beams.

  17. Single-stranded DNA Binding by the Helix-Hairpin-Helix Domain of XPF Protein Contributes to the Substrate Specificity of the ERCC1-XPF Protein Complex.

    PubMed

    Das, Devashish; Faridounnia, Maryam; Kovacic, Lidija; Kaptein, Robert; Boelens, Rolf; Folkers, Gert E

    2017-02-17

    The nucleotide excision repair protein complex ERCC1-XPF is required for incision of DNA upstream of DNA damage. Functional studies have provided insights into the binding of ERCC1-XPF to various DNA substrates. However, because no structure for the ERCC1-XPF-DNA complex has been determined, the mechanism of substrate recognition remains elusive. Here we biochemically characterize the substrate preferences of the helix-hairpin-helix (HhH) domains of XPF and ERCC-XPF and show that the binding to single-stranded DNA (ssDNA)/dsDNA junctions is dependent on joint binding to the DNA binding domain of ERCC1 and XPF. We reveal that the homodimeric XPF is able to bind various ssDNA sequences but with a clear preference for guanine-containing substrates. NMR titration experiments and in vitro DNA binding assays also show that, within the heterodimeric ERCC1-XPF complex, XPF specifically recognizes ssDNA. On the other hand, the HhH domain of ERCC1 preferentially binds dsDNA through the hairpin region. The two separate non-overlapping DNA binding domains in the ERCC1-XPF heterodimer jointly bind to an ssDNA/dsDNA substrate and, thereby, at least partially dictate the incision position during damage removal. Based on structural models, NMR titrations, DNA-binding studies, site-directed mutagenesis, charge distribution, and sequence conservation, we propose that the HhH domain of ERCC1 binds to dsDNA upstream of the damage, and XPF binds to the non-damaged strand within a repair bubble.

  18. FANCI protein binds to DNA and interacts with FANCD2 to recognize branched structures.

    PubMed

    Yuan, Fenghua; El Hokayem, Jimmy; Zhou, Wen; Zhang, Yanbin

    2009-09-04

    In this study, we report that the purified wild-type FANCI (Fanconi anemia complementation group I) protein directly binds to a variety of DNA substrates. The DNA binding domain roughly encompasses residues 200-1000, as suggested by the truncation study. When co-expressed in insect cells, a small fraction of FANCI forms a stable complex with FANCD2 (Fanconi anemia complementation group D2). Intriguingly, the purified FANCI-FANCD2 complex preferentially binds to the branched DNA structures when compared with either FANCI or FANCD2 alone. Co-immunoprecipitation with purified proteins indicates that FANCI interacts with FANCD2 through its C-terminal amino acid 1001-1328 fragment. Although the C terminus of FANCI is dispensable for direct DNA binding, it seems to be involved in the regulation of DNA binding activity. This notion is further enhanced by two C-terminal point mutations, R1285Q and D1301A, which showed differentiated DNA binding activity. We also demonstrate that FANCI forms discrete nuclear foci in HeLa cells in the absence or presence of exogenous DNA damage. The FANCI foci are colocalized perfectly with FANCD2 and partially with proliferating cell nuclear antigen irrespective of mitomycin C treatment. An increased number of FANCI foci form and become resistant to Triton X extraction in response to mitomycin C treatment. Our data suggest that the FANCI-FANCD2 complex may participate in repair of damaged replication forks through its preferential recognition of branched structures.

  19. Chimeric proteins for detection and quantitation of DNA mutations, DNA sequence variations, DNA damage and DNA mismatches

    DOEpatents

    McCutchen-Maloney, Sandra L.

    2002-01-01

    Chimeric proteins having both DNA mutation binding activity and nuclease activity are synthesized by recombinant technology. The proteins are of the general formula A-L-B and B-L-A where A is a peptide having DNA mutation binding activity, L is a linker and B is a peptide having nuclease activity. The chimeric proteins are useful for detection and identification of DNA sequence variations including DNA mutations (including DNA damage and mismatches) by binding to the DNA mutation and cutting the DNA once the DNA mutation is detected.

  20. The prion protein binds thiamine.

    PubMed

    Perez-Pineiro, Rolando; Bjorndahl, Trent C; Berjanskii, Mark V; Hau, David; Li, Li; Huang, Alan; Lee, Rose; Gibbs, Ebrima; Ladner, Carol; Dong, Ying Wei; Abera, Ashenafi; Cashman, Neil R; Wishart, David S

    2011-11-01

    Although highly conserved throughout evolution, the exact biological function of the prion protein is still unclear. In an effort to identify the potential biological functions of the prion protein we conducted a small-molecule screening assay using the Syrian hamster prion protein [shPrP(90-232)]. The screen was performed using a library of 149 water-soluble metabolites that are known to pass through the blood-brain barrier. Using a combination of 1D NMR, fluorescence quenching and surface plasmon resonance we identified thiamine (vitamin B1) as a specific prion ligand with a binding constant of ~60 μM. Subsequent studies showed that this interaction is evolutionarily conserved, with similar binding constants being seen for mouse, hamster and human prions. Various protein construct lengths, both with and without the unstructured N-terminal region in the presence and absence of copper, were examined. This indicates that the N-terminus has no influence on the protein's ability to interact with thiamine. In addition to thiamine, the more biologically abundant forms of vitamin B1 (thiamine monophosphate and thiamine diphosphate) were also found to bind the prion protein with similar affinity. Heteronuclear NMR experiments were used to determine thiamine's interaction site, which is located between helix 1 and the preceding loop. These data, in conjunction with computer-aided docking and molecular dynamics, were used to model the thiamine-binding pharmacophore and a comparison with other thiamine binding proteins was performed to reveal the common features of interaction.

  1. Projecting global tropical cyclone economic damages with validation of tropical cyclone economic damage model

    NASA Astrophysics Data System (ADS)

    Iseri, Y.; Iwasaki, A.; Miyazaki, C.; Kanae, S.

    2014-12-01

    Tropical cyclones (TCs) sometimes cause serious damages to human society and thus possible changes of TC properties in the future have been concerned. In fact, the Fifth Assessment Report (AR5) by IPCC (Intergovernmental Panel on Climate Change) mentions likely increasing in intensity and rain rate of TCs. In addition, future change of socioeconomic condition (e.g. population growth) might worsen TC impacts in the future. Thereby, in this study, we developed regression models to estimate economic damages by TCs (hereafter TC damage model), and employed those models to project TC economic damages under several future climate and socioeconomic scenarios. We developed the TC damage models for each of 4 regions; western North Pacific, North American, North Indian, and Southern Hemisphere. The inputs for TC damage model are tropical cyclone central pressure, populations in the area exposed by tropical cyclone wind, and GDP (Gross Domestic Product) per capita. The TC damage models we firstly developed tended to overestimate very low damages and also underestimate very high damages. Thereby we modified structure of TC damage models to improve model performance, and then executed extensive validation of the model. The modified model presented better performance in estimating very low and high TC damages. After the modification and validation of the model, we determined the structure of TC damage models and projected TC economic damages. The result indicated increase in TC economic damage in global scale, while TC economic damage against world GDP would decrease in the future, which result is consistent with previous study.

  2. Sulfur and selenium antioxidants: challenging radical scavenging mechanisms and developing structure-activity relationships based on metal binding.

    PubMed

    Zimmerman, Matthew T; Bayse, Craig A; Ramoutar, Ria R; Brumaghim, Julia L

    2015-04-01

    Because sulfur and selenium antioxidants can prevent oxidative damage, numerous animal and clinical trials have investigated the ability of these compounds to prevent the oxidative stress that is an underlying cause of cardiovascular disease, Alzheimer's disease, and cancer, among others. One of the most common sources of oxidative damage is metal-generated hydroxyl radical; however, very little research has focused on determining the metal-binding abilities and structural attributes that affect oxidative damage prevention by sulfur and selenium compounds. In this review, we describe our ongoing investigations into sulfur and selenium antioxidant prevention of iron- and copper-mediated oxidative DNA damage. We determined that many sulfur and selenium compounds inhibit Cu(I)-mediated DNA damage and that DNA damage prevention varies dramatically when Fe(II) is used in place of Cu(I) to generate hydroxyl radical. Oxidation potentials of the sulfur or selenium compounds do not correlate with their ability to prevent DNA damage, highlighting the importance of metal coordination rather than reactive oxygen species scavenging as an antioxidant mechanism. Additional gel electrophoresis, mass spectrometry, and UV-visible studies confirmed sulfur and selenium antioxidant binding to Cu(I) and Fe(II). Ultimately, our studies established that both the hydroxyl-radical-generating metal ion and the chemical environment of the sulfur or selenium significantly affect DNA damage prevention and that metal coordination is an essential mechanism for these antioxidants.

  3. [Damage from passive tobacco smoking].

    PubMed

    Bartkowiak, Z

    1995-01-01

    The author presents data on the biological casualties and consequences of tobacco-smoking. Smoking is the most dangerous addiction in the scale of the world and in Poland. It causes numerous premature decrease and tobacco-dependent sickness. The author characterises the spread of this addiction in Poland concentrating on the problem of the passive smoking harmfulness. Non-smokers, children and youth, embryo and foetus during the pregnancy are exposed to the passive smoking. The experimental examinations of animals and the analysis of the lateral stream of the tobacco smoke confirm not the least, but rather the greater damage of the passive smoking than the active one. The mechanisms of acting of the tobacco smoke on the passive smokers' body and the health consequences are discussed. The manners, means and activities that are useful for the health protection of non-smokers against the tobacco smoke and the ways of the smoking prevention are described.

  4. Fatigue Damage in Composite Materials

    NASA Astrophysics Data System (ADS)

    Revuelta, D.; Miravete, A.

    2002-02-01

    The phenomenon of fatigue is critical for designing structures including elements made of composite materials. The accurate prediction of the life and fatigue resistance of laminated composites is one of the subjects of inquiry in materials science. The ability of predicting the life of laminates is important for designing, operation, and safety analysis of a composite structure under specific conditions. To predict reliably the life of structures, it is necessary to know the mechanisms of cyclic deformation and damage. It is also necessary to develop a qualitative theory of fatigue failure that should be based on the concepts of solids mechanics. Developing such a theory requires to evaluate the microscopic parameters and the macroscopic variables of the material at the level of a laminate and the structure and to determine exactly the load modes acting on the system.

  5. Nanofoams Response to Radiation Damage

    SciTech Connect

    Fu, Engang; Serrano De Caro, Magdalena; Wang, Yongqiang; Nastasi, Michael; Zepeda-Ruiz, Luis; Bringa, Eduardo M.; Baldwin, Jon K.; Caro, Jose A.

    2012-07-30

    Conclusions of this presentation are: (1) np-Au foams were successfully synthesized by de-alloying process; (2) np-Au foams remain porous structure after Ne ion irradiation to 1 dpa; (3) SFTs were observed in irradiated np-Au foams with highest and intermediate flux, while no SFTs were observed with lowest flux; (4) SFTs were observed in irradiated np-Au foams at RT, whereas no SFTs were observed at LNT irradiation; (5) The diffusivity of vacancies in Au at RT is high enough so that the vacancies have enough time to agglomerate and thus collapse. As a result, SFTs were formed; (6) The high flux created much more damage/time, vacancies don't have enough time to diffuse or recombine. As a result, SFTs were formed.

  6. Wireless Damage Location Sensing System

    NASA Technical Reports Server (NTRS)

    Woodard, Stanley E. (Inventor); Taylor, Bryant Douglas (Inventor)

    2012-01-01

    A wireless damage location sensing system uses a geometric-patterned wireless sensor that resonates in the presence of a time-varying magnetic field to generate a harmonic response that will experience a change when the sensor experiences a change in its geometric pattern. The sensing system also includes a magnetic field response recorder for wirelessly transmitting the time-varying magnetic field and for wirelessly detecting the harmonic response. The sensing system compares the actual harmonic response to a plurality of predetermined harmonic responses. Each predetermined harmonic response is associated with a severing of the sensor at a corresponding known location thereof so that a match between the actual harmonic response and one of the predetermined harmonic responses defines the known location of the severing that is associated therewith.

  7. Gear Damage Detection Using Oil Debris Analysis

    NASA Technical Reports Server (NTRS)

    Dempsey, Paula J.

    2001-01-01

    The purpose of this paper was to verify, when using an oil debris sensor, that accumulated mass predicts gear pitting damage and to identify a method to set threshold limits for damaged gears. Oil debris data was collected from 8 experiments with no damage and 8 with pitting damage in the NASA Glenn Spur Gear Fatigue Rig. Oil debris feature analysis was performed on this data. Video images of damage progression were also collected from 6 of the experiments with pitting damage. During each test, data from an oil debris sensor was monitored and recorded for the occurrence of pitting damage. The data measured from the oil debris sensor during experiments with damage and with no damage was used to identify membership functions to build a simple fuzzy logic model. Using fuzzy logic techniques and the oil debris data, threshold limits were defined that discriminate between stages of pitting wear. Results indicate accumulated mass combined with fuzzy logic analysis techniques is a good predictor of pitting damage on spur gears.

  8. Rapid Approaches Towards DNA Damage Analysis

    PubMed Central

    Crews, N.; Paidipalli, Manasa

    2013-01-01

    The severe effects of DNA damage on human health have led to a tremendous amount of research being focused. Owing to the importance of damage detection, different approaches for the detection and quantification of the damaged DNA will be presented. In this work, we have modeled DNA damage using well-known mutagens: UV radiation to create photoproducts and restriction enzyme digestion to create double strand breaks. We will show that quantitative PCR (qPCR), a widely known measure of detecting the presence of the target DNA can be used to quantify photoproducts/intramolecular DNA damage. Our results indicate that a comparison of the initial concentration available in the undamaged and the damaged samples can be used to reveal the effect of damaged DNA in its amplification. By analyzing multiple regions using this technique, their relative susceptibility to damage can be measured. We also show that high-resolution melting analysis (HRMA), a measure of the bond energy between two DNA strands, can be used to quantify double strand breaks. The strand breaks resulted in a change in the overall distribution of the bond energy thus causing variations in the melting profile. HRMA has also been examined to compare DNA damage resulting from UV-A, UV-B and UV-C irradiation. The evaluation techniques demonstrated can potentially be extended to various types of DNA damage.

  9. Biophysical characterizations of human mitochondrial transcription factor A and its binding to tumor suppressor p53

    PubMed Central

    Wong, Tuck Seng; Rajagopalan, Sridharan; Freund, Stefan M.; Rutherford, Trevor J.; Andreeva, Antonina; Townsley, Fiona M.; Petrovich, Miriana; Fersht, Alan R.

    2009-01-01

    Human mitochondrial transcription factor A (TFAM) is a multi-functional protein, involved in different aspects of maintaining mitochondrial genome integrity. In this report, we characterized TFAM and its interaction with tumor suppressor p53 using various biophysical methods. DNA-free TFAM is a thermally unstable protein that is in equilibrium between monomers and dimers. Self-association of TFAM is modulated by its basic C-terminal tail. The DNA-binding ability of TFAM is mainly contributed by its first HMG-box, while the second HMG-box has low-DNA-binding capability. We also obtained backbone resonance assignments from the NMR spectra of both HMG-boxes of TFAM. TFAM binds primarily to the N-terminal transactivation domain of p53, with a Kd of 1.95 ± 0.19 μM. The C-terminal regulatory domain of p53 provides a secondary binding site for TFAM. The TFAM–p53-binding interface involves both TAD1 and TAD2 sub-domains of p53. Helices α1 and α2 of the HMG-box constitute the main p53-binding region. Since both TFAM and p53 binds preferentially to distorted DNA, the TFAM–p53 interaction is implicated in DNA damage and repair. In addition, the DNA-binding mechanism of TFAM and biological relevance of the TFAM–p53 interaction are discussed. PMID:19755502

  10. Water binding in legume seeds

    NASA Technical Reports Server (NTRS)

    Vertucci, C. W.; Leopold, A. C.

    1987-01-01

    The physical status of water in seeds has a pivotal role in determining the physiological reactions that can take place in the dry state. Using water sorption isotherms from cotyledon and axis tissue of five leguminous seeds, the strength of water binding and the numbers of binding sites have been estimated using van't Hoff analyses and the D'Arcy/Watt equation. These parameters of water sorption are calculated for each of the three regions of water binding and for a range of temperatures. Water sorption characteristics are reflective of the chemical composition of the biological materials as well as the temperature at which hydration takes place. Changes in the sorption characteristics with temperature and hydration level may suggest hydration-induced structural changes in cellular components.

  11. Galectin-3-Binding and Metastasis

    PubMed Central

    Nangia-Makker, Pratima; Balan, Vitaly; Raz, Avraham

    2013-01-01

    i. Summary Galectin-3 is a member of a family of carbohydrate-binding proteins. It is present in the nucleus, the cytoplasm and also extracellular matrix of many normal and neoplastic cell types. Arrays of reports show an upregulation of this protein in transformed and metastatic cell lines (1, 2). Moreover, in many human carcinomas, an increased expression of galectin-3 correlates with progressive tumor stages (3–6). Several lines of analysis have demonstrated that the galectins participate in cell-cell and cell-matrix interactions by recognizing and binding complimentary glycoconjugates and thereby play a crucial role in normal and pathological processes. Elevated expression of the protein is associated with an increased capacity for anchorage-independent growth, homotypic aggregation, and tumor cell lung colonization (7–9). In this chapter we describe the methods of purification of galectin-3 from transformed E. coli and some of the commonly used functional assays for analyzing galectin-3 binding. PMID:22674139

  12. Computational Prediction of RNA-Binding Proteins and Binding Sites

    PubMed Central

    Si, Jingna; Cui, Jing; Cheng, Jin; Wu, Rongling

    2015-01-01

    Proteins and RNA interaction have vital roles in many cellular processes such as protein synthesis, sequence encoding, RNA transfer, and gene regulation at the transcriptional and post-transcriptional levels. Approximately 6%–8% of all proteins are RNA-binding proteins (RBPs). Distinguishing these RBPs or their binding residues is a major aim of structural biology. Previously, a number of experimental methods were developed for the determination of protein–RNA interactions. However, these experimental methods are expensive, time-consuming, and labor-intensive. Alternatively, researchers have developed many computational approaches to predict RBPs and protein–RNA binding sites, by combining various machine learning methods and abundant sequence and/or structural features. There are three kinds of computational approaches, which are prediction from protein sequence, prediction from protein structure, and protein-RNA docking. In this paper, we review all existing studies of predictions of RNA-binding sites and RBPs and complexes, including data sets used in different approaches, sequence and structural features used in several predictors, prediction method classifications, performance comparisons, evaluation methods, and future directions. PMID:26540053

  13. Optimal Battery Charging for Damage Mitigation

    NASA Technical Reports Server (NTRS)

    Hartley, Tom T.; Lorenzo, Carl F.

    2003-01-01

    Our control philosophy is to charge the NiH2 cell in such a way that the damage incurred during the charging period is minimized, thus extending its cycle life. This requires nonlinear dynamic model of NiH2 cell and a damage rate model. We must do this first. This control philosophy is generally considered damage mitigating control or life-extending control. This presentation covers how NiH2 cells function, electrode behavior, an essentialized model, damage mechanisms for NiH2 batteries, battery continuum damage modeling, and battery life models. The presentation includes graphs and a chart illustrating how charging a NiH2 battery with different voltages and currents affects damages the battery and affects its life. The presentation concludes with diagrams of control system architectures for tracking battery recharging.

  14. DNA Damage Signals and Space Radiation Risk

    NASA Technical Reports Server (NTRS)

    Cucinotta, Francis A.

    2011-01-01

    Space radiation is comprised of high-energy and charge (HZE) nuclei and protons. The initial DNA damage from HZE nuclei is qualitatively different from X-rays or gamma rays due to the clustering of damage sites which increases their complexity. Clustering of DNA damage occurs on several scales. First there is clustering of single strand breaks (SSB), double strand breaks (DSB), and base damage within a few to several hundred base pairs (bp). A second form of damage clustering occurs on the scale of a few kbp where several DSB?s may be induced by single HZE nuclei. These forms of damage clusters do not occur at low to moderate doses of X-rays or gamma rays thus presenting new challenges to DNA repair systems. We review current knowledge of differences that occur in DNA repair pathways for different types of radiation and possible relationships to mutations, chromosomal aberrations and cancer risks.

  15. Structural Damage Detection Using Virtual Passive Controllers

    NASA Technical Reports Server (NTRS)

    Lew, Jiann-Shiun; Juang, Jer-Nan

    2001-01-01

    This paper presents novel approaches for structural damage detection which uses the virtual passive controllers attached to structures, where passive controllers are energy dissipative devices and thus guarantee the closed-loop stability. The use of the identified parameters of various closed-loop systems can solve the problem that reliable identified parameters, such as natural frequencies of the open-loop system may not provide enough information for damage detection. Only a small number of sensors are required for the proposed approaches. The identified natural frequencies, which are generally much less sensitive to noise and more reliable than the identified natural frequencies, are used for damage detection. Two damage detection techniques are presented. One technique is based on the structures with direct output feedback controllers while the other technique uses the second-order dynamic feedback controllers. A least-squares technique, which is based on the sensitivity of natural frequencies to damage variables, is used for accurately identifying the damage variables.

  16. Synthetic heparin-binding growth factor analogs

    DOEpatents

    Pena, Louis A.; Zamora, Paul; Lin, Xinhua; Glass, John D.

    2007-01-23

    The invention provides synthetic heparin-binding growth factor analogs having at least one peptide chain that binds a heparin-binding growth factor receptor, covalently bound to a hydrophobic linker, which is in turn covalently bound to a non-signaling peptide that includes a heparin-binding domain. The synthetic heparin-binding growth factor analogs are useful as soluble biologics or as surface coatings for medical devices.

  17. Probing the Dynamic Interaction between Damaged DNA and a Cellular Responsive Protein Using a Piezoelectric Mass Biosensor.

    PubMed

    Jin, Yulong; Xie, Yunfeng; Wu, Kui; Huang, Yanyan; Wang, Fuyi; Zhao, Rui

    2017-03-15

    The binding events between damaged DNA and recognition biomolecules are of great interest for understanding the activity of DNA-damaging drugs and the related DNA repair networks. Herein, a simple and sensitive sensor system was tailored for real-time probing of the dynamic molecular recognition between cisplatin-damaged-DNA (cisPt-DNA) and a cellular responsive protein, high-mobility-group box 1 (HMGB1). By integration of flow injection analysis (FIA) with quartz crystal microbalance (QCM), the interaction time-course of cisPt-DNA and HMGB1 domain A (HMGB1a) was investigated. The highly specific sensing interface was carefully designed and fabricated using cisPt-DNA as recognition element. A hybrid self-assembled monolayer consisting of cysteamine and mercaptohexanol was introduced to resist nonspecific adsorption. The calculated kinetic parameters (kass and kdiss) and the dissociation constant (KD) demonstrated the rapid recognition and tight binding of HMGB1a toward cisPt-DNA. Molecular docking was employed to simulate the complex formed by cisPt-DNA and HMGB1a. The tight binding of such a DNA-damage responsive complex is appealing for the downstream molecular recognition event related to the resistance to DNA repair. This continuous-flow QCM biosensor is an ideal tool for studying specific interactions between drug-damaged-DNAs and their recognition proteins in a physiological-relevant environment, and will provide a potential sensor platform for rapid screening and evaluating metal anticancer drugs.

  18. Assessment and control of structural damage

    NASA Technical Reports Server (NTRS)

    Jeong, G. D.; Stubbs, N.; Yao, J. T. P.

    1988-01-01

    The objective of this paper is to summarize and review several investigations on the assessment and control of structural damage in civil engineering. Specifically, the definition of structural damage is discussed. A candidate method for the evaluation of damage is then reviewed and demonstrated. Various ways of implementing passive and active control of civil engineering structures are next summarized. Finally, the possibility of applying expert systems is discussed.

  19. A Combat Battle Damage Assessor Expert System,

    DTIC Science & Technology

    1984-05-01

    Next, the requirements should receive just enough repair to allow for the battle damage assessor software it to be flown to a rear area for complete are...R’ D-Ai48 898 A COMBAT BATTLE DAMAGE ASSESSOR EXPERT SYSTEM(U) AIR 1/i FORCE WRIGHT AERONAUTICAL LABS WRIGHT-PATTERSON AFB ON D E NELSON MAY 84 SBI...8217.c.sw-x -: "--..... . .. .-. :;.-...... .............. .. . .. . . .. . .. . . r74 IV A COMBAT BATTLE DAMAGE ASSESSOR EXPERT SYSTEM DAL E. NELSON

  20. An Experimental Investigation of Damage Resistances and Damage Tolerance of Composite Materials

    NASA Technical Reports Server (NTRS)

    Prabhakaran, R.

    2003-01-01

    The project included three lines of investigation, aimed at a better understanding of the damage resistance and damage tolerance of pultruded composites. The three lines of investigation were: (i) measurement of permanent dent depth after transverse indentation at different load levels, and correlation with other damage parameters such as damage area (from x-radiography) and back surface crack length, (ii) estimation of point stress and average stress characteristic dimensions corresponding to measured damage parameters, and (iii) an attempt to measure the damage area by a reflection photoelastic technique. All the three lines of investigation were pursued.

  1. Evidence against the nuclear in situ binding of arsenicals-oxidative stress theory of arsenic carcinogenesis

    SciTech Connect

    Kitchin, Kirk T. Wallace, Kathleen

    2008-10-15

    A large amount of evidence suggests that arsenicals act via oxidative stress in causing cancer in humans and experimental animals. It is possible that arsenicals could bind in situ close to nuclear DNA followed by Haber-Weiss type oxidative DNA damage. Therefore, we tested this hypothesis by using radioactive {sup 73}As labeled arsenite and vacuum filtration methodology to determine the binding affinity and capacity of {sup 73}As arsenite to calf thymus DNA and Type 2A unfractionated histones, histone H3, H4 and horse spleen ferritin. Arsenicals are known to release redox active Fe from ferritin. At concentrations up to about 1 mM, neither DNA nor any of the three proteins studied, Type II-A histones, histone H3, H4 or ferritin, bound radioactive arsenite in a specific manner. Therefore, it appears highly unlikely that initial in situ binding of trivalent arsenicals, followed by in situ oxidative DNA damage, can account for arsenic's carcinogenicity. This experimental evidence (lack of arsenite binding to DNA, histone Type II-A and histone H3, H4) does not rule out other possible oxidative stress modes of action for arsenic such as (a) diffusion of longer lived oxidative stress molecules, such as H{sub 2}O{sub 2} into the nucleus and ensuing oxidative damage, (b) redox chemistry by unbound arsenicals in the nucleus, or (c) arsenical-induced perturbations in Fe, Cu or other metals which are already known to oxidize DNA in vitro and in vivo.

  2. TNFα-Damaged-HUVECs Microparticles Modify Endothelial Progenitor Cell Functional Activity

    PubMed Central

    Luna, Carlos; Carmona, Andrés; Alique, Matilde; Carracedo, Julia; Ramirez, Rafael

    2015-01-01

    Endothelial progenitor cells (EPCs) have an important role in the maintenance of vascular integrity and homeostasis. While there are many studies that explain EPCs mechanisms action, there are few studies that demonstrate how they interact with other emerging physiological elements such as Endothelial Microparticles (EMPs). EMPs are membranous structures with a size between 100 and 1000 nm that act as molecular information transporter in biological systems and are known as an important elements in develop different pathologies; moreover a lot of works explains that are novel biomarkers. To elucidate these interactions, we proposed an in vitro model of endothelial damage mediated by TNFalpha, in which damaged EMPs and EPCs are in contact to assess EPCs functional effects. We have observed that damaged EMPs can modulate several EPCs classic factors as colony forming units (CFUs), contribution to repair a physically damaged endothelium (wound healing), binding to mature endothelium, and co-adjuvants to the formation of new vessels in vitro (angiogenesis). All of these in a dose-dependent manner. Damaged EMPs at a concentration of 103 MPs/ml have an activating effect of these capabilities, while at concentrations of 105 MPs/ml these effects are attenuated or reduced. This in vitro model helps explain that in diseases where there is an imbalance between these two elements (EPCs and damaged EMPs), the key cellular elements in the regeneration and maintenance of vascular homeostasis (EPCs) are not fully functional, and could explain, at least in part, endothelial dysfunction associated in various pathologies. PMID:26733886

  3. Independent mechanisms recruit the cohesin loader protein NIPBL to sites of DNA damage

    PubMed Central

    Bot, Christopher; Pfeiffer, Annika; Giordano, Fosco; Dantuma, Nico P.; Ström, Lena

    2017-01-01

    ABSTRACT NIPBL is required to load the cohesin complex on to DNA. While the canonical role of cohesin is to couple replicated sister chromatids together until the onset of mitosis, it also promotes tolerance to DNA damage. Here, we show that NIPBL is recruited to DNA damage throughout the cell cycle via independent mechanisms, influenced by type of damage. First, the heterochromatin protein HP1γ (also known as CBX3) recruits NIPBL to DNA double-strand breaks (DSBs) through the corresponding HP1-binding motif within the N-terminus. By contrast, the C-terminal HEAT repeat domain is unable to recruit NIPBL to DSBs but independently targets NIPBL to laser microirradiation-induced DNA damage. Each mechanism is dependent on the RNF8 and RNF168 ubiquitylation pathway, while the recruitment of the HEAT repeat domain requires further ATM or ATR activity. Thus, NIPBL has evolved a sophisticated response to damaged DNA that is influenced by the form of damage, suggesting a highly dynamic role for NIPBL in maintaining genomic stability. PMID:28167679

  4. Independent mechanisms recruit the cohesin loader protein NIPBL to sites of DNA damage.

    PubMed

    Bot, Christopher; Pfeiffer, Annika; Giordano, Fosco; Manjeera, Dharani E; Dantuma, Nico P; Ström, Lena

    2017-03-15

    NIPBL is required to load the cohesin complex on to DNA. While the canonical role of cohesin is to couple replicated sister chromatids together until the onset of mitosis, it also promotes tolerance to DNA damage. Here, we show that NIPBL is recruited to DNA damage throughout the cell cycle via independent mechanisms, influenced by type of damage. First, the heterochromatin protein HP1γ (also known as CBX3) recruits NIPBL to DNA double-strand breaks (DSBs) through the corresponding HP1-binding motif within the N-terminus. By contrast, the C-terminal HEAT repeat domain is unable to recruit NIPBL to DSBs but independently targets NIPBL to laser microirradiation-induced DNA damage. Each mechanism is dependent on the RNF8 and RNF168 ubiquitylation pathway, while the recruitment of the HEAT repeat domain requires further ATM or ATR activity. Thus, NIPBL has evolved a sophisticated response to damaged DNA that is influenced by the form of damage, suggesting a highly dynamic role for NIPBL in maintaining genomic stability.

  5. SUMO-mediated regulation of DNA damage repair and responses

    PubMed Central

    Sarangi, Prabha; Zhao, Xiaolan

    2015-01-01

    Sumoylation plays important roles during DNA damage repair and responses. Recent broad-scope and substrate-based studies have shed light on the regulation and significance of sumoylation during these processes. An emerging paradigm is that sumoylation of many DNA metabolism proteins is controlled by DNA engagement. Such “on-site modification” can explain low substrate modification levels and has important implications in sumoylation mechanisms and effects. New studies also suggest that sumoylation can regulate a process through an ensemble effect or via major substrates. Additionally, we describe new trends in the functional effects of sumoylation, such as bi-directional changes in biomolecule binding and multi-level coordination with other modifications. These emerging themes and models will stimulate our thinking and research in sumoylation and genome maintenance. PMID:25778614

  6. Structural damage assessment as an identification problem

    NASA Technical Reports Server (NTRS)

    Hajela, Prabhat; Soeiro, F. J.

    1989-01-01

    Damage assessment of structural assemblies is treated as an identification problem. A brief review of identification methods is first presented with particular focus on the output error approach. The use of numerical optimization methods in identifying the location and extent of damage in structures is studied. The influence of damage on eigenmode shapes and static displacements is explored as a means of formulating a measure of damage in the structure. Preliminary results obtained in this study are presented and special attention is directed at the shortcomings associated with the nonlinear programming approach to solving the optimization problem.

  7. DNA damage response, redox status and hematopoiesis.

    PubMed

    Weiss, Cary N; Ito, Keisuke

    2014-01-01

    The ability of hematopoietic stem cells (HSCs) to self-renew and differentiate into progenitors is essential for homeostasis of the hematopoietic system. The longevity of HSCs makes them vulnerable to accumulating DNA damage, which may be leukemogenic or result in senescence and cell death. Additionally, the ability of HSCs to self-renew and differentiate allows DNA damage to spread throughout the hematologic system, leaving the organism vulnerable to disease. In this review we discuss cell fate decisions made in the face of DNA damage and other cellular stresses, and the role of reactive oxygen species in the long-term maintenance of HSCs and their DNA damage response.

  8. Method for assaying clustered DNA damages

    DOEpatents

    Sutherland, Betsy M.

    2004-09-07

    Disclosed is a method for detecting and quantifying clustered damages in DNA. In this method, a first aliquot of the DNA to be tested for clustered damages with one or more lesion-specific cleaving reagents under conditions appropriate for cleavage of the DNA to produce single-strand nicks in the DNA at sites of damage lesions. The number average molecular length (Ln) of double stranded DNA is then quantitatively determined for the treated DNA. The number average molecular length (Ln) of double stranded DNA is also quantitatively determined for a second, untreated aliquot of the DNA. The frequency of clustered damages (.PHI..sub.c) in the DNA is then calculated.

  9. CHARACTERIZATION OF THERMALLY DAMAGED LX-17

    SciTech Connect

    Hsu, P C

    2007-07-11

    Thermal damage was applied to LX-17 at 190 C for several hours. The damaged LX-17 samples, after cooled down to room temperature, were characterized for their material properties (density, porosity, permeability, moduli), safety, and performance. Weight losses upon thermal exposure were insignificant (< 0.1% wt.). The damaged LX-17 samples expanded, resulting in a bulk density reduction of 4.3%. Subsequent detonation measurements (cylinder tests) were conducted on the thermally-damaged LX-17 samples. The results showed that the fractions of damaged LX-17 reacted were slightly lower than those of pristine LX-17. The thermally damaged LX-17 had a detonation velocity of 7.315 mm/{micro}s, lower than that (7.638 mm/{micro}s) of pristine LX-17. Detonation energy density for the damaged LX-17 was 5.08 kJ/cm{sup 3}, about 9.0% lower than the detonation energy density of 5.50 kJ/cm{sup 3} for the pristine LX-17. The break-out curves showed reaction zone lengths for pristine LX-17 and damaged LX-17 were similar but the damaged samples had ragged detonation fronts.

  10. Quantifying pulsed laser induced damage to graphene

    SciTech Connect

    Currie, Marc; Caldwell, Joshua D.; Bezares, Francisco J.; Robinson, Jeremy; Anderson, Travis; Chun, Hayden; Tadjer, Marko

    2011-11-21

    As an emerging optical material, graphene's ultrafast dynamics are often probed using pulsed lasers yet the region in which optical damage takes place is largely uncharted. Here, femtosecond laser pulses induced localized damage in single-layer graphene on sapphire. Raman spatial mapping, SEM, and AFM microscopy quantified the damage. The resulting size of the damaged area has a linear correlation with the optical fluence. These results demonstrate local modification of sp{sup 2}-carbon bonding structures with optical pulse fluences as low as 14 mJ/cm{sup 2}, an order-of-magnitude lower than measured and theoretical ablation thresholds.

  11. Allosteric Dynamic Control of Binding

    PubMed Central

    Sumbul, Fidan; Acuner-Ozbabacan, Saliha Ece; Haliloglu, Turkan

    2015-01-01

    Proteins have a highly dynamic nature and there is a complex interrelation between their structural dynamics and binding behavior. By assuming various conformational ensembles, they perform both local and global fluctuations to interact with other proteins in a dynamic infrastructure adapted to functional motion. Here, we show that there is a significant association between allosteric mutations, which lead to high-binding-affinity changes, and the hinge positions of global modes, as revealed by a large-scale statistical analysis of data in the Structural Kinetic and Energetic Database of Mutant Protein Interactions (SKEMPI). We further examined the mechanism of allosteric dynamics by conducting studies on human growth hormone (hGH) and pyrin domain (PYD), and the results show how mutations at the hinge regions could allosterically affect the binding-site dynamics or induce alternative binding modes by modifying the ensemble of accessible conformations. The long-range dissemination of perturbations in local chemistry or physical interactions through an impact on global dynamics can restore the allosteric dynamics. Our findings suggest a mechanism for the coupling of structural dynamics to the modulation of protein interactions, which remains a critical phenomenon in understanding the effect of mutations that lead to functional changes in proteins. PMID:26338442

  12. Cellulose binding domain fusion proteins

    SciTech Connect

    Shoseyov, Oded; Shpiegl, Itai; Goldstein, Marc A.; Doi, Roy H.

    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.

  13. Cellulose binding domain fusion proteins

    DOEpatents

    Shoseyov, O.; Yosef, K.; Shpiegl, I.; Goldstein, M.A.; Doi, R.H.

    1998-02-17

    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.

  14. 14-3-3 proteins, FHA domains and BRCT domains in the DNA damage response.

    PubMed

    Mohammad, Duaa H; Yaffe, Michael B

    2009-09-02

    The DNA damage response depends on the concerted activity of protein serine/threonine kinases and modular phosphoserine/threonine-binding domains to relay the damage signal and recruit repair proteins. The PIKK family of protein kinases, which includes ATM/ATR/DNA-PK, preferentially phosphorylate Ser-Gln sites, while their basophilic downstream effecter kinases, Chk1/Chk2/MK2 preferentially phosphorylate hydrophobic-X-Arg-X-X-Ser/Thr-hydrophobic sites. A subset of tandem BRCT domains act as phosphopeptide binding modules that bind to ATM/ATR/DNA-PK substrates after DNA damage. Conversely, 14-3-3 proteins interact with substrates of Chk1/Chk2/MK2. FHA domains have been shown to interact with substrates of ATM/ATR/DNA-PK and CK2. In this review we consider how substrate phosphorylation together with BRCT domains, FHA domains and 14-3-3 proteins function to regulate ionizing radiation-induced nuclear foci and help to establish the G(2)/M checkpoint. We discuss the role of MDC1 a molecular scaffold that recruits early proteins to foci, such as NBS1 and RNF8, through distinct phosphodependent interactions. In addition, we consider the role of 14-3-3 proteins and the Chk2 FHA domain in initiating and maintaining cell cycle arrest.

  15. Carbon dots based FRET for the detection of DNA damage.

    PubMed

    Kudr, Jiri; Richtera, Lukas; Xhaxhiu, Kledi; Hynek, David; Heger, Zbynek; Zitka, Ondrej; Adam, Vojtech

    2017-02-09

    Here, we aimed our attention at the synthesis of carbon dots (C-dots) with the ability to interact with DNA to suggest an approach for the detection of DNA damage. Primarily, C-dots modified with amine moieties were synthesized using the one-step microwave pyrolysis of citric acid in the presence of diethylenetriamine. The C-dots showed strong photoluminescence with a quantum yield of 4%. In addition, the C-dots (2.8±0.8nm) possessed a good colloidal stability and exhibited a positive surface charge (ζ=36mV) at a neutral pH. An interaction study of the C-dots and the DNA fragment of λ bacteriophage was performed, and the DNA binding resulted in changes to the photoluminescent and absorption properties of the C-dots. A binding of the C-dots to DNA was also observed as a change to DNA electrophoretic mobility and a decreased ability to intercalate ethidium bromide (EtBr). Moreover, the Förster (or fluorescence) resonance energy transfer (FRET) between the C-dots and EtBr was studied, in which the C-dots serve as an excitation energy donor and the EtBr serves as an acceptor. When DNA was damaged using ultraviolet (UV) radiation (λ=254nm) and hydroxyl radicals, the intensity of the emitted photoluminescence at 612nm significantly decreased. The concept was proved on analysis of the genomic DNA from PC-3 cells and DNA isolated from melanoma tissues.

  16. Protein binding assay for hyaluronate

    SciTech Connect

    Lacy, B.E.; Underhill, C.B.

    1986-11-01

    A relatively quick and simple assay for hyaluronate was developed using the specific binding protein, hyaluronectin. The hyaluronectin was obtained by homogenizing the brains of Sprague-Dawley rats, and then centrifuging the homogenate. The resulting supernatant was used as a source of crude hyaluronectin. In the binding assay, the hyaluronectin was mixed with (/sup 3/H)hyaluronate, followed by an equal volume of saturated (NH/sub 4/)/sub 2/SO/sub 4/, which precipitated the hyaluronectin and any (/sup 3/H)hyaluronate associated with it, but left free (/sup 3/H)hyaluronate in solution. The mixture was then centrifuged, and the amount of bound (/sup 3/H)hyaluronate in the precipitate was determined. Using this assay, the authors found that hyaluronectin specifically bound hyaluronate, since other glycosaminoglycans failed to compete for the binding protein. In addition, the interaction between hyaluronectin and hyaluronate was of relatively high affinity, and the size of the hyaluronate did not appear to substantially alter the amount of binding. To determine the amount of hyaluronate in an unknown sample, they used a competition assay in which the binding of a set amount of (/sup 3/H)hyaluronate was blocked by the addition of unlabeled hyaluronate. By comparing the degree of competition of the unknown samples with that of known amounts of hyaluronate, it was possible to determine the amount of hyaluronate in the unknowns. They have found that this method is sensitive to 1 ..mu..g or less of hyaluronate, and is unaffected by the presence of proteins.

  17. Spectrometry researches on interaction and sonodynamic damage of riboflavin (RF) to bovine serum albumin (BSA).

    PubMed

    Wang, Zhiqiu; Li, Jushi; Wang, Jun; Zou, Mingming; Wang, Siyu; Li, Ying; Kong, Yumei; Xia, Lixin

    2012-02-15

    In this paper, the riboflavin (RF) was used to study the interaction and sonodynamic damage to bovine serum albumin (BSA) by fluorescence and UV-vis spectroscopy. The results showed that the RF could efficiently bind to BSA in aqueous solution. Under ultrasonic irradiation, the RF could obviously damage the BSA. In addition, synchronous fluorescence spectroscopy revealed that the RF showed more accessible to tryptophan (Trp) residues than to tyrosine (Tyr) residues. Also, it damaged Trp residues more seriously than Tyr residues under ultrasonic irradiation. At last, the generation of reactive oxygen species (ROS) in sonodynamic process was estimated by the method of Oxidation-Extraction Spectrometry (OES). And then, several radical scavengers were used to determine the kind of ROS. It was found that at least the singlet oxygen ((1)O(2)) and hydroxyl radicals (*OH) were generated.

  18. Earthquake damage to underground facilities

    SciTech Connect

    Pratt, H.R.; Stephenson, D.E.; Zandt, G.; Bouchon, M.; Hustrulid, W.A.

    1980-01-01

    In order to assess the seismic risk for an underground facility, a data base was established and analyzed to evaluate the potential for seismic disturbance. Substantial damage to underground facilities is usually the result of displacements primarily along pre-existing faults and fractures, or at the surface entrance to these facilities. Evidence of this comes from both earthquakes and large explosions. Therefore, the displacement due to earthquakes as a function of depth is important in the evaluation of the hazard to underground facilities. To evaluate potential displacements due to seismic effects of block motions along pre-existing or induced fractures, the displacement fields surrounding two types of faults were investigated. Analytical models were used to determine relative displacements of shafts and near-surface displacement of large rock masses. Numerical methods were used to determine the displacement fields associated with pure strike-slip and vertical normal faults. Results are presented as displacements for various fault lengths as a function of depth and distance. This provides input to determine potential displacements in terms of depth and distance for underground facilities, important for assessing potential sites and design parameters.

  19. Lung oxidative damage by hypoxia.

    PubMed

    Araneda, O F; Tuesta, M

    2012-01-01

    One of the most important functions of lungs is to maintain an adequate oxygenation in the organism. This organ can be affected by hypoxia facing both physiological and pathological situations. Exposure to this condition favors the increase of reactive oxygen species from mitochondria, as from NADPH oxidase, xanthine oxidase/reductase, and nitric oxide synthase enzymes, as well as establishing an inflammatory process. In lungs, hypoxia also modifies the levels of antioxidant substances causing pulmonary oxidative damage. Imbalance of redox state in lungs induced by hypoxia has been suggested as a participant in the changes observed in lung function in the hypoxic context, such as hypoxic vasoconstriction and pulmonary edema, in addition to vascular remodeling and chronic pulmonary hypertension. In this work, experimental evidence that shows the implied mechanisms in pulmonary redox state by hypoxia is reviewed. Herein, studies of cultures of different lung cells and complete isolated lung and tests conducted in vivo in the different forms of hypoxia, conducted in both animal models and humans, are described.

  20. Preliminary damage and intensity survey

    USGS Publications Warehouse

    Brewer, L.R.

    1992-01-01

    A major earthquake struck the Mojave Desert region in southern California at about 5 a.m. local time on June 28th, 1992. Seismologists located the epicenter of the magnitude 7.6 (Ms) earthquake 5-10km southwest of Landers, a small community about 150km east of Los Angeles. the earthquake shook a wide area of southern California, southern Nevada, western Arizona, and extreme northwest Mexico. High-rise buildings swayed in cities as far north as Boise, Idaho, and as far east as Albuquerque, New Mexico, and Denver, Colorado. Standing waves called seiches disturbed the surface of lakes and bays as far away as Washington State and the Gulf Coast of Texas. In United States, the Landers shock was felt over a contiguous land area approximately 103.600km2. Approximately 3 hrs after the Landers earthquake, a magntude 6.7 (Ms) aftershock struck an area near Big Bear Lake, California, less than 50 km from the main-shcok epicenter. Within its epicentral area this afterhoskc caused consideralbe damage.   

  1. DNA damage by various radiations

    NASA Astrophysics Data System (ADS)

    Hasegawa, K.; Yoshioka, H.; Yoshioka, H.

    1997-01-01

    In an attempt to shed light on the influence of tritiated water on DNA we have investigated the post-irradiation damage with a simple plasmid DNA, pBR322 and pUC18. The survival of covalently closed circular (CCC) DNA form was directly followed by agarose gel electrophoresis. The survival percentage of DNA in tritiated water was almost the same as with the irradiation with X-rays at the same absorbed dose. For irradiation with γ-rays, on the other hand, the decay rate was larger than those observed with both tritiated water and X-rays. The percentages of breakage for DNA in tritiated water, X-rays and γ-rays were found to be 34, 38 and 33% at 100 Gy of absorbed dose. The effect of dose rate was not observed for irradiation with tritiated water, X-rays and γ-rays. In order to study protection of DNA against radiation, we investigated the protecting effect of tea catechin which is the main component of (-)-epigallocatechin gallate (EGCg). The protection mechanism for DNA against radiation-induced scission has been studied using ESR spin-trapping method.

  2. Lung Oxidative Damage by Hypoxia

    PubMed Central

    Araneda, O. F.; Tuesta, M.

    2012-01-01

    One of the most important functions of lungs is to maintain an adequate oxygenation in the organism. This organ can be affected by hypoxia facing both physiological and pathological situations. Exposure to this condition favors the increase of reactive oxygen species from mitochondria, as from NADPH oxidase, xanthine oxidase/reductase, and nitric oxide synthase enzymes, as well as establishing an inflammatory process. In lungs, hypoxia also modifies the levels of antioxidant substances causing pulmonary oxidative damage. Imbalance of redox state in lungs induced by hypoxia has been suggested as a participant in the changes observed in lung function in the hypoxic context, such as hypoxic vasoconstriction and pulmonary edema, in addition to vascular remodeling and chronic pulmonary hypertension. In this work, experimental evidence that shows the implied mechanisms in pulmonary redox state by hypoxia is reviewed. Herein, studies of cultures of different lung cells and complete isolated lung and tests conducted in vivo in the different forms of hypoxia, conducted in both animal models and humans, are described. PMID:22966417

  3. A novel role of the chromokinesin Kif4A in DNA damage response

    PubMed Central

    Wu, Guikai; Zhou, Longen; Khidr, Lily; Guo, Xuning Emily; Kim, Wankee; Lee, Young Mi; Krasieva, Tatiana; Chen, Phang-Lang

    2011-01-01

    Chromokinesins are microtubule-motor molecules that possess chromatin binding activity and are important for mitotic and meiotic regulation. The chromokinesin-member Kif4A is unique in that it localizes to nucleus during interphase of the cell cycle. Kif4 deletion by gene targeting in mouse embryonic cells was known to associate with DNA damage response. However, its precise role in DNA damage or repair pathway is not clear. Here we report that Kif4A associates with BRCA2 in a biochemical identification and that the interaction is mediated by the Kif4A C-terminal cargo-binding domain and BRCA2 C-terminal conserved region. Upon nucleus-specific laser micro-irradiation, Kif4A was rapidly recruited to sites of DNA damage. Significantly, the depletion of Kif4A from cells by shRNA impaired the ionizing-radiation induced foci (IRIF) formation of Rad51, both quantitatively and qualitatively. In contrast, the IRIF of γ-H2AX or NBS1 was largely intact. Moreover, Kif4A knockdown rendered cells hypersensitive to ionizing radiation in a colonogenic survival assay. We further demonstrated that Kif4A deficiency led to significantly decreased homologous recombination in an I-SceI endonuclease induced in vivo recombination assay. Together, our results suggest a novel role for a chromokinesin family member in the DNA damage response by modulating the BRCA2/Rad51 pathway. PMID:18604178

  4. A novel role of the chromokinesin Kif4A in DNA damage response.

    PubMed

    Wu, Guikai; Zhou, Longen; Khidr, Lily; Guo, Xuning Emily; Kim, Wankee; Lee, Young Mi; Krasieva, Tatiana; Chen, Phang-Lang

    2008-07-01

    Chromokinesins are microtubule-motor molecules that possess chromatin binding activity and are important for mitotic and meiotic regulation. The chromokinesin-member Kif4A is unique in that it localizes to nucleus during interphase of the cell cycle. Kif4 deletion by gene targeting in mouse embryonic cells was known to associate with DNA damage response. However, its precise role in DNA damage or repair pathway is not clear. Here we report that Kif4A associates with BRCA2 in a biochemical identification and that the interaction is mediated by the Kif4A C-terminal cargo-binding domain and BRCA2 C-terminal conserved region. Upon nucleus-specific laser micro-irradiation, Kif4A was rapidly recruited to sites of DNA damage. Significantly, the depletion of Kif4A from cells by shRNA impaired the ionizing-radiation induced foci (IRIF) formation of Rad51, both quantitatively and qualitatively. In contrast, the IRIF of gamma-H2AX or NBS1 was largely intact. Moreover, Kif4A knockdown rendered cells hypersensitive to ionizing radiation in a colonogenic survival assay. We further demonstrated that Kif4A deficiency led to significantly decreased homologous recombination in an I-SceI endonuclease induced in vivo recombination assay. Together, our results suggest a novel role for a chromokinesin family member in the DNA damage response by modulating the BRCA2/Rad51 pathway.

  5. Controlling the response to DNA damage by the APC/C-Cdh1.

    PubMed

    de Boer, H Rudolf; Guerrero Llobet, S; van Vugt, Marcel A T M

    2016-03-01

    Proper cell cycle progression is safeguarded by the oscillating activities of cyclin/cyclin-dependent kinase complexes. An important player in the regulation of mitotic cyclins is the anaphase-promoting complex/cyclosome (APC/C), a multi-subunit E3 ubiquitin ligase. Prior to entry into mitosis, the APC/C remains inactive, which allows the accumulation of mitotic regulators. APC/C activation requires binding to either the Cdc20 or Cdh1 adaptor protein, which sequentially bind the APC/C and facilitate targeting of multiple mitotic regulators for proteasomal destruction, including Securin and Cyclin B, to ensure proper chromosome segregation and mitotic exit. Emerging data have indicated that the APC/C, particularly in association with Cdh1, also functions prior to mitotic entry. Specifically, the APC/C-Cdh1 is activated in response to DNA damage in G2 phase cells. These observations are in line with in vitro and in vivo genetic studies, in which cells lacking Cdh1 expression display various defects, including impaired DNA repair and aberrant cell cycle checkpoints. In this review, we summarize the current literature on APC/C regulation in response to DNA damage, the functions of APC/C-Cdh1 activation upon DNA damage, and speculate how APC/C-Cdh1 can control cell fate in the context of persistent DNA damage.

  6. Femtosecond damage threshold of multilayer metal films

    NASA Astrophysics Data System (ADS)

    Ibrahim, Wael M. G.; Elsayed-Ali, Hani E.; Shinn, Michelle D.; Bonner, Carl E.

    2003-05-01

    With the availability of terawatt laser systems with subpicosecond pulses, laser damage to optical components has become the limiting factor for further increases in the output peak power. Evaluation of different material structures in accordance to their suitability for high-power laser systems is essential. Multi-shot damage experiments, using 110 fs laser pulses at 800 nm, on polycrystalline single layer gold films and multi-layer (gold-vanadium, and gold-titanium) films were conducted. The laser incident fluence was varied, in both cases, from 0.1 to 0.6 J/cm2. No evidence of surface damage was apparent in the gold sample up to a fluence of 0.3 J/cm2. The multilayer sample experienced the onset of surface damage at the lowest fluence value used of 0.1 J/cm2. Damage results are in contrast with the time resolved ultrafast thermoreflectivity measurements that revealed a reduction of the thermoreflectivity signal for the multilayer films. This decrease in the thermoreflectivity signal signifies a reduction in the surface electron temperature that should translate in a lower lattice temperature at the later stage. Hence, one should expect a higher damage threshold for the multilayer samples. Comparison of the experimental results with the predictions of the Two-Temperature Model (TTM) is presented. The damage threshold of the single layer gold film corresponds to the melting threshold predicted by the model. In contrast to the single layer gold film, the multi-layer sample damaged at almost one third the damage threshold predicted by the TTM model. Possible damage mechanisms leading to the early onset of damage for the multilayer films are discussed.

  7. Deciphering the Binding between Nupr1 and MSL1 and Their DNA-Repairing Activity

    PubMed Central

    Doménech, Rosa; Pantoja-Uceda, David; Gironella, Meritxell; Santoro, Jorge; Velázquez-Campoy, Adrián; Neira, José L.; Iovanna, Juan L.

    2013-01-01

    The stress protein Nupr1 is a highly basic, multifunctional, intrinsically disordered protein (IDP). MSL1 is a histone acetyl transferase-associated protein, known to intervene in the dosage compensation complex (DCC). In this work, we show that both Nupr1 and MSL1 proteins were recruited and formed a complex into the nucleus in response to DNA-damage, which was essential for cell survival in reply to cisplatin damage. We studied the interaction of Nupr1 and MSL1, and their binding affinities to DNA by spectroscopic and biophysical methods. The MSL1 bound to Nupr1, with a moderate affinity (2.8 µM) in an entropically-driven process. MSL1 did not bind to non-damaged DNA, but it bound to chemically-damaged-DNA with a moderate affinity (1.2 µM) also in an entropically-driven process. The Nupr1 protein bound to chemically-damaged-DNA with a slightly larger affinity (0.4 µM), but in an enthalpically-driven process. Nupr1 showed different interacting regions in the formed complexes with Nupr1 or DNA; however, they were always disordered (“fuzzy”), as shown by NMR. These results underline a stochastic description of the functionality of the Nupr1 and its other interacting partners. PMID:24205110

  8. Temporal relationship of serum markers and tissue damage during acute intestinal ischemia/reperfusion

    PubMed Central

    la Garza, Francisco Javier Guzmán-de; Ibarra-Hernández, Juan Manuel; Cordero-Pérez, Paula; Villegas-Quintero, Pablo; Villarreal-Ovalle, Claudia Ivette; Torres-González, Liliana; Oliva-Sosa, Norma Edith; Alarcón-Galván, Gabriela; Fernández-Garza, Nancy Esthela; Muñoz-Espinosa, Linda Elsa; Cámara-Lemarroy, Carlos Rodrigo; Carrillo-Arriaga, José Gerardo

    2013-01-01

    OBJECTIVE: It is essential to identify a serological marker of injury in order to study the pathophysiology of intestinal ischemia reperfusion. In this work, we studied the evolution of several serological markers after intestinal ischemia reperfusion injury in rats. The markers of non-specific cell damage were aspartate aminotransferase, alanine aminotransaminase, and lactic dehydrogenase, the markers of inflammation were tumor necrosis factor alpha, interleukin-6, and interleukin-1 beta, and the markers of intestinal mucosal damage were intestinal fatty acid binding protein and D-lactate. We used Chiús classification to grade the histopathological damage. METHODS: We studied 35 Wistar rats divided into groups according to reperfusion time. The superior mesenteric artery was clamped for 30 minutes, and blood and biopsies were collected at 1, 3, 6, 12, 24, and 48 hours after reperfusion. We plotted the mean ± standard deviation and compared the baseline and maximum values for each marker using Student's t-test. RESULTS: The maximum values of interleukin-1 beta and lactic dehydrogenase were present before the maximal histopathological damage. The maximum tumor necrosis factor alpha and D-lactate expressions coincided with histopathological damage. Alanine aminotransaminase and aspartate aminotransferase had a maximum expression level that increased following the histopathological damage. The maximum expressions of interluken-6 and intestinal fatty acid binding protein were not significantly different from the Sham treated group. CONCLUSION: For the evaluation of injury secondary to acute intestinal ischemia reperfusion with a 30 minute ischemia period, we recommend performing histopathological grading, quantification of D-lactate, which is synthesized by intestinal bacteria and is considered an indicator of mucosal injury, and quantification of tumor necrosis factor alpha as indicators of acute inflammation three hours after reperfusion. PMID:23917671

  9. Gilvocarcin V, a Photodynamic DNA Damaging Agent Of Unusual Potency

    NASA Astrophysics Data System (ADS)

    Elespuru, Rosalie K.; Look, Sally A.

    1988-02-01

    Gilvocarcin V (GV) is a planar, aromatic DNA-intercalating C-glycoside isolated as a natural product antibiotic. In the presence of UVA or visible radiation, it becomes a DNA damaging agent at low doses in both bacterial and mammalian cells. In mice treated without regard to light exposure, GV exhibited antitumor activity at high doses, with little accompanying toxicity. Wavelength-dependence studies showed that lambda prophage induction profiles were similar to (part of) the absorption spectrum of GV, with a peak near 400 nm. However, significant induction at a higher wavelength (546 nm), was observed at relatively high (e.g. 1 μg/m1) concentrations of GV. The DNA damaging activity of GV was dependent on both the concentration of antibiotic and the fluence of radiation in a reciprocal manner. Mutagenesis and DNA binding experiments suggest a preference for interaction with AT-rich regions of DNA, but multiple modes of interaction seem likely. The presence of different C-glycosides on the gilvocarcin V chromophore may alter the pharmacological properties of the molecule, but photoactivation appears to be independent of these groups. The therapeutic possibilities of gilvocarcins remain largely unexplored; the demonstrated potency of these compounds when activated, the reciprocity effect, possibility of structural variation, and apparent lack of toxicity in mammalian systems are properties which could be exploited in therapeutic development.

  10. Earthquake hazard and damage on traditional rural structures in Turkey

    NASA Astrophysics Data System (ADS)

    Korkmaz, H. H.; Korkmaz, S. Z.; Donduren, M. S.

    2010-03-01

    During the last earthquakes in Turkey, reinforced concrete structures in the cities and masonry structures in the rural part were exposed to damage and failure. Masonry houses such as earthen, brick and stone structures are composed of building blocks with weak inter-binding action which have low tension capacity. Bending and shear forces generate tensile stresses which cannot be well tolerated. In this paper, the performance of masonry structures during recent earthquakes in Turkey is discussed with illustrative photographs taken after earthquakes. The followings are the main weakness in the materials and unreinforced masonry constructions and other reasons for the extensive damage of masonry buildings. Very low tensile and shear strength particularly with poor mortar, brittle behaviour in tension as well as compression, stress concentration at corners of windows and doors, overall unsymmetry in plan and elevation of building, unsymmetry due to imbalance in the sizes and positions of walls and openings in the walls, defects in construction such as use of substandard materials, unfilled joints between bricks, not-plump walls, improper bonding between walls at right angles etc.

  11. The Yeast Copper Response Is Regulated by DNA Damage

    PubMed Central

    Dong, Kangzhen; Addinall, Stephen G.; Lydall, David

    2013-01-01

    Copper is an essential but potentially toxic redox-active metal, so the levels and distribution of this metal are carefully regulated to ensure that it binds to the correct proteins. Previous studies of copper-dependent transcription in the yeast Saccharomyces cerevisiae have focused on the response of genes to changes in the exogenous levels of copper. We now report that yeast copper genes are regulated in response to the DNA-damaging agents methyl methanesulfonate (MMS) and hydroxyurea by a mechanism(s) that requires the copper-responsive transcription factors Mac1 and AceI, copper superoxide dismutase (Sod1) activity, and the Rad53 checkpoint kinase. Furthermore, in copper-starved yeast, the response of the Rad53 pathway to MMS is compromised due to a loss of Sod1 activity, consistent with the model that yeast imports copper to ensure Sod1 activity and Rad53 signaling. Crucially, the Mac1 transcription factor undergoes changes in its redox state in response to changing levels of copper or MMS. This study has therefore identified a novel regulatory relationship between cellular redox, copper homeostasis, and the DNA damage response in yeast. PMID:23959798

  12. 7 CFR 51.317 - Damage.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ..., CERTIFICATION, AND STANDARDS) United States Standards for Grades of Apples Definitions § 51.317 Damage. “Damage... appearance, or the edible or shipping quality of the apple. In addition, specific defect measurements are based on an apple three inches in diameter. Corresponding smaller or larger areas would be allowed...

  13. 7 CFR 51.317 - Damage.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ..., CERTIFICATION, AND STANDARDS) United States Standards for Grades of Apples Definitions § 51.317 Damage. “Damage... appearance, or the edible or shipping quality of the apple. In addition, specific defect measurements are based on an apple three inches in diameter. Corresponding smaller or larger areas would be allowed...

  14. Detection of insect damage in almonds

    NASA Astrophysics Data System (ADS)

    Kim, Soowon; Schatzki, Thomas F.

    1999-01-01

    Pinhole insect damage in natural almonds is very difficult to detect on-line. Further, evidence exists relating insect damage to aflatoxin contamination. Hence, for quality and health reasons, methods to detect and remove such damaged nuts are of great importance in this study, we explored the possibility of using x-ray imaging to detect pinhole damage in almonds by insects. X-ray film images of about 2000 almonds and x-ray linescan images of only 522 pinhole damaged almonds were obtained. The pinhole damaged region appeared slightly darker than non-damaged region in x-ray negative images. A machine recognition algorithm was developed to detect these darker regions. The algorithm used the first order and the second order information to identify the damaged region. To reduce the possibility of false positive results due to germ region in high resolution images, germ detection and removal routines were also included. With film images, the algorithm showed approximately an 81 percent correct recognition ratio with only 1 percent false positives whereas line scan images correctly recognized 65 percent of pinholes with about 9 percent false positives. The algorithms was very fast and efficient requiring only minimal computation time. If implemented on line, theoretical throughput of this recognition system would be 66 nuts/second.

  15. [Mechanisms of electromagnetic radiation damaging male reproduction].

    PubMed

    Xue, Lei; Chen, Hao-Yu; Wang, Shui-Ming

    2012-08-01

    More and more evidence from over 50 years of researches on the effects of electromagnetic radiation on male reproduction show that a certain dose of electromagnetic radiation obviously damages male reproduction, particularly the structure and function of spermatogenic cells. The mechanisms of the injury may be associated with energy dysmetabolism, lipid peroxidation, abnormal expressions of apoptosis-related genes and proteins, and DNA damage.

  16. Effects of rain damage on wilting forages

    Technology Transfer Automated Retrieval System (TEKTRAN)

    One of the most common problems faced by hay or silage producers is how to manage production schedules around unfavorable weather. Inevitably, some wilting forage crops are damaged by unexpected rainfall events each year, and producers often inquire about the effects of unexpected rain damage, and w...

  17. Brain Damage in Deaf Vocational Rehabilitation Clients.

    ERIC Educational Resources Information Center

    Getz, Marc; Vernon, McCay

    1986-01-01

    Screening of 54 deaf vocational clients by the Bender-Gestalt and other tests indicated the likely presence of significantly more brain damage than among the hearing population with a particularly high correlation between low IQ and brain damage in the deaf population. (DB)

  18. 7 CFR 51.2852 - Serious damage.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... specific defects shall be considered as serious damage: (a) Watery scales when more than the equivalent of two entire outer fleshy scales are affected by an off-colored, watersoaked condition. The off-color... fleshy scales, or when cuts seriously damage the appearance of the onion; and, (f) Dry sunken areas...

  19. 7 CFR 51.2852 - Serious damage.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... specific defects shall be considered as serious damage: (a) Watery scales when more than the equivalent of two entire outer fleshy scales are affected by an off-colored, watersoaked condition. The off-color... fleshy scales, or when cuts seriously damage the appearance of the onion; and, (f) Dry sunken areas...

  20. Sorting of fungal-damaged white sorghum

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A high-speed, color image-based sorting machine was modified to separate white sorghum with symptoms of fungal damage. Most of the sorghum tested was typically white, but over 27% of the bulk contained grains with fungal damage of various degrees, from severe to very slight. Grains with slight fun...

  1. 7 CFR 51.2932 - Damage.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... Standards for Grades of Apricots Definitions § 51.2932 Damage. Damage means any specific defect defined in... the apricot. The dimensions given for these defects are based on an apricot with a diameter of 17/8 inches (4.8 cm). Correspondingly larger or smaller dimensions are allowed on larger or smaller...

  2. 7 CFR 51.2932 - Damage.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... Standards for Grades of Apricots Definitions § 51.2932 Damage. Damage means any specific defect defined in... the apricot. The dimensions given for these defects are based on an apricot with a diameter of 17/8 inches (4.8 cm). Correspondingly larger or smaller dimensions are allowed on larger or smaller...

  3. 7 CFR 51.2932 - Damage.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... Standards for Grades of Apricots Definitions § 51.2932 Damage. Damage means any specific defect defined in... the apricot. The dimensions given for these defects are based on an apricot with a diameter of 17/8 inches (4.8 cm). Correspondingly larger or smaller dimensions are allowed on larger or smaller...

  4. 25 CFR 169.13 - Other damages.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 25 Indians 1 2010-04-01 2010-04-01 false Other damages. 169.13 Section 169.13 Indians BUREAU OF INDIAN AFFAIRS, DEPARTMENT OF THE INTERIOR LAND AND WATER RIGHTS-OF-WAY OVER INDIAN LANDS § 169.13 Other damages. In addition to the consideration for a grant of right-of-way provided for by the provisions...

  5. 7 CFR 51.3748 - Damage.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... Standards for Grades of Honey Dew and Honey Ball Type Melons Definitions § 51.3748 Damage. Damage means any..., or the edible or marketing quality of the melon. (a) The following specific defects shall be... pressure of the weight of other melons or from lidding of the crate; (2) Yellow spots; (3) Superficial...

  6. 7 CFR 51.3748 - Damage.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... Standards for Grades of Honey Dew and Honey Ball Type Melons Definitions § 51.3748 Damage. Damage means any..., or the edible or marketing quality of the melon. (a) The following specific defects shall be... pressure of the weight of other melons or from lidding of the crate; (2) Yellow spots; (3) Superficial...

  7. 7 CFR 51.3748 - Damage.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... Standards for Grades of Honey Dew and Honey Ball Type Melons Definitions § 51.3748 Damage. Damage means any..., or the edible or marketing quality of the melon. (a) The following specific defects shall be... pressure of the weight of other melons or from lidding of the crate; (2) Yellow spots; (3) Superficial...

  8. 7 CFR 51.3159 - Serious damage.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 7 Agriculture 2 2010-01-01 2010-01-01 false Serious damage. 51.3159 Section 51.3159 Agriculture... Standards for Grades of Nectarines Definitions § 51.3159 Serious damage. “Serious damage” means any specific...; (3) When there are more than two indentations; (4) When the aggregate area of indentations...

  9. Repair of radiation damage in mammalian cells

    SciTech Connect

    Setlow, R.B.

    1981-01-01

    The responses, such as survival, mutation, and carcinogenesis, of mammalian cells and tissues to radiation are dependent not only on the magnitude of the damage to macromolecular structures - DNA, RNA, protein, and membranes - but on the rates of macromolecular syntheses of cells relative to the half-lives of the damages. Cells possess a number of mechanisms for repairing damage to DNA. If the repair systems are rapid and error free, cells can tolerate much larger doses than if repair is slow or error prone. It is important to understand the effects of radiation and the repair of radiation damage because there exist reasonable amounts of epidemiological data that permits the construction of dose-response curves for humans. The shapes of such curves or the magnitude of the response will depend on repair. Radiation damage is emphasized because: (a) radiation dosimetry, with all its uncertainties for populations, is excellent compared to chemical dosimetry; (b) a number of cancer-prone diseases are known in which there are defects in DNA repair and radiation results in more chromosomal damage in cells from such individuals than in cells from normal individuals; (c) in some cases, specific radiation products in DNA have been correlated with biological effects, and (d) many chemical effects seem to mimic radiation effects. A further reason for emphasizing damage to DNA is the wealth of experimental evidence indicating that damages to DNA can be initiating events in carcinogenesis.

  10. 7 CFR 51.586 - Serious damage.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 2 2014-01-01 2014-01-01 false Serious damage. 51.586 Section 51.586 Agriculture..., CERTIFICATION, AND STANDARDS) United States Standards for Celery Definitions § 51.586 Serious damage. Serious... point 11/2 inches above the point of attachment to the base and the first node, or between a point...

  11. Research on fire hose couplings damages

    NASA Astrophysics Data System (ADS)

    Babut, C.; Ungureanu, N.; Ungureanu, M.

    2017-01-01

    This paper presents a part of a research on the damaging of fire hose coupling. These are important components of the hydraulic systems used by firefighters to deliver one or more suppression agents into the fire. The operating regime parameters, working environment and general functioning conditions during fire suppression operations may lead to malfunctions and/or damaging of the couplings.

  12. 7 CFR 29.2513 - Damage.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ...-Cured Tobacco (u.s. Types 22, 23, and Foreign Type 96) § 29.2513 Damage. The effect of mold, must, rot, black rot, or other fungus or bacterial diseases which attack tobacco in its cured state. Tobacco having the odor of mold, must, or rot is considered damaged. (See Rule 20, § 29.2636.)...

  13. 7 CFR 29.2513 - Damage.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ...-Cured Tobacco (u.s. Types 22, 23, and Foreign Type 96) § 29.2513 Damage. The effect of mold, must, rot, black rot, or other fungus or bacterial diseases which attack tobacco in its cured state. Tobacco having the odor of mold, must, or rot is considered damaged. (See Rule 20, § 29.2636.)...

  14. 7 CFR 29.2513 - Damage.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ...-Cured Tobacco (u.s. Types 22, 23, and Foreign Type 96) § 29.2513 Damage. The effect of mold, must, rot, black rot, or other fungus or bacterial diseases which attack tobacco in its cured state. Tobacco having the odor of mold, must, or rot is considered damaged. (See Rule 20, § 29.2636.)...

  15. 7 CFR 29.2513 - Damage.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ...-Cured Tobacco (u.s. Types 22, 23, and Foreign Type 96) § 29.2513 Damage. The effect of mold, must, rot, black rot, or other fungus or bacterial diseases which attack tobacco in its cured state. Tobacco having the odor of mold, must, or rot is considered damaged. (See Rule 20, § 29.2636.)...

  16. 7 CFR 29.2513 - Damage.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ...-Cured Tobacco (u.s. Types 22, 23, and Foreign Type 96) § 29.2513 Damage. The effect of mold, must, rot, black rot, or other fungus or bacterial diseases which attack tobacco in its cured state. Tobacco having the odor of mold, must, or rot is considered damaged. (See Rule 20, § 29.2636.)...

  17. Mechanical Properties of Shock-Damaged Rocks

    NASA Technical Reports Server (NTRS)

    He, Hongliang; Ahrens, T. J.

    1994-01-01

    Stress-strain tests were performed both on shock-damaged gabbro and limestone. The effective Young's modulus decreases with increasing initial damage parameter value, and an apparent work-softening process occurs prior to failure. To further characterize shock-induced microcracks, the longitudinal elastic wave velocity behavior of shock-damaged gabbro in the direction of compression up to failure was measured using an acoustic transmission technique under uniaxial loading. A dramatic increase in velocity was observed for the static compressive stress range of 0-50 MPa. Above that stress range, the velocity behavior of lightly damaged (D(sub 0) less than 0.1) gabbro is almost equal to unshocked gabbro. The failure strength of heavily-damaged (D(sub 0) greater than 0.1) gabbro is approx. 100-150 MPa, much lower than that of lightly damaged and unshocked gabbros (approx. 230-260 MPa). Following Nur's theory, the crack shape distribution was analyzed. The shock-induced cracks in gabbro appear to be largely thin penny-shaped cracks with c/a values below 5 x 10(exp -4). Moreover, the applicability of Ashby and Sammis's theory relating failure strength and damage parameter of shock-damaged rocks was examined and was found to yield a good estimate of the relation of shock-induced deficit in elastic modulus with the deficit in compressive strength.

  18. Damage-Sensitivity Correlations in Explosives

    DTIC Science & Technology

    2014-07-25

    Electron beam Sample X - Ray Tube Detector Rotating Stage • Pristine PBX for current study, relatively free of voids (grey circles) Damage of... X - Ray Computed Microtomography (XCMT) was conducted on the damaged explosive and the results analyzed using a MATLAB routine which quantified

  19. Eigenproperties of suspension bridges with damage

    NASA Astrophysics Data System (ADS)

    Materazzi, Annibale Luigi; Ubertini, Filippo

    2011-12-01

    The vertical vibration of suspension bridges with a damage in the main cables is studied using a continuum formulation. Starting from a model for damaged suspended cables recently proposed in the literature, an improved expression for the dynamic increment of cable tension is derived. The nonlinear equation of motion of the damaged bridge is obtained by extending this model to include the stiffening girder. The linear undamped modal eigenproperties are then extracted, in closed-form, from the linearized equation of motion, thus generalizing to the presence of an arbitrary damage the expressions known from the literature for undamaged suspension bridges. The linear dynamics of the damaged bridge reveals to be completely described by means of the same two non-dimensional parameters that govern the linear dynamics of undamaged bridges and which account for the mechanical characteristics of both the main cable and the girder, with the addition of three non-dimensional parameters characterizing damage intensity, position and extent. After presenting the mathematical formulation, a parametric analysis is conducted with the purpose of investigating the sensitivity of natural frequencies and mode shapes to damage, which, in fact, is a crucial point concerning damage detection applications using inverse methods. All through the paper, systematic comparisons with finite element simulations are presented for the purpose of model validation.

  20. 7 CFR 51.2657 - Serious damage.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... Standards for Grades for Sweet Cherries 1 Definitions § 51.2657 Serious damage. Serious damage means any... healed; (d) Cracks which are not well healed; and, (e) Pulled stems with skin or flesh of cherry torn or which causes the cherry to leak. [36 FR 8502, May 5, 1971; 36 FR 9061, May 19, 1971. Redesignated at...

  1. Damage Identification with Linear Discriminant Operators

    SciTech Connect

    Farrar, C.R.; Nix, D.A.; Duffey, T.A.; Cornwell, P.J.; Pardoen, G.C.

    1999-02-08

    This paper explores the application of statistical pattern recognition and machine learning techniques to vibration-based damage detection. First, the damage detection process is described in terms of a problem in statistical pattern recognition. Next, a specific example of a statistical-pattern-recognition-based damage detection process using a linear discriminant operator, ''Fisher's Discriminant'', is applied to the problem of identifying structural damage in a physical system. Accelerometer time histories are recorded from sensors attached to the system as that system is excited using a measured input. Linear Prediction Coding (LPC) coefficients are utilized to convert the accelerometer time-series data into multi-dimensional samples representing the resonances of the system during a brief segment of the time series. Fisher's discriminant is then used to find the linear projection of the LPC data distributions that best separates data from undamaged and damaged systems. The method i s applied to data from concrete bridge columns as the columns are progressively damaged. For this case, the method captures a clear distinction between undamaged and damaged vibration profiles. Further, the method assigns a probability of damage that can be used to rank systems in order of priority for inspection.

  2. 7 CFR 51.901 - Damage.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... whole, or the marketing quality of the stems. (a) The following shall be considered as damage to the... following shall be considered as damage to stems: (1) Active powdery mildew or any other disease when present on the stems to the extent that it detracts from the appearance of the bunch or when scars...

  3. 7 CFR 51.901 - Damage.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... whole, or the marketing quality of the stems. (a) The following shall be considered as damage to the... following shall be considered as damage to stems: (1) Active powdery mildew or any other disease when present on the stems to the extent that it detracts from the appearance of the bunch or when scars...

  4. 7 CFR 51.901 - Damage.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... whole, or the marketing quality of the stems. (a) The following shall be considered as damage to the... following shall be considered as damage to stems: (1) Active powdery mildew or any other disease when present on the stems to the extent that it detracts from the appearance of the bunch or when scars...

  5. 7 CFR 51.3157 - Damage.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... Standards for Grades of Nectarines Definitions § 51.3157 Damage. “Damage” means any specific defect defined... marketing quality of the fruit. The following specific defects shall be considered as damage: (a) Growth... one-fourth inch in length if outside of the stem cavity; (b) Heat injury, sprayburn or sunburn:...

  6. FANCI binds branched DNA and is monoubiquitinated by UBE2T-FANCL.

    PubMed

    Longerich, Simonne; San Filippo, Joseph; Liu, Dongqing; Sung, Patrick

    2009-08-28

    FANCI is integral to the Fanconi anemia (FA) pathway of DNA damage repair. Upon the occurrence of DNA damage, FANCI becomes monoubiquitinated on Lys-523 and relocalizes to chromatin, where it functions with monoubiquitinated FANCD2 to facilitate DNA repair. We show that FANCI and its C-terminal fragment possess a DNA binding activity that prefers branched structures. We also demonstrate that FANCI can be ubiquitinated on Lys-523 by the UBE2T-FANCL pair in vitro. These findings should facilitate future efforts directed at elucidating molecular aspects of the FA pathway.

  7. A Disease-Causing Variant in PCNA Disrupts a Promiscuous Protein Binding Site.

    PubMed

    Duffy, Caroline M; Hilbert, Brendan J; Kelch, Brian A

    2016-03-27

    The eukaryotic DNA polymerase sliding clamp, proliferating cell nuclear antigen or PCNA, is a ring-shaped protein complex that surrounds DNA to act as a sliding platform for increasing processivity of cellular replicases and for coordinating various cellular pathways with DNA replication. A single point mutation, Ser228Ile, in the human PCNA gene was recently identified to cause a disease whose symptoms resemble those of DNA damage and repair disorders. The mutation lies near the binding site for most PCNA-interacting proteins. However, the structural consequences of the S228I mutation are unknown. Here, we describe the structure of the disease-causing variant, which reveals a large conformational change that dramatically transforms the binding pocket for PCNA client proteins. We show that the mutation markedly alters the binding energetics for some client proteins, while another, p21(CIP1), is only mildly affected. Structures of the disease variant bound to peptides derived from two PCNA partner proteins reveal that the binding pocket can adjust conformation to accommodate some ligands, indicating that the binding site is dynamic and pliable. Our work has implications for the plasticity of the binding site in PCNA and reveals how a disease mutation selectively alters interactions to a promiscuous binding site that is critical for DNA metabolism.

  8. The activation of a specific DNA binding protein by neutron irradiation

    SciTech Connect

    Teale, B.; Singh, S.; Cohen, D.

    1995-08-30

    The purpose of this investigation was to determine whether the quality of ionizing radiation is critical for activation of a radiation-specific DNA binding protein. We have previously shown that after exposing Epstein Barr virus-transformed lymphoblastoid cells to ionizing radiation, a specific DNA binding factor appears in the nucleus apparently as a result of translocation from the cytoplasm. This protein binds to a number of different genomic sequences and a consensus motif has been identified. Because the protein was not activated by UV light, it was of interest whether high linear energy transfer (LET) radiation was capable of activation. We describe here the activation of a specific DNA binding protein by high LET neutron radiation. The protein binds a region adjacent to and overlapping with the distal repeat within a 179 base-pair fragment of the well-characterized Simian Virus (SV40) bidirectional promoter/enhancer element. The appearance of the DNA binding activity was dose dependent and reached a maximum level by 90 min postirradiation. A reduction in DNA binding activity was evident at later times after irradiation. The specific nature of this response and the rapidity of activation may indicate a pivotal role for this protein in repair or in some other aspect of the cellular response to radiation damage. 22 refs., 4 figs.

  9. Current pulse effects on cylindrical damage experiments

    SciTech Connect

    Kaul, Ann M; Rousculp, Christopher L

    2009-01-01

    A series of joint experiments between LANL and VNIIEF use a VNIIEF-designed helical generator to provide currents for driving a LANL-designed cylindrical spallation experimental load. Under proper driving conditions, a cylindrical configuration allows for a natural recollection of the damaged material. In addition, the damaged material is able to come to a complete stop due to its strength, avoiding application of further forces. Thus far, experiments have provided data about failure initiation of a well-characterized material (aluminum) in a cylindrical geometry, behavior of material recollected after damage from pressures in the damage initiation regime, and behavior of material recollected after complete failure. In addition to post-shot collection of the damaged target material for subsequent metallographic analysis, dynamic in-situ experimental diagnostics include velocimetry and transverse radial radiography. This paper will focus on the effects of tailoring the driving current pulse to obtain the desired data.

  10. Comprehensive model of damage accumulation in silicon

    SciTech Connect

    Mok, K. R. C.; Benistant, F.; Jaraiz, M.; Rubio, J. E.; Castrillo, P.; Pinacho, R.; Srinivasan, M. P.

    2008-01-01

    Ion implantation induced damage accumulation is crucial to the simulation of silicon processing. We present a physically based damage accumulation model, implemented in a nonlattice atomistic kinetic Monte Carlo simulator, that can simulate a diverse range of interesting experimental observations. The model is able to reproduce the ion-mass dependent silicon amorphous-crystalline transition temperature of a range of ions from C to Xe, the amorphous layer thickness for a range of amorphizing implants, the superlinear increase in damage accumulation with dose, and the two-layered damage distribution observed along the path of a high-energy ion. In addition, this model is able to distinguish between dynamic annealing and post-cryogenic implantation annealing, whereby dynamic annealing is more effective in removing damage than post-cryogenic implantation annealing at the same temperature.

  11. Surviving the breakup: the DNA damage checkpoint.

    PubMed

    Harrison, Jacob C; Haber, James E

    2006-01-01

    In response to even a single chromosomal double-strand DNA break, cells enact the DNA damage checkpoint. This checkpoint triggers cell cycle arrest, providing time for the cell to repair damaged chromosomes before entering mitosis. This mechanism helps prevent the segregation of damaged or mutated chromosomes and thus promotes genomic stability. Recent work has elucidated the molecular mechanisms underlying several critical steps in checkpoint activation, notably the recruitment of the upstream checkpoint kinases of the ATM and ATR families to different damaged DNA structures and the molecular events through which these kinases activate their effectors. Chromatin modification has emerged as one important component of checkpoint activation and maintenance. Following DNA repair, the checkpoint pathway is inactivated in a process termed recovery. A related but genetically distinct process, adaptation, controls cell cycle re-entry in the face of unrepairable damage.

  12. Tissue damage detection by osmotic surveillance.

    PubMed

    Enyedi, Balázs; Kala, Snigdha; Nikolich-Zugich, Tijana; Niethammer, Philipp

    2013-09-01

    How tissue damage is detected to induce inflammatory responses is unclear. Most studies have focused on damage signals released by cell breakage and necrosis. Whether tissues use other cues in addition to cell lysis to detect that they are damaged is unknown. We find that osmolarity differences between interstitial fluid and the external environment mediate rapid leukocyte recruitment to sites of tissue damage in zebrafish by activating cytosolic phospholipase a2 (cPLA2) at injury sites. cPLA2 initiates the production of non-canonical arachidonate metabolites that mediate leukocyte chemotaxis through a 5-oxo-ETE receptor (OXE-R). Thus, tissues can detect damage through direct surveillance of barrier integrity, with cell swelling probably functioning as a pro-inflammatory intermediate in the process.

  13. Ozone damage detection in cantaloupe plants

    NASA Technical Reports Server (NTRS)

    Gausman, H. W.; Escobar, D. E.; Rodriguez, R. R.; Thomas, C. E.; Bowen, R. L.

    1978-01-01

    Ozone causes up to 90 percent of air pollution injury to vegetation in the United States; excess ozone affects plant growth and development and can cause undetected decrease in yields. Laboratory and field reflectance measurements showed that ozone-damaged cantaloupe (Cucumis melo L.) leaves had lower water contents and higher reflectance than did nondamaged leaves. Cantaloupe plants which were lightly, severely, and very severely ozone-damaged were distinguishable from nondamaged plants by reflectance measurements in the 1.35- to 2.5 micron near-infrared water absorption waveband. Ozone-damaged leaf areas were detected photographically 16 h before the damage was visible. Sensors are available for use with aircraft and spacecraft that possibly could be used routinely to detect ozone-damaged crops.

  14. Local wavefield velocity imaging for damage evaluation

    NASA Astrophysics Data System (ADS)

    Chia, Chen Ciang; Gan, Chia Sheng; Mustapha, F.

    2017-02-01

    Ultrasonic Propagation Imaging or Acoustic Wavefield Imaging has been widely used to evaluate structural damages and internal features. Inspecting complete wavefield time history for damage identification is tedious and error-prone. A more effective way is by extracting damage-related information into a single image. A wavefield velocity imaging method that maps the local estimates of group or phase velocity is proposed. Actual velocity values rather than arbitrarily-scaled intensities are mapped, enabling damage sizing without the need of supervised training or inspecting wavefield propagation video. Performance of the proposed method was tested by inspecting a 100 mm by 100 mm area of a 2 mm thick stainless steel specimen. Local phase velocity maps of A0 mode showed a half-thickness hole of 2 mm diameter as significant change in local phase velocity from the nominal 2 m/ms. Full width at half maximum of relevant velocity profiles proved the accuracy and consistency of the damage sizing.

  15. Cavitation damage in blood clots under HIFU

    NASA Astrophysics Data System (ADS)

    Weiss, Hope; Ahadi, Golnaz; Hoelscher, Thilo; Szeri, Andrew

    2010-11-01

    High Intensity Focused Ultrasound (HIFU) has been shown to accelerate thrombolysis, the dissolution of blood clots, in vitro and in vivo, for treatment of ischemic stroke. Cavitation in sonothrombolysis is thought to play an important role, although the mechanisms are not fully understood. The damage to a blood clot associated with bubble collapses in a HIFU field is studied. The region of damage caused by a bubble collapse on the fibrin network of the blood clot exposed to HIFU is estimated, and compared with experimental assessment of the damage. The mechanical damage to the network caused by a bubble is probed using two independent approaches, a strain based method and an energy based method. Immunoflourescent fibrin staining is used to assess the region of damage experimentally.

  16. Beta-lactam-fosfomycin antagonism involving modification of penicillin-binding protein 3 in Pseudomonas aeruginosa.

    PubMed Central

    Reguera, J A; Baquero, F; Berenguer, J; Martinez-Ferrer, M; Martinez, J L

    1990-01-01

    Antagonism between fosfomycin and antipseudomonal penicillins, cefotaxime, and ceftriaxone was observed in Pseudomonas aeruginosa RYC212. Fosfomycin, a non-beta-lactam antibiotic that acts on bacterial cell wall synthesis, decreased the expression of penicillin-binding protein 3 and induced beta-lactamase. The antagonistic effect was reduced in the presence of high concentrations of the beta-lactamase inhibitor tazobactam or in fosfomycin-resistant mutants. We suggest that products resulting from fosfomycin cell wall damage could interact with a system that regulates penicillin-binding protein and beta-lactamase production. Images PMID:2127343

  17. Temporal binding of interval markers

    PubMed Central

    Derichs, Christina; Zimmermann, Eckart

    2016-01-01

    How we estimate the passage of time is an unsolved mystery in neuroscience. Illusions of subjective time provide an experimental access to this question. Here we show that time compression and expansion of visually marked intervals result from a binding of temporal interval markers. Interval markers whose onset signals were artificially weakened by briefly flashing a whole-field mask were bound in time towards markers with a strong onset signal. We explain temporal compression as the consequence of summing response distributions of weak and strong onset signals. Crucially, temporal binding occurred irrespective of the temporal order of weak and strong onset markers, thus ruling out processing latencies as an explanation for changes in interval duration judgments. If both interval markers were presented together with a mask or the mask was shown in the temporal interval center, no compression occurred. In a sequence of two intervals, masking the middle marker led to time compression for the first and time expansion for the second interval. All these results are consistent with a model view of temporal binding that serves a functional role by reducing uncertainty in the final estimate of interval duration. PMID:27958311

  18. Anion binding in biological systems

    NASA Astrophysics Data System (ADS)

    Feiters, Martin C.; Meyer-Klaucke, Wolfram; Kostenko, Alexander V.; Soldatov, Alexander V.; Leblanc, Catherine; Michel, Gurvan; Potin, Philippe; Küpper, Frithjof C.; Hollenstein, Kaspar; Locher, Kaspar P.; Bevers, Loes E.; Hagedoorn, Peter-Leon; Hagen, Wilfred R.

    2009-11-01

    We compare aspects of biological X-ray absorption spectroscopy (XAS) studies of cations and anions, and report on some examples of anion binding in biological systems. Brown algae such as Laminaria digitata (oarweed) are effective accumulators of I from seawater, with tissue concentrations exceeding 50 mM, and the vanadate-containing enzyme haloperoxidase is implicated in halide accumulation. We have studied the chemical state of iodine and its biological role in Laminaria at the I K edge, and bromoperoxidase from Ascophyllum nodosum (knotted wrack) at the Br K edge. Mo is essential for many forms of life; W only for certain archaea, such as Archaeoglobus fulgidus and the hyperthermophilic archaeon Pyrococcus furiosus, and some bacteria. The metals are bound and transported as their oxo-anions, molybdate and tungstate, which are similar in size. The transport protein WtpA from P. furiosus binds tungstate more strongly than molybdate, and is related in sequence to Archaeoglobus fulgidus ModA, of which a crystal structure is known. We have measured A. fulgidus ModA with tungstate at the W L3 (2p3/2) edge, and compared the results with the refined crystal structure. XAS studies of anion binding are feasible even if only weak interactions are present, are biologically relevant, and give new insights in the spectroscopy.

  19. Significance of "high probability/low damage" versus "low probability/high damage" flood events

    NASA Astrophysics Data System (ADS)

    Merz, B.; Elmer, F.; Thieken, A. H.

    2009-06-01

    The need for an efficient use of limited resources fosters the application of risk-oriented design in flood mitigation. Flood defence measures reduce future damage. Traditionally, this benefit is quantified via the expected annual damage. We analyse the contribution of "high probability/low damage" floods versus the contribution of "low probability/high damage" events to the expected annual damage. For three case studies, i.e. actual flood situations in flood-prone communities in Germany, it is shown that the expected annual damage is dominated by "high probability/low damage" events. Extreme events play a minor role, even though they cause high damage. Using typical values for flood frequency behaviour, flood plain morphology, distribution of assets and vulnerability, it is shown that this also holds for the general case of river floods in Germany. This result is compared to the significance of extreme events in the public perception. "Low probability/high damage" events are more important in the societal view than it is expressed by the expected annual damage. We conclude that the expected annual damage should be used with care since it is not in agreement with societal priorities. Further, risk aversion functions that penalise events with disastrous consequences are introduced in the appraisal of risk mitigation options. It is shown that risk aversion may have substantial implications for decision-making. Different flood mitigation decisions are probable, when risk aversion is taken into account.

  20. The FHA domain determines Drosophila Chk2/Mnk localization to key mitotic structures and is essential for early embryonic DNA damage responses.

    PubMed

    Takada, Saeko; Collins, Eric R; Kurahashi, Kayo

    2015-05-15

    DNA damage responses, including mitotic centrosome inactivation, cell-cycle delay in mitosis, and nuclear dropping from embryo cortex, maintain genome integrity in syncytial Drosophila embryos. A conserved signaling kinase, Chk2, known as Mnk/Loki, is essential for the responses. Here we demonstrate that functional EGFP-Mnk expressed from a transgene localizes to the nucleus, centrosomes, interkinetochore/centromere region, midbody, and pseudocleavage furrows without DNA damage and in addition forms numerous foci/aggregates on mitotic chromosomes upon DNA damage. We expressed EGFP-tagged Mnk deletion or point mutation variants and investigated domain functions of Mnk in vivo. A triple mutation in the phosphopeptide-binding site of the forkhead-associated (FHA) domain disrupted normal Mnk localization except to the nucleus. The mutation also disrupted Mnk foci formation on chromosomes upon DNA damage. FHA mutations and deletion of the SQ/TQ-cluster domain (SCD) abolished Mnk transphosphorylations and autophosphorylations, indicative of kinase activation after DNA damage. A potent NLS was found at the C-terminus, which is required for normal Mnk function. We propose that the FHA domain in Mnk plays essential dual functions in mediating embryonic DNA damage responses by means of its phosphopeptide-binding ability: activating Mnk in the nucleus upon DNA damage and recruiting Mnk to multiple subcellular structures independently of DNA damage.

  1. The FHA domain determines Drosophila Chk2/Mnk localization to key mitotic structures and is essential for early embryonic DNA damage responses

    PubMed Central

    Takada, Saeko; Collins, Eric R.; Kurahashi, Kayo

    2015-01-01

    DNA damage responses, including mitotic centrosome inactivation, cell-cycle delay in mitosis, and nuclear dropping from embryo cortex, maintain genome integrity in syncytial Drosophila embryos. A conserved signaling kinase, Chk2, known as Mnk/Loki, is essential for the responses. Here we demonstrate that functional EGFP-Mnk expressed from a transgene localizes to the nucleus, centrosomes, interkinetochore/centromere region, midbody, and pseudocleavage furrows without DNA damage and in addition forms numerous foci/aggregates on mitotic chromosomes upon DNA damage. We expressed EGFP-tagged Mnk deletion or point mutation variants and investigated domain functions of Mnk in vivo. A triple mutation in the phosphopeptide-binding site of the forkhead-associated (FHA) domain disrupted normal Mnk localization except to the nucleus. The mutation also disrupted Mnk foci formation on chromosomes upon DNA damage. FHA mutations and deletion of the SQ/TQ-cluster domain (SCD) abolished Mnk transphosphorylations and autophosphorylations, indicative of kinase activation after DNA damage. A potent NLS was found at the C-terminus, which is required for normal Mnk function. We propose that the FHA domain in Mnk plays essential dual functions in mediating embryonic DNA damage responses by means of its phosphopeptide-binding ability: activating Mnk in the nucleus upon DNA damage and recruiting Mnk to multiple subcellular structures independently of DNA damage. PMID:25808488

  2. Subsurface damage detection and damage mechanism analysis of chemical-mechanical polished optics

    NASA Astrophysics Data System (ADS)

    Ye, Hui; Yang, Wei; Bi, Guo; Yang, Ping; Guo, Yinbiao

    2014-09-01

    Detection of the subsurface damage depth in optical elements has significance on the subsequent material removal amount and improving element surface quality. The paper focuses on the subsurface damage of chemical-mechanical polished K9 specimen, and analyses the chemical-mechanical polishing mechanism and the cause of subsurface damage. A most suitable etchant is chosen and the step-by-step etching method is applied to measure the subsurface damage depth. A microscope is used to detect the damage morphology and the variation trend at different depth. Research shows that the subsurface damage caused by chemical-mechanical polishing is Hertz scratch, and the scratch quantity below surface presents a variation of zero-more-less-disappeared. The K9 specimen is polished for 3 min under the pressure of 2.5 Kgf and the spindle speed of 43139 r/min, thus resulting in a subsurface damage depth 15.3μm.

  3. Damage Tolerant Microstructures for Shock Environments

    NASA Astrophysics Data System (ADS)

    Cerreta, Ellen; Dennis-Koller, Darcie; Escobedo, Juan Pablo; Fensin, Saryu; Valone, Steve; Trujillo, Carl; Bronkhorst, Curt; Lebensohn, Ricardo

    While dynamic failure, due to shock loading, has been studied for many years, our current ability to predict and simulate evolving damage during dynamic loading remains limited. One reason for this is due to the lack of understanding for the linkages between process-induced as well as evolved microstructure and damage. To this end, the role of microstructure on the early stages of dynamic damage has been studied in high purity Ta and Cu. This work, which utilizes plate-impact experiments to interrogate these effects, has recently been extended to a subset to Cu-alloys (Cu-Pb, Cu-Nb, and Cu-Ag). These multi-length scale studies, have identified a number of linkages between damage nucleation and growth and microstructural features such as: grain boundary types, grain boundary orientation with respect to loading direction, grain orientation, and bi-metal interfaces. A combination of modeling and simulation techniques along with experimental observation has been utilized to examine the mechanisms for the ductile damage processes such as nucleation, growth and coalescence. This work has identified differing features of importance for damage nucleation in high purity and alloyed materials, lending insight into features of concern for mitigating shock induced damage in more complicated alloy systems.

  4. Flat Surface Damage Detection System (FSDDS)

    NASA Technical Reports Server (NTRS)

    Williams, Martha; Lewis, Mark; Gibson, Tracy; Lane, John; Medelius, Pedro; Snyder, Sarah; Ciarlariello, Dan; Parks, Steve; Carrejo, Danny; Rojdev, Kristina

    2013-01-01

    The Flat Surface Damage Detection system (FSDDS} is a sensory system that is capable of detecting impact damages to surfaces utilizing a novel sensor system. This system will provide the ability to monitor the integrity of an inflatable habitat during in situ system health monitoring. The system consists of three main custom designed subsystems: the multi-layer sensing panel, the embedded monitoring system, and the graphical user interface (GUI). The GUI LABVIEW software uses a custom developed damage detection algorithm to determine the damage location based on the sequence of broken sensing lines. It estimates the damage size, the maximum depth, and plots the damage location on a graph. Successfully demonstrated as a stand alone technology during 2011 D-RATS. Software modification also allowed for communication with HDU avionics crew display which was demonstrated remotely (KSC to JSC} during 2012 integration testing. Integrated FSDDS system and stand alone multi-panel systems were demonstrated remotely and at JSC, Mission Operations Test using Space Network Research Federation (SNRF} network in 2012. FY13, FSDDS multi-panel integration with JSC and SNRF network Technology can allow for integration with other complementary damage detection systems.

  5. Radiation damage in barium fluoride detector materials

    SciTech Connect

    Levey, P.W.; Kierstead, J.A.; Woody, C.L.

    1988-01-01

    To develop radiation hard detectors, particularly for high energy physics studies, radiation damage is being studied in BaF/sub 2/, both undoped and doped with La, Ce, Nd, Eu, Gd and Tm. Some dopants reduce radiation damage. In La doped BaF/sub 2/ they reduce the unwanted long lifetime luminescence which interferes with the short-lived fluorescence used to detect particles. Radiation induced coloring is being studied with facilities for making optical measurements before, during and after irradiation with /sup 60/C0 gamma rays. Doses of 10/sup 6/ rad, or less, create only ionization induced charge transfer effects since lattice atom displacement damage is negligible at these doses. All crystals studied exhibit color center formation, between approximately 200 and 800 nm, during irradiation and color center decay after irradiation. Thus only measurements made during irradiation show the total absorption present in a radiation field. Both undoped and La doped BaF/sub 2/ develop damage at minimum detectable levels in the UV---which is important for particle detectors. For particle detector applications these studies must be extended to high dose irradiations with particles energetic enough to cause lattice atom displacement damage. In principle, the reduction in damage provided by dopants could apply to other applications requiring radiation damage resistant materials.

  6. Intelligent-based Structural Damage Detection Model

    SciTech Connect

    Lee, Eric Wai Ming; Yu, K.F.

    2010-05-21

    This paper presents the application of a novel Artificial Neural Network (ANN) model for the diagnosis of structural damage. The ANN model, denoted as the GRNNFA, is a hybrid model combining the General Regression Neural Network Model (GRNN) and the Fuzzy ART (FA) model. It not only retains the important features of the GRNN and FA models (i.e. fast and stable network training and incremental growth of network structure) but also facilitates the removal of the noise embedded in the training samples. Structural damage alters the stiffness distribution of the structure and so as to change the natural frequencies and mode shapes of the system. The measured modal parameter changes due to a particular damage are treated as patterns for that damage. The proposed GRNNFA model was trained to learn those patterns in order to detect the possible damage location of the structure. Simulated data is employed to verify and illustrate the procedures of the proposed ANN-based damage diagnosis methodology. The results of this study have demonstrated the feasibility of applying the GRNNFA model to structural damage diagnosis even when the training samples were noise contaminated.

  7. Nucleotides of transcription factor binding sites exert interdependent effects on the binding affinities of transcription factors

    PubMed Central

    Bulyk, Martha L.; Johnson, Philip L. F.; Church, George M.

    2002-01-01

    We can determine the effects of many possible sequence variations in transcription factor binding sites using microarray binding experiments. Analysis of wild-type and mutant Zif268 (Egr1) zinc fingers bound to microarrays containing all possible central 3 bp triplet binding sites indicates that the nucleotides of transcription factor binding sites cannot be treated independently. This indicates that the current practice of characterizing transcription factor binding sites by mutating individual positions of binding sites one base pair at a time does not provide a true picture of the sequence specificity. Similarly, current bioinformatic practices using either just a consensus sequence, or even mononucleotide frequency weight matrices to provide more complete descriptions of transcription factor binding sites, are not accurate in depicting the true binding site specificities, since these methods rely upon the assumption that the nucleotides of binding sites exert independent effects on binding affinity. Our results stress the importance of complete reference tables of all possible binding sites for comparing protein binding preferences for various DNA sequences. We also show results suggesting that microarray binding data using particular subsets of all possible binding sites can be used to extrapolate the relative binding affinities of all possible full-length binding sites, given a known binding site for use as a starting sequence for site preference refinement. PMID:11861919

  8. Role of beer lipid-binding proteins in preventing lipid destabilization of foam.

    PubMed

    Cooper, D J; Husband, F A; Mills, E N C; Wilde, P J

    2002-12-18

    The negative effect of fatty acids on the foam stability of beer has been assessed. Long-chain fatty acids are far more damaging than short-chain fatty acids on the foam stability of beer at the concentrations employed. Polypeptides have been isolated from an all malt beer by hydrophobic interaction chromatography. Using this technique five groups of polypeptides were isolated, group 1 being the least hydrophobic and group 5 the most hydrophobic, all of which exhibited similar polypeptide compositions by SDS-PAGE. All five hydrophobic polypeptide groups bound [(14)C]linoleic acid; however, group 5, the most hydrophobic group, bound the most linoleic acid. Groups 1 and 5 were titrated with cis-parinaric acid (CPA) to produce binding curves, which were compared with a binding curve obtained for bovine serum albumin (BSA). Groups 1 and 5 both produced binding curves that saturated at approximately 5.5 microM and 4 microM CPA and had association constants (K(a)) of 6.27 x 10(7) and 1.62 x 10(7) M(-1), respectively. In comparison, BSA produced a binding curve that saturated at 6 microM CPA and had a K(a) of 3.95 x 10(7) M(-1). Further investigation has shown that group 1 is pH sensitive and group 5 pH insensitive with respect to lipid binding. The lipid-binding activity of group 5 was also shown to be unaffected by ethanol concentration. Linoleic acid (5 microM) when added to beer resulted in unstable foam. Group 5 was added to the lipid-damaged beer and was shown to restore the foam stability to values that were obtained for the control beer. It has therefore been demonstrated that proteins isolated from beer have a lipid-binding capacity and that they can convey a degree of protection against lipid-induced foam destabilization.

  9. Is post-transcriptional stabilization, splicing and translation of selective mRNAs a key to the DNA damage response?

    PubMed Central

    2011-01-01

    In response to DNA damage, cells activate a complex, kinase-based signaling network that consists of two components—a rapid phosphorylation-driven signaling cascade that results in immediate inhibition of Cdk/cyclin complexes to arrest the cell cycle along with recruitment of repair machinery to damaged DNA, followed by a delayed transcriptional response that promotes cell cycle arrest through the induction of Cdk inhibitors, such as p21. In recent years a third layer of complexity has emerged that involves post-transcriptional control of mRNA stability, splicing and translation as a critical part of the DNA damage response. Here, we describe recent work implicating DNA damage-dependent modification of RNA-binding proteins that are responsible for some of these mRNA effects, highlighting recent work on post-transcriptional regulation of the cell cycle checkpoint protein/apoptosis inducer Gadd45α by the checkpoint kinase MAPKAP Kinase-2. PMID:21173571

  10. Tissue damage thresholds during therapeutic electrical stimulation

    NASA Astrophysics Data System (ADS)

    Cogan, Stuart F.; Ludwig, Kip A.; Welle, Cristin G.; Takmakov, Pavel

    2016-04-01

    Objective. Recent initiatives in bioelectronic modulation of the nervous system by the NIH (SPARC), DARPA (ElectRx, SUBNETS) and the GlaxoSmithKline Bioelectronic Medicines effort are ushering in a new era of therapeutic electrical stimulation. These novel therapies are prompting a re-evaluation of established electrical thresholds for stimulation-induced tissue damage. Approach. In this review, we explore what is known and unknown in published literature regarding tissue damage from electrical stimulation. Main results. For macroelectrodes, the potential for tissue damage is often assessed by comparing the intensity of stimulation, characterized by the charge density and charge per phase of a stimulus pulse, with a damage threshold identified through histological evidence from in vivo experiments as described by the Shannon equation. While the Shannon equation has proved useful in assessing the likely occurrence of tissue damage, the analysis is limited by the experimental parameters of the original studies. Tissue damage is influenced by factors not explicitly incorporated into the Shannon equation, including pulse frequency, duty cycle, current density, and electrode size. Microelectrodes in particular do not follow the charge per phase and charge density co-dependence reflected in the Shannon equation. The relevance of these factors to tissue damage is framed in the context of available reports from modeling and in vivo studies. Significance. It is apparent that emerging applications, especially with microelectrodes, will require clinical charge densities that exceed traditional damage thresholds. Experimental data show that stimulation at higher charge densities can be achieved without causing tissue damage, suggesting that safety parameters for microelectrodes might be distinct from those defined for macroelectrodes. However, these increased charge densities may need to be justified by bench, non-clinical or clinical testing to provide evidence of device

  11. Tissue damage thresholds during therapeutic electrical stimulation

    PubMed Central

    Cogan, Stuart F; Ludwig, Kip A; Welle, Cristin G; Takmakov, Pavel

    2017-01-01

    Objective Recent initiatives in bioelectronic modulation of the nervous system by the NIH (SPARC), DARPA (ElectRx, SUBNETS) and the GlaxoSmithKline Bioelectronic Medicines effort are ushering in a new era of therapeutic electrical stimulation. These novel therapies are prompting a re-evaluation of established electrical thresholds for stimulation-induced tissue damage. Approach In this review, we explore what is known and unknown in published literature regarding tissue damage from electrical stimulation. Main results For macroelectrodes, the potential for tissue damage is often assessed by comparing the intensity of stimulation, characterized by the charge density and charge per phase of a stimulus pulse, with a damage threshold identified through histological evidence from in vivo experiments as described by the Shannon equation. While the Shannon equation has proved useful in assessing the likely occurrence of tissue damage, the analysis is limited by the experimental parameters of the original studies. Tissue damage is influenced by factors not explicitly incorporated into the Shannon equation, including pulse frequency, duty cycle, current density, and electrode size. Microelectrodes in particular do not follow the charge per phase and charge density co-dependence reflected in the Shannon equation. The relevance of these factors to tissue damage is framed in the context of available reports from modeling and in vivo studies. Significance It is apparent that emerging applications, especially with microelectrodes, will require clinical charge densities that exceed traditional damage thresholds. Experimental data show that stimulation at higher charge densities can be achieved without causing tissue damage, suggesting that safety parameters for microelectrodes might be distinct from those defined for macroelectrodes. However, these increased charge densities may need to be justified by bench, non-clinical or clinical testing to provide evidence of device safety

  12. Cooperative Ligand Binding to Linear Chain Molecules

    ERIC Educational Resources Information Center

    Applequist, Jon

    1977-01-01

    Summarizes the Ising model of ligand binding as it applies to cooperative binding to long chain molecules. Also presents some illustrations which help to visualize the connection between the interaction parameters and the shape of the binding isotherm. (Author/MR)

  13. Erythropoietin binding protein from mammalian serum

    DOEpatents

    Clemons, G.K.

    1997-04-29

    Purified mammalian erythropoietin binding-protein is disclosed, and its isolation, identification, characterization, purification, and immunoassay are described. The erythropoietin binding protein can be used for regulation of erythropoiesis by regulating levels and half-life of erythropoietin. A diagnostic kit for determination of level of erythropoietin binding protein is also described. 11 figs.

  14. Feature-Based Binding and Phase Theory

    ERIC Educational Resources Information Center

    Antonenko, Andrei

    2012-01-01

    Current theories of binding cannot provide a uniform account for many facts associated with the distribution of anaphors, such as long-distance binding effects and the subject-orientation of monomorphemic anaphors. Further, traditional binding theory is incompatible with minimalist assumptions. In this dissertation I propose an analysis of…

  15. Erythropoietin binding protein from mammalian serum

    DOEpatents

    Clemons, Gisela K.

    1997-01-01

    Purified mammalian erythropoietin binding-protein is disclosed, and its isolation, identification, characterization, purification, and immunoassay are described. The erythropoietin binding protein can be used for regulation of erythropoiesis by regulating levels and half-life of erythropoietin. A diagnostic kit for determination of level of erythropoietin binding protein is also described.

  16. Damage Detection Using Holography and Interferometry

    NASA Technical Reports Server (NTRS)

    Decker, Arthur J.

    2003-01-01

    This paper reviews classical approaches to damage detection using laser holography and interferometry. The paper then details the modern uses of electronic holography and neural-net-processed characteristic patterns to detect structural damage. The design of the neural networks and the preparation of the training sets are discussed. The use of a technique to optimize the training sets, called folding, is explained. Then a training procedure is detailed that uses the holography-measured vibration modes of the undamaged structures to impart damage-detection sensitivity to the neural networks. The inspections of an optical strain gauge mounting plate and an International Space Station cold plate are presented as examples.

  17. Fatigue damage analysis under variable amplitude cycling

    NASA Technical Reports Server (NTRS)

    Leis, B. N.; Forte, T. P.

    1983-01-01

    This paper explores the suitability of a recently proposed mean stress parameter and introduces a nonlinear damage accumulation procedure. Data covering a range of positive and negative stress ratios from +0.6 to -2.66, for several aluminum alloys and steels, are assembled and shown to be well correlated by a simple damage parameter. A nonlinear damage accumulation postulate is advanced to replace the usual linear procedure. Results of critical experiments performed to assess the suitability of the postulate are introduced and shown to support a non-linear criterion. The implications of this work related to variable amplitude life prediction are discussed.

  18. Aging processes, DNA damage, and repair.

    PubMed

    Gilchrest, B A; Bohr, V A

    1997-04-01

    The second triennial FASEB Summer Research Conference on "Clonal Senescence and Differentiation" (August 17-22, 1996) focused on the interrelationships between aging processes and DNA damage and repair. The attendees represented a cross section of senior and junior investigators working in fields ranging from classic cellular gerontology to yeast and nematode models of aging to basic mechanisms of DNA damage and repair. The meeting opened with a keynote address by Dr. Bruce Ames that emphasized the documented relationships between oxidative damage, cancer, and aging. This was followed by eight platform sessions, one poster discussion, one featured presentation, and an after-dinner address. The following sections highlight the key points discussed.

  19. [Liability in Anaesthesiology: theory of disproportionate damage].

    PubMed

    Galán Gutiérrez, J C; Galán Cortés, J C

    2013-10-01

    An analysis is made of the controversial application of the theory of disproportionate damage in the anaesthetic act, due to the high inherent risk, and regardless of the seriousness and importance of the surgery being performed. The existence of a disproportionate damage, that is, damage not foreseen nor accountable within the framework of the professional performance of the anaesthetist, does not by itself determine the existence of liability on the part of the anaesthetist, but the demand from the professionals themselves for a coherent explanation of the serious disagreement between the initial risk implied by their actions and the final consequence produced.

  20. Control of radiation damage in the TEM.

    PubMed

    Egerton, R F

    2013-04-01

    The problem of electron-beam damage in the transmission electron microscope is reviewed, with an emphasis on radiolysis processes in soft materials and organic specimens. Factors that determine the dose-limited resolution are identified for three different operational modes: bright-field scattering-contrast, phase-contrast and dark-field microscopy. Methods of reducing radiation damage are discussed, including low-dose techniques, cooling or encapsulating the specimen, and the choice of imaging mode, incident-beam diameter and incident-electron energy. Further experiments are suggested as a means of obtaining a better understanding and control of electron-beam damage.

  1. CFD Script for Rapid TPS Damage Assessment

    NASA Technical Reports Server (NTRS)

    McCloud, Peter

    2013-01-01

    This grid generation script creates unstructured CFD grids for rapid thermal protection system (TPS) damage aeroheating assessments. The existing manual solution is cumbersome, open to errors, and slow. The invention takes a large-scale geometry grid and its large-scale CFD solution, and creates a unstructured patch grid that models the TPS damage. The flow field boundary condition for the patch grid is then interpolated from the large-scale CFD solution. It speeds up the generation of CFD grids and solutions in the modeling of TPS damages and their aeroheating assessment. This process was successfully utilized during STS-134.

  2. War damages and reconstruction of Peruca dam

    SciTech Connect

    Nonveiller, E.; Rupcic, J. |; Sever, Z.

    1999-04-01

    The paper describes the heavy damages caused by blasting in the Peruca rockfill dam in Croatia in January 1993. Complete collapse of the dam by overtopping was prevented through quick action of the dam owner by dumping clayey gravel on the lowest sections of the dam crest and opening the bottom outlet of the reservoir, thus efficiently lowering the water level. After the damages were sufficiently established and alternatives for restoration of the dam were evaluated, it was decided to construct a diaphragm wall through the damaged core in the central dam part as the impermeable dam element and to rebuild the central clay core at the dam abutments. Reconstruction works are described.

  3. NONDESTRUCTIVE EVALUATION (NDE) OF DAMAGED STRUCTURAL CERAMICS

    SciTech Connect

    Brennan, R. E.; Green, W. H.; Sands, J. M.; Yu, J. H.

    2009-03-03

    A combination of destructive and nondestructive testing methods was utilized to evaluate the impact velocity and energy conditions that caused fracture in alumina structural ceramics. Drop tower testing was used for low velocity impact with a high mass indenter and fragment simulating projectile testing was used for high velocity impact with a low mass projectile. The damaged samples were nondestructively evaluated using digital radiography and ultrasound C-scan imaging. The bulk damage detected by these techniques was compared to surface damage observed by visual inspection.

  4. E2F-7 couples DNA damage-dependent transcription with the DNA repair process.

    PubMed

    Zalmas, Lykourgos-Panagiotis; Coutts, Amanda S; Helleday, Thomas; La Thangue, Nicholas B

    2013-09-15

    The cellular response to DNA damage, mediated by the DNA repair process, is essential in maintaining the integrity and stability of the genome. E2F-7 is an atypical member of the E2F family with a role in negatively regulating transcription and cell cycle progression under DNA damage. Surprisingly, we found that E2F-7 makes a transcription-independent contribution to the DNA repair process, which involves E2F-7 locating to and binding damaged DNA. Further, E2F-7 recruits CtBP and HDAC to the damaged DNA, altering the local chromatin environment of the DNA lesion. Importantly, the E2F-7 gene is a target for somatic mutation in human cancer and tumor-derived mutant alleles encode proteins with compromised transcription and DNA repair properties. Our results establish that E2F-7 participates in 2 closely linked processes, allowing it to directly couple the expression of genes involved in the DNA damage response with the DNA repair machinery, which has relevance in human malignancy.

  5. Fibrinogen-induced perivascular microglial clustering is required for the development of axonal damage in neuroinflammation

    PubMed Central

    Davalos, Dimitrios; Kyu Ryu, Jae; Merlini, Mario; Baeten, Kim M.; Le Moan, Natacha; Petersen, Mark A.; Deerinck, Thomas J.; Smirnoff, Dimitri S.; Bedard, Catherine; Hakozaki, Hiroyuki; Gonias Murray, Sara; Ling, Jennie B.; Lassmann, Hans; Degen, Jay L.; Ellisman, Mark H.; Akassoglou, Katerina

    2012-01-01

    Blood-brain barrier disruption, microglial activation and neurodegeneration are hallmarks of multiple sclerosis. However, the initial triggers that activate innate immune responses and their role in axonal damage remain unknown. Here we show that the blood protein fibrinogen induces rapid microglial responses toward the vasculature and is required for axonal damage in neuroinflammation. Using in vivo two-photon microscopy, we demonstrate that microglia form perivascular clusters before myelin loss or paralysis onset and that, of the plasma proteins, fibrinogen specifically induces rapid and sustained microglial responses in vivo. Fibrinogen leakage correlates with areas of axonal damage and induces reactive oxygen species release in microglia. Blocking fibrin formation with anticoagulant treatment or genetically eliminating the fibrinogen binding motif recognized by the microglial integrin receptor CD11b/CD18 inhibits perivascular microglial clustering and axonal damage. Thus, early and progressive perivascular microglial clustering triggered by fibrinogen leakage upon blood-brain barrier disruption contributes to axonal damage in neuroinflammatory disease. PMID:23187627

  6. SUMOylation of xeroderma pigmentosum group C protein regulates DNA damage recognition during nucleotide excision repair

    PubMed Central

    Akita, Masaki; Tak, Yon-Soo; Shimura, Tsutomu; Matsumoto, Syota; Okuda-Shimizu, Yuki; Shimizu, Yuichiro; Nishi, Ryotaro; Saitoh, Hisato; Iwai, Shigenori; Mori, Toshio; Ikura, Tsuyoshi; Sakai, Wataru; Hanaoka, Fumio; Sugasawa, Kaoru

    2015-01-01

    The xeroderma pigmentosum group C (XPC) protein complex is a key factor that detects DNA damage and initiates nucleotide excision repair (NER) in mammalian cells. Although biochemical and structural studies have elucidated the interaction of XPC with damaged DNA, the mechanism of its regulation in vivo remains to be understood in more details. Here, we show that the XPC protein undergoes modification by small ubiquitin-related modifier (SUMO) proteins and the lack of this modification compromises the repair of UV-induced DNA photolesions. In the absence of SUMOylation, XPC is normally recruited to the sites with photolesions, but then immobilized profoundly by the UV-damaged DNA-binding protein (UV-DDB) complex. Since the absence of UV-DDB alleviates the NER defect caused by impaired SUMOylation of XPC, we propose that this modification is critical for functional interactions of XPC with UV-DDB, which facilitate the efficient damage handover between the two damage recognition factors and subsequent initiation of NER. PMID:26042670

  7. Posttranscriptional regulation of gene expression-adding another layer of complexity to the DNA damage response.

    PubMed

    Boucas, Jorge; Riabinska, Arina; Jokic, Mladen; Herter-Sprie, Grit S; Chen, Shuhua; Höpker, Katja; Reinhardt, H Christian

    2012-01-01

    In response to DNA damage, cells activate a complex, kinase-based signaling network to arrest the cell cycle and allow time for DNA repair, or, if the extend of damage is beyond repair capacity, induce apoptosis. This signaling network, which is collectively referred to as the DNA damage response (DDR), is primarily thought to consist of two components-a rapid phosphorylation-driven signaling cascade that results in immediate inhibition of Cdk/cyclin complexes and a delayed transcriptional response that promotes a prolonged cell cycle arrest through the induction of Cdk inhibitors, such as p21. In recent years a third layer of complexity has emerged that involves potent posttranscriptional regulatory mechanisms that control the cellular response to DNA damage. Although much has been written on the relevance of the DDR in cancer and on the post-transcriptional role of microRNAs (miRs) in cancer, the post-transcriptional regulation of the DDR by non-coding RNAs and RNA-binding proteins (RBPs) still remains elusive in large parts. Here, we review the recent developments in this exciting new area of research in the cellular response to genotoxic stress. We put specific emphasis on the role of RBPs and the control of their function through DNA damage-activated protein kinases.

  8. Combating wind erosion of sandy soils and crop damage in the coastal deserts: Wind tunnel experiments

    NASA Astrophysics Data System (ADS)

    Genis, Arthur; Vulfson, Leonid; Ben-Asher, Jiftah

    2013-06-01

    In the western Negev desert of Israel frequent sandstorms cause heavy damage to young lettuce, carrot, peanut and potato plants during the planting season. The damage of plants is based mainly on the mechanical impact of saltating sand particles, which causes irreversible injuries to the plant leaves. Current agro-technique measures taken to prevent wind damage to crop in Israel are based on high frequency irrigation. Although the high-frequency irrigation helps bind soil particles together by forming a soil crust, it is associated with the large waste of water, which is not practical under the arid conditions. Application of polyacrylamide (PAM) as a chemical stabilizer has proved to be effective for prevention of soil erosion, saving irrigation water and a stable growth of plants in the early stages. Although the technique of PAM application is not yet used commercially in Israel, the preliminary studies suggested that it might have the potential to reduce the damage to the plant leaves by sandstorms, providing both environmental and agricultural benefits. In this study the effectiveness of PAM for preventing sandstorms in the western Negev was also investigated. Optimal concentration and volume of PAM solution per hectare of bare sandy soil were determined. For this purpose a wind tunnel was used to determine wind velocities of the first and continuous detachment of particles. The ability of PAM application to minimize the damage of plants by sandstorms was experimentally verified using image analysis tools.

  9. Oleandrin induces DNA damage responses in cancer cells by suppressing the expression of Rad51

    PubMed Central

    Bao, Zhengqiang; Tian, Baoping; Wang, Xiaohui; Feng, Hanrong; Liang, Ye; Chen, Zhihua; Li, Wen; Shen, Huahao; Ying, Songmin

    2016-01-01

    Oleandrin is a monomeric compound extracted from leaves and seeds of Nerium oleander. It had been reported that oleandrin could effectively inhibit the growth of human cancer cells. However, the specific mechanisms of the oleandrin-induced anti-tumor effects remain largely unclear. Genomic instability is one of the main features of cancer cells, it can be the combined effect of DNA damage and tumour-specific DNA repair defects. DNA damage plays important roles during tumorigenesis. In fact, most of the current chemotherapy agents were designed to kill cancer cells by inducing DNA damage. In this study, we found that oleandrin was effective to induce apoptosis in cancer cells, and cause rapid DNA damage response, represented by nuclear RPA (Replication Protein A, a single strand DNA binding protein) and γH2AX(a marker for DNA double strand breaks) foci formation. Interestingly, expression of RAD51, a key protein involved in homologous recombination (HR), was suppressed while XRCC1 was up-regulated in oleandrin treated cancer cells. These results suggested that XRCC1 may play a predominant role in repairing oleandrin-induced DNA damage. Collectively, oleandrin may be a potential anti-tumor agent by suppressing the expression of Rad51. PMID:27449097

  10. Ku70 Serine 155 mediates Aurora B inhibition and activation of the DNA damage response

    PubMed Central

    Fell, Victoria L.; Walden, Elizabeth A.; Hoffer, Sarah M.; Rogers, Stephanie R.; Aitken, Amelia S.; Salemi, Louisa M.; Schild-Poulter, Caroline

    2016-01-01

    The Ku heterodimer (Ku70/Ku80) is the central DNA binding component of the classical non-homologous end joining (NHEJ) pathway that repairs DNA double-stranded breaks (DSBs), serving as the scaffold for the formation of the NHEJ complex. Here we show that Ku70 is phosphorylated on Serine 155 in response to DNA damage. Expression of Ku70 bearing a S155 phosphomimetic substitution (Ku70 S155D) in Ku70-deficient mouse embryonic fibroblasts (MEFs) triggered cell cycle arrest at multiple checkpoints and altered expression of several cell cycle regulators in absence of DNA damage. Cells expressing Ku70 S155D exhibited a constitutive DNA damage response, including ATM activation, H2AX phosphorylation and 53BP1 foci formation. Ku70 S155D was found to interact with Aurora B and to have an inhibitory effect on Aurora B kinase activity. Lastly, we demonstrate that Ku and Aurora B interact following ionizing radiation treatment and that Aurora B inhibition in response to DNA damage is dependent upon Ku70 S155 phosphorylation. This uncovers a new pathway where Ku may relay signaling to Aurora B to enforce cell cycle arrest in response to DNA damage. PMID:27849008

  11. Hydrogen sulfide induces oxidative damage to RNA and DNA in a sulfide-tolerant marine invertebrate.

    PubMed

    Joyner-Matos, Joanna; Predmore, Benjamin L; Stein, Jenny R; Leeuwenburgh, Christiaan; Julian, David

    2010-01-01

    Hydrogen sulfide acts as an environmental toxin across a range of concentrations and as a cellular signaling molecule at very low concentrations. Despite its toxicity, many animals, including the mudflat polychaete Glycera dibranchiata, are periodically or continuously exposed to sulfide in their environment. We tested the hypothesis that a broad range of ecologically relevant sulfide concentrations induces oxidative stress and oxidative damage to RNA and DNA in G. dibranchiata. Coelomocytes exposed in vitro to sulfide (0-3 mmol L(-1) for 1 h) showed dose-dependent increases in oxidative stress (as 2',7'-dichlorofluorescein fluorescence) and superoxide production (as dihydroethidine fluorescence). Coelomocytes exposed in vitro to sulfide (up to 0.73 mmol L(-1) for 2 h) also acquired increased oxidative damage to RNA (detected as 8-oxo-7,8-dihydroguanosine) and DNA (detected as 8-oxo-7,8-dihydro-2'-deoxyguanosine). Worms exposed in vivo to sulfide (0-10 mmol L(-1) for 24 h) acquired elevated oxidative damage to RNA and DNA in both coelomocytes and body wall tissue. While the consequences of RNA and DNA oxidative damage are poorly understood, oxidatively damaged deoxyguanosine bases preferentially bind thymine, causing G-T transversions and potentially causing heritable point mutations. This suggests that sulfide can be an environmental mutagen in sulfide-tolerant invertebrates.

  12. The effect of Pot1 binding on the repair of thymine analogs in a telomeric DNA sequence.

    PubMed

    Theruvathu, Jacob A; Darwanto, Agus; Hsu, Chia Wei; Sowers, Lawrence C

    2014-08-01

    Telomeric DNA can form duplex regions or single-stranded loops that bind multiple proteins, preventing it from being processed as a DNA repair intermediate. The bases within these regions are susceptible to damage; however, mechanisms for the repair of telomere damage are as yet poorly understood. We have examined the effect of three thymine (T) analogs including uracil (U), 5-fluorouracil (5FU) and 5-hydroxymethyluracil (5hmU) on DNA-protein interactions and DNA repair within the GGTTAC telomeric sequence. The replacement of T with U or 5FU interferes with Pot1 (Pot1pN protein of Schizosaccharomyces pombe) binding. Surprisingly, 5hmU substitution only modestly diminishes Pot1 binding suggesting that hydrophobicity of the T-methyl group likely plays a minor role in protein binding. In the GGTTAC sequence, all three analogs can be cleaved by DNA glycosylases; however, glycosylase activity is blocked if Pot1 binds. An abasic site at the G or T positions is cleaved by the endonuclease APE1 when in a duplex but not when single-stranded. Abasic site formation thermally destabilizes the duplex that could push a damaged DNA segment into a single-stranded loop. The inability to enzymatically cleave abasic sites in single-stranded telomere regions would block completion of the base excision repair cycle potentially causing telomere attrition.

  13. Modeling laser damage to the retina

    NASA Astrophysics Data System (ADS)

    Clark, Clifton D.

    This dissertation presents recent progress in several areas related to modeling laser damage to the retina. In Chapter 3, we consider the consequences of using the Arrhenius damage model to predict the damage thresholds of multiple pulse, or repetitive pulse, exposures. We have identified a few fundamental trends associated with the multiple pulse damage predictions made by the Arrhenius model. These trends differ from what would be expected by non-thermal mechanisms, and could prove useful in differentiating thermal and non-thermal damage. Chapter 4 presents a new rate equation damage model hypothesized to describe photochemical damage. The model adds a temperature dependent term to the simple rate equation implied by the principle of reciprocity that is characteristic of photochemical damage thresholds. A recent damage threshold study, conducted in-vitro, has revealed a very sharp transition between thermal and photochemical damage threshold trends. For the wavelength used in the experiment (413 nm), thermal damage thresholds were observed at exposure levels that were twice the expected photochemical damage threshold, based on the traditional understanding of photochemical damage. Our model accounts for this observed trend by introducing a temperature dependent quenching, or repair, rate to the photochemical damage rate. For long exposures that give a very small temperature rise, the model reduces to the principle of reciprocity. Near the transition region between thermal and photochemical damage, the model allows the damage threshold to be set by thermal mechanisms, even at exposure above the reciprocity exposure. In Chapter 5, we describe a retina damage model that includes thermal lensing in the eye by coupling beam propagation and heat transfer models together. Thermal lensing has recently been suggested as a contributing factor to the large increase in measured retinal damage thresholds in the near infrared. The transmission of the vitreous decreases

  14. The Verrucomicrobia LexA-Binding Motif: Insights into the Evolutionary Dynamics of the SOS Response

    PubMed Central

    Erill, Ivan; Campoy, Susana; Kılıç, Sefa; Barbé, Jordi

    2016-01-01

    The SOS response is the primary bacterial mechanism to address DNA damage, coordinating multiple cellular processes that include DNA repair, cell division, and translesion synthesis. In contrast to other regulatory systems, the composition of the SOS genetic network and the binding motif of its transcriptional repressor, LexA, have been shown to vary greatly across bacterial clades, making it an ideal system to study the co-evolution of transcription factors and their regulons. Leveraging comparative genomics approaches and prior knowledge on the core SOS regulon, here we define the binding motif of the Verrucomicrobia, a recently described phylum of emerging interest due to its association with eukaryotic hosts. Site directed mutagenesis of the Verrucomicrobium spinosum recA promoter confirms that LexA binds a 14 bp palindromic motif with consensus sequence TGTTC-N4-GAACA. Computational analyses suggest that recognition of this novel motif is determined primarily by changes in base-contacting residues of the third alpha helix of the LexA helix-turn-helix DNA binding motif. In conjunction with comparative genomics analysis of the LexA regulon in the Verrucomicrobia phylum, electrophoretic shift assays reveal that LexA binds to operators in the promoter region of DNA repair genes and a mutagenesis cassette in this organism, and identify previously unreported components of the SOS response. The identification of tandem LexA-binding sites generating instances of other LexA-binding motifs in the lexA gene promoter of Verrucomicrobia species leads us to postulate a novel mechanism for LexA-binding motif evolution. This model, based on gene duplication, successfully addresses outstanding questions in the intricate co-evolution of the LexA protein, its binding motif and the regulatory network it controls. PMID:27489856

  15. Nonphysiological binding of ethylene by plants.

    PubMed

    Abeles, F B

    1984-03-01

    Ethylene binding to seedling tissue of Vicia faba, Phaseolus vulgaris, Glycine max, and Triticum aestivum was demonstrated by determining transit time required for ethylene to move through a glass tube filled with seedling tissue. Transit time for ethylene was greater than that for methane indicating that these tissues had an affinity for ethylene. However, the following observations suggest that the binding was not physiological. Inhibitors of ethylene action such as Ag(+) ions and CO(2) did not decrease binding. Mushrooms which have no known sites of ethylene action also demonstrated ethylene binding. The binding of acetylene, propylene, ethylene, propane, and ethane more closely followed their solubility in water than any known physiological activity.

  16. Synthetic heparin-binding factor analogs

    DOEpatents

    Pena, Louis A.; Zamora, Paul O.; Lin, Xinhua; Glass, John D.

    2010-04-20

    The invention provides synthetic heparin-binding growth factor analogs having at least one peptide chain, and preferably two peptide chains branched from a dipeptide branch moiety composed of two trifunctional amino acid residues, which peptide chain or chains bind a heparin-binding growth factor receptor and are covalently bound to a non-signaling peptide that includes a heparin-binding domain, preferably by a linker, which may be a hydrophobic linker. The synthetic heparin-binding growth factor analogs are useful as pharmaceutical agents, soluble biologics or as surface coatings for medical devices.

  17. Characterization of a DNA-damage-recognition protein from F9 teratocarcinoma cells, which is inducible by retinoic acid and cyclic AMP.

    PubMed

    Chao, C C; Sun, N K; Lin-Chao, S

    1993-02-15

    A nuclear protein that recognizes u.v.-damaged DNA was detected in extracts from murine F9 embryonic stem cells using a DNA-binding assay. The nuclear-protein-binding activity was increased in cells after treatment with retinoic acid/dibutyryl cyclic AMP (dbcAMP), with optimum induction at 6 days. In vitro treatment of nuclear extracts with agents that affect protein conformation (such as urea, Nonidet P40 and Ca2+) slightly modulated the damage-recognition activity. Furthermore, treatment of nuclear extracts with phosphatase dramatically inhibited the binding activity. In addition, damaged-DNA recognition of the nuclear extracts was effectively inhibited by damaged double- and single-stranded DNA. The expression of the nuclear protein with similar characteristics was abundant in HeLa cells and was increased in drug- or u.v.-resistant cells. The findings suggest that the recognition of a u.v.-DNA adduct is modulated, at least in part, by an activity that is induced during retinoic acid/dbcAMP-induced differentiation. These results also imply that the identified damage-recognition protein may be important for the sensitivity or resistance of mammalian cells to DNA damage.

  18. 7 CFR 51.586 - Serious damage.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ..., CERTIFICATION, AND STANDARDS) United States Standards for Celery Definitions § 51.586 Serious damage. Serious... the appearance, or the edible or shipping quality of the celery stalk or the general appearance of...

  19. Thermodynamical Framework for Ductile Damage and Plasticity

    NASA Astrophysics Data System (ADS)

    César de Sá, José M. A.; Andrade, Filipe X. C.; Andrade Pires, Francisco M.

    2010-05-01

    Many models employed for the prediction of plastic deformation rely exclusively on elastoplastic theories, disregarding significant effects of internal degradation [1]. Constitutive models based on the Continuum Damage Mechanics theory provide more realistic predictions since damage is taken into account as an internal variable. In the present contribution, Lemaire's model for ductile damage [2] is questioned under the assumption of the principle of maximum inelastic dissipation [3]. The model is enhanced with a nonlocal formulation where the damage variable is spatially averaged by means of an integral operator [4]. Thermodynamical admissibility of the nonlocal model is checked by applying the global version of the Clausius-Duhem inequality [5]. Results from numerical analysis show that the constitutive model is insensitive to spatial discretization.

  20. 7 CFR 29.2263 - Damage.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    .... The effect of mold, must, rot, black rot, or other fungous or bacterial diseases which attack tobacco in its cured state. Tobacco having the odor of mold, must, or rot is considered damaged. (See Rule...

  1. 7 CFR 29.6011 - Damage.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... INSPECTION Standards Definitions § 29.6011 Damage. The effect of mold, must, rot, black rot or other fungus or bacterial diseases which attack tobacco in its cured state. Tobacco having the odor of mold,...

  2. 7 CFR 29.3514 - Damage.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... Type 95) § 29.3514 Damage. The effect of mold, must, rot, black rot, or other fungus or bacterial diseases which attack tobacco in its cured state. Tobacco having the odor of mold, must, or rot...

  3. 7 CFR 29.6011 - Damage.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... INSPECTION Standards Definitions § 29.6011 Damage. The effect of mold, must, rot, black rot or other fungus or bacterial diseases which attack tobacco in its cured state. Tobacco having the odor of mold,...

  4. 7 CFR 29.1012 - Damage.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... Type 92) § 29.1012 Damage. The effect of mold, must, rot, black rot, or other fungus or bacterial diseases which attack tobacco in its cured state. Tobacco having the odor of mold, must, or rot...

  5. 7 CFR 29.2263 - Damage.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    .... The effect of mold, must, rot, black rot, or other fungous or bacterial diseases which attack tobacco in its cured state. Tobacco having the odor of mold, must, or rot is considered damaged. (See Rule...

  6. 7 CFR 30.17 - Damage.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... Damage. The effect of mold, must, rot, black rot, or other fungous or bacterial diseases which attack tobacco in its cured state. Any tobacco having the odor of mold, must, or rot shall be included in...

  7. 7 CFR 29.3514 - Damage.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... Type 95) § 29.3514 Damage. The effect of mold, must, rot, black rot, or other fungus or bacterial diseases which attack tobacco in its cured state. Tobacco having the odor of mold, must, or rot...

  8. 7 CFR 30.17 - Damage.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... Damage. The effect of mold, must, rot, black rot, or other fungous or bacterial diseases which attack tobacco in its cured state. Any tobacco having the odor of mold, must, or rot shall be included in...

  9. 7 CFR 29.1012 - Damage.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... Type 92) § 29.1012 Damage. The effect of mold, must, rot, black rot, or other fungus or bacterial diseases which attack tobacco in its cured state. Tobacco having the odor of mold, must, or rot...

  10. Brain Training for Cancer Survivors' Nerve Damage

    MedlinePlus

    ... html Brain Training for Cancer Survivors' Nerve Damage Neurofeedback seems to offers relief from chemo-induced pain, ... brain waves with a type of training called neurofeedback seems to help cancer survivors ease symptoms of ...

  11. Radiation damage due to electromagnetic showers

    SciTech Connect

    Rakhno, Igor; Mokhov, Nikolai; Striganov, Sergei; /Fermilab

    2008-05-01

    Radiation-induced damage due to atomic displacements is essential to correctly predict the behavior of materials in nuclear reactors and at charged-particle accelerators. Traditionally the damage due to hadrons was of major interest. The recent increased interest in high-energy lepton colliders gave rise to the problem of prediction of radiation damage due to electromagnetic showers in a wide energy range--from a few hundred keV and up to a few hundred GeV. The report describes results of an electron- and positron-induced displacement cross section evaluation. It is based on detailed lepton-nucleus cross sections, realistic nuclear form-factors and a modified Kinchin-Pease damage model. Numerical data on displacement cross sections for various target nuclei is presented.

  12. 7 CFR 51.908 - Serious damage.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... Standards for Grades of Table Grapes (European or Vinifera Type) 1 Definitions § 51.908 Serious damage..., affected by decay or waterberry, or affected by heat or freezing. Grapes which show healed cracks at...

  13. 7 CFR 51.3157 - Damage.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ..., CERTIFICATION, AND STANDARDS) United States Standards for Grades of Nectarines Definitions § 51.3157 Damage... stem cavity; or, (5) When more than one-fourth inch in length if outside of the stem cavity; (b)...

  14. Radiation damage of gallium arsenide production cells

    NASA Technical Reports Server (NTRS)

    Mardesich, N.; Garlick, G. F. J.

    1987-01-01

    High-efficiency gallium arsenide cells, made by the liquid epitaxy method (LPE), have been irradiated with 1-MeV electrons up to fluences of 10 to the 16th e/sq cm. Measurements have been made of cell spectral response and dark and light-excited current-voltage characteristics and analyzed using computer-based models to determine underlying parameters such as damage coefficients. It is possible to use spectral response to sort out damage effects in the different cell component layers. Damage coefficients are similar to other reported in the literature for the emitter and buffer (base). However, there is also a damage effect in the window layer and possibly at the window emitter interface similar to that found for proton-irradiated liquid-phase epitaxy-grown cells. Depletion layer recombination is found to be less than theoretically expected at high fluence.

  15. Gallium arsenide solar cell radiation damage study

    NASA Technical Reports Server (NTRS)

    Maurer, R. H.; Herbert, G. A.; Kinnison, J. D.; Meulenberg, A.

    1989-01-01

    A thorough analysis has been made of electron- and proton- damaged GaAs solar cells suitable for use in space. It is found that, although some electrical parametric data and spectral response data are quite similar, the type of damage due to the two types of radiation is different. An I-V analysis model shows that electrons damage the bulk of the cell and its currents relatively more, while protons damage the junction of the cell and its voltages more. It is suggested that multiple defects due to protons in a strong field region such as a p/n junction cause the greater degradation in cell voltage, whereas the individual point defects in the quasi-neutral minority-carrier-diffusion regions due to electrons cause the greater degradation in cell current and spectral response.

  16. 7 CFR 51.2960 - Damage.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... of the rest of the shell or the majority of shells in the lot, or darker discoloration covering a... considered as damaged; and, (g) Insects when an insect or insect fragment, web or frass is present inside...

  17. Atomic oxygen damage characterization by photothermal scanning

    NASA Technical Reports Server (NTRS)

    Williams, A. W.; Wood, N. J.; Zakaria, A. B.

    1993-01-01

    In this paper we use a photothermal imaging technique to characterize the damage caused to an imperfectly coated gold-coated Kapton sample exposed to successively increased fluences of atomic oxygen in a laboratory atomic source.

  18. Structural damage identification using mathematical optimization techniques

    NASA Technical Reports Server (NTRS)

    Shen, Mo-How Herman

    1991-01-01

    An identification procedure is proposed to identify damage characteristics (location and size of the damage) from dynamic measurements. This procedure was based on minimization of the mean-square measure of difference between measurement data (natural frequencies and mode shapes) and the corresponding predictions obtained from the computational model. The procedure is tested for simulated damage in the form of stiffness changes in a simple fixed free spring mass system and symmetric cracks in a simply supported Bernoulli Euler beam. It is shown that when all the mode information is used in the identification procedure it is possible to uniquely determine the damage properties. Without knowing the complete set of modal information, a restricted region in the initial data space has been found for realistic and convergent solution from the identification process.

  19. 7 CFR 29.3514 - Damage.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... Type 95) § 29.3514 Damage. The effect of mold, must, rot, black rot, or other fungus or bacterial diseases which attack tobacco in its cured state. Tobacco having the odor of mold, must, or rot...

  20. 7 CFR 30.17 - Damage.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... Damage. The effect of mold, must, rot, black rot, or other fungous or bacterial diseases which attack tobacco in its cured state. Any tobacco having the odor of mold, must, or rot shall be included in...