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Sample records for t4 lysozyme vectorially

  1. An improved 96-well turbidity assay for T4 lysozyme activity

    Science.gov (United States)

    2015-05-13

    Additional information T4L is an enzyme found in T4 phage that is tolerant of mutations and amenable to crystallization. Extensive mutagenesis and...pnas.0912009106. [16] W.A. Baase, L. Liu, D.E. Tronrud, B.W. Matthews, Lessons from the lysozyme of phage T4 , Protein Sci. 19 (2010) 631–641. [17] D... phage T4 , Cold Spring Harb. Symp. Quant. Biol. 26 (1961) 25–30. [21] D.W. Heinz, W.A. Baase, B.W. Matthews, Folding and function of a T4 lysozyme

  2. Expression of T4 Lysozyme Gene (gene e) in Streptococcus ...

    African Journals Online (AJOL)

    GREGO

    2007-04-16

    Apr 16, 2007 ... cloning and inserting Streptococcal replication origin of pTRW10 vector into pL1. pL2 plasmid isolated from E. coli was introduced into S. salivarius subsp. thermophilus and Lactococcus lactis cells by electro-transformation. The lysozyme enzymes expressing by these bacteria were found to be active on.

  3. Expression of T4 Lysozyme Gene (gene e) in Streptococcus ...

    African Journals Online (AJOL)

    pL2 plasmid isolated from E. coli was introduced into S. salivarius subsp. thermophilus and Lactococcus lactis cells by electro-transformation. The lysozyme enzymes expressing by these bacteria were found to be active on Micrococcus luteus cells and thereby preventing their growth on assay plates. Thermostability of ...

  4. The Effects of the Recombinant CCR5 T4 Lysozyme Fusion Protein on HIV-1 Infection.

    Directory of Open Access Journals (Sweden)

    Qingwen Jin

    Full Text Available Insertion of T4 lysozyme (T4L into the GPCR successfully enhanced GPCR protein stability and solubilization. However, the biological functions of the recombinant GPCR protein have not been analyzed.We engineered the CCR5-T4L mutant and expressed and purified the soluble recombinant protein using an E.coli expression system. The antiviral effects of this recombinant protein in THP-1 cell lines, primary human macrophages, and PBMCs from different donors were investigated. We also explored the possible mechanisms underlying the observed antiviral effects.Our data showed the biphasic inhibitory and promotion effects of different concentrations of soluble recombinant CCR5-T4L protein on R5 tropic human immunodeficiency virus-1 (HIV-1 infection in THP-1 cell lines, human macrophages, and PBMCs from clinical isolates. We demonstrated that soluble recombinant CCR5-T4L acts as a HIV-1 co-receptor, interacts with wild type CCR5, down-regulates the surface CCR5 expression in human macrophages, and interacts with CCL5 to inhibit macrophage migration. Using binding assays, we further determined that recombinant CCR5-T4L and [125I]-CCL5 compete for the same binding site on wild type CCR5.Our results suggest that recombinant CCR5-T4L protein marginally promotes HIV-1 infection at low concentrations and markedly inhibits infection at higher concentrations. This recombinant protein may be helpful in the future development of anti-HIV-1 therapeutic agents.

  5. The Effects of the Recombinant CCR5 T4 Lysozyme Fusion Protein on HIV-1 Infection.

    Science.gov (United States)

    Jin, Qingwen; Chen, Hong; Wang, Xingxia; Zhao, Liandong; Xu, Qingchen; Wang, Huijuan; Li, Guanyu; Yang, Xiaofan; Ma, Hongming; Wu, Haoquan; Ji, Xiaohui

    2015-01-01

    Insertion of T4 lysozyme (T4L) into the GPCR successfully enhanced GPCR protein stability and solubilization. However, the biological functions of the recombinant GPCR protein have not been analyzed. We engineered the CCR5-T4L mutant and expressed and purified the soluble recombinant protein using an E.coli expression system. The antiviral effects of this recombinant protein in THP-1 cell lines, primary human macrophages, and PBMCs from different donors were investigated. We also explored the possible mechanisms underlying the observed antiviral effects. Our data showed the biphasic inhibitory and promotion effects of different concentrations of soluble recombinant CCR5-T4L protein on R5 tropic human immunodeficiency virus-1 (HIV-1) infection in THP-1 cell lines, human macrophages, and PBMCs from clinical isolates. We demonstrated that soluble recombinant CCR5-T4L acts as a HIV-1 co-receptor, interacts with wild type CCR5, down-regulates the surface CCR5 expression in human macrophages, and interacts with CCL5 to inhibit macrophage migration. Using binding assays, we further determined that recombinant CCR5-T4L and [125I]-CCL5 compete for the same binding site on wild type CCR5. Our results suggest that recombinant CCR5-T4L protein marginally promotes HIV-1 infection at low concentrations and markedly inhibits infection at higher concentrations. This recombinant protein may be helpful in the future development of anti-HIV-1 therapeutic agents.

  6. Metal ions-binding T4 lysozyme as an intramolecular protein purification tag compatible with X-ray crystallography.

    Science.gov (United States)

    Boura, Evzen; Baumlova, Adriana; Chalupska, Dominika; Dubankova, Anna; Klima, Martin

    2017-06-01

    Phage T4 lysozyme is a well folded and highly soluble protein that is widely used as an insertion tag to improve solubility and crystallization properties of poorly behaved recombinant proteins. It has been used in the fusion protein strategy to facilitate crystallization of various proteins including multiple G protein-coupled receptors, lipid kinases, or sterol binding proteins. Here, we present a structural and biochemical characterization of its novel, metal ions-binding mutant (mbT4L). We demonstrate that mbT4L can be used as a purification tag in the immobilized-metal affinity chromatography and that, in many respects, it is superior to the conventional hexahistidine tag. In addition, structural characterization of mbT4L suggests that mbT4L can be used as a purification tag compatible with X-ray crystallography. © 2017 The Protein Society.

  7. Metal ions-binding T4 lysozyme as an intramolecular protein purification tag compatible with X-ray crystallography

    Czech Academy of Sciences Publication Activity Database

    Bouřa, Evžen; Bäumlová, Adriana; Chalupská, Dominika; Dubánková, Anna; Klíma, Martin

    2017-01-01

    Roč. 26, č. 6 (2017), s. 1116-1123 ISSN 0961-8368 R&D Projects: GA ČR(CZ) GJ17-07058Y; GA MŠk LO1302 Institutional support: RVO:61388963 Keywords : phage T4 * lysozyme * endolysin * histidine tag * protein purification * crystal structure Subject RIV: CE - Biochemistry OBOR OECD: Biochemistry and molecular biology Impact factor: 2.523, year: 2016

  8. Synthesis of 1,2-Azaborines and the Preparation of Their Protein Complexes with T4 Lysozyme Mutants.

    Science.gov (United States)

    Lee, Hyelee; Liu, Shih-Yuan

    2017-03-25

    We describe a general synthesis of 1,2-azaborines using standard air-free techniques and protein complex preparation with T4 lysozyme mutants by vapor diffusion. Oxygen- and moisture-sensitive compounds are prepared and isolated under an inert atmosphere (N2) using either a vacuum gas manifold or a glove box. As an example of azaborine synthesis, we demonstrate the synthesis and purification of the volatile N-H-B-ethyl-1,2-azaborine by a five-step sequence involving distillation and column chromatography for the isolation of products. T4 lysozyme mutants L99A and L99A/M102Q are expressed with Escherichia coli RR1 strain. Standard protocols for chemical cell lysis followed by purification using carboxymethyl ion exchange column affords protein of sufficiently high purity for crystallization. Protein crystallization is performed in various concentrations of precipitant at different pH ranges using the hanging drop vapor diffusion method. Complex preparation with the small molecules is carried out by vapor diffusion method under an inert atmosphere. X-ray diffraction analysis of the crystal complex provides unambiguous structural evidence of binding interactions between the protein binding site and 1,2-azaborines.

  9. Functional fusions of T4 lysozyme in the third intracellular loop of a G protein-coupled receptor identified by a random screening approach in yeast.

    Science.gov (United States)

    Mathew, Elizabeth; Ding, Fa-Xiang; Naider, Fred; Dumont, Mark E

    2013-01-01

    The insertion of a stable soluble protein into loops of transmembrane proteins has proved to be a successful approach for enhancing their stabilities and crystallization, and may also be useful in contexts where the inserted proteins can modulate or report on the activities of membrane proteins. While the use of T4 lysozyme to replace portions of the third intracellular loops of G protein-coupled receptors (GPCRs) has allowed determination of the structures of members of this important class of receptors, the creation of such fusion proteins generally leads to loss of signaling function of the resulting fusion protein, since the third intracellular loops of GPCRs play critical roles in their interactions with G proteins. We describe here a random screening approach allowing insertion of T4 lysozyme into diverse positions in the third loop of the yeast α-pheromone receptor, a GPCR encoded by the yeast STE2 gene. Insertions were accompanied by varying extents of deletion or duplication of the loop. A set of phenotypic screens allow detection of potentially rare variant receptors that are expressed, bind to agonist and are capable of signal transduction via activation of the cognate G protein. A large fraction of screened full-length receptor variants containing at least partial duplications of the loop on either side of the inserted T4 lysozyme retain the ability to activate the downstream signaling pathway in response to binding of ligand. However, we were unable to identify any receptors with truncated C-termini that retain significant signaling function in the presence of inserted T4 lysozyme. Our results establish the feasibility of creating functional receptors containing insertions of T4 lysozyme in their third intracellular loops.

  10. Interaction-component analysis of the effects of urea and its alkylated derivatives on the structure of T4-lysozyme

    Science.gov (United States)

    Yamamori, Yu; Matubayasi, Nobuyuki

    2017-06-01

    The effects of urea and its alkylated derivatives on the structure of T4-lysozyme were analyzed from the standpoint of energetics. Molecular dynamics simulations were conducted with explicit solvent, and the energy-representation method was employed to compute the free energy of transfer of the protein from pure-water solvent to the mixed solvents of water with urea, methylurea, 1,1-dimethylurea, and isopropylurea. Through the decomposition of the transfer free energy into the cosolvent and water contributions, it was observed that the former is partially cancelled by the latter and governs the total free energy of transfer. To determine the interaction component responsible for the transfer energetics, the correlations of the transfer free energy were also examined against the change in the solute-solvent interaction energy upon transfer and the corresponding changes in the electrostatic, van der Waals, and excluded-volume components. It was then found over the set of protein structures ranging from native to (partially) unfolded ones that the transfer free energy changes in parallel with the van der Waals component even when the cosolvent is alkylated. The electrostatic and excluded-volume components play minor roles in the structure modification of the protein, and the denaturing ability of alkylurea is brought by the van der Waals interaction.

  11. Dynamics of xenon binding inside the hydrophobic cavity of pseudo-wild-type bacteriophage T4 lysozyme explored through xenon-based NMR spectroscopy.

    Science.gov (United States)

    Desvaux, Hervé; Dubois, Lionel; Huber, Gaspard; Quillin, Michael L; Berthault, Patrick; Matthews, Brian W

    2005-08-24

    Wild-type bacteriophage T4 lysozyme contains a hydrophobic cavity with binding properties that have been extensively studied by X-ray crystallography and NMR. In the present study, the monitoring of 1H chemical shift variations under xenon pressure enables the determination of the noble gas binding constant (K = 60.2 M(-1)). Although the interaction site is highly localized, dipolar cross-relaxation effects between laser-polarized xenon and nearby protons (SPINOE) are rather poor. This is explained by the high value of the xenon-proton dipolar correlation time (0.8 ns), much longer than the previously reported values for xenon in medium-size proteins. This indicates that xenon is highly localized within the protein cavity, as confirmed by the large chemical shift difference between free and bound xenon. The exploitation of the xenon line width variation vs xenon pressure and protein concentration allows the extraction of the exchange correlation time between free and bound xenon. Comparison to the exchange experienced by protein protons indicates that the exchange between the open and closed conformations of T4 lysozyme is not required for xenon binding.

  12. Probing the folded state and mechanical unfolding pathways of T4 lysozyme using all-atom and coarse-grained molecular simulation

    Energy Technology Data Exchange (ETDEWEB)

    Zheng, Wenjun, E-mail: wjzheng@buffalo.edu; Glenn, Paul [Department of Physics, University at Buffalo, Buffalo, New York 14260 (United States)

    2015-01-21

    The Bacteriophage T4 Lysozyme (T4L) is a prototype modular protein comprised of an N-terminal and a C-domain domain, which was extensively studied to understand the folding/unfolding mechanism of modular proteins. To offer detailed structural and dynamic insights to the folded-state stability and the mechanical unfolding behaviors of T4L, we have performed extensive equilibrium and steered molecular dynamics simulations of both the wild-type (WT) and a circular permutation (CP) variant of T4L using all-atom and coarse-grained force fields. Our all-atom and coarse-grained simulations of the folded state have consistently found greater stability of the C-domain than the N-domain in isolation, which is in agreement with past thermostatic studies of T4L. While the all-atom simulation cannot fully explain the mechanical unfolding behaviors of the WT and the CP variant observed in an optical tweezers study, the coarse-grained simulations based on the Go model or a modified elastic network model (mENM) are in qualitative agreement with the experimental finding of greater unfolding cooperativity in the WT than the CP variant. Interestingly, the two coarse-grained models predict different structural mechanisms for the observed change in cooperativity between the WT and the CP variant—while the Go model predicts minor modification of the unfolding pathways by circular permutation (i.e., preserving the general order that the N-domain unfolds before the C-domain), the mENM predicts a dramatic change in unfolding pathways (e.g., different order of N/C-domain unfolding in the WT and the CP variant). Based on our simulations, we have analyzed the limitations of and the key differences between these models and offered testable predictions for future experiments to resolve the structural mechanism for cooperative folding/unfolding of T4L.

  13. Proton NMR measurements of bacteriophage T4 lysozyme aided by 15N isotopic labeling: structural and dynamic studies of larger proteins

    International Nuclear Information System (INIS)

    McIntosh, L.P.; Griffey, R.H.; Muchmore, D.C.; Nielson, C.P.; Redfield, A.G.; Dahlquist, F.W.

    1987-01-01

    A strategy for resolution and assignment of single proton resonances in proteins of molecular mass up to at least 40 kDa is presented. This approach is based on 15 N (or 13 C) labeling of selected residues in a protein. The resonances from protons directly bonded to labeled atoms are detected in a two-dimensional 1H- 15 N (or 13 C) spectrum. The nuclear Overhauser effects from isotopically tagged protons are selectively observed in one-dimensional isotope-directed measurements. Using this approach, we have observed approximately 160 resonances from 15 N-bonded protons in the backbone and sidechains of uniformly 15 N-labeled T4 lysozyme (molecular mass = 18.7 kDa). Partial proton-deuterium exchange can be used to simplify the 1H- 15 N spectrum of this protein. These resonances are identified by amino acid class using selective incorporation of 15 N-labeled amino acids and are assigned to specific residues by mutational substitution, multiple 15 N and 13 C labeling, and isotope-directed nuclear Overhauser effect measurements. For example, using a phenyl[ 15 N]alanine-labeled lysozyme variant containing two consecutive phenylalanine residues in an alpha-helical region, we observe an isotope-directed nuclear Overhauser effect from the amide proton of Phe-66 to that of Phe-67

  14. Off-resonance R1ρ relaxation outside of the fast exchange limit: An experimental study of a cavity mutant of T4 lysozyme

    International Nuclear Information System (INIS)

    Korzhnev, Dmitry M.; Orekhov, Vladislav Yu.; Dahlquist, Frederick W.; Kay, Lewis E.

    2003-01-01

    An 15 N off-resonance R 1ρ spin relaxation study of an L99A point mutant of T4 lysozyme is presented. Previous CPMG-based relaxation dispersion studies of exchange in this protein have established that the molecule interconverts between a populated ground state and an excited state (3.4%) with an exchange rate constant of 1450 s -1 at 25 deg. C. It is shown that for the majority of residues in this protein the offset dependence of the R 1ρ relaxation rates cannot be well fit using models which are only valid in the fast exchange regime. In contrast, a recently derived expression by Trott and Palmer (J. Magn. Reson., 154, 157-160, 2002) which is valid over a wider window of exchange than other relations, is shown to fit the data well. Values of (signed) chemical shift differences between exchanging sites have been extracted and are in reasonable agreement with shift differences measured using CPMG methods. A set of simulations is presented which help establish the exchange regimes that are best suited to analysis by off-resonance R 1ρ techniques

  15. Neutron structure of the T26H mutant of T4 phage lysozyme provides insight into the catalytic activity of the mutant enzyme and how it differs from that of wild type.

    Science.gov (United States)

    Hiromoto, Takeshi; Meilleur, Flora; Shimizu, Rumi; Shibazaki, Chie; Adachi, Motoyasu; Tamada, Taro; Kuroki, Ryota

    2017-10-01

    T4 phage lysozyme is an inverting glycoside hydrolase that degrades the murein of bacterial cell walls by cleaving the β-1,4-glycosidic bond. The substitution of the catalytic Thr26 residue to a histidine converts the wild type from an inverting to a retaining enzyme, which implies that the original general acid Glu11 can also act as an acid/base catalyst in the hydrolysis. Here, we have determined the neutron structure of the perdeuterated T26H mutant to clarify the protonation states of Glu11 and the substituted His26, which are key in the retaining reaction. The 2.09-Å resolution structure shows that the imidazole group of His26 is in its singly protonated form in the active site, suggesting that the deprotonated Nɛ2 atom of His26 can attack the anomeric carbon of bound substrate as a nucleophile. The carboxyl group of Glu11 is partially protonated and interacts with the unusual neutral state of the guanidine moiety of Arg145, as well as two heavy water molecules. Considering that one of the water-binding sites has the potential to be occupied by a hydronium ion, the bulk solvent could be the source for the protonation of Glu11. The respective protonation states of Glu11 and His26 are consistent with the bond lengths determined by an unrestrained refinement of the high-resolution X-ray structure of T26H at 1.04-Å resolution. The detail structural information, including the coordinates of the deuterium atoms in the active site, provides insight into the distinctively different catalytic activities of the mutant and wild type enzymes. © 2017 The Authors Protein Science published by Wiley Periodicals, Inc. on behalf of The Protein Society.

  16. Thyroxine (T4) Test

    Science.gov (United States)

    ... Too much or too little T4 can indicate thyroid disease . The T4 hormone comes in two forms: Free ... is used to evaluate thyroid function and diagnose thyroid disease. Why do I need a thyroxine test? Thyroid ...

  17. t4 Workshop Report

    Science.gov (United States)

    Silbergeld, Ellen K.; Contreras, Elizabeth Q.; Hartung, Thomas; Hirsch, Cordula; Hogberg, Helena; Jachak, Ashish C.; Jordan, William; Landsiedel, Robert; Morris, Jeffery; Patri, Anil; Pounds, Joel G.; de Vizcaya Ruiz, Andrea; Shvedova, Anna; Tanguay, Robert; Tatarazako, Norihasa; van Vliet, Erwin; Walker, Nigel J.; Wiesner, Mark; Wilcox, Neil; Zurlo, Joanne

    2014-01-01

    Summary In October 2010, a group of experts met as part of the transatlantic think tank for toxicology (t4) to exchange ideas about the current status and future of safety testing of nanomaterials. At present, there is no widely accepted path forward to assure appropriate and effective hazard identification for engineered nanomaterials. The group discussed needs for characterization of nanomaterials and identified testing protocols that incorporate the use of innovative alternative whole models such as zebrafish or C. elegans, as well as in vitro or alternative methods to examine specific functional pathways and modes of action. The group proposed elements of a potential testing scheme for nanomaterials that works towards an integrated testing strategy, incorporating the goals of the NRC report Toxicity Testing in the 21st Century: A Vision and a Strategy by focusing on pathways of toxic response, and utilizing an evidence-based strategy for developing the knowledge base for safety assessment. Finally, the group recommended that a reliable, open, curated database be developed that interfaces with existing databases to enable sharing of information. PMID:21993959

  18. A Vectorial Semantics Approach to Personality Assessment

    OpenAIRE

    Neuman, Yair; Cohen, Yochai

    2014-01-01

    Personality assessment and, specifically, the assessment of personality disorders have traditionally been indifferent to computational models. Computational personality is a new field that involves the automatic classification of individuals' personality traits that can be compared against gold-standard labels. In this context, we introduce a new vectorial semantics approach to personality assessment, which involves the construction of vectors representing personality dimensions and disorders...

  19. Crystallization of Hevamine, an Enzyme with Lysozyme/Chitinase Activity from Hevea brasiliensis Latex

    NARCIS (Netherlands)

    ROZEBOOM, HJ; BUDIANI, A; BEINTEMA, JJ

    1990-01-01

    Hevamine, an enzyme with both lysozyme and chitinase activity, was isolated and purified from Hevea brasiliensis (rubber tree) latex. The enzyme (molecular weight 29,000) is homologous to certain “pathogenesis-related” proteins from plants, but not to hen egg-white or phage T4 lysozyme. To

  20. Colorectal adenomas produce lysozyme.

    Science.gov (United States)

    Rubio, C A

    2003-01-01

    Lysozyme is an innate non-immunologic antibacterial enzyme produced by the Paneth cells of the upper intestinal tract. Lysozyme is not normally secreted in the lower intestinal tract. Previous reports indicate, however, that lysozyme may be secreted by colorectal neoplasias. The aim was to audit lysozyme expression in colorectal diseases including neoplasias. For that purpose, sections were stained with lysozyme (Muramidase), Ki67 (MIB1) and CD 68. Intense lysozyme overexpression (+++) was compared among 177 colorectal tissues: 35 having normal mucosa, 20 regenerative mucosa in inflammatory bowel disease (IBD), 2 inflammatory polyps, 3 collagenous colitis, 2 melanosis coli, 21 hyperplastic polyps, 42 tubular adenomas, 9 serrated adenomas, 30 villous adenomas and 13 invasive carcinomas. Intense lysozyme overexpression (+++) was found in 9.5% of the hyperplastic polyps, in 97.6% of the tubular adenomas, in 88.9% of the serrated adenomas, in 93.3% of the villous adenomas, in 76.9% of the carcinomas, but in none of the other tissues investigated. Neoplastic colorectal cells may acquire the capacity to produce lysozyme. The presence of that enzyme may not be a haphazard, capricious event in mutated colorectal epithelial cells but part of a more elaborate molecular behavior, not necessarily antibacterial. Recently, it was demonstrated that patients having lysozyme-secreting breast carcinomas were associated with a favorable prognosis. Whether lysozyme expression has any bearing on the biological behavior of colorectal carcinomas remains to be elucidated. Lysozyme overexpression (+++) also occurred in 2 of the 21 hyperplastic polyps, suggesting that intense lysozyme production might herald a possible dysplastic evolution in some hyperplastic polyps.

  1. Vectorial optical fields fundamentals and applications

    CERN Document Server

    2014-01-01

    Polarization is a vector nature of light that plays an important role in optical science and engineering. While existing textbook treatments of light assume beams with spatially homogeneous polarization, there is an increasing interest in vectorial optical fields with spatially engineered states of polarization. New effects and phenomena have been predicted and observed for light beams with these unconventional polarization states. This edited review volume aims to provide a comprehensive overview and summarize the latest developments in this important emerging field of optics. This book will cover the fundamentals including mathematical and physical descriptions, experimental generation, manipulation, focusing, propagation, and the applications of the engineered vectorial optical fields in focal field engineering, plasmonic focusing and optical antenna, single molecular imaging, optical tweezers/trapping, as well as optical measurements and instrumentations. Readership: Students, professionals, post-graduat...

  2. 216-T-4 interim stabilization final report

    International Nuclear Information System (INIS)

    Smith, D.L.

    1996-01-01

    This report provides a general description of the activities performed for the interim stabilization of the 216-T-4-1 ditch, 216-T-4-2 ditch, and 216-T-4-2 pond. Interim stabilization was required to reduce the amount of surface-contaminated acres and to minimize the migration of radioactive contamination. Work associated with the 216-T4-1 ditch and 216-T-4-2 pond was performed by the Radiation Area Remedial Action (RARA) Project. Work associated with the 216-T-4-2 ditch was done concurrently but was funded by Westinghouse Hanford Company (WHC) Tank Waste Remediation Systems (TWRS)

  3. Evidence of vectorial photoelectric effect on Copper

    International Nuclear Information System (INIS)

    Pedersoli, E.; Banfi, F.; Ressel, B.; Pagliara, S.; Giannetti, C.; Galimberti, G.; Lidia, S.; Corlett, J.; Ferrini, G.; Parmigiani, F.

    2005-01-01

    Quantum efficiency (QE) measurements of single photon photoemission from a Cu(111) single crystal and a Cu polycrystal photocathodes, irradiated by 150 fs-6.28 eV laser pulses, are reported over a broad range of incidence angle, both in s and p polarizations. The maximum QE (≅4x10 -4 ) for polycrystalline Cu is obtained in p polarization at an angle of incidence θ=65 deg. . We observe a QE enhancement in p polarization which cannot be explained in terms of optical absorption, a phenomenon known as vectorial photoelectric effect. Issues concerning surface roughness and symmetry considerations are addressed. An explanation in terms of nonlocal conductivity tensor is proposed

  4. Gamma-T4 hybrid bacteriophage carrying the thymidine kinase gene of bacteriophage T4.

    OpenAIRE

    Mileham, A J; Murray, N E; Revel, H R

    1984-01-01

    Among 32 lambda-T4 recombinant phages selected for growth on a thymidylate synthetase-deficient (thyA) host, 2 were shown to carry the T4 thymidine kinase (tk) gene. The lambda-T4tk phages contain two T4 HindIII DNA fragments (2.0 and 1.5 kilobases) that hybridize to restriction fragments of T4 DNA, encompassing the tk locus at 60 kilobases on the T4 map. The T4tk insert compensates for the simultaneous host deficiencies of thymidine kinase and thymidylate synthetase in a thymidine kinase-def...

  5. Transcription of the T4 late genes

    OpenAIRE

    Geiduschek, E Peter; Kassavetis, George A

    2010-01-01

    Abstract This article reviews the current state of understanding of the regulated transcription of the bacteriophage T4 late genes, with a focus on the underlying biochemical mechanisms, which turn out to be unique to the T4-related family of phages or significantly different from other bacterial systems. The activator of T4 late transcription is the gene 45 protein (gp45), the sliding clamp of the T4 replisome. Gp45 becomes topologically linked to DNA through the action of its clamp-loader, ...

  6. Transcription of the T4 late genes

    Directory of Open Access Journals (Sweden)

    Kassavetis George A

    2010-10-01

    Full Text Available Abstract This article reviews the current state of understanding of the regulated transcription of the bacteriophage T4 late genes, with a focus on the underlying biochemical mechanisms, which turn out to be unique to the T4-related family of phages or significantly different from other bacterial systems. The activator of T4 late transcription is the gene 45 protein (gp45, the sliding clamp of the T4 replisome. Gp45 becomes topologically linked to DNA through the action of its clamp-loader, but it is not site-specifically DNA-bound, as other transcriptional activators are. Gp45 facilitates RNA polymerase recruitment to late promoters by interacting with two phage-encoded polymerase subunits: gp33, the co-activator of T4 late transcription; and gp55, the T4 late promoter recognition protein. The emphasis of this account is on the sites and mechanisms of actions of these three proteins, and on their roles in the formation of transcription-ready open T4 late promoter complexes.

  7. A Vectorial Semantics Approach to Personality Assessment

    Science.gov (United States)

    Neuman, Yair; Cohen, Yochai

    2014-04-01

    Personality assessment and, specifically, the assessment of personality disorders have traditionally been indifferent to computational models. Computational personality is a new field that involves the automatic classification of individuals' personality traits that can be compared against gold-standard labels. In this context, we introduce a new vectorial semantics approach to personality assessment, which involves the construction of vectors representing personality dimensions and disorders, and the automatic measurements of the similarity between these vectors and texts written by human subjects. We evaluated our approach by using a corpus of 2468 essays written by students who were also assessed through the five-factor personality model. To validate our approach, we measured the similarity between the essays and the personality vectors to produce personality disorder scores. These scores and their correspondence with the subjects' classification of the five personality factors reproduce patterns well-documented in the psychological literature. In addition, we show that, based on the personality vectors, we can predict each of the five personality factors with high accuracy.

  8. A systematic method for analysing the protein hydration structure of T4 lysozyme

    Czech Academy of Sciences Publication Activity Database

    Kysilka, Jiří; Vondrášek, Jiří

    2013-01-01

    Roč. 26, č. 10 (2013), s. 479-487 ISSN 0952-3499 R&D Projects: GA ČR GAP208/10/0725; GA MŠk(CZ) LH11020 Grant - others:GA ČR(CZ) GAP302/10/0427 Institutional support: RVO:61388963 Keywords : protein hydration structure * water * X-ray crystallography * cluster algorithm * interaction enthalpy Subject RIV: CE - Biochemistry Impact factor: 2.337, year: 2013

  9. Lysozymes in the animal kingdom

    Indian Academy of Sciences (India)

    ... variation exists in their catalytic mechanisms and the genomic organization of their genes. Regarding their biological role, the widely recognized function of lysozymes is their contribution to antibacterial defence but, additionally, some lysozymes (belonging to different types) are known to function as digestive enzymes.

  10. Dimensionality reduction for 3D vectorial optical scattering problems

    NARCIS (Netherlands)

    Ivanova, Alyona; Stoffer, Remco; Hammer, Manfred

    2010-01-01

    The spatial dimensionality of vectorial 3D frequency domain optical scattering problems is reduced by means of a global expansion of the field in one direction in slab modes of some reference slice(s). A variational formalism yields the equations in the other two directions. These coupled partial

  11. Vectorial diffraction of extreme ultraviolet light and ultrashort light pulses

    NARCIS (Netherlands)

    Nugrowati, A.M.

    2008-01-01

    In this thesis, we present applications in optics involving the diffraction theory of light for two advanced technologies. We have used a rigorous vectorial diffraction method to model: (i) the imaging of mask structures in extreme ultraviolet lithography, and (ii) ultrashort pulse propagation

  12. Occupational asthma induced by inhaled egg lysozyme.

    Science.gov (United States)

    Bernstein, J A; Kraut, A; Bernstein, D I; Warrington, R; Bolin, T; Warren, C P; Bernstein, I L

    1993-02-01

    A 26-year-old man employed in a company which manufactured hen egg white derived lysozyme for use in the pharmaceutical industry was evaluated for occupational asthma. The worker began to experience immediate-onset asthmatic symptoms two months after starting to work with egg lysozyme powder. The work process involved the production of approximately 1,000 kg of purified dried lysozyme powder per week. Prick skin testing was positive to egg lysozyme (50 mg/ml) and other egg protein components, but negative to whole egg white and egg yolk reagents. Serum specific IgE to egg lysozyme was documented. Decrements in serial peak expiratory flow rates were associated with lysozyme exposure at work. A specific bronchoprovocation challenge to lysozyme powder was positive demonstrating an isolated immediate asthmatic response (48 percent decrease from baseline FEV1). This is the first reported case of lysozyme-induced asthma specifically caused by inhalational exposure to egg lysozyme.

  13. Evaluation of UMELISA® T4 NEONATAL and UMELISA® T4 using polystyrene plates coated with anti-thyroxine (T4) monoclonal antibodies.

    Science.gov (United States)

    Castells, Elisa M; González, Ernesto C; Pérez, Pedro L; Del Río, Lesley; Tejeda, Yileidis; Perea, Yenitse; Martín, Odalys; Espinosa, Maryeris; Rivero, Jose A; Frómeta, Amarilys

    2018-01-01

    Congenital hypothyroidism is one of the most common preventable causes of mental retardation. The Center of Immunoassay has developed the UMELISA® T4 NEONATAL and UMELISA® T4 to determine neonatal T4 levels in dried blood and serum samples. Both reagent kits use the same polystyrene plates coated with anti-thyroxine (T4) polyclonal antibodies as solid phase. This work shows the re-standardization of the UMELISA® T4 NEONATAL and UMELISA® T4 using plates coated with anti-T4 monoclonal antibodies (T4Mabs). Polystyrene plates of the modified assays were firstly coated with polyclonal IgG sheep-anti-mouse IgG for 18 hours. T4Mabs were added to the plates and incubated for 2 hours at room temperature. Different performance parameters were evaluated and correlation studies with the commercial kits done. Using polystyrene plates coated with T4Mabs increases the slope of the calibration curve in the clinical interest zone. The assay conjugates work twice diluted in respect to the ones of the commercial kits. Recovery percentages (90.8-110.7 for UMELISA® T4 NEONATAL and 92.1-109.3 for UMELISA® T4) and intra (7.2-7.6 for UMELISA® T4 NEONATAL and 6.9-7.2 for UMELISA® T4) and inter (7.4-8.5 for UMELISA® T4 NEONATAL and 7.1-8.5 for UMELISA® T4) coefficients of variation were similar to the ones described for the commercial kits. Limits of detection and quantification were 9.0 and 21.1 nmol/L for UMELISA® T4 NEONATAL, and 8.9 and 20.5 nmol/L for UMELISA® T4, respectively. The results also showed high overall concordance between assays (n = 244, r = 0.92, ρc = 0.91 for UMELISA® T4 NEONATAL and n = 492, r = 0.92, ρc = 0.9 for UMELISA® T4). The analytical sensibility of UMELISA® T4 NEONATAL and UMELISA® T4 is improved by using polystyrene plates coated with T4Mabs, without affecting the precision and accuracy of the results. T4: L-Thyroxine; ELISA: Enzyme-linked immunosorbent assay; SUMA: Ultra Micro Analytic System; UMELISA: Ultramicro enzyme

  14. An age-structured extension to the vectorial capacity model.

    Directory of Open Access Journals (Sweden)

    Vasiliy N Novoseltsev

    Full Text Available Vectorial capacity and the basic reproductive number (R(0 have been instrumental in structuring thinking about vector-borne pathogen transmission and how best to prevent the diseases they cause. One of the more important simplifying assumptions of these models is age-independent vector mortality. A growing body of evidence indicates that insect vectors exhibit age-dependent mortality, which can have strong and varied affects on pathogen transmission dynamics and strategies for disease prevention.Based on survival analysis we derived new equations for vectorial capacity and R(0 that are valid for any pattern of age-dependent (or age-independent vector mortality and explore the behavior of the models across various mortality patterns. The framework we present (1 lays the groundwork for an extension and refinement of the vectorial capacity paradigm by introducing an age-structured extension to the model, (2 encourages further research on the actuarial dynamics of vectors in particular and the relationship of vector mortality to pathogen transmission in general, and (3 provides a detailed quantitative basis for understanding the relative impact of reductions in vector longevity compared to other vector-borne disease prevention strategies.Accounting for age-dependent vector mortality in estimates of vectorial capacity and R(0 was most important when (1 vector densities are relatively low and the pattern of mortality can determine whether pathogen transmission will persist; i.e., determines whether R(0 is above or below 1, (2 vector population growth rate is relatively low and there are complex interactions between birth and death that differ fundamentally from birth-death relationships with age-independent mortality, and (3 the vector exhibits complex patterns of age-dependent mortality and R(0 ∼ 1. A limiting factor in the construction and evaluation of new age-dependent mortality models is the paucity of data characterizing vector mortality

  15. Advanced vectorial simulation of VCSELs with nano structures invited paper

    DEFF Research Database (Denmark)

    Chung, Il-Sug; Mørk, Jesper

    2009-01-01

    The single-mode properties and design issues of three vertical-cavity surface-emitting laser (VCSEL) structures incorporating nano structures are rigorously investigated. Nano structuring enables to deliver selective pumping or loss to the fundamental mode as well as stabilizing the output polari...... polarization state. Comparison of three vectorial simulation methods reveals that the modal expansion method is suitable for treating the nano structured VCSEL designs....

  16. A computational definition of the notion of vectorial space

    OpenAIRE

    Arrighi, Pablo; Dowek, Gilles

    2009-01-01

    We usually define an algebraic structure by a set, some operations defined on this set and some propositions that the algebraic structure must validate. In some cases, we can replace these propositions by an algorithm on terms constructed upon these operations that the algebraic structure must validate. We show in this note that this is the case for the notions of vectorial space and bilinear operation. KEYWORDS: Rewrite system, vector space, bilinear operation, tensorial product, semantics, ...

  17. Anaerobic Metabolism in T4 Acanthamoeba Genotype.

    Science.gov (United States)

    Alves, Daniella de Sousa Mendes Moreira; Alves, Luciano Moreira; da Costa, Tatiane Luiza; de Castro, Ana Maria; Vinaud, Marina Clare

    2017-06-01

    Members of the genus Acanthamoeba are of the most common protozoa that has been isolated from a variety of environment and affect immunocompromised individuals, causing granulomatous amoebic encephalitis and skin lesions. Acanthamoeba, in immunocompetent patients, may cause a keratitis related to corneal microtrauma. These free-living amoebas easily adapt to the host environment and wield metabolic pathways such as the energetic and respiratory ones in order to maintain viability for long periods. The energetic metabolism of cysts and trophozoites remains mostly unknown. There are a few reports on the energetic metabolism of these organisms as they are mitochondriate eukaryotes and some studies under aerobic conditions showing that Acanthamoeba hydrolyzes glucose into pyruvate via glycolysis. The aim of this study was to detect the energetic metabolic pathways with emphasis on anaerobic metabolism in trophozoites of three isolates of Acanthamoeba sp belonging to the T4 genotype. Two samples were collected in the environment and one was a clinical sample. The evaluation of these microorganisms proceeded as follows: rupture of trophozoites (7.5 × 10 3 parasites/ml) and biochemical analysis with high performance liquid chromatography and spectrophotometry. The anaerobic glycolysis was identified through the detection of glucose, pyruvate, and lactate. The protein catabolism was identified through the detection of fumarate, urea, and creatinine. The fatty acid oxidation was identified through the detection of acetate, beta-hydroxybutyrate, and propionate. The detected substances are the result of the consumption of energy reserves such as glycogen and lipids. The anaerobic glycolysis and protein catabolism pathways were observed in all three isolates: one clinical and two environmental. This study represents the first report of energetic pathways used by trophozoites from different isolates of the T4 genotype Acanthamoeba.

  18. Vectorial role of some dermanyssoid mites (Acari, Mesostigmata, Dermanyssoidea

    Directory of Open Access Journals (Sweden)

    Valiente Moro C.

    2005-06-01

    Full Text Available Among transmissible diseases, vectorial diseases represent a major problem for public health. In the group of acarina, while ticks are the most commonly implicated vectors, other arthropods and notably Dermanyssoidea are also involved in the transmission of pathogenic agents. Since the role of this superfamily is at present largely unknown, we have reviewed the vectorial role of these mites in the appearance, survival and propagation of pathogens. Various authors have shown that Dermanyssoidea are implicated in the transmission of both bacteria (Salmonella, Spirocheta, Rickettsia or Pasteurella and viruses (equine encephalitis viruses, West Nile virus, Fowl pox virus, the virus causing Newcastle disease and tick borne encephalitis viruses or hantaviruses. Finally, some authors have also shown their role in the transmission of some protozoa and filaria. As the vectorial character of such mites has been more clearly demonstrated (Dermanyssus gallinae, Ornithonyssus bacoti and Allodermanyssus sanguineus, it would be interesting to continue studies to better understand the role of this superfamily in the epidemiology of certain zoonoses.

  19. Vectorial analysis of the collimated beam of a small Gaussian source

    Science.gov (United States)

    Cao, Changqing; Wang, Ting; Zeng, Xiaodong; Feng, Zhejun; Zhang, Wenrui; Zhang, Xiaobing; Chen, Kun

    2018-03-01

    A vectorial analysis method to describe the collimated beam is proposed, the formulas of the intensity distribution and divergence angles represented in terms of Bessel functions are derived, and the propagation properties such as the vectorial structure of the collimated field and the shape of the beam spot are discussed in detail. Omitting the vectorial nature of the collimated beam can cause an error of 7.6% in determining the intensity distribution on the optical axis of the collimated beam.

  20. Complex coacervates of hyaluronic acid and lysozyme

    DEFF Research Database (Denmark)

    Water, Jorrit J.; Schack, Malthe M.; Velazquez-Campoy, Adrian

    2014-01-01

    stoichiometry was determined using solution depletion and isothermal titration calorimetry. The binding stoichiometry of lysozyme to hyaluronic acid (870 kDa) determined by solution depletion was found to be 225.9 ± 6.6 mol, or 0.1 bound lysozyme molecules per hyaluronic acid monomer. This corresponded well...... with that obtained by isothermal titration calorimetry of 0.09 bound lysozyme molecules per hyaluronic acid monomer. The complexation did not alter the secondary structure of lysozyme measured by Fourier-transform infrared spectroscopy overlap analysis and had no significant impact on the Tm of lysozyme determined...

  1. Sweetness characterization of recombinant human lysozyme.

    Science.gov (United States)

    Matano, Mami; Nakajima, Kana; Kashiwagi, Yutaka; Udaka, Shigezo; Maehashi, Kenji

    2015-10-01

    Lysozyme, a bacteriolytic enzyme, is widely distributed in nature and is a component of the innate immune system. It is established that chicken egg lysozyme elicits sweetness. However, the sweetness of human milk lysozyme, which is vital for combating microbial infections of the gastrointestinal tract of breast-fed infants, has not been characterized. This study aimed to assess the elicitation of sweetness using recombinant mammalian lysozymes expressed in Pichia pastoris. Recombinant human lysozyme (h-LZ) and other mammalian lysozymes of mouse, dog, cat and bovine milk elicited similar sweetness as determined using a sensory test, whereas bovine stomach lysozyme (bs-LZ) did not. Assays of cell cultures showed that h-LZ activated the human sweet taste receptor hT1R2/hT1R3, whereas bs-LZ did not. Point mutations confirmed that the sweetness of h-LZ was independent of enzyme activity and substrate-binding sites, although acidic amino acid residues of bs-LZ played a significant role in diminishing sweetness. Therefore, we conclude that elicitation of sweetness is a ubiquitous function among all lysozymes including mammalian lysozymes. These findings may provide novel insights into the biological implications of T1R2/T1R3-activation by mammalian lysozyme in the oral cavity and gastrointestinal tract. However, the function of lysozyme within species lacking the functional sweet taste receptor gene, such as cat, is currently unknown. Copyright © 2015 Elsevier Inc. All rights reserved.

  2. T-4G Simulator and T-4 Ground Training Devices in USAF Undergraduate Pilot Training.

    Science.gov (United States)

    Woodruff, Robert R.; Smith, James F.

    The objective of the project was to investigate the utility of using an A/F37A-T4G T-37 flight simulator within the context of Air Force undergraduate pilot training. Twenty-one subjects, selected from three undergraduate pilot training classes, were given contact flight training in a TP4G/EPT simulator before going to T-37 aircraft for further…

  3. Álgebra vectorial y geometría euclidiana

    OpenAIRE

    Salazar Caicedo, José Alonso

    1992-01-01

    Las leyes del álgebra vectorial asociadas al conjunto de los segmentos dirigidos del plano (o del espacio), en conexión con un producto interior, permiten demostrar una gran variedad de proposiciones y teoremas de la geometría clásica euclidiana, utilizando procedimientos y técnicas relativamente simples. Desde un punto de vista didáctico y pedagógico, surge el problema de examinar hasta qué punto es posible recuperar gran parte del arsenal de ideas fructíferas que proporcionaban a otras g...

  4. Fully vectorial highly nonparaxial beam close to the waist.

    Science.gov (United States)

    Chaumet, Patrick C

    2006-12-01

    I use the angular spectrum representation to compute exactly the Gaussian beam close to the waist (w(0)) in the case of a highly nonparaxial field (w(0)lambda). The computation is done in the vectorial case for a polarized Gaussian beam. In the area of the waist, the contribution of the propagating and evanescent waves is discussed. Moreover, the Gaussian wave is developed in terms of a series, which permits one to get analytical expressions for both propagating and evanescent waves when the observation is close to the waist.

  5. Quality control of radioimmunoassay reagents for T4

    International Nuclear Information System (INIS)

    Wayan Radiatning, S.

    1987-01-01

    Quality control of radioimmunoassay reagents for T4. A program of quality control testing has been carried out for 125 I-T4, T4 standards and T4 antisera. 125 I-labelled T4 has been tested for its specific activity, radiochemical purity using a Sephadex G-25 column, immunological activity, based on the immunological reaction between labelled antigen and excess T4 antibody, and non-specific binding. The useful shelf-life of the labelled compound was determined by monitoring the decrease in radiochemical purity and immunological activity, and the increase in non-specific binding. T4 standards were calibrated by means of T4 RIA kit manufactured by DPC (Diagnostic Products Corporation). A test on parallelism was also performed using hyperthyroid sera. T4 antisera were evaluated with respect to titre, avidity and specifity. The test results on 125 I-T4 show a specific activity varying between 1830-2020 uCi/ug, a radiochemical purity above 90%, an immunological more than 80% and a non-specific binding of less than 5%. The standard curve for T4 was found to coincide well with the standard curve of the DPC kit and parallel with the curve for hyperthyroid sera. The titre of T4 antisera obtained was 1:300, the avidity was about 4.8 x 10 7 and the cross-reaction for T3 was 1.6%. It can be concluded from the experimental results, that the 125 I-T4, T4 standards and T4 antisera prepared meet the requirements for the manufacture of T4 kits. (author). 5 refs.; 14 figs

  6. The generalized vectorial laws of reflection and refraction

    International Nuclear Information System (INIS)

    Bhattacharjee, Pramode Ranjan

    2005-01-01

    This paper discloses two important discoveries. These are: (i) discovery of ambiguity in the well-established laws of reflection and refraction of light which have been in regular use for many years, and (ii) discovery of generalized vectorial laws of reflection and refraction of light. The existing definitions of angle of incidence, angle of reflection and angle of refraction are considered first. Each of these definitions is found to be ambiguous, not in compliance with the fundamental definition of angle in geometry. Two typical questions (one in the case of reflection and the other for refraction) have been addressed, which cannot be dealt with by using the existing laws of reflection and refraction of light. Thus, the existing laws of reflection and refraction of light seem to be ambiguous in respect of generality and their validity in a broad sense is questionable. With a view to removing the ambiguities, proper definitions of the above three angles are given first and then the statement of the generalized vectorial law of reflection (as well as that of refraction) has been offered

  7. Vectorial diffraction properties of THz vortex Bessel beams.

    Science.gov (United States)

    Wu, Zhen; Wang, Xinke; Sun, Wenfeng; Feng, Shengfei; Han, Peng; Ye, Jiasheng; Yu, Yue; Zhang, Yan

    2018-01-22

    A vortex Bessel beam combines the merits of an optical vortex and a Bessel beam, including a spiral wave front and a non-diffractive feature, which has immense application potentials in optical trapping, optical fabrication, optical communications, and so on. Here, linearly and circularly polarized vortex Bessel beams in the terahertz (THz) frequency range are generated by utilizing a THz quarter wave plate, a spiral phase plate, and Teflon axicons with different opening angles. Taking advantage of a THz focal-plane imaging system, vectorial diffraction properties of the THz vortex Bessel beams are comprehensively characterized and discussed, including the transverse (Ex, Ey) and longitudinal (Ez) polarization components. The experimental phenomena are accurately simulated by adopting the vectorial Rayleigh diffraction integral. By varying the opening angle of the axicon, the characteristic parameters of these THz vortex Bessel beams are exhibited and compared, including the light spot size, the diffraction-free range, and the phase evolution process. This work provides the precise experimental and theoretical bases for the comprehension and application of a THz vortex Bessel beam.

  8. Sustained Benefit Lasting One Year from T4 Instead of T3-T4 Sympathectomy for Isolated Axillary Hyperhidrosis

    Science.gov (United States)

    Munia, Marco Antonio S.; Wolosker, Nelson; Kaufmann, Paulo; de Campos, José Ribas Milanes; Puech-Leão, Pedro

    2008-01-01

    INTRODUCTION Level T4 video-assisted thoracoscopic sympathectomy proved superior to T3-T4 treatment for controlling axillary hyperhidrosis at the initial and six-month follow-ups of these patients. OBJECTIVE To compare the results of two levels of sympathectomy (T3-T4 vs. T4) for treating axillary sudoresis over one year of follow-up. METHODS Sixty-four patients with axillary hyperhidrosis were randomized to denervation of T3-T4 or T4 alone and followed prospectively. All patients were examined preoperatively and were followed postoperatively for one year. Axillary hyperhidrosis treatment was evaluated, along with the presence, location, and severity of compensatory hyperhidrosis and self-reported quality of life. RESULTS According to patient reports after one year, all cases of axillary hyperhidrosis were successfully treated by surgery. There were no instances of treatment failure. After six months, compensatory hyperhidrosis was present in 27 patients of the T3-T4 group (87.1%) and in 16 patients of the T4 group (48.5%). After one year, all T3-T4 patients experienced some degree of compensatory hyperhidrosis, compared to only 14 patients in the T4 group (42.4%). In addition, compensatory hyperhidrosis was less severe in the T4 patients (p hyperhidrosis, but the T4 group showed milder compensatory hyperhidrosis and greater patient satisfaction at the one-year follow-up. PMID:19060999

  9. Far-field divergence of a vectorial plane wave diffracted by a circular aperture from the vectorial structure

    International Nuclear Information System (INIS)

    Zhou Guo-Quan

    2011-01-01

    Based on the vectorial structure of an electromagnetic wave, the analytical and concise expressions for the TE and TM terms of a vectorial plane wave diffracted by a circular aperture are derived in the far-field. The expressions of the energy flux distributions of the TE term, the TM term and the diffracted plane wave are also presented. The ratios of the power of the TE and TM terms to that of the diffracted plane wave are examined in the far-field. In addition, the far-field divergence angles of the TE term, the TM term and the diffracted plane wave, which are related to the energy flux distribution, are investigated. The different energy flux distributions of the TE and TM terms result in the discrepancy of their divergence angles. The influences of the linearly polarized angle and the radius of the circular aperture on the far-field divergence angles of the TE term, the TM term and the diffracted plane wave are discussed in detail. This research may promote the recognition of the optical propagation through a circular aperture. (electromagnetism, optics, acoustics, heat transfer, classical mechanics, and fluid dynamics)

  10. Scientist prepare Lysozyme Protein Crystal

    Science.gov (United States)

    1996-01-01

    Dan Carter and Charles Sisk center a Lysozyme Protein crystal grown aboard the USML-2 shuttle mission. Protein isolated from hen egg-white and functions as a bacteriostatic enzyme by degrading bacterial cell walls. First enzyme ever characterized by protein crystallography. It is used as an excellent model system for better understanding parameters involved in microgravity crystal growth experiments. The goal is to compare kinetic data from microgravity experiments with data from laboratory experiments to study the equilibrium.

  11. Evolution of the mammalian lysozyme gene family

    Science.gov (United States)

    2011-01-01

    Background Lysozyme c (chicken-type lysozyme) has an important role in host defense, and has been extensively studied as a model in molecular biology, enzymology, protein chemistry, and crystallography. Traditionally, lysozyme c has been considered to be part of a small family that includes genes for two other proteins, lactalbumin, which is found only in mammals, and calcium-binding lysozyme, which is found in only a few species of birds and mammals. More recently, additional testes-expressed members of this family have been identified in human and mouse, suggesting that the mammalian lysozyme gene family is larger than previously known. Results Here we characterize the extent and diversity of the lysozyme gene family in the genomes of phylogenetically diverse mammals, and show that this family contains at least eight different genes that likely duplicated prior to the diversification of extant mammals. These duplicated genes have largely been maintained, both in intron-exon structure and in genomic context, throughout mammalian evolution. Conclusions The mammalian lysozyme gene family is much larger than previously appreciated and consists of at least eight distinct genes scattered around the genome. Since the lysozyme c and lactalbumin proteins have acquired very different functions during evolution, it is likely that many of the other members of the lysozyme-like family will also have diverse and unexpected biological properties. PMID:21676251

  12. Evolution of the mammalian lysozyme gene family

    Directory of Open Access Journals (Sweden)

    Biegel Jason M

    2011-06-01

    Full Text Available Abstract Background Lysozyme c (chicken-type lysozyme has an important role in host defense, and has been extensively studied as a model in molecular biology, enzymology, protein chemistry, and crystallography. Traditionally, lysozyme c has been considered to be part of a small family that includes genes for two other proteins, lactalbumin, which is found only in mammals, and calcium-binding lysozyme, which is found in only a few species of birds and mammals. More recently, additional testes-expressed members of this family have been identified in human and mouse, suggesting that the mammalian lysozyme gene family is larger than previously known. Results Here we characterize the extent and diversity of the lysozyme gene family in the genomes of phylogenetically diverse mammals, and show that this family contains at least eight different genes that likely duplicated prior to the diversification of extant mammals. These duplicated genes have largely been maintained, both in intron-exon structure and in genomic context, throughout mammalian evolution. Conclusions The mammalian lysozyme gene family is much larger than previously appreciated and consists of at least eight distinct genes scattered around the genome. Since the lysozyme c and lactalbumin proteins have acquired very different functions during evolution, it is likely that many of the other members of the lysozyme-like family will also have diverse and unexpected biological properties.

  13. Lysis of lysis-inhibited bacteriophage T4-infected cells.

    OpenAIRE

    Abedon, S T

    1992-01-01

    T4 bacteriophage (phage)-infected cells show a marked increase in latent-period length, called lysis inhibition, upon adsorption of additional T4 phages (secondary adsorption). Lysis inhibition is a complex phenotype requiring the activity of at least six T4 genes. Two basic mysteries surround our understanding of the expression of lysis inhibition: (i) the mechanism of initiation (i.e., how secondary adsorption leads to the expression of lysis inhibition) and (ii) the mechanism of lysis (i.e...

  14. Reduction of product platform complexity by vectorial Euclidean algorithm

    Energy Technology Data Exchange (ETDEWEB)

    Navarrete, Israel Aguilera; Guzman, Alejandro A. Lozano [Cerro Blanco, Santiago de Queretaro (Mexico)

    2013-11-15

    In traditional machine, equipment and devices design, technical solutions are practically independent, thus increasing designs cost and complexity. Overcoming this situation has been tackled just using designer's experience. In this work, a product platform complexity reduction is presented based on a matrix representation of technical solutions versus product properties. This matrix represents the product platform. From this matrix, the Euclidean distances among technical solutions are obtained. Thus, the vectorial distances among technical solutions are identified in a new matrix of order of the number of technical solutions identified. This new matrix can be reorganized in groups with a hierarchical structure, in such a way that modular design of products is now more tractable. As a result of this procedure, the minimum vector distances are found thus being possible to identify the best technical solutions for the design problem raised. Application of these concepts is shown with two examples.

  15. A new family of lysozyme inhibitors contributing to lysozyme tolerance in gram-negative bacteria.

    Directory of Open Access Journals (Sweden)

    Lien Callewaert

    2008-03-01

    Full Text Available Lysozymes are ancient and important components of the innate immune system of animals that hydrolyze peptidoglycan, the major bacterial cell wall polymer. Bacteria engaging in commensal or pathogenic interactions with an animal host have evolved various strategies to evade this bactericidal enzyme, one recently proposed strategy being the production of lysozyme inhibitors. We here report the discovery of a novel family of bacterial lysozyme inhibitors with widespread homologs in gram-negative bacteria. First, a lysozyme inhibitor was isolated by affinity chromatography from a periplasmic extract of Salmonella Enteritidis, identified by mass spectrometry and correspondingly designated as PliC (periplasmic lysozyme inhibitor of c-type lysozyme. A pliC knock-out mutant no longer produced lysozyme inhibitory activity and showed increased lysozyme sensitivity in the presence of the outer membrane permeabilizing protein lactoferrin. PliC lacks similarity with the previously described Escherichia coli lysozyme inhibitor Ivy, but is related to a group of proteins with a common conserved COG3895 domain, some of them predicted to be lipoproteins. No function has yet been assigned to these proteins, although they are widely spread among the Proteobacteria. We demonstrate that at least two representatives of this group, MliC (membrane bound lysozyme inhibitor of c-type lysozyme of E. coli and Pseudomonas aeruginosa, also possess lysozyme inhibitory activity and confer increased lysozyme tolerance upon expression in E. coli. Interestingly, mliC of Salmonella Typhi was picked up earlier in a screen for genes induced during residence in macrophages, and knockout of mliC was shown to reduce macrophage survival of S. Typhi. Based on these observations, we suggest that the COG3895 domain is a common feature of a novel and widespread family of bacterial lysozyme inhibitors in gram-negative bacteria that may function as colonization or virulence factors in bacteria

  16. A New Family of Lysozyme Inhibitors Contributing to Lysozyme Tolerance in Gram-Negative Bacteria

    Science.gov (United States)

    Callewaert, Lien; Aertsen, Abram; Deckers, Daphne; Vanoirbeek, Kristof G. A.; Vanderkelen, Lise; Van Herreweghe, Joris M.; Masschalck, Barbara; Nakimbugwe, Dorothy; Robben, Johan; Michiels, Chris W.

    2008-01-01

    Lysozymes are ancient and important components of the innate immune system of animals that hydrolyze peptidoglycan, the major bacterial cell wall polymer. Bacteria engaging in commensal or pathogenic interactions with an animal host have evolved various strategies to evade this bactericidal enzyme, one recently proposed strategy being the production of lysozyme inhibitors. We here report the discovery of a novel family of bacterial lysozyme inhibitors with widespread homologs in gram-negative bacteria. First, a lysozyme inhibitor was isolated by affinity chromatography from a periplasmic extract of Salmonella Enteritidis, identified by mass spectrometry and correspondingly designated as PliC (periplasmic lysozyme inhibitor of c-type lysozyme). A pliC knock-out mutant no longer produced lysozyme inhibitory activity and showed increased lysozyme sensitivity in the presence of the outer membrane permeabilizing protein lactoferrin. PliC lacks similarity with the previously described Escherichia coli lysozyme inhibitor Ivy, but is related to a group of proteins with a common conserved COG3895 domain, some of them predicted to be lipoproteins. No function has yet been assigned to these proteins, although they are widely spread among the Proteobacteria. We demonstrate that at least two representatives of this group, MliC (membrane bound lysozyme inhibitor of c-type lysozyme) of E. coli and Pseudomonas aeruginosa, also possess lysozyme inhibitory activity and confer increased lysozyme tolerance upon expression in E. coli. Interestingly, mliC of Salmonella Typhi was picked up earlier in a screen for genes induced during residence in macrophages, and knockout of mliC was shown to reduce macrophage survival of S. Typhi. Based on these observations, we suggest that the COG3895 domain is a common feature of a novel and widespread family of bacterial lysozyme inhibitors in gram-negative bacteria that may function as colonization or virulence factors in bacteria interacting with

  17. T4 phages against Escherichia coli diarrhea: potential and problems.

    Science.gov (United States)

    Denou, Emmanuel; Bruttin, Anne; Barretto, Caroline; Ngom-Bru, Catherine; Brüssow, Harald; Zuber, Sophie

    2009-05-25

    A combination of in vitro and in vivo experiments with comparative phage genomics was used for the rational design of a phage cocktail against E. coli diarrhea. Orally applied T4 coliphages representing three different subgroups (T4-, RB49- and JS98-like phages) had no negative impact on the murine gut microbiota. T4 phages were found with high titers in the cecum and colon and lower titers in the small intestine, but were not detected in the blood, liver or spleen. No adverse effects were observed after one-month exposure to phage nor were serum anti-T4 antibodies detected. T4 phages belonging to the same subgroup showed closely related genomes that differed by 12 (phage JS10 vs. JS98 reference) to 17 (phage JSE vs. RB49 reference) insertion/deletions mostly representing single small ORFs. Bioinformatic analysis did not reveal undesired genes in the T4 genomes. Sequence variability was seen over the tail fibre genes, but the variability did not correlate with phage host range. The investigated T4 phages were not only species- but also strain-specific, necessitating the use of phage cocktails consisting of 10 and 16 T4 phage isolates to cover half to two thirds of E. coli strains representing the five main pathotypes isolated from diarrhea patients.

  18. Invertebrate lysozymes: Diversity and distribution, molecular ...

    Indian Academy of Sciences (India)

    This review describes the current knowledge on i-type lysozymes, outlining their distribution, molecular mechanism and in vivo function taking the representative from Venerupis philippinarum (formerly Tapes japonica) (Vp-ilys) as a model. In addition, invertebrate g-type and ch-type (chalaropsis) lysozymes, which have ...

  19. Complex coacervation of lysozyme and heparin

    DEFF Research Database (Denmark)

    van de Weert, Marco; Andersen, Mia Bendix; Frokjaer, Sven

    2004-01-01

    To characterize complex coacervates/flocculates of lysozyme and heparin in terms of binding stoichiometry and to determine the effect of complexation on protein structure and stability.......To characterize complex coacervates/flocculates of lysozyme and heparin in terms of binding stoichiometry and to determine the effect of complexation on protein structure and stability....

  20. Hydrophobic nano-carrier for lysozyme adsorption

    Indian Academy of Sciences (India)

    2016-08-26

    Aug 26, 2016 ... For this purpose, adsorption conditions wereoptimized and maximum lysozyme binding capacity was found to be 278.8 mg g−1 polymer in pH 7.0 phosphate buffer at 25∘C. Desorption and reusability properties of the nanoparticles were investigated and lysozyme adsorption efficiency did not change ...

  1. Fluorescence Studies of Lysozyme Nucleation

    Science.gov (United States)

    Pusey, Marc L.; Smith, Lori

    1998-01-01

    Fluorescence is one of the most powerful tools available for the study of macromolecules. For example, fluorescence can be used to study self association through methods such as anisotropy (the rotational rate of the molecule in solution), quenching (the accessibility of a bound probe to the bulk solution), and resonance energy transfer (measurement of the distance between two species). Fluorescence can also be used to study the local environment of the probe molecules, and the changes in that environment which accompany crystal nucleation and growth. However fluorescent techniques have been very much underutilized in macromolecular growth studies. One major advantage is that the fluorescent species generally must be at low concentration, typically ca 10-5 to 10-6 M. Thus one can study a very wide range of solution conditions, ranging from very high to very low protein concentration, he latter of which are not readily accessible to scattering techniques. We have prepared a number of fluorescent derivatives of chicken egg white lysozyme (CEWL). Fluorescent probes have been attached to two different sites, ASP 101 and the N-terrninal amine, with a sought for use in different lines of study. Preliminary resonance energy transfer studies have been -carried out using pyrene acetic acid (Ex 340 mn, Em 376 nm) lysozyme as a donor and cascade blue (Ex 377 run, Em 423 nm) labeled lysozyme as an acceptor. The emission of both the pyrene and cascade blue probes was followed as a function of the salt protein concentrations. The data show an increase in cascade blue and a concomitant decrease in the pyrene fluorescence as either the salt or protein concentrations are increased, suggesting that the two species are approaching each other close enough for resonance energy transfer to occur. This data can be analyzed to measure the distance between the probe molecules and, knowing their locations on the protein molecule their distances from and orientations with respect to each

  2. Studies on biotransformation of lysozyme, 4

    International Nuclear Information System (INIS)

    Yuzuriha, Teruaki; Katayama, Kouichi; Tsutsumi, Junzo

    1978-01-01

    The radioimmunoassay of hen egg-white lysozyme (hen lysozyme) was investigated and evaluated in order to examine the biotransformation of the enzyme in men. 125 I-labeled hen lysozyme was prepared and separation of the free from the bound form of the enzyme was performed by dextran-treated charcoal method in comparison with two-antibody method. From the result, it was indicated that dextran-treated charcoal method was useful from the viewpoint of precision, reproducibility and simplicity. The stability of hen lysozyme labeled was determined by the degree of radioactivity bound to antibody. The recovery of radioactivity bound to antibody decreased with the time lapse after labeling and it was found that the recovery rate was restored by dialysis. In this radioimmunoassay system, the competitive binding of labeled and unlabeled hen lysozyme to antibody was examined, and 0.3 to 5.0 ng/ml of the enzyme were detectable. It was found that human lysozyme had an influence on the radioimmunoassay system with over 2 μg/ml. But, the influence did not disturb the assay of hen lysozyme in human serum. Additionally, since it was found that the ratio of the bound form to the free increased with human serum, the assay of hen lysozyme in human serum was carried out using a calibration curve of the enzyme added to untreated human serum. Thus, human serum levels after oral administration of hen lysozyme were measured by the radioimmunoassay in comparison with the determination method by lytic activity. This radioimmunoassay system was found to be appropriate for the determination of hen lysozyme in human serum. (auth.)

  3. A vectorial description of electromagnetic scattering by large bodies of spherical shape

    International Nuclear Information System (INIS)

    Bourrely, C.; Lemaire, T.; Chiappetta, P.; Centre National de la Recherche Scientifique, 13 - Marseille

    1989-10-01

    We present a new method to obtain a vectorial solution of Helmholtz equation for large homogeneous scatterers having a cylindrical symmetry and a shape approximately spherical. Limitations of the method for arbitrarily shaped particles are discussed

  4. Identifying Orbital Angular Momentum of Vectorial Vortices with Pancharatnam Phase and Stokes Parameters.

    Science.gov (United States)

    Zhang, Dengke; Feng, Xue; Cui, Kaiyu; Liu, Fang; Huang, Yidong

    2015-07-10

    In this work, an explicit formula is deduced for identifying the orbital angular moment (OAM) of vectorial vortex with space-variant state of polarization (SOP). Different to scalar vortex, the OAM of vectorial vortex can be attributed to two parts: 1. the azimuthal gradient of Pancharatnam phase; 2. the product between the azimuthal gradient of orientation angle of SOP and relevant solid angle on the Poincaré sphere. With our formula, a geometrical description for OAM of light beams can be achieved under the framework of the traditional Poincaré sphere. Numerical simulations for two types of vectorial vortices have been carried on to confirm our presented formula as well as demonstrate the geometrical description of OAM. Furthermore, this work would pave the way for precise characterization of OAM charge of vectorial vortices.

  5. Structural and Mechanistic Studies of Pesticin, a Bacterial Homolog of Phage Lysozymes*

    Science.gov (United States)

    Patzer, Silke I.; Albrecht, Reinhard; Braun, Volkmar; Zeth, Kornelius

    2012-01-01

    Yersinia pestis produces and secretes a toxin named pesticin that kills related bacteria of the same niche. Uptake of the bacteriocin is required for activity in the periplasm leading to hydrolysis of peptidoglycan. To understand the uptake mechanism and to investigate the function of pesticin, we combined crystal structures of the wild type enzyme, active site mutants, and a chimera protein with in vivo and in vitro activity assays. Wild type pesticin comprises an elongated N-terminal translocation domain, the intermediate receptor binding domain, and a C-terminal activity domain with structural analogy to lysozyme homologs. The full-length protein is toxic to bacteria when taken up to the target site via the outer or the inner membrane. Uptake studies of deletion mutants in the translocation domain demonstrate their critical size for import. To further test the plasticity of pesticin during uptake into bacterial cells, the activity domain was replaced by T4 lysozyme. Surprisingly, this replacement resulted in an active chimera protein that is not inhibited by the immunity protein Pim. Activity of pesticin and the chimera protein was blocked through introduction of disulfide bonds, which suggests unfolding as the prerequisite to gain access to the periplasm. Pesticin, a muramidase, was characterized by active site mutations demonstrating a similar but not identical residue pattern in comparison with T4 lysozyme. PMID:22593569

  6. Re-initiation repair in bacteriophage T4

    International Nuclear Information System (INIS)

    Cupido, M.

    1981-01-01

    Irradiation of bacteriophage T4 with ultraviolet light induces the formation of pyrimidine dimers in its DNA. These dimers hamper replication of DNA and, to a lesser extent, transcription of DNA after its infection of bacteria. A number of pathways enable phage T4 to multiply dimer-containing DNA. One of these pathways has been named replication repair and is described in this thesis. The properties of two phage strains, unable to perform replication repair, have been studied to obtain a picture of the repair process. The mutations in these strains that affect replication repair have been located on the genomic map of T4. (Auth.)

  7. Vectorial Ekeland Variational Principles and Inclusion Problems in Cone Quasi-Uniform Spaces

    Directory of Open Access Journals (Sweden)

    Jiang Zhu

    2012-01-01

    principle, vectorial quasiequilibrium principle are obtained. Also, several other important principles in nonlinear analysis are extended to cone quasi-uniform spaces. The results of this paper extend, generalize, and improve the corresponding results for Ekeland's variational principles of the directed vectorial perturbation type and other generalizations of Ekeland's variational principles in the setting of F-type topological space and quasi-metric spaces in the literatures. Even in usual real metric spaces, some of our results are new.

  8. A Vectorial Capacity Product to Monitor Changing Malaria Transmission Potential in Epidemic Regions of Africa

    OpenAIRE

    Pietro Ceccato; Christelle Vancutsem; Robert Klaver; James Rowland; Stephen J. Connor

    2012-01-01

    Rainfall and temperature are two of the major factors triggering malaria epidemics in warm semi-arid (desert-fringe) and high altitude (highland-fringe) epidemic risk areas. The ability of the mosquitoes to transmit Plasmodium spp. is dependent upon a series of biological features generally referred to as vectorial capacity. In this study, the vectorial capacity model (VCAP) was expanded to include the influence of rainfall and temperature variables on malaria transmission potential. Data ...

  9. Identifying Orbital Angular Momentum of Vectorial Vortices with Pancharatnam Phase and Stokes Parameters

    OpenAIRE

    Zhang, Dengke; Feng, Xue; Cui, Kaiyu; Liu, Fang; Huang, Yidong

    2015-01-01

    In this work, an explicit formula is deduced for identifying the orbital angular moment (OAM) of vectorial vortex with space-variant state of polarization (SOP). Different to scalar vortex, the OAM of vectorial vortex can be attributed to two parts: 1. the azimuthal gradient of Pancharatnam phase; 2. the product between the azimuthal gradient of orientation angle of SOP and relevant solid angle on the Poincar? sphere. With our formula, a geometrical description for OAM of light beams can be a...

  10. Influence of polymers on lysozyme molecules association

    Directory of Open Access Journals (Sweden)

    Gromovoy T. Yu

    2011-12-01

    Full Text Available Aim. Study of lysozyme molecules behaviour at immobilization in gelatin and carboxymethyl cellulose sodium salt solutions by matrix-assisted laser desorption/ionization (MALDI. Methods. Determination of the activity of lysozyme, both free and entrapped in gelatin and carboxymethyl cellulose sodium salt (Na-CMC solutions, was conducted by bacteriolytic method. The enzyme interaction with polymers was confirmed by viscometry and mass-spectrometry methods. Results. The occurrence of lysozyme associates in aqueous solution in monomeric and oligomeric forms was shown. A non-valent interaction of the enzyme with solutions of polymers results in the dissociation of oligomeric associates into subunits, which depends on the support nature and mass ratio of lysozyme to polymer. The quantitative retention of immobilized lysozyme hydrolytic activity was established, which favours obtaining mucoadhesive film forms with bacteriolytic action. Conclusions. The lysozyme immobilization by non-valent interactions in gelatin solution and Na-CMC solutions causes dissociation of the enzyme oligomeric structures; a stronger lysozyme coupling with NaCMC was noted.

  11. A vectorial approach to determine frozen orbital conditions

    Science.gov (United States)

    Circi, Christian; Condoleo, Ennio; Ortore, Emiliano

    2017-06-01

    Taking into consideration a probe moving in an elliptical orbit around a celestial body, the possibility of determining conditions which lead to constant values on average of all the orbit elements has been investigated here, considering the influence of the planetary oblateness and the long-term effects deriving from the attraction of several perturbing bodies. To this end, three equations describing the variation of orbit eccentricity, apsidal line and angular momentum unit vector have been first retrieved, starting from a vectorial expression of the Lagrange planetary equations and considering for the third-body perturbation the gravity-gradient approximation, and then exploited to demonstrate the feasibility of achieving the above-mentioned goal. The study has led to the determination of two families of solutions at constant mean orbit elements, both characterised by a co-planarity condition between the eccentricity vector, the angular momentum and a vector resulting from the combination of the orbital poles of the perturbing bodies. As a practical case, the problem of a probe orbiting the Moon has been faced, taking into account the temporal evolution of the perturbing poles of the Sun and Earth, and frozen solutions at argument of pericentre 0^{circ } or 180^{circ } have been found.

  12. Acanthamoeba T4 genotype associated with keratitis infections in Tunisia.

    Science.gov (United States)

    Dendana, F; Sellami, H; Trabelsi, H; Neji, S; Cheikhrouhou, F; Makni, F; Ayadi, A

    2013-01-01

    Acanthamoeba keratitis (AK) is a sight-threatening infection. We report five cases of AK diagnosed from 2005 to 2009 in the Laboratory of Parasitology-Mycology at Habib Bourguiba Sfax Hospital, Tunisia. All were associated with improper care of contact lenses (rinsing of contact lenses with tap water and inappropriate cleaning) and lens storage. The patients displayed different clinical presentations: corneal inflammation, corneal ulceration, and corneal abscess. The diagnosis was made after direct examination, culture, and polymerase chain reaction amplification with specific primers. The genotype classification was based on the highly variable DF3 region in the 18S rRNA gene. This is the first study characterizing Acanthamoeba genotype in Tunisia and North Africa. All Acanthamoeba isolates were associated to the T4 genotype. Three different DF3 sequence types were related to AK infections T4/10, T4/15, and T4/16.

  13. The diversity and evolution of the T4-type bacteriophages.

    Science.gov (United States)

    Desplats, Carine; Krisch, Henry M

    2003-05-01

    Recent studies suggest that viruses are the most numerous entities in the biosphere; bacteriophages, the viruses that infect Eubacteria and Archaea, constitute a substantial fraction of this population. In spite of their ubiquity, the vast majority of phages in the environment have never been studied and nothing is known about them. For the last 10 years our research has focused on an extremely widespread group of phages, the T4-type. It has now become evident that phage T4 has a myriad of relatives in nature that differ significantly in their host range. The genomes of all these phages have homology to the T4 genes that determine virion morphology. Although phylogenetically related, these T4-type phages can be subdivided into four groups that are increasingly distant from T4: the T-evens, the pseudo T-evens, the schizo T-evens and the exo T-evens. Genomic comparisons between the various T4-type phages and T4 indicate that these genomes share homology not only for virion structural components but also for most of the essential genes involved in the T4 life cycle. This suggests that horizontal transmission of the genetic information may have played a less general role in the evolution of these phages than has been supposed. Nevertheless, we have identified several regions of the T4-type genome, such as the segment containing the tail fiber genes that exhibit evidence of extensive modular shuffling during evolution. The T4-type genomes appear to be a mosaic containing a large and fixed group of essential genes as well as highly variable set of non-essential genes. These non-essential genes are probably important for the adaptation of these phages to their particular life-style. Furthermore, swapping autonomous domains within the essential proteins may slightly modify their function(s) and contribute to the adaptive ability of the T4-type phage family. Regulatory sequences also display considerable evolutionary plasticity and this too may facilitate the adaptation of

  14. A New Family of Lysozyme Inhibitors Contributing to Lysozyme Tolerance in Gram-Negative Bacteria

    OpenAIRE

    Callewaert, Lien; Aertsen, Abram; Deckers, Daphne; Vanoirbeek, Kristof G. A.; Vanderkelen, Lise; Van Herreweghe, Joris M.; Masschalck, Barbara; Nakimbugwe, Dorothy; Robben, Johan; Michiels, Chris W.

    2008-01-01

    Lysozymes are ancient and important components of the innate immune system of animals that hydrolyze peptidoglycan, the major bacterial cell wall polymer. Bacteria engaging in commensal or pathogenic interactions with an animal host have evolved various strategies to evade this bactericidal enzyme, one recently proposed strategy being the production of lysozyme inhibitors. We here report the discovery of a novel family of bacterial lysozyme inhibitors with widespread homologs in gram-negative...

  15. Genetic diversity among five T4-like bacteriophages.

    Science.gov (United States)

    Nolan, James M; Petrov, Vasiliy; Bertrand, Claire; Krisch, Henry M; Karam, Jim D

    2006-05-23

    Bacteriophages are an important repository of genetic diversity. As one of the major constituents of terrestrial biomass, they exert profound effects on the earth's ecology and microbial evolution by mediating horizontal gene transfer between bacteria and controlling their growth. Only limited genomic sequence data are currently available for phages but even this reveals an overwhelming diversity in their gene sequences and genomes. The contribution of the T4-like phages to this overall phage diversity is difficult to assess, since only a few examples of complete genome sequence exist for these phages. Our analysis of five T4-like genomes represents half of the known T4-like genomes in GenBank. Here, we have examined in detail the genetic diversity of the genomes of five relatives of bacteriophage T4: the Escherichia coli phages RB43, RB49 and RB69, the Aeromonas salmonicida phage 44RR2.8t (or 44RR) and the Aeromonas hydrophila phage Aeh1. Our data define a core set of conserved genes common to these genomes as well as hundreds of additional open reading frames (ORFs) that are nonconserved. Although some of these ORFs resemble known genes from bacterial hosts or other phages, most show no significant similarity to any known sequence in the databases. The five genomes analyzed here all have similarities in gene regulation to T4. Sequence motifs resembling T4 early and late consensus promoters were observed in all five genomes. In contrast, only two of these genomes, RB69 and 44RR, showed similarities to T4 middle-mode promoter sequences and to the T4 motA gene product required for their recognition. In addition, we observed that each phage differed in the number and assortment of putative genes encoding host-like metabolic enzymes, tRNA species, and homing endonucleases. Our observations suggest that evolution of the T4-like phages has drawn on a highly diverged pool of genes in the microbial world. The T4-like phages harbour a wealth of genetic material that has

  16. Genetic diversity among five T4-like bacteriophages

    Directory of Open Access Journals (Sweden)

    Bertrand Claire

    2006-05-01

    Full Text Available Abstract Background Bacteriophages are an important repository of genetic diversity. As one of the major constituents of terrestrial biomass, they exert profound effects on the earth's ecology and microbial evolution by mediating horizontal gene transfer between bacteria and controlling their growth. Only limited genomic sequence data are currently available for phages but even this reveals an overwhelming diversity in their gene sequences and genomes. The contribution of the T4-like phages to this overall phage diversity is difficult to assess, since only a few examples of complete genome sequence exist for these phages. Our analysis of five T4-like genomes represents half of the known T4-like genomes in GenBank. Results Here, we have examined in detail the genetic diversity of the genomes of five relatives of bacteriophage T4: the Escherichia coli phages RB43, RB49 and RB69, the Aeromonas salmonicida phage 44RR2.8t (or 44RR and the Aeromonas hydrophila phage Aeh1. Our data define a core set of conserved genes common to these genomes as well as hundreds of additional open reading frames (ORFs that are nonconserved. Although some of these ORFs resemble known genes from bacterial hosts or other phages, most show no significant similarity to any known sequence in the databases. The five genomes analyzed here all have similarities in gene regulation to T4. Sequence motifs resembling T4 early and late consensus promoters were observed in all five genomes. In contrast, only two of these genomes, RB69 and 44RR, showed similarities to T4 middle-mode promoter sequences and to the T4 motA gene product required for their recognition. In addition, we observed that each phage differed in the number and assortment of putative genes encoding host-like metabolic enzymes, tRNA species, and homing endonucleases. Conclusion Our observations suggest that evolution of the T4-like phages has drawn on a highly diverged pool of genes in the microbial world. The T4

  17. Modelling polarization dependent absorption: The vectorial Lambert-Beer law

    Science.gov (United States)

    Franssens, G.

    2014-07-01

    The scalar Lambert-Beer law, describing the absorption of unpolarized light travelling through a linear non-scattering medium, is simple, well-known, and mathematically trivial. However, when we take the polarization of light into account and consider a medium with polarization dependent absorption, we now need a Vectorial Lambert-Beer Law (VLBL) to quantify this interaction. Such a generalization of the scalar Lambert-Beer law appears not to be readily available. A careful study of this topic reveals that it is not a trivial problem. We will see that the VLBL is not and cannot be a straightforward vectorized version of its scalar counterpart. The aim of the work is to present the general form of the VLBL and to explain how it arises. A reasonable starting point to derive the VLBL is the Vectorial Radiative Transfer Equation (VRTE), which models the absorption and scattering of (partially) polarized light travelling through a linear medium. When we turn off scattering, the VRTE becomes an infinitesimal model for the VLBL holding in the medium. By integrating this equation, we expect to find the VLBL. Surprisingly, this is not the end of the story. It turns out that light propagation through a medium with polarization-dependent absorption is mathematically not that trivial. The trickiness behind the VLBL can be understood in the following terms. The matrix in the VLBL, relating any input Stokes vector to the corresponding output Stokes vector, must necessarily be a Mueller matrix. The subset of invertible Mueller matrices forms a Lie group. It is known that this Lie group contains the ortho-chronous Lorentz group as a subgroup. The group manifold of this subgroup has a (well-known) non-trivial topology. Consequently, the manifold of the Lie group of Mueller matrices also has (at least the same, but likely a more general) non-trivial topology (the full extent of which is not yet known). The type of non-trivial topology, possessed by the manifold of (invertible

  18. Comparative genomics of the T4-Like Escherichia coli phage JS98: implications for the evolution of T4 phages.

    Science.gov (United States)

    Chibani-Chennoufi, Sandra; Canchaya, Carlos; Bruttin, Anne; Brüssow, Harald

    2004-12-01

    About 130 kb of sequence information was obtained from the coliphage JS98 isolated from the stool of a pediatric diarrhea patient in Bangladesh. The DNA shared up to 81% base pair identity with phage T4. The most conserved regions between JS98 and T4 were the structural genes, but their degree of conservation was not uniform. The head genes showed the highest sequence conservation, followed by the tail, baseplate, and tail fiber genes. Many tail fiber genes shared only protein sequence identity. Except for the insertion of endonuclease genes in T4 and gene 24 duplication in JS98, the structural gene maps of the two phages were colinear. The receptor-recognizing tail fiber proteins gp37 and gp38 were only distantly related to T4, but shared up to 83% amino acid identity to other T6-like phages, suggesting lateral gene transfer. A greater degree of variability was seen between JS98 and T4 over DNA replication and DNA transaction genes. While most of these genes came in the same order and shared up to 76% protein sequence identity, a few rearrangements, insertions, and replacements of genes were observed. Many putative gene insertions in the DNA replication module of T4 were flanked by intron-related endonuclease genes, suggesting mobile DNA elements. A hotspot of genome diversification was located downstream of the DNA polymerase gene 43 and the DNA binding gene 32. Comparative genomics of 100-kb genome sequence revealed that T4-like phages diversify more by the accumulation of point mutations and occasional gene duplication events than by modular exchanges.

  19. Electrostatic interactions in protein adsorption probed by comparing lysozyme and succinylated lysozyme

    NARCIS (Netherlands)

    Veen, van der M.; Norde, W.; Cohen Stuart, M.A.

    2004-01-01

    The influence of electrostatic interactions on protein adsorption was studied by comparing the adsorption of lysozyme and succinylated lysozyme at silica surfaces. The succinylation affects the charge of the protein, but also the stability. Although changes in stability can have an influence on

  20. Seymour Benzer and T4 rII

    Indian Academy of Sciences (India)

    Home; Journals; Resonance – Journal of Science Education; Volume 13; Issue 10. Seymour Benzer and T4 rII - Running the Map into the Ground. R Jayaraman. General Article Volume 13 Issue 10 October 2008 pp 898-908. Fulltext. Click here to view fulltext PDF. Permanent link:

  1. T-4G Methodology: Undergraduate Pilot Training T-37 Phase.

    Science.gov (United States)

    Woodruff, Robert R.; And Others

    The report's brief introduction describes the application of T-4G methodology to the T-37 instrument phase of undergraduate pilot training. The methodology is characterized by instruction in trainers, proficiency advancement, a highly structured syllabus, the training manager concept, early exposure to instrument training, and hands-on training.…

  2. Structure and assembly of bacteriophage T4 head

    Directory of Open Access Journals (Sweden)

    Black Lindsay W

    2010-12-01

    Full Text Available Abstract The bacteriophage T4 capsid is an elongated icosahedron, 120 nm long and 86 nm wide, and is built with three essential proteins; gp23*, which forms the hexagonal capsid lattice, gp24*, which forms pentamers at eleven of the twelve vertices, and gp20, which forms the unique dodecameric portal vertex through which DNA enters during packaging and exits during infection. The past twenty years of research has greatly elevated the understanding of phage T4 head assembly and DNA packaging. The atomic structure of gp24 has been determined. A structural model built for gp23 using its similarity to gp24 showed that the phage T4 major capsid protein has the same fold as that found in phage HK97 and several other icosahedral bacteriophages. Folding of gp23 requires the assistance of two chaperones, the E. coli chaperone GroEL and the phage coded gp23-specific chaperone, gp31. The capsid also contains two non-essential outer capsid proteins, Hoc and Soc, which decorate the capsid surface. The structure of Soc shows two capsid binding sites which, through binding to adjacent gp23 subunits, reinforce the capsid structure. Hoc and Soc have been extensively used in bipartite peptide display libraries and to display pathogen antigens including those from HIV, Neisseria meningitides, Bacillus anthracis, and FMDV. The structure of Ip1*, one of the components of the core, has been determined, which provided insights on how IPs protect T4 genome against the E. coli nucleases that degrade hydroxymethylated and glycosylated T4 DNA. Extensive mutagenesis combined with the atomic structures of the DNA packaging/terminase proteins gp16 and gp17 elucidated the ATPase and nuclease functional motifs involved in DNA translocation and headful DNA cutting. Cryo-EM structure of the T4 packaging machine showed a pentameric motor assembled with gp17 subunits on the portal vertex. Single molecule optical tweezers and fluorescence studies showed that the T4 motor packages

  3. Mobile DNA elements in T4 and related phages.

    Science.gov (United States)

    Edgell, David R; Gibb, Ewan A; Belfort, Marlene

    2010-10-28

    Mobile genetic elements are common inhabitants of virtually every genome where they can exert profound influences on genome structure and function in addition to promoting their own spread within and between genomes. Phage T4 and related phage have long served as a model system for understanding the molecular mechanisms by which a certain class of mobile DNA, homing endonucleases, promote their spread. Homing endonucleases are site-specific DNA endonucleases that initiate mobility by introducing double-strand breaks at defined positions in genomes lacking the endonuclease gene, stimulating repair and recombination pathways that mobilize the endonuclease coding region. In phage T4, homing endonucleases were first discovered as encoded within the self-splicing td, nrdB and nrdD introns of T4. Genomic data has revealed that homing endonucleases are extremely widespread in T-even-like phage, as evidenced by the astounding fact that ~11% of the T4 genome encodes homing endonuclease genes, with most of them located outside of self-splicing introns. Detailed studies of the mobile td intron and its encoded endonuclease, I-TevI, have laid the foundation for genetic, biochemical and structural aspects that regulate the mobility process, and more recently have provided insights into regulation of homing endonuclease function. Here, we summarize the current state of knowledge regarding T4-encoded homing endonucleases, with particular emphasis on the td/I-TevI model system. We also discuss recent progress in the biology of free-standing endonucleases, and present areas of future research for this fascinating class of mobile genetic elements.

  4. Mobile DNA elements in T4 and related phages

    Directory of Open Access Journals (Sweden)

    Belfort Marlene

    2010-10-01

    Full Text Available Abstract Mobile genetic elements are common inhabitants of virtually every genome where they can exert profound influences on genome structure and function in addition to promoting their own spread within and between genomes. Phage T4 and related phage have long served as a model system for understanding the molecular mechanisms by which a certain class of mobile DNA, homing endonucleases, promote their spread. Homing endonucleases are site-specific DNA endonucleases that initiate mobility by introducing double-strand breaks at defined positions in genomes lacking the endonuclease gene, stimulating repair and recombination pathways that mobilize the endonuclease coding region. In phage T4, homing endonucleases were first discovered as encoded within the self-splicing td, nrdB and nrdD introns of T4. Genomic data has revealed that homing endonucleases are extremely widespread in T-even-like phage, as evidenced by the astounding fact that ~11% of the T4 genome encodes homing endonuclease genes, with most of them located outside of self-splicing introns. Detailed studies of the mobile td intron and its encoded endonuclease, I-TevI, have laid the foundation for genetic, biochemical and structural aspects that regulate the mobility process, and more recently have provided insights into regulation of homing endonuclease function. Here, we summarize the current state of knowledge regarding T4-encoded homing endonucleases, with particular emphasis on the td/I-TevI model system. We also discuss recent progress in the biology of free-standing endonucleases, and present areas of future research for this fascinating class of mobile genetic elements.

  5. Formas armónicas con valores en un fibrado vectorial e inmersiones de variedades riemannianas

    OpenAIRE

    Llauce Santamaría, Edwin Edilberto

    2014-01-01

    El propósito de este trabajo es discutir la aplicación de la teoría de las formas armónicas con valores en un fibrado vectorial y su relación con las inmersiones en una variedad riemanniana. Sea M una variedad riemanniana y E un fibrado vectorial riemanniano sobre M, entonces podemos definir de manera natural el operador laplaciano en las formas diferenciales con valores en E y expresaremos el producto escalar ⟨θ, θ⟩, donde θ es una p-forma con valores en E, en términos de la curvatura y la d...

  6. Genomic organization and evolution of ruminant lysozyme c genes

    OpenAIRE

    IRWIN, David M

    2015-01-01

    Ruminant stomach lysozyme is a long established model of adaptive gene evolution. Evolution of stomach lysozyme function required changes in the site of expression of the lysozyme c gene and changes in the enzymatic properties of the enzyme. In ruminant mammals, these changes were associated with a change in the size of the lysozyme c gene family. The recent release of near complete genome sequences from several ruminant species allows a more complete examination of the evolution and diversif...

  7. LPS-Activated Monocytes Are Unresponsive to T4 Phage and T4-Generated Escherichia coli Lysate.

    Science.gov (United States)

    Bocian, Katarzyna; Borysowski, Jan; Zarzycki, Michał; Wierzbicki, Piotr; Kłosowska, Danuta; Weber-Dąbrowska, Beata; Korczak-Kowalska, Grażyna; Górski, Andrzej

    2016-01-01

    A growing body of data shows that bacteriophages can interact with different kinds of immune cells. The objective of this study was to investigate whether T4 bacteriophage and T4-generated Escherichia coli lysate affect functions of monocytes, the key population of immune cells involved in antibacterial immunity. To that end, we evaluated how T4 and E. coli lysate influence the expression of main costimulatory molecules including CD40, CD80 and CD86, TLR2, TLR4 on monocytes, as well as the production of IL-6 and IL-12 in cultures of peripheral blood mononuclear cells (PBMCs). Separate experiments were performed on unactivated and LPS-activated PBMCs cultures. Both studied preparations significantly increased the percentage of CD14(+)CD16(-)CD40(+) and CD14(+)CD16(-)CD80(+) monocytes in unactivated PBMCs cultures, as well as the concentration of IL-6 and IL-12 in culture supernates. However, neither purified T4 nor E. coli lysate had any significant effect on monocytes in LPS-activated PBMCs cultures. We conclude that LPS-activated monocytes are unresponsive to phages and products of phage-induced lysis of bacteria. This study is highly relevant to phage therapy because it suggests that in patients with infections caused by Gram-negative bacteria the administration of phage preparations to patients and lysis of bacteria by phages are not likely to overly stimulate monocytes.

  8. LPS-activated monocytes are unresponsive to T4 phage and T4-generated Escherichia coli lysate

    Directory of Open Access Journals (Sweden)

    Katarzyna Bocian

    2016-08-01

    Full Text Available A growing body of data shows that bacteriophages can interact with different kinds of immune cells. The objective of this study was to investigate whether T4 bacteriophage and T4-generated Escherichia coli lysate affect functions of monocytes, the key population of immune cells involved in antibacterial immunity. To that end we evaluated how T4 and E. coli lysate influence the expression of main costimulatory molecules including CD40, CD80 and CD86, TLR2, TLR4 on monocytes, as well as the production of IL-6 and IL-12 in cultures of peripheral blood mononuclear cells (PBMCs. Separate experiments were performed on unactivated and LPS-activated PBMCs cultures. Both studied preparations significantly increased the percentage of CD14+CD16-CD40+ and CD14+CD16-CD80+ monocytes in unactivated PBMCs cultures, as well as the concentration of IL-6 and IL-12 in culture supernates. However, neither purified T4 nor E. coli lysate had any significant effect on monocytes in LPS-activated PBMCs cultures. We conclude that LPS-activated monocytes are unresponsive to phages and products of phage-induced lysis of bacteria. This study is highly relevant to phage therapy because it suggests that in patients with infections caused by Gram-negative bacteria the administration of phage preparations to patients and lysis of bacteria by phages are not likely to overly stimulate monocytes.

  9. Crystallization of lysozyme from lysozyme - ovalbumin mixtures: Separation potential and crystal growth kinetics

    Science.gov (United States)

    Maosoongnern, Somchai; Flood, Chalongsri; Flood, Adrian E.; Ulrich, Joachim

    2017-07-01

    Lysozyme was successfully separated from mixtures of lysozyme and ovalbumin by crystallization. The purity of the lysozyme product is more than 98%, the remaining activity is greater than 97%, and the yields of the crystal products were greater than 80%. The experimental conditions used were varied to study the effect of the operating parameters on the growth kinetics of lysozyme crystal and the separation ability of the process. The growth rates of lysozyme are second order with respect to the relative supersaturation. Therefore the growth kinetics of the crystallization process is controlled by the surface integration mechanism. The calculated growth rate constants were 5.4×10-6 cm/h and 2.5×10-6 cm/h for the crystallization process at 20 °C and 10 °C, respectively. There is no significant effect of the ovalbumin impurity up to the concentration of 67.5% ovalbumin (based on total protein) on the growth kinetics of lysozyme. Changing the NaCl concentration from 4% to 3% had no effect on the growth kinetics of lysozyme, although this does change the solubility and therefore the yield. The calculated activation energy was 53.08 kJ/mol which supports the hypothesis that the crystallization process is controlled by the surface integration mechanism.

  10. Microarray analysis of gene expression during bacteriophage T4 infection.

    Science.gov (United States)

    Luke, Kimberly; Radek, Agnes; Liu, XiuPing; Campbell, John; Uzan, Marc; Haselkorn, Robert; Kogan, Yakov

    2002-08-01

    Genomic microarrays were used to examine the complex temporal program of gene expression exhibited by bacteriophage T4 during the course of development. The microarray data confirm the existence of distinct early, middle, and late transcriptional classes during the bacteriophage replicative cycle. This approach allows assignment of previously uncharacterized genes to specific temporal classes. The genomic expression data verify many promoter assignments and predict the existence of previously unidentified promoters.

  11. Genetical studies with radiation sensitive mutants of bacteriophage T4

    International Nuclear Information System (INIS)

    Boyle, J.M.

    This thesis is concerned with a study of the properties of radiation sensitive mutants of bacteriophage T4. An introduction is presented which reviews the current concepts of radiation repair mechanisms, and their relationship to genetic recombination in bacteria and phage T4. Following the description of materials and methods, the results section is presented in three parts. Part I deals with the isolation and purification of a new radiation sensitive mutant of T4, called y. The properties of y are compared with those of two previously isolated radiation sensitive mutants, v 1 and x. Part II describes the properties of y under three complex radiobiological conditions, namely multiplicity reactivation, depression of viability and the Luria-Latarjet experiment. In Part III, complementation and mapping data are presented, which show that y, x, and v 1 are mutants of separate cistrons and unlinked in mapping experiments. The wild allele in each case is dominant. The sizes of cistrons y, x, and v are 3.2, 6.8, and 1.6% of the total chromosome respectively. The properties of recombinants v 1 x, v 1 y, and xy are described. In the discussion the possible mode of action of y is discussed. (author)

  12. Thermophysical properties of lysozyme (protein) solutions

    Science.gov (United States)

    Liu, Jiaching; Yang, Wen-Jei

    1992-01-01

    Thermophysical properties of protein solutions composed of the lysozyme crystals with a 0.1 M sodium acetate and 5 percent NaCl solution as the buffer (pH = 4.0) are determined. The properties being measured include specific heat, thermal conductivity, dynamic viscosity, and surface tension. The protein concentrations are varied. Thermal diffusivity is calculated using the measured results. The purpose of the research is to measure thermophysical properties of lysozyme solutions which would serve as the data bank for controlling and modeling the crystal growth process on earth as well as in space.

  13. Lysozyme Resistance in Streptococcus suis Is Highly Variable and Multifactorial

    Science.gov (United States)

    Wichgers Schreur, Paul J.; van Weeghel, Christian; Rebel, Johanna M. J.; Smits, Mari A.; van Putten, Jos P. M.; Smith, Hilde E.

    2012-01-01

    Background Streptococcus suis is an important infectious agent for pigs and occasionally for humans. The host innate immune system plays a key role in preventing and eliminating S. suis infections. One important constituent of the innate immune system is the protein lysozyme, which is present in a variety of body fluids and immune cells. Lysozyme acts as a peptidoglycan degrading enzyme causing bacterial lysis. Several pathogens have developed mechanisms to evade lysozyme-mediated killing. In the present study we compared the lysozyme sensitivity of various S. suis isolates and investigated the molecular basis of lysozyme resistance for this pathogen. Results The lysozyme minimal inhibitory concentrations of a wide panel of S. suis isolates varied between 0.3 to 10 mg/ml. By inactivating the oatA gene in a serotype 2 and a serotype 9 strain, we showed that OatA-mediated peptidoglycan modification partly contributes to lysozyme resistance. Furthermore, inactivation of the murMN operon provided evidence that additional peptidoglycan crosslinking is not involved in lysozyme resistance in S. suis. Besides a targeted approach, we also used an unbiased approach for identifying factors involved in lysozyme resistance. Based on whole genome comparisons of a lysozyme sensitive strain and selected lysozyme resistant derivatives, we detected several single nucleotide polymorphisms (SNPs) that were correlated with the lysozyme resistance trait. Two SNPs caused defects in protein expression of an autolysin and a capsule sugar transferase. Analysis of specific isogenic mutants, confirmed the involvement of autolysin activity and capsule structures in lysozyme resistance of S. suis. Conclusions This study shows that lysozyme resistance levels are highly variable among S. suis isolates and serotypes. Furthermore, the results show that lysozyme resistance in S. suis can involve different mechanisms including OatA-mediated peptidolycan modification, autolysin activity and capsule

  14. Backbone 1H, 13C, and 15N resonance assignments for lysozyme from bacteriophage lambda

    OpenAIRE

    Di Paolo, Alexandre; Duval, Val?rie; Matagne, Andr?; Redfield, Christina

    2010-01-01

    Lysozyme from lambda bacteriophage (? lysozyme) is an 18 kDa globular protein displaying some of the structural features common to all lysozymes; in particular, ? lysozyme consists of two structural domains connected by a helix, and has its catalytic residues located at the interface between these two domains. An interesting feature of ? lysozyme, when compared to the well-characterised hen egg-white lysozyme, is its lack of disulfide bridges; this makes ? lysozyme an interesting system for s...

  15. Oblique incidence of semi-guided waves on rectangular slab waveguide discontinuities: A vectorial QUEP solver

    NARCIS (Netherlands)

    Hammer, Manfred

    2015-01-01

    The incidence of thin-film-guided, in-plane unguided waves at oblique angles on straight discontinuities of dielectric slab waveguides, an early problem of integrated optics, is being re-considered. The 3-D frequency domain Maxwell equations reduce to a parametrized inhomogeneous vectorial problem

  16. Assessment of vectorial total variation penalties on realistic dual-energy CT data.

    Science.gov (United States)

    Rigie, David S; Sanchez, Adrian A; La Rivière, Patrick J

    2017-04-21

    Vectorial extensions of total variation have recently been developed for regularizing the reconstruction and denoising of multi-channel images, such as those arising in spectral computed tomography. Early studies have focused mainly on simulated, piecewise-constant images whose structure may favor total-variation penalties. In the current manuscript, we apply vectorial total variation to real dual-energy CT data of a whole turkey in order to determine if the same benefits can be observed in more complex images with anatomically realistic textures. We consider the total nuclear variation ([Formula: see text]) as well as another vectorial total variation based on the Frobenius norm ([Formula: see text]) and standard channel-by-channel total variation ([Formula: see text]). We performed a series of 3D TV denoising experiments comparing the three TV variants across a wide range of smoothness parameter settings, optimizing each regularizer according to a very-high-dose 'ground truth' image. Consistent with the simulation studies, we find that both vectorial TV variants achieve a lower error than the channel-by-channel TV and are better able to suppress noise while preserving actual image features. In this real data study, the advantages are subtler than in the previous simulation study, although the [Formula: see text] penalty is found to have clear advantages over either [Formula: see text] or [Formula: see text] when comparing material images formed from linear combinations of the denoised energy images.

  17. Dynamical breakdown of chiral symmetry in vectorial theories: QED and QCD

    International Nuclear Information System (INIS)

    Garcia, J.C.M.

    1987-01-01

    Using a variational approach for the Effective Potential for composite operators we dicuss the dynamical breakdown of chiral symmetry in two vectorial theories: Quantum Electrodynamics (QED) and Quantum Chromodynamics (QCD). We study the energetic aspects of the problem calculating the Effective Potential with the asymptotic nonperturbative solutions of the Schwinger-Dyson equation for the fermion selfenergy. (author) [pt

  18. Solvent freeze out crystallization of lysozyme from a lysozyme-ovalbumin mixture

    Energy Technology Data Exchange (ETDEWEB)

    Diaz Borbon, V.; Ulrich, J. [Martin-Luther-Universitaet Halle-Wittenberg, Zentrum fuer Ingenieurwissenschaft, Verfahrenstechnik/TVT, 06099 Halle Saale (Germany)

    2012-05-15

    Hen egg white lysozyme (HEWL) crystallization conditions from an ovalbumin-lysozyme mixture were found by screening tests and further located in pseudo-phase diagrams. This information was used to set up the initial conditions for the solvent freeze out (SFO) process. The process uses the freezing of ice to create the supersaturation for the proteins to crystallize out of the solution. The crystallization of HEWL (15 mg/mL) out of a lysozyme-ovalbumin mixture (1.7 mg/mL) is carried out by SFO. Under the reported conditions, a crystallization yield of 69 % was obtained. A mean crystal size of 77.8 {mu}m was enhanced in a crystallization time of 15.1 h. The lysozyme nature of the crystals is proven by SDS PAGE and enzymatic activity tests. (copyright 2012 WILEY-VCH Verlag GmbH and Co. KGaA, Weinheim) (orig.)

  19. 1H NMR studies of human lysozyme: Spectral assignment and comparison with hen lysozyme

    International Nuclear Information System (INIS)

    Redfield, C.; Dobson, C.M.

    1990-01-01

    Complete main-chain (NH and αCH) 1 H NMR assignments are reported for the 130 residues of human lysozyme, along with extensive assignments for side-chain protons. Analysis of 2-D NOESY experiments shows that the regions of secondary structure for human lysozyme in solution are essentially identical with those found previously in a similar study of hen lysozyme and are in close accord with the structure of the protein reported previously from x-ray diffraction studies in the crystalline state. Comparison of the chemical shifts, spin-spin coupling constants, and hydrogen exchange behavior are also consistent with closely similar structures for the two proteins in solution. In a number of cases specific differences in the NMR parameters between hen and human lysozymes can be correlated with specific differences observed in the crystal structures

  20. T4 genes in the marine ecosystem: studies of the T4-like cyanophages and their role in marine ecology.

    Science.gov (United States)

    Clokie, Martha R J; Millard, Andrew D; Mann, Nicholas H

    2010-10-28

    From genomic sequencing it has become apparent that the marine cyanomyoviruses capable of infecting strains of unicellular cyanobacteria assigned to the genera Synechococcus and Prochlorococcus are not only morphologically similar to T4, but are also genetically related, typically sharing some 40-48 genes. The large majority of these common genes are the same in all marine cyanomyoviruses so far characterized. Given the fundamental physiological differences between marine unicellular cyanobacteria and heterotrophic hosts of T4-like phages it is not surprising that the study of cyanomyoviruses has revealed novel and fascinating facets of the phage-host relationship. One of the most interesting features of the marine cyanomyoviruses is their possession of a number of genes that are clearly of host origin such as those involved in photosynthesis, like the psbA gene that encodes a core component of the photosystem II reaction centre. Other host-derived genes encode enzymes involved in carbon metabolism, phosphate acquisition and ppGpp metabolism. The impact of these host-derived genes on phage fitness has still largely to be assessed and represents one of the most important topics in the study of this group of T4-like phages in the laboratory. However, these phages are also of considerable environmental significance by virtue of their impact on key contributors to oceanic primary production and the true extent and nature of this impact has still to be accurately assessed.

  1. T4 genes in the marine ecosystem: studies of the T4-like cyanophages and their role in marine ecology

    Directory of Open Access Journals (Sweden)

    Millard Andrew D

    2010-10-01

    Full Text Available Abstract From genomic sequencing it has become apparent that the marine cyanomyoviruses capable of infecting strains of unicellular cyanobacteria assigned to the genera Synechococcus and Prochlorococcus are not only morphologically similar to T4, but are also genetically related, typically sharing some 40-48 genes. The large majority of these common genes are the same in all marine cyanomyoviruses so far characterized. Given the fundamental physiological differences between marine unicellular cyanobacteria and heterotrophic hosts of T4-like phages it is not surprising that the study of cyanomyoviruses has revealed novel and fascinating facets of the phage-host relationship. One of the most interesting features of the marine cyanomyoviruses is their possession of a number of genes that are clearly of host origin such as those involved in photosynthesis, like the psbA gene that encodes a core component of the photosystem II reaction centre. Other host-derived genes encode enzymes involved in carbon metabolism, phosphate acquisition and ppGpp metabolism. The impact of these host-derived genes on phage fitness has still largely to be assessed and represents one of the most important topics in the study of this group of T4-like phages in the laboratory. However, these phages are also of considerable environmental significance by virtue of their impact on key contributors to oceanic primary production and the true extent and nature of this impact has still to be accurately assessed.

  2. Interaction mechanism between 4-aminoantipyrine and the enzyme lysozyme

    International Nuclear Information System (INIS)

    Teng Yue; Ji Fanying; Li Chao; Yu Zehua; Liu Rutao

    2011-01-01

    4-Aminoantipyrine (AAP) is scarcely administered as a kind of analgesic drug because of the side effect. The residue of AAP in the environment poses a potential threat to human health. To evaluate the toxicity of AAP at the protein level, the effects of AAP on lysozyme were investigated using spectroscopic and molecular docking methods. Addition of AAP effectively quenched the intrinsic fluorescence of lysozyme. Static quenching of lysozyme by AAP revealed the formation of complex. After the inner filter effect was eliminated, the number of binding sites, the binding constant and the thermodynamic parameters were measured, and indicated that AAP can spontaneously bind with lysozyme through hydrophobic interactions with one binding site. Molecular docking results revealed that AAP bound into the enzyme active site and interacted with the Trp 62 and Trp 63 residues of lysozyme, which illustrated that the lysozyme activity was inhibited by AAP, in accordance with the results of the lysozyme activity experiment. Furthermore, the binding of AAP can result in demonstrable change of the conformation of lysozyme. This work is helpful for clarifying the molecular toxic mechanism of AAP in vivo. - Highlights: → This work established the binding mode of AAP with lysozyme in molecular level. → Mechanism was explored by multiple spectroscopic and molecular docking methods. → AAP can inhibit lysozyme activity and induce conformational changes in lysozyme.

  3. Human lysozyme has fungicidal activity against nasal fungi.

    Science.gov (United States)

    Woods, Charmaine M; Hooper, David N; Ooi, Eng H; Tan, Lor-Wai; Carney, A Simon

    2011-01-01

    The cationic antimicrobial peptide lysozyme is the most prevalent innate immune protein in nasal secretions but there is a paucity of research regarding its role in paranasal sinus disease. Lysozyme is generally regarded as an antibacterial agent; however, some data suggest activity toward yeast. This study was designed to determine if lysozyme displays fungicidal activity toward fungi commonly identified in patients with chronic rhinosinusitis (CRS) or fungal sinusitis. Using a colony-forming unit assay the fungicidal activity of lysozyme (0, 0.5, 5, and 50 micromolar; 0- to 7-hour treatment) was tested against strains of Aspergillus fumigatus, the yeast Candida albicans, and other fungi commonly identified in mucin of patients with CRS. Fungi cultured directly from the mucin of two CRS patients were also tested to determine if they were resistant to the fungicidal activity of lysozyme. The fungicidal effect of lysozyme was both concentration and time dependent. After 7-hour treatment lysozyme (5 micromolar) had >80% fungicidal activity against A. fumigatus, Penicillium sp., Acremonium sp., C. albicans, and Candida parapsilosis. The fungicidal activity of lysozyme toward Alternaria alternata could not be determined. Lysozyme was also fungicidal toward the clinical isolates A. fumigatus and Aspergillus terreus cultured from the mucin of CRS patients. Lysozyme displays fungicidal activity toward many fungi commonly identified in patients with CRS, as well as clinical fungi isolates cultured from the mucin of CRS patients. Additional studies are required to determine the regulation of lysozyme in CRS.

  4. Vectorial field propagation through high NA objectives using polarized Gaussian beam decomposition

    Science.gov (United States)

    Worku, N.; Gross, H.

    2017-08-01

    Scalar fields can be propagated through non-paraxial systems using the Gaussian beam decomposition method. However, for high NA objectives, this scalar treatment is not sufficient to correctly describe the electromagnetic fields inside the focal region due to high ray bendings, which result in a significant change in the polarization state of light. To model these vectorial effects, the Gaussian beam decomposition method has to be extended to include the polarization state of light. In this work we have combined it with the three dimensional polarization ray tracing in order to propagate vectorial fields through high NA optical systems. During the Gaussian beam decomposition, the polarization state of each individual beamlet is represented by a polarization vector [𝐸𝑥, 𝐸𝑦, 𝐸𝑧 ] associated with its central ray. Individual Gaussian beams are then propagated through the system using the complex ray tracing method. The effect of the optical system on the polarization state of each beam is computed by applying the three dimensional polarization ray tracing of the corresponding central rays. Finally the individual beams are superposed coherently in the plane of interest resulting in the complete vectorial field. We apply the proposed method to compute the vectorial field inside the focal region of a high NA microscope objective lens and compare our result to the vectorial Debye integral method. We find that the Gaussian beam decomposition method overcomes serious limitations of algorithms relying on Fourier transforms, i.e. the field sampling requirements are less critical in high NA focusing and in the presence of large aberrations. However, sharp edges in the amplitude profile are difficult to handle as they should be replaced with smooth Gaussian edge.

  5. Hydrophobic nano-carrier for lysozyme adsorption

    Indian Academy of Sciences (India)

    adsorption–desorption cycles by using same nanoparticles. 2.2e Assay of lysozyme activity: A simple spectrophoto- metric method from Shugar [38] was used for the activ- ity tests. For this, Micrococcus lysodeikticus suspension was prepared by mixing 9 g of M. lysodeikticus with 30 ml of carbonate buffer (pH 9, 100 mM).

  6. Regular arrangement of periodates bound to lysozyme

    Czech Academy of Sciences Publication Activity Database

    Ondráček, Jan; Weiss, M.S.; Brynda, Jiří; Fiala, J.; Jursík, F.; Řezáčová, Pavlína; Jenner, L.B.; Sedláček, Juraj

    2005-01-01

    Roč. 61, Pt9 (2005), s. 1181-1189 ISSN 0907-4449 Institutional research plan: CEZ:AV0Z50520514 Keywords : hen egg white lysozyme * periodate * epitaxy Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.401, year: 2005

  7. Aptamer-Based Electrochemical Sensing of Lysozyme

    Directory of Open Access Journals (Sweden)

    Alina Vasilescu

    2016-06-01

    Full Text Available Protein analysis and quantification are required daily by thousands of laboratories worldwide for activities ranging from protein characterization to clinical diagnostics. Multiple factors have to be considered when selecting the best detection and quantification assay, including the amount of protein available, its concentration, the presence of interfering molecules, as well as costs and rapidity. This is also the case for lysozyme, a 14.3-kDa protein ubiquitously present in many organisms, that has been identified with a variety of functions: antibacterial activity, a biomarker of several serious medical conditions, a potential allergen in foods or a model of amyloid-type protein aggregation. Since the design of the first lysozyme aptamer in 2001, lysozyme became one of the most intensively-investigated biological target analytes for the design of novel biosensing concepts, particularly with regards to electrochemical aptasensors. In this review, we discuss the state of the art of aptamer-based electrochemical sensing of lysozyme, with emphasis on sensing in serum and real samples.

  8. Lysozyme Uptake by Oxidized Starch Polymer Microgels

    NARCIS (Netherlands)

    Li, Yuan; de Vries, Renko; Kleijn, Mieke; Slaghek, Ted; Timmermans, Johan; Stuart, Martien Cohen; Norde, Willem

    2010-01-01

    With the aim of determining suitable conditions for uptake and release of globular proteins on microgels, we studied the interaction between phosphated, highly cross-linked, negatively charged oxidized potato starch polymer (OPSP) microgel particles and lysozyme from hen eggs. Our microgel shows a

  9. Lysozyme uptake by oxidized starch polymer microgels

    NARCIS (Netherlands)

    Li, Y.; Vries, de R.J.; Kleijn, J.M.; Slaghek, T.M.; Timmermans, J.; Cohen Stuart, M.A.; Norde, W.

    2010-01-01

    With the aim of determining suitable conditions for uptake and release of globular proteins on microgels, we studied the interaction between phosphated, highly cross-linked, negatively charged oxidized potato starch polymer (OPSP) microgel particles and lysozyme from hen eggs. Our microgel shows a

  10. Apoptosis in Acanthamoeba castellanii belonging to the T4 genotype.

    Science.gov (United States)

    Baig, Abdul M; Lalani, Salima; Khan, Naveed A

    2017-07-01

    Here we describe features of apoptosis in unicellular Acanthamoeba castellanii belonging to the T4 genotype. When exposed to apoptosis-inducing compounds such as doxorubicin, A. castellanii trophozoites exhibited cell shrinkage and membrane blebbing as observed microscopically, DNA fragmentation using agarose gel electrophoresis, and phosphatidylserine (PS) externalization using annexin V immunostaining. Overall, these findings suggest the existence of apoptosis in A. castellanii possibly mediated by intrinsic apoptotic cascade. Further research in this field could provide avenues to selectively induce apoptosis in A. castellanii by triggering intrinsic apoptotic cascade. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  11. The EFTTRA-T4 experiment on americium transmutation

    CERN Document Server

    Konings, R J M; Dassel, G; Pijlgroms, B J; Somers, J; Toscano, E

    2000-01-01

    In the EFTTRA-T4 experiment the irradiation behaviour of a target containing americium dispersed in MgAl sub 2 O sub 4 was studied. Pellets containing 10-12 wt% sup 2 sup 4 sup 1 Am were fabricated by the infiltration method. However, it was found that the americium, intended to be present as AmO sub 2 sub - sub x , formed a compound, probably AmAlO sub 3 , during sintering. The T4 target was irradiated in the High Flux Reactor (HFR) Petten from August 1996 to January 1998 (358.4 fpd's). Post-test burn-up calculations indicated that the sup 2 sup 4 sup 1 Am concentration is reduced to 4% of the initial value at the end of the irradiation. The fraction of the initial americium atoms that were fissioned is 28%. Non-destructive and destructive examinations of the target indicated that swelling of the target pellets occurred. This is attributed to accumulation of helium, produced by alpha decay of sup 2 sup 4 sup 2 Cm that occurs in the transmutation scheme of sup 2 sup 4 sup 1 Am.

  12. The Antimicrobial Peptide Lysozyme Is Induced after Multiple Trauma

    Directory of Open Access Journals (Sweden)

    Tim Klüter

    2014-01-01

    Full Text Available The antimicrobial peptide lysozyme is an important factor of innate immunity and exerts high potential of antibacterial activity. In the present study we evaluated the lysozyme expression in serum of multiple injured patients and subsequently analyzed their possible sources and signaling pathways. Expression of lysozyme was examined in blood samples of multiple trauma patients from the day of trauma until 14 days after trauma by ELISA. To investigate major sources of lysozyme, its expression and regulation in serum samples, different blood cells, and tissue samples were analysed by ELISA and real-time PCR. Neutrophils and hepatocytes were stimulated with cytokines and supernatant of Staphylococcus aureus. The present study demonstrates the induction and release of lysozyme in serum of multiple injured patients. The highest lysozyme expression of all tested cells and tissues was detected in neutrophils. Stimulation with trauma-related factors such as interleukin-6 and S. aureus induced lysozyme expression. Liver tissue samples of patients without trauma show little lysozyme expression compared to neutrophils. After stimulation with bacterial fragments, lysozyme expression of hepatocytes is upregulated significantly. Toll-like receptor 2, a classic receptor of Gram-positive bacterial protein, was detected as a possible target for lysozyme induction.

  13. Lysozyme Photochemistry as a Function of Temperature. The Protective Effect of Nanoparticles on Lysozyme Photostability.

    Science.gov (United States)

    Oliveira Silva, Catarina; Petersen, Steffen B; Pinto Reis, Catarina; Rijo, Patrícia; Molpeceres, Jesús; Vorum, Henrik; Neves-Petersen, Maria Teresa

    2015-01-01

    The presence of aromatic residues and their close spatial proximity to disulphide bridges makes hen egg white lysozyme labile to UV excitation. UVB induced photo-oxidation of tryptophan and tyrosine residues leads to photochemical products, such as, kynurenine, N-formylkynurenine and dityrosine and to the disruption of disulphide bridges in proteins. We here report that lysozyme UV induced photochemistry is modulated by temperature, excitation power, illumination time, excitation wavelength and by the presence of plasmonic quencher surfaces, such as gold, and by the presence of natural fluorescence quenchers, such as hyaluronic acid and oleic acid. We show evidence that the photo-oxidation effects triggered by 295 nm at 20°C are reversible and non-reversible at 10°C, 25°C and 30°C. This paper provides evidence that the 295 nm damage threshold of lysozyme lies between 0.1 μW and 0.3 μW. Protein conformational changes induced by temperature and UV light have been detected upon monitoring changes in the fluorescence emission spectra of lysozyme tryptophan residues and SYPRO® Orange. Lysozyme has been conjugated onto gold nanoparticles, coated with hyaluronic acid and oleic acid (HAOA). Steady state and time resolved fluorescence studies of free and conjugated lysozyme onto HAOA gold nanoparticles reveals that the presence of the polymer decreased the rate of the observed photochemical reactions and induced a preference for short fluorescence decay lifetimes. Size and surface charge of the HAOA gold nanoparticles have been determined by dynamic light scattering and zeta potential measurements. TEM analysis of the particles confirms the presence of a gold core surrounded by a HAOA matrix. We conclude that HAOA gold nanoparticles may efficiently protect lysozyme from the photochemical effects of UVB light and this nanocarrier could be potentially applied to other proteins with clinical relevance. In addition, this study confirms that the temperature plays a

  14. Lysozymes and lysozyme-like proteins from the fall armyworm, Spodoptera frugiperda.

    Science.gov (United States)

    Chapelle, Michael; Girard, Pierre-Alain; Cousserans, François; Volkoff, Nathalie-Anne; Duvic, Bernard

    2009-12-01

    Lysozyme is an important component of the insect non-specific immune response against bacteria that is characterized by its ability to break down bacterial cell-walls. By searching an EST database from the fall armyworm, Spodoptera frugiperda (Negre et al., 2006), we identified five sequences encoding proteins of the lysozyme family. The deduced protein sequences corresponded to three classical c-type lysozymes Sf-Lys1, Sf-Lys2 and Sf-Lys3, and two lysozyme-like proteins, Sf-LLP1 and Sf-LLP2. Sf-Lys1 was purified from the hemolymph of Escherichia coli-challenged S. frugiperda larvae. The mature protein had a molecular mass of 13.975 Da with an isoelectric point of 8.77 and showed 98.3% and 96.7% identity with lysozymes from Spodoptera litura and Spodoptera exigua, respectively. As the other insect lysozymes, Sf-Lys1 was active against gram positive bacteria such as Micrococcus luteus but also induced a slight permeabilization of the inner membrane of E. coli. Genes encoding these five Sf-Lys or Sf-LLPs were differentially up-regulated in three immune-competent tissues (hemocytes, fat body and gut) after challenges with non-pathogenic bacteria, E. coli and M. luteus, or entomopathogenic bacterium, Photorhabdus luminescens. Sf-Lys1 and Sf-Lys2 were mainly induced in fat body in the presence of E. coli or P. luminescens. Sf-Lys3, which had an acidic isoelectric point, was found to be the most up-regulated of all five Sf-Lys or Sf-LLPs in hemocytes and gut after challenge with P. luminescens. More molecular data are now available to investigate differences in physiological functions of these different members of the lysozyme superfamily.

  15. Quantum thetas on noncommutative T4 from embeddings into lattice

    International Nuclear Information System (INIS)

    Chang-Young, Ee; Kim, Hoil

    2007-01-01

    In this paper, we investigate the theta vector and quantum theta function over noncommutative T 4 from the embedding of RxZ 2 . Manin has constructed the quantum theta functions from the lattice embedding into vector space (x finite group). We extend Manin's construction of the quantum theta function to the embedding of vector space x lattice case. We find that the holomorphic theta vector exists only over the vector space part of the embedding, and over the lattice part we can only impose the condition for the Schwartz function. The quantum theta function built on this partial theta vector satisfies the requirement of the quantum theta function. However, two subsequent quantum translations from the embedding into the lattice part are nonadditive, contrary to the additivity of those from the vector space part

  16. Pathogenic assays of acanthamoeba belonging to the t4 genotype.

    Directory of Open Access Journals (Sweden)

    Hamed Mirjalali

    2013-12-01

    Full Text Available Acanthamoeba genus is introduced as opportunistic and cosmopolitan parasite. Monkey and wistar rat are appropriate models for experimental study on Acanthamoeba infection. In this study Acanthamoeba spp. were isolated from hot spring (HS, windows dust (WD and a corneal sample of keratitis patient (KP and their pathogenicity surveyed by in vitro and in vivo tests.Isolates of Acanthamoeba were cultivated axenically for 12 months in PYG medium. Overall, 30 wistar rats, in 6 equal groups were used for developing experimental Acanthamoeba keratitis (AK and Granulomatous Amoebic Encephalitis (GAE. The Keratitis and Granulomatous Encephalitis experiments were performed by intrastromal and intranasal inoculation of Acanthamoeba cysts, respectively. Pathogenicity of the three isolates was also evaluated by in vitro test using osmotolerance and temperature tolerance assays. Identification of genotypes were performed by PCR technique and sequencing.None of the isolates could perform AK and GAE in wistar rats, although all isolates were described as T4 genotype. Isolates obtained from KP and WD could grow only in 30 °C, but not in 37 °C and 40 °C. On the other hand, HS isolate grew in 30 °C and 37 °C but not in 40 °C. Moreover, all of isolate grew in 0.5 M mannitol but not in 1 M and 1.5 M.T4 isolates with a long-term axenic culture and different factors related to host and parasite may play role in pathogenicity of these free-living amoebae.

  17. Repair of psoralen plus near ultraviolet light damage in bacteriophage T4

    International Nuclear Information System (INIS)

    Zerler, B.R.; Wallace, S.S.

    1979-01-01

    The sensitivity of bacteriophage T4 to psoralen plus NUV was investigated using a series of T4 repair defective mutants. The recombinational repair deficient mutants T4x, T4y and T4w were more sensitive than wild-type while T4v, an endonuclease V mutant, exhibited the same sensitivity as wild-type. However, endonuclease V appeared to initiate abortive repair in the absence of the x and presumably y gene products. Host repair gene products were shown not to be involved in the repair of psoralen plus NUV damages in T4. (author)

  18. Preparation of lysozyme molecularly imprinted polymers and purification of lysozyme from egg white.

    Science.gov (United States)

    Wang, Xuejiao; Dong, Shaohua; Bai, Quan

    2014-06-01

    Molecular imprinting as a promising and facile separation technique has received much attention because of its high selectivity for target molecules. In this study, lysozyme molecularly imprinted polymers (Lys-MIPs) were successfully prepared by the entrapment method with lysozyme as the template molecule, acrylamide as the functional monomer and N,N-methylenebisacrylamide as the cross-linker. The removal of the template lysozyme from the molecularly imprinted polymers was investigated in detail by two methods. The synthesized Lys-MIPs were characterized by scanning electron microscopy and Fourier transform-infrared, and the adsorption capacity, selectivity and reproducibility of the Lys-MIPs were also evaluated. The maximum adsorption capacity reached 94.8 mg/g, which is twice that of nonmolecularly imprinted polymers, and satisfactory selectivity and reproducibility were achieved. Using the Lys-MIP column, lysozyme could be separated completely from egg white, with purity close to 100% and mass recovery of 98.2%. This illustrated that the synthesized Lys-MIPs had high specific recognition and selectivity to the template lysozyme when they were applied to a mixture of protein standards and a real sample. Copyright © 2014 John Wiley & Sons, Ltd.

  19. Vectorial near-field imaging of a GaN based photonic crystal cavity

    International Nuclear Information System (INIS)

    La China, F.; Intonti, F.; Caselli, N.; Lotti, F.; Vinattieri, A.; Gurioli, M.; Vico Triviño, N.; Carlin, J.-F.; Butté, R.; Grandjean, N.

    2015-01-01

    We report a full optical deep sub-wavelength imaging of the vectorial components of the electric local density of states for the confined modes of a modified GaN L3 photonic crystal nanocavity. The mode mapping is obtained with a scanning near-field optical microscope operating in a resonant forward scattering configuration, allowing the vectorial characterization of optical passive samples. The optical modes of the investigated cavity emerge as Fano resonances and can be probed without the need of embedded light emitters or evanescent light coupling into the nanocavity. The experimental maps, independently measured in the two in-plane polarizations, turn out to be in excellent agreement with numerical predictions

  20. Vectorial laws of refraction and reflection using the cross product and dot product.

    Science.gov (United States)

    Tkaczyk, Eric R

    2012-03-01

    We demonstrate that published vectorial laws of reflection and refraction of light based solely on the cross product do not, in general, uniquely determine the direction of the reflected and refracted waves without additional information. This is because the cross product does not have a unique inverse operation, which is explained in this Letter in linear algebra terms. However, a vector is in fact uniquely determined if both the cross product (vector product) and dot product (scalar product) with a known vector are specified, which can be written as a single equation with a left-invertible matrix. It is thus possible to amend the vectorial laws of reflection and refraction to incorporate both the cross and dot products for a complete specification with unique solution. This enables highly efficient, unambiguous computation of reflected and refracted wave vectors from the incident wave and surface normal. © 2012 Optical Society of America

  1. Calculation of vectorial three-dimensional transfer functions in large-angle focusing systems.

    Science.gov (United States)

    Schönle, Andreas; Hell, Stefan W

    2002-10-01

    The optical transfer function (OTF) is used in describing imaging systems in the Fourier domain. So far the calculation of the OTF of a large-aperture imaging system has been difficult because the vectorial nature of light breaks the cylindrical symmetry of the pupil function. We derive a simple line integral solution for calculating the vectorial three-dimensional OTF. We further extend this approach to imaging through a planar interface of two media with mismatched refractive indices. In general, our formalism allows for calculation of the Fourier transform of any product of two arbitrary vector components of the electromagnetic field. Arbitrary phase or amplitude modifications of the pupil function can be taken into account.

  2. Interaction of magnetic nanoparticles with lysozyme amyloid fibrils

    International Nuclear Information System (INIS)

    Gdovinová, Veronika; Tomašovičová, Natália; Batko, Ivan; Batková, Marianna; Balejčíková, Lucia; Garamus, Vasyl M.; Petrenko, Viktor I.; Avdeev, Mikhail V.; Kopčanský, Peter

    2017-01-01

    This work is devoted to the structural study of complex solutions of magnetic nanoparticles with lysozyme amyloid fibrils due to possible ordering of such system by applying the external magnetic field. The interaction of magnetic nanoparticles with amyloid fibrils has been followed by atomic force microscopy and small-angle X-ray scattering. It has been observed that magnetic nanoparticles (MNPs) adsorb to lysozyme amyloid fibrils. It was found that MNPs alter amyloids structures, namely the diameter of lysozyme amyloid fibrils is increased whereas the length of fibrils is decreased. In the same time MNPs do not change the helical pitch significantly. - Highlights: • Solution of MNPs with lysozyme amyloid fibrils was characterized by AFM and SAXS. • MNPs adsorb to lysozyme amyloid fibrils. • Diameter and size of lysozyme amyloid fibrils change due to doping with MNPs.

  3. Recognition of lysozyme using surface imprinted bacterial cellulose nanofibers.

    Science.gov (United States)

    Saylan, Yeşeren; Tamahkar, Emel; Denizli, Adil

    2017-11-01

    Here, we developed the lysozyme imprinted bacterial cellulose (Lyz-MIP/BC) nanofibers via the surface imprinting strategy that was designed to recognize lysozyme. This study includes the molecular imprinting method onto the surface of bacterial cellulose nanofibers in the presence of lysozyme by metal ion coordination, as well as further characterizations methods FTIR, SEM and contact angle measurements. The maximum lysozyme adsorption capacity of Lyz-MIP/BC nanofibers was found to be 71 mg/g. The Lyz-MIP/BC nanofibers showed high selectivity for lysozyme towards bovine serum albumin and cytochrome c. Overall, the Lyz-MIP/BC nanofibers hold great potential for lysozyme recognition due to the high binding capacity, significant selectivity and excellent reusability.

  4. Interaction of magnetic nanoparticles with lysozyme amyloid fibrils

    Energy Technology Data Exchange (ETDEWEB)

    Gdovinová, Veronika [Institute of Experimental Physics SAS, Watsonova 47, 040 01 Košice (Slovakia); Frank Laboratory of Neutron Physics, Joint Institute for Nuclear Research, Joliot-Curie 6, 141980 Dubna, Moscow Region (Russian Federation); Tomašovičová, Natália, E-mail: nhudak@saske.sk [Institute of Experimental Physics SAS, Watsonova 47, 040 01 Košice (Slovakia); Frank Laboratory of Neutron Physics, Joint Institute for Nuclear Research, Joliot-Curie 6, 141980 Dubna, Moscow Region (Russian Federation); Batko, Ivan; Batková, Marianna; Balejčíková, Lucia [Institute of Experimental Physics SAS, Watsonova 47, 040 01 Košice (Slovakia); Garamus, Vasyl M. [Helmholtz-Zentrum Geesthacht: Zentrum fr Material, und Kstenforschung GmbH, Max-Plank-Strae 1, Geesthacht 216502 (Germany); Petrenko, Viktor I. [Frank Laboratory of Neutron Physics, Joint Institute for Nuclear Research, Joliot-Curie 6, 141980 Dubna, Moscow Region (Russian Federation); Physics Department, Taras Shevchenko Kyiv National University, Volodymyrska Street 64, 01601 Kyiv (Ukraine); Avdeev, Mikhail V. [Frank Laboratory of Neutron Physics, Joint Institute for Nuclear Research, Joliot-Curie 6, 141980 Dubna, Moscow Region (Russian Federation); Kopčanský, Peter [Institute of Experimental Physics SAS, Watsonova 47, 040 01 Košice (Slovakia)

    2017-06-01

    This work is devoted to the structural study of complex solutions of magnetic nanoparticles with lysozyme amyloid fibrils due to possible ordering of such system by applying the external magnetic field. The interaction of magnetic nanoparticles with amyloid fibrils has been followed by atomic force microscopy and small-angle X-ray scattering. It has been observed that magnetic nanoparticles (MNPs) adsorb to lysozyme amyloid fibrils. It was found that MNPs alter amyloids structures, namely the diameter of lysozyme amyloid fibrils is increased whereas the length of fibrils is decreased. In the same time MNPs do not change the helical pitch significantly. - Highlights: • Solution of MNPs with lysozyme amyloid fibrils was characterized by AFM and SAXS. • MNPs adsorb to lysozyme amyloid fibrils. • Diameter and size of lysozyme amyloid fibrils change due to doping with MNPs.

  5. Assessment of vectorial total variation penalties on realistic dual-energy CT data

    Science.gov (United States)

    Rigie, David S.; Sanchez, Adrian A.; La Rivière, Patrick J.

    2017-04-01

    Vectorial extensions of total variation have recently been developed for regularizing the reconstruction and denoising of multi-channel images, such as those arising in spectral computed tomography. Early studies have focused mainly on simulated, piecewise-constant images whose structure may favor total-variation penalties. In the current manuscript, we apply vectorial total variation to real dual-energy CT data of a whole turkey in order to determine if the same benefits can be observed in more complex images with anatomically realistic textures. We consider the total nuclear variation (\\text{T}{{\\text{V}}\\text{N}} ) as well as another vectorial total variation based on the Frobenius norm (\\text{T}{{\\text{V}}\\text{F}} ) and standard channel-by-channel total variation (\\text{T}{{\\text{V}}\\text{S}} ). We performed a series of 3D TV denoising experiments comparing the three TV variants across a wide range of smoothness parameter settings, optimizing each regularizer according to a very-high-dose ‘ground truth’ image. Consistent with the simulation studies, we find that both vectorial TV variants achieve a lower error than the channel-by-channel TV and are better able to suppress noise while preserving actual image features. In this real data study, the advantages are subtler than in the previous simulation study, although the \\text{T}{{\\text{V}}\\text{N}} penalty is found to have clear advantages over either \\text{T}{{\\text{V}}\\text{S}} or \\text{T}{{\\text{V}}\\text{F}} when comparing material images formed from linear combinations of the denoised energy images.

  6. Transmisión vectorial y congénita del Trypanosoma cruzi en Las Lomitas, Formosa Vectorial and congenital transmission of Trypanosoma cruzi in Las Lomitas, Formosa

    Directory of Open Access Journals (Sweden)

    Sergio Sosa-Estani

    2009-10-01

    Full Text Available La enfermedad de Chagas causada por el Trypanosoma cruzi es una causa importante de morbimortalidad en Latino América. El objetivo de este estudio fue describir las tasas de infestación en las viviendas de cuatro comunidades aborígenes de Las Lomitas (Región del Gran Chaco, Formosa, Argentina; la tasa de infección en la población infantil residente en las mismas, en donantes de sangre y en mujeres embarazadas que asistieron al Hospital de Las Lomitas y la tasa de infección congénita de niños nacidos de mujeres infectadas durante el período de estudio. La tasa de infestación en 172 viviendas evaluadas en 2006 alcanzó el 32%. La prevalencia de infección en 445 personas fue de 17.5% y en menores de 5 años de edad fue 8.6%. La tasa de infección en donantes de sangre alcanzó a 18.6% y en mujeres embarazadas fue 29.1%. La tasa de infección considerada congénita en 47 niños nacidos de mujeres infectadas residentes en viviendas bajo vigilancia fue de 17.0%. El estudio mostró, al momento de su inicio, índices compatibles con transmisión vectorial activa. Después del control vectorial con insecticidas, la tasa de infestación se redujo a 3.3%. El sistema de salud local incorporó procedimientos de prevención primaria y secundaria para evitar nuevos casos e instaurar el tratamiento de la población infectada.Chagas disease, caused by Trypanosoma cruzi, is a major cause of morbidity and mortality in Latin America. The objective of this study was to describe the rate of infestation in four aboriginal communities in Las Lomitas (Great Chaco Region, Formosa, Argentina; the rate of infection in children residing in these communities, in blood donors and in pregnant women who received care at the Hospital Las Lomitas, as well as the rate of congenital infection in children born to women infected during the study period. The rate of infestation of 172 households evaluated in 2006 reached 32%. Prevalence of infection among 445 people was 17

  7. Third party annotation gene data set of eutherian lysozyme genes

    Directory of Open Access Journals (Sweden)

    Marko Premzl

    2014-12-01

    Full Text Available The eutherian comparative genomic analysis protocol annotated most comprehensive eutherian lysozyme gene data set. Among 209 potential coding sequences, the third party annotation gene data set of eutherian lysozyme genes included 116 complete coding sequences that first described seven major gene clusters. As one new framework of future experiments, the present integrated gene annotations, phylogenetic analysis and protein molecular evolution analysis proposed new classification and nomenclature of eutherian lysozyme genes.

  8. Preferential binding of lysozyme to elastic fibres in pulmonary emphysema

    OpenAIRE

    Shteyngart, B.; Chaiwiriyakul, S.; Wong, J.; Cantor, J.

    1998-01-01

    BACKGROUND—Lysozyme is increased in inflammatory reactions and is a component of the extracellular matrix, but its possible role in lung diseases such as emphysema and interstitial fibrosis has not been investigated.
METHODS—To characterise differences in lysozyme content among normal, emphysematous, and fibrotic human lungs, tissue sections obtained from necropsy specimens were immunostained with rabbit polyclonal anti-human lysozyme antibody using the labelled streptavi...

  9. Pyroelectricity in globular protein lysozyme films

    Science.gov (United States)

    Stapleton, A.; Noor, M. R.; Haq, E. U.; Silien, C.; Soulimane, T.; Tofail, S. A. M.

    2018-03-01

    Pyroelectricity is the ability of certain non-centrosymmetric materials to generate an electric charge in response to a change in temperature and finds use in a range of applications from burglar alarms to thermal imaging. Some biological materials also exhibit pyroelectricity but the examples of the effect are limited to fibrous proteins, polypeptides, and tissues and organs of animals and plants. Here, we report pyroelectricity in polycrystalline aggregate films of lysozyme, a globular protein.

  10. Comparison of the microbicidal and muramidase activities of mouse lysozyme M and P.

    OpenAIRE

    Markart, Philipp; Faust, Nicole; Graf, Thomas; Na, Cheng-Lun; Weaver, Timothy E; Akinbi, Henry T

    2004-01-01

    Lysozyme is one of the most abundant antimicrobial proteins in the airspaces of the lung. Mice express two lysozyme genes, lysozyme M and P, but only the M enzyme is detected in abundance in lung tissues. Disruption of the lysozyme M locus significantly increased bacterial burden and mortality following intratracheal infection with a Gram-negative bacterium. Unexpectedly, significant lysozyme enzyme activity (muramidase activity) was detected in the airspaces of uninfected lysozyme M-/- mice,...

  11. Laser ablation of the protein lysozyme

    DEFF Research Database (Denmark)

    Schou, Jørgen; Canulescu, Stela; Amoruso, Salvatore

    Lysozyme is a well-known protein, which is used in food processing because of its bactericidal properties. The mass (14307 amu) is in the range in which it easily can be monitored by mass spectrometric methods, for example by MALDI (Matrix assisted laser desorption ionization). We have recently...... to a substrate as intact molecules by the violent laser impact ( up to 50 mJ/pulse) has not yet been understood. One issue is that up to 150 ng/pulse is removed by the laser, and much of the material is ejected from the target in relatively large chunks. We have explored as well the excitation mechanics by laser...... impact. Samples of pressed lysozyme prepared in the same manner as in ns-experiments have been irradiated at 527 nm with >>300-fs pulses and at a similar fluence as in ns ablation. Even though the pulse energy was much smaller, there was a considerable ablation weight loss of lysozyme from each shot...

  12. Evaluation of oriented lysozyme immobilized with monoclonal antibody

    Science.gov (United States)

    Aoyagi, Satoka; Okada, Keigo; Shigyo, Ayako; Man, Naoki; Karen, Akiya

    2008-12-01

    The orientation of a lysozyme immobilized with a monoclonal antibody was evaluated based on determination of the uppermost surface structure using time-of-flight secondary ion mass spectrometry (TOF-SIMS). Specific peaks of the oriented lysozyme immobilized with monoclonal anti-lysozyme antibody were obtained in comparison with reference samples, non-oriented immobilized lysozyme and immobilized anti-lysozyme antibody. All samples were freeze-dried before TOF-SIMS measurement, and then each sample was measured using TOF-SIMS with a bismuth cluster ion source. TOF-SIMS spectra were analyzed to select peaks specific to the oriented immobilized lysozyme as well as to identify their chemical formula and ensemble of amino acids. The possible chemical formulae of the lysozyme fragments were then investigated with an element matching program and a residue matching program. The results from TOF-SIMS spectra analysis were compared to the amino acid sequence of the lysozyme and its three-dimensional structure registered in the protein data bank. Finally, the fragment-ion-generating regions of the oriented immobilized lysozyme were determined based on the suggested residues and the three-dimensional structure.

  13. Reversible aggregation of lysozyme in a biodegradable amphiphilic multiblock copolymer.

    Science.gov (United States)

    van de Weert, Marco; van Dijkhuizen-Radersma, Riemke; Bezemer, Jeroen M; Hennink, Wim E; Crommelin, Daan J A

    2002-07-01

    Lysozyme-loaded poly(ethylene glycol terephthalate)-poly(butylene terephthalate) (PEGT/PBT) films were prepared using a water-in-oil emulsification solvent evaporation method. Infrared spectroscopic analysis of the dried films indicated the presence of non-covalent lysozyme aggregates in the polymer matrix. The use of methanol to enhance the drying rate of the films increased the relative amount of aggregates. Surprisingly, quantitative in-vitro release of fully active, non-aggregated lysozyme was observed, indicating that lysozyme forms reversible aggregates during encapsulation in PEGT/PBT films.

  14. Effect of alpha particles on bacteriophage T4Br(+)

    International Nuclear Information System (INIS)

    Leonteva, G.A.; Akoev, I.G.; Grigorev, A.E.

    1983-01-01

    The effects of heavy particle radiation, which is believed to be responsible for the high relative biological effectiveness (RBE) of space hadrons, on bacteriophages are investigated. Dry film cultures of bacteriophage T4 were irradiated with 5.3 MeV Po-210 alpha particles to doses from 5 to 60 Gray, and compared with cultures irradiated by Co-60 gamma radiation. Examination of the exponential dose-response curves for bacteriophage survival indicates an RBE of 4.68 for the alpha particles. The r-mutation frequency per 10,000 surviving phages is found to peak at 7.1 at doses between 65 and 85 Gray for gamma radiation, however it declines steadily from a level of 10.2 per 10,000 survivors with increasing dose of alpha radiation. Comparison of the mutation frequencies at the same levels of lethality and the spectra of mutations produced by the two types of radiation indicates alpha and gamma radiation to differ as well in the mechanisms of mutation production. It is concluded that the observed high RBE of space hadrons cannot be explained by the presence of high-energy particles in the secondary radiation. 13 references

  15. Partial characterization of Acanthamoeba castellanii (T4 genotype) DNase activity.

    Science.gov (United States)

    Iqbal, Junaid; Panjwani, Shamvil; Siddiqui, Ruqaiyyah; Khan, Naveed Ahmed

    2015-02-01

    The deoxyribonuclease (DNase) activities of Acanthamoeba castellanii belonging to the T4 genotype were investigated. Using zymographic assays, the DNase activities had approximate molecular masses of 25 and 35 kDa. A. castellanii DNases exhibited activity at wide-ranging temperature of up to 60 °C and at pH ranging from 4 to 9. The DNases activities were unaffected by proteinase-K treatment, divalent cations such as Ca(++), Cu(++), Mg(++), and Zn(++), or divalent cation chelating agent ethylenediaminetetraacetic acid (EDTA) or sodium dodecyl sulfate (SDS). The non-reliance on divalent cations and homology data suggests that A. castellanii DNases belong to the class of eukaryotic lysosomal DNase II but exhibit robust properties. The DNases activity in A. castellanii interfered with the genomic DNA extraction. Extraction methods involving EDTA, SDS, and proteinase-K resulted in low yield of genomic DNA. On the other hand, these methods resulted in high yield of genomic DNA from human cells suggesting the robust nature of A. castellanii DNases that are unaffected by reagents normally used in blocking eukaryotic DNases. In contrast, the use of chaotropic agent such as guanidine thiocyanate improved the yield of genomic DNA from A. castellanii cells significantly. Further purification and characterization of Acanthamoeba DNases is needed to study their non-classic distinct properties and to determine their role in the biology, cellular differentiation, cell cycle progression, and arrest of Acanthamoeba.

  16. [Friedrich Mauz: T4 assessor and military psychiatrist].

    Science.gov (United States)

    Silberzahn-Jandt, G; Schmuhl, H-W

    2012-03-01

    Friedrich Mauz is one of the medical perpetrators of the second tier whose biography is difficult to comprehend. Autobiographies from three different political systems exist - Weimar Republic, the Third Reich, and postwar Germany in which he constantly reinvented himself. While after 1933 he suddenly emphasized his participation in the civil war turmoil during the early period of the Weimar Republic and his patriotism, he then depicted himself after 1945 as an apolitical person characterized by Württemberg pietism who inwardly rejected the Nazi State but had found himself prepared to accept "all sorts of humiliating concessions." He claimed that he had always remained true to his scientific code of conduct and had distanced himself from psychiatric genetics. In point of fact, Mauz was among those exonerated in the denazification trial in 1946 and was able to pursue his career in the Federal Republic of Germany. However, if the sources are read against the grain, a different picture emerges. Mauz's career stalled in the 1930s, not because he had been politically offensive, but because his scientific work was flimsy and considered lacking originality, particularly since he had chosen constitution research and psychotherapy as his main fields of interest, which were overshadowed by research in genetic psychiatry in the 1930s. Mauz tendered his services to the Nazi policy of genetic health, served as a medical assessor in proceedings based on the "Law for the Prevention of Genetically Diseased Offspring," permitted himself to be recruited for the T4 program as a medical expert, even participated in the deliberations on a future "Law on Euthanasia," and as a consulting psychiatrist for the German Armed Forces contributed to military medicine.

  17. Influence of lysozyme complexation with purified Aldrich humic acid on lysozyme activity

    NARCIS (Netherlands)

    Li, Y.; Tan, W.F.; Wang, M.X.; Liu, F.; Weng, L.P.; Norde, W.; Koopal, L.K.

    2012-01-01

    Humic acid is an important component of dissolved organic matter and in two previous papers it has been shown that purified Aldrich humic acid (PAHA) forms strong complexes with the oppositely charged protein lysozyme (LSZ). The complexation and aggregation of enzymes with humic acids may lead to

  18. Immobilization of lysozyme on cotton fabrics; synthesis, characterication, and activity

    Science.gov (United States)

    The antimicrobial activity of lysozyme derives from the hydrolysis of the bacterial cell wall polysaccharide at the glycosidic bond that links N-acetyl-glucosamine and N-acetyl-muramic acid. Maintaining the activity of lysozyme while bound to a cellulose substrate is a goal toward developing enzyme...

  19. Co-option of bacteriophage lysozyme genes by bivalve genomes.

    Science.gov (United States)

    Ren, Qian; Wang, Chunyang; Jin, Min; Lan, Jiangfeng; Ye, Ting; Hui, Kaimin; Tan, Jingmin; Wang, Zheng; Wyckoff, Gerald J; Wang, Wen; Han, Guan-Zhu

    2017-01-01

    Eukaryotes have occasionally acquired genetic material through horizontal gene transfer (HGT). However, little is known about the evolutionary and functional significance of such acquisitions. Lysozymes are ubiquitous enzymes that degrade bacterial cell walls. Here, we provide evidence that two subclasses of bivalves (Heterodonta and Palaeoheterodonta) acquired a lysozyme gene via HGT, building on earlier findings. Phylogenetic analyses place the bivalve lysozyme genes within the clade of bacteriophage lysozyme genes, indicating that the bivalves acquired the phage-type lysozyme genes from bacteriophages, either directly or through intermediate hosts. These bivalve lysozyme genes underwent dramatic structural changes after their co-option, including intron gain and fusion with other genes. Moreover, evidence suggests that recurrent gene duplication occurred in the bivalve lysozyme genes. Finally, we show the co-opted lysozymes exhibit a capacity for antibacterial action, potentially augmenting the immune function of related bivalves. This represents an intriguing evolutionary strategy in the eukaryote-microbe arms race, in which the genetic materials of bacteriophages are co-opted by eukaryotes, and then used by eukaryotes to combat bacteria, using a shared weapon against a common enemy. © 2017 The Authors.

  20. Adsorption of lysozyme unto silica and polystyrene surfaces in ...

    African Journals Online (AJOL)

    The adsorption capacity of lysozyme (chicken egg white) from aqueous solutions unto silica and polystyrene interfaces was studied at varying lysozyme concentrations and ionic strength. The studies revealed an increase in adsorption capacity with increase in concentration and with maximum adsorption densities of 1.34 ...

  1. Analysis of lysozyme in cheese by immunocapture mass spectrometry.

    Science.gov (United States)

    Schneider, Nadine; Becker, Cord-Michael; Pischetsrieder, Monika

    2010-01-15

    The enzyme lysozyme is used as a preservative to prevent late blowing of ripened cheese, caused by Clostridium tyrobutyricum. Since the enzyme is extracted from hen egg white, lysozyme has to be declared on food product labels as a potential allergen. Here, a method is reported that combines immunocapture purification and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) analysis for the detection of lysozyme in cheese samples. Cheese extracts were treated with magnetic particles coated with a monoclonal antibody directed against lysozyme. After immunocapture purification, lysozyme was detected by MALDI-TOF-MS. The limit of detection of the assay was about 5mg/kg lysozyme in cheese. The method reliably distinguished between cheese samples which had been produced with and without lysozyme. Thus, the novel assay allows the reliable, sensitive, and specific detection of lysozyme in a food matrix. The assay could be easily adapted to other target peptides and proteins in complex food matrices and, therefore, has a broad application potential, e.g. for the analysis of allergens. 2009 Elsevier B.V. All rights reserved.

  2. Adaptive functional diversification of lysozyme in insectivorous bats.

    Science.gov (United States)

    Liu, Yang; He, Guimei; Xu, Huihui; Han, Xiuqun; Jones, Gareth; Rossiter, Stephen J; Zhang, Shuyi

    2014-11-01

    The role of gene duplication in generating new genes and novel functions is well recognized and is exemplified by the digestion-related protein lysozyme. In ruminants, duplicated chicken-type lysozymes facilitate the degradation of symbiotic bacteria in the foregut. Chicken-type lysozyme has also been reported to show chitinase-like activity, yet no study has examined the molecular evolution of lysozymes in species that specialize on eating insects. Insectivorous bats number over 900 species, and lysozyme expression in the mouths of some of these species is associated with the ingestion of insect cuticle, suggesting a chitinase role. Here, we show that chicken-type lysozyme has undergone multiple duplication events in a major family of insect-eating bats (Vespertilionidae) and that new duplicates have undergone molecular adaptation. Examination of duplicates from two insectivorous bats-Pipistrellus abramus and Scotophilus kuhlii-indicated that the new copy was highly expressed in the tongue, whereas the other one was less tissue-specific. Functional assays applied to pipistrelle lysozymes confirmed that, of the two copies, the tongue duplicate was more efficient at breaking down glycol chitin, a chitin derivative. These results suggest that the evolution of lysozymes in vespertilionid bats has likely been driven in part by natural selection for insectivory. © The Author 2014. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  3. Antimicrobial activity of lysozyme with special relevance to milk ...

    African Journals Online (AJOL)

    This review discusses the antimicrobial activity of lysozyme with special emphasis on milk's lysozyme, and attempts to shed some light on the recent advances elucidating the mechanism of its antimicrobial activity against sensitive microorganisms as well as the means used by some bacteria to resist such an activity.

  4. Salt-specific effects in lysozyme solutions

    OpenAIRE

    T. Janc; M. Kastelic; M. Bončina; V. Vlachy

    2016-01-01

    The effects of additions of low-molecular-mass salts on the properties of aqueous lysozyme solutions are examined by using the cloud-point temperature, $T_{cloud}$, measurements. Mixtures of protein, buffer, and simple salt in water are studied at pH=6.8 (phosphate buffer) and pH=4.6 (acetate buffer). We show that an addition of buffer in the amount above $I_{buffer} = 0.6$ mol dm$^{-3}$ does not affect the $T_{cloud}$ values. However, by replacing a certain amount of the buffer electrolyte b...

  5. Lysozyme, intradermal diffusion of india ink and peritoneal capillary permeability in mice.

    Science.gov (United States)

    Bianchi, C; Goi, A; Ronchini, A

    1982-04-01

    Hen egg white lysozyme injected intracutaneously into mice inhibits the intradermal diffusion of India Ink. The effect of lysozyme persists in presence of hyaluronidase. Lysozyme injected i.p. into mice inhibits acetic acid (i.p.) induced leakage of Pontamine Sky Blue into the abdominal cavity. The data suggest that lysozyme might be tried for applications outside the microbiological field.

  6. Transfer Scheme Evaluation Model for a Transportation Hub based on Vectorial Angle Cosine

    Directory of Open Access Journals (Sweden)

    Li-Ya Yao

    2014-07-01

    Full Text Available As the most important node in public transport network, efficiency of a transport hub determines the entire efficiency of the whole transport network. In order to put forward effective transfer schemes, a comprehensive evaluation index system of urban transport hubs’ transfer efficiency was built, evaluation indexes were quantified, and an evaluation model of a multi-objective decision hub transfer scheme was established based on vectorial angle cosine. Qualitative and quantitative analysis on factors affecting transfer efficiency is conducted, which discusses the passenger satisfaction, transfer coordination, transfer efficiency, smoothness, economy, etc. Thus, a new solution to transfer scheme utilization was proposed.

  7. Biological cycle and preliminary data on vectorial competence of Triatoma boliviana in laboratory conditions.

    Science.gov (United States)

    Durán, Pamela; Siñani, Edda; Depickère, Stéphanie

    2014-12-01

    With more than 140 potential vectors of Chagas disease, it is important to better know the biology and especially the vectorial capacity of the triatomine species which live in the surroundings of human dwellings. In Bolivia where 17 triatomine species are reported, the principal vector is Triatoma infestans. In some valleys of the department of La Paz where T. infestans is not present, a new species (Triatoma boliviana) was described in 2007. This species lives in a sylvatic environment not far away from the dwellings, and occasionally some individuals are found inside the houses. This study was carried out to describe the biological cycle of T. boliviana and to determine its vectorial competence. The development of a cohort of 95 nymphs of first instar (N1) was followed through nymphal instars and adult stage until death in laboratory (22°C). They were fed twice a week on an immobilized mouse. The median egg-to-adult development time was 8.4 months. The mortality by nymphal instar was lower than 7% except for N1 (67%) and N5 (18%). All nymph instars needed at least two feedings to molt (until six feedings for N5). The differentiation of a nymph into a female or a male could not be detected until the fifth instar for which the food intake was greater for a nymph developing into a female. Adults fed about once a week. The adult life span was around 400 days. The fecundity was 4.2 eggs/female/week, with a hatching rate of 50% and a hatching time of 39 days. In the same conditions, T. infestans showed a similar fecundity but a greater hatching rate and hatching time. A trial for rearing the adults at a higher temperature (26°C) showed a drastic fall in the fecundity and in the hatching rate. The vectorial competence was analyzed for fifth instars and adults by three parameters: the ability to feed on human beings, the capacity to be infected by T. cruzi and the postfeeding defecation delay. Results showed a relatively high vectorial competence: (1) insects fed

  8. Derivada parcial de un campo vectorial respecto de una variable escalar

    OpenAIRE

    Macho Ortiz, Andrés

    2014-01-01

    Derivar un campo vectorial respecto una variable escalar (por ejemplo el tiempo o la frecuencia) no es nada evidente. Hay un momento en que nos encontramos con la necesidad de derivar los vectores unitarios de la base del sistema de coordenadas respecto de esa variable escalar. Si trabajamos con coordenadas cilíndricas o esféricas dicha derivada no tiene por qué ser nula y hay que calcularla, que es lo difícil del asunto. Para poder calcularla habrá que hacer un cálculo tedioso qu...

  9. Mosaicos raster de cartografía vectorial: procedimiento automatizado de creación

    OpenAIRE

    Manso Callejo, Miguel Ángel; Moreno Marimbaldo, Francisco J.; Jiménez, Sergio; Pozo Ortego, Isaac

    2008-01-01

    En este documento se presenta una metodología, formada por un conjunto de tareas susceptibles de ser automatizadas, para producir un mosaico raster a partir de cartografía vectorial teselada. Como banco de prueba de la metodología se ha utilizado el Mapa Topográfico Nacional a escala 1:25.000 (MTN-25). La metodología propuesta implica: la eliminación de los elementos ajenos a la cartografía (cartela, etc.), la conversión de coordenadas proyectadas a geográficas, la creación de un mapa índice ...

  10. Production of lysozyme and lysozyme-superoxide dismutase dimers bound by a ditryptophan cross-link in carbonate radical-treated lysozyme.

    Science.gov (United States)

    Paviani, Verônica; Queiroz, Raphael F; Marques, Emerson F; Di Mascio, Paolo; Augusto, Ohara

    2015-12-01

    Despite extensive investigation of the irreversible oxidations undergone by proteins in vitro and in vivo, the products formed from the oxidation of Trp residues remain incompletely understood. Recently, we characterized a ditryptophan cross-link produced by the recombination of hSOD1-tryptophanyl radicals generated from attack of the carbonate radical produced during the bicarbonate-dependent peroxidase activity of the enzyme. Here, we examine whether the ditryptophan cross-link is produced by the attack of the carbonate radical on proteins other than hSOD1. To this end, we treated hen egg white lysozyme with photolytically and enzymatically generated carbonate radical. The radical yields were estimated and the lysozyme modifications were analyzed by SDS-PAGE, western blot, enzymatic activity and MS/MS analysis. Lysozyme oxidation by both systems resulted in its inactivation and dimerization. Lysozyme treated with the photolytic system presented monomers oxidized to hydroxy-tryptophan at Trp(28) and Trp(123) and N-formylkynurenine at Trp(28), Trp(62) and Trp(123). Lysozyme treated with the enzymatic system rendered monomers oxidized to N-formylkynurenine at Trp(28). The dimers were characterized as lysozyme-Trp(28)-Trp(28)-lysozyme and lysozyme-Trp(28)-Trp(32)-hSOD1. The results further demonstrate that the carbonate radical is prone to causing biomolecule cross-linking and hence, may be a relevant player in pathological mechanisms. The possibility of exploring the formation of ditryptophan cross-links as a carbonate radical biomarker is discussed. Copyright © 2015 Elsevier Inc. All rights reserved.

  11. Study of a photo-induced lysozyme-riboflavin bond

    International Nuclear Information System (INIS)

    Ferrer, I.; Silva, E.

    1985-01-01

    Irradiation of lysozyme in the presence of riboflavin results in the formation of a lysozyme-riboflavin adduct. Reduction and carboxymethylation of the four disulfide bonds as well as the chemical modification of the Tyr residues and the photochemical alteration of the His residue in lysozyme, do not affect the formation of the photo-induced lysozyme-riboflavin bond. When the lysozyme-riboflavin adduct was subjected to mild acid hydrolysis and ion exchange chromatography, the retention of a compound containing 14 C-riboflavin was observed. Free 14 C-ribboflavin, on the contrary is not retained by the column. The photo-oxidation of free Trp in the presence of 14 C-riboflavin, gave a compound which bound to the ion exchange resin like the above-mentioned derivative. The photo-oxidation of the Trp residues in lysozyme and in peptides obtained from lysozyme showed very high quantum yields, and these values were directly related to the incorporation of 14 C-riboflavin in these samples. (orig.)

  12. [Purification and characterization of a lysozyme from a marine microorganism].

    Science.gov (United States)

    Zou, Yan-Li; Sun, Mi; Wang, Yue-Jun

    2005-05-01

    A novel lysozyme was purified from a marine microorganism and its major characteristics were studied. Cell-free supernatant was prepared by centrifugation of culture broth, ultrafiltration using a hollow fiber (molecular weight cut off, 50kD) and concentration using a hollow fiber (molecular weight cut off, 10kD). The crude lysozyme was purified 34.7 fold to electrophoretic homogeneity with a recovery of 24.1% by CM-Sepharose FF cationic-exchange and Sephadex G-100 gel chromatography. The relative molecular weight of this lysozyme was determined as about 39 kD. The optimum pH and temperature towards Micrococcus lysodleikticus were pH 8.0 and 35 degrees C respectively, and the enzyme was stable at temperature below 50 degrees C and pH 5.0 - 10.0. The lysozyme activity was slightly enhanced by Zn2+ and Cu2+ and slightly inhibited by Mn2+ and Ag+. The lysozyme showed good compatibility to many common chemical agents such as EDTA (0.1%) and KH2 PO4 (1.0%). The lysozyme had broad-spectrum against many bacteria, including a number of pathogens, which were resistant to egg-white lysozyme.

  13. Lysozyme loading and release from Se doped hydroxyapatite nanoparticles.

    Science.gov (United States)

    Wang, Yanhua; Hao, Hang; Zhang, Shengmin

    2016-04-01

    Element-substituted hydroxyapatite (HA) based nanocomposites have become a promising therapeutic material for improving bone defect repair. Selenium substituted HA nanoparticles can both induce apoptosis of bone tumor cells and enhance osteointegration. However, the effect of selenite ions on the proteins in combination with the HA nanoparticles remains to be elucidated. Here, we investigated the influence of selenium doping concentration on the loading and release of lysozyme (LSM) as a model protein drug. The selenium substituted HA-LSM composites with different doping concentrations were synthesized and characterized. The subsequent delivery of lysozyme was studied in a phosphate buffer solution (PBS). We found that selenium substituted HA-LSM composites with Se:P=10% showed the highest amount of lysozyme loading (41.7%), whereas the amount of lysozyme loaded in undoped HA nanoparticles was the lowest (34.1%). The doped selenium interacts with lysozyme molecules, which leads to the increase of β-sheet and unordered, and the decrease of self-association, α-helix and β-turns in protein structures. Moreover, selenium addition significantly slows the protein release from HA-LSM composites. The composites with Se:P=10% release lysozyme at the slightly slower rate among the samples with different Se doping concentrations. It also shows that the released lysozyme retains most of its enzymatic activity. Copyright © 2016 Elsevier B.V. All rights reserved.

  14. Effect of secondary structure on the interactions of peptide T4 LYS (11-36) in mixtures of aqueous sodium chloride and 2,2,2,-Trifluoroethanol

    Energy Technology Data Exchange (ETDEWEB)

    Anderson, Camille O.; Spiegelberg, Susanne; Prausnitz, John M.; Blanch, Harvey W.

    2001-10-01

    The potential of mean force for protein-protein interactions is key to the development of a statistical-mechanical model for salt-induced protein precipitation and crystallization, and for understanding certain disease states, including cataract formation and {beta}-amyloid pathology in Alzheimer's disease. Fluorescence anisotropy provides a method for quantitative characterization of intermolecular interactions due to reversible association. Monomer-dimer equilibria for the peptide T4 LYS(11-36) were studied by fluorescence anisotropy. This peptide, derived from the {beta}-sheet region of the T4 lysozyme molecule, has the potential to form amyloid fibrils. 2,2,2-trifluoroethanol (TFE) induces a change in peptide secondary structure, and was used in aqueous solutions at concentrations from 0 to 50% (v/v) at 25 and 37 C to examine the role of peptide conformation on peptide-peptide interactions. The association constant for dimerization increased with rising TFE concentration and with falling temperature. The peptide-peptide potential of mean force was computed from these association constants. Circular-dichroism measurements showed that the secondary structure of the peptide plays an important role in these strong attractive interactions due to intermolecular hydrogen-bond formation and hydrophobic interactions.

  15. Expression and prognostic significance of lysozyme in male breast cancer

    International Nuclear Information System (INIS)

    Serra, Carlos; Baltasar, Aniceto; Medrano, Justo; Vizoso, Francisco; Alonso, Lorena; Rodríguez, Juan C; González, Luis O; Fernández, María; Lamelas, María L; Sánchez, Luis M; García-Muñiz, José L

    2002-01-01

    Lysozyme, one of the major protein components of human milk that is also synthesized by a significant percentage of breast carcinomas, is associated with lesions that have a favorable outcome in female breast cancer. Here we evaluate the expression and prognostic value of lysozyme in male breast cancer (MBC). Lysozyme expression was examined by immunohistochemical methods in a series of 60 MBC tissue sections and in 15 patients with gynecomastia. Staining was quantified using the HSCORE (histological score) system, which considers both the intensity and the percentage of cells staining at each intensity. Prognostic value of lysozyme was retrospectively evaluated by multivariate analysis taking into account conventional prognostic factors. Lysozyme immunostaining was negative in all cases of gynecomastia. A total of 27 of 60 MBC sections (45%) stained positively for this protein, but there were clear differences among them with regard to the intensity and percentage of stained cells. Statistical analysis showed that lysozyme HSCORE values in relation to age, tumor size, nodal status, histological grade, estrogen receptor status, metastasis and histological type did not increase the statistical significance. Univariate analysis confirmed that both nodal involvement and lysozyme values were significant predictors of short-term relapse-free survival. Multivariate analysis, according to Cox's regression model, also showed that nodal status and lysozyme levels were significant independent indicators of short-term relapse-free survival. Tumor expression of lysozyme is associated with lesions that have an unfavorable outcome in male breast cancer. This milk protein may be a new prognostic factor in patients with breast cancer

  16. Separation of lysozyme using superparamagnetic carboxymethyl chitosan nanoparticles.

    Science.gov (United States)

    Sun, Jun; Su, Yujie; Rao, Shengqi; Yang, Yanjun

    2011-08-01

    Functionalized Fe(3)O(4) nanoparticles conjugated with polyethylene glycol (PEG) and carboxymethyl chitosan (CM-CTS) were developed and used as a novel magnetic absorbing carrier for the separation and purification of lysozyme from the aqueous solution and chicken egg white, respectively. The morphology of magnetic CM-CTS nanoparticles was observed by transmission electron microscope (TEM). It was found that the diameter of superparamagnetic carboxymethyl chitosan nanoparticles (Fe(3)O(4) (PEG+CM-CTS)) was about 15 nm, and could easily aggregate by a magnet when suspending in the aqueous solution. The adsorption capacity of lysozyme onto the superparamagnetic Fe(3)O(4) (PEG+CM-CTS) nanoparticles was determined by changing the medium pH, temperature, ionic strength and the concentration of lysozyme. The maximum adsorption loading reached 256.4 mg/g. Due to the small diameter, the adsorption equilibrium of lysozyme onto the nanoparticles reached very quickly within 20 min. The adsorption equilibrium of lysozyme onto the superparamagnetic nanoparticles fitted well with the Langmuir model. The nanoparticles were stable when subjected to six repeated adsorption-elution cycles. Separation and purification were monitored by determining the lysozyme activity using Micrococcus lysodeikticus as substrate. The lysozyme was purified from chicken egg white in a single step had higher purity, as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Considering that the superparamagnetic nanoparticles possess the advantages of high efficiency, cost-effectiveness and excellent binding of a larger amount of lysozyme and easier separation from the reaction system, thus this type of superparamagnetic nanoparticles would bring advantages to the conventional separation techniques of lysozyme from chicken egg white. Copyright © 2011 Elsevier B.V. All rights reserved.

  17. Demonstration of a vectorial optical field generator with adaptive close loop control

    Science.gov (United States)

    Chen, Jian; Kong, Lingjiang; Zhan, Qiwen

    2017-12-01

    We experimentally demonstrate a vectorial optical field generator (VOF-Gen) with an adaptive close loop control. The close loop control capability is illustrated with the calibration of polarization modulation of the system. To calibrate the polarization ratio modulation, we generate 45° linearly polarized beam and make it propagate through a linear analyzer whose transmission axis is orthogonal to the incident beam. For the retardation calibration, circularly polarized beam is employed and a circular polarization analyzer with the opposite chirality is placed in front of the CCD as the detector. In both cases, the close loop control automatically changes the value of the corresponding calibration parameters in the pre-set ranges to generate the phase patterns applied to the spatial light modulators and records the intensity distribution of the output beam by the CCD camera. The optimized calibration parameters are determined corresponding to the minimum total intensity in each case. Several typical kinds of vectorial optical beams are created with and without the obtained calibration parameters, and the full Stokes parameter measurements are carried out to quantitatively analyze the polarization distribution of the generated beams. The comparisons among these results clearly show that the obtained calibration parameters could remarkably improve the accuracy of the polarization modulation of the VOF-Gen, especially for generating elliptically polarized beam with large ellipticity, indicating the significance of the presented close loop in enhancing the performance of the VOF-Gen.

  18. A vectorial capacity product to monitor changing malaria transmission potential in epidemic regions of Africa

    Science.gov (United States)

    Ceccato, Pietro; Vancutsem, Christelle; Klaver, Robert; Rowland, James; Connor, Stephen J.

    2012-01-01

    Rainfall and temperature are two of the major factors triggering malaria epidemics in warm semi-arid (desert-fringe) and high altitude (highland-fringe) epidemic risk areas. The ability of the mosquitoes to transmit Plasmodium spp. is dependent upon a series of biological features generally referred to as vectorial capacity. In this study, the vectorial capacity model (VCAP) was expanded to include the influence of rainfall and temperature variables on malaria transmission potential. Data from two remote sensing products were used to monitor rainfall and temperature and were integrated into the VCAP model. The expanded model was tested in Eritrea and Madagascar to check the viability of the approach. The analysis of VCAP in relation to rainfall, temperature and malaria incidence data in these regions shows that the expanded VCAP correctly tracks the risk of malaria both in regions where rainfall is the limiting factor and in regions where temperature is the limiting factor. The VCAP maps are currently offered as an experimental resource for testing within Malaria Early Warning applications in epidemic prone regions of sub-Saharan Africa. User feedback is currently being collected in preparation for further evaluation and refinement of the VCAP model.

  19. A vectorial capacity product to monitor changing malaria transmission potential in epidemic regions of Africa.

    Science.gov (United States)

    Ceccato, Pietro; Vancutsem, Christelle; Klaver, Robert; Rowland, James; Connor, Stephen J

    2012-01-01

    Rainfall and temperature are two of the major factors triggering malaria epidemics in warm semi-arid (desert-fringe) and high altitude (highland-fringe) epidemic risk areas. The ability of the mosquitoes to transmit Plasmodium spp. is dependent upon a series of biological features generally referred to as vectorial capacity. In this study, the vectorial capacity model (VCAP) was expanded to include the influence of rainfall and temperature variables on malaria transmission potential. Data from two remote sensing products were used to monitor rainfall and temperature and were integrated into the VCAP model. The expanded model was tested in Eritrea and Madagascar to check the viability of the approach. The analysis of VCAP in relation to rainfall, temperature and malaria incidence data in these regions shows that the expanded VCAP correctly tracks the risk of malaria both in regions where rainfall is the limiting factor and in regions where temperature is the limiting factor. The VCAP maps are currently offered as an experimental resource for testing within Malaria Early Warning applications in epidemic prone regions of sub-Saharan Africa. User feedback is currently being collected in preparation for further evaluation and refinement of the VCAP model.

  20. A Vectorial Capacity Product to Monitor Changing Malaria Transmission Potential in Epidemic Regions of Africa

    Directory of Open Access Journals (Sweden)

    Pietro Ceccato

    2012-01-01

    Full Text Available Rainfall and temperature are two of the major factors triggering malaria epidemics in warm semi-arid (desert-fringe and high altitude (highland-fringe epidemic risk areas. The ability of the mosquitoes to transmit Plasmodium spp. is dependent upon a series of biological features generally referred to as vectorial capacity. In this study, the vectorial capacity model (VCAP was expanded to include the influence of rainfall and temperature variables on malaria transmission potential. Data from two remote sensing products were used to monitor rainfall and temperature and were integrated into the VCAP model. The expanded model was tested in Eritrea and Madagascar to check the viability of the approach. The analysis of VCAP in relation to rainfall, temperature and malaria incidence data in these regions shows that the expanded VCAP correctly tracks the risk of malaria both in regions where rainfall is the limiting factor and in regions where temperature is the limiting factor. The VCAP maps are currently offered as an experimental resource for testing within Malaria Early Warning applications in epidemic prone regions of sub-Saharan Africa. User feedback is currently being collected in preparation for further evaluation and refinement of the VCAP model.

  1. Direct observation of temperature-driven magnetic symmetry transitions by vectorial resolved MOKE magnetometry.

    Science.gov (United States)

    Luis F Cuñado, Jose; Pedrosa, Javier; Ajejas, Fernando; Perna, Paolo; Miranda, Rodolfo; Camarero, Julio

    2017-10-11

    Angle- and temperature-dependent vectorial magnetometry measurements are necessary to disentangle the effective magnetic symmetry in magnetic nanostructures. Here we present a detailed study on an Fe(1 0 0) thin film system with competing collinear biaxial (four-fold symmetry) and uniaxial (two-fold) magnetic anisotropies, carried out with our recently developed full angular/broad temperature range/vectorial-resolved magneto-optical Kerr effect magnetometer, named TRISTAN. The data give direct views on the angular and temperature dependence of the magnetization reversal pathways, from which characteristic axes, remanences, critical fields, domain wall types, and effective magnetic symmetry are obtained. In particular, although the remanence shows four-fold angular symmetry for all investigated temperatures (15 K-400 K), the critical fields show strong temperature and angular dependencies and the reversal mechanism changes for specific angles at a given (angle-dependent) critical temperature, showing signatures of an additional collinear two-fold symmetry. This symmetry-breaking is more relevant as temperature increases to room temperature. It originates from the competition between two anisotropy contributions with different symmetry and temperature evolution. The results highlight the importance of combining temperature and angular studies, and the need to look at different magnetic parameters to unravel the underlying magnetic symmetries and temperature evolutions of the symmetry-breaking effects in magnetic nanostructures.

  2. Full-vectorial propagation model and modified effective mode area of four-wave mixing in straight waveguides

    DEFF Research Database (Denmark)

    Guo, Kai; Friis, Søren Michael Mørk; Christensen, Jesper Bjerge

    2017-01-01

    We derive from Maxwell's equations full-vectorial nonlinear propagation equations of four-wave mixing valid in straight semiconductor-on-insulator waveguides. Special attention is given to the resulting effective mode area, which takes a convenient form known from studies in photonic crystal fibers......, but has not been introduced in the context of integrated waveguides. We show that the difference between our full-vectorial effective mode area and the scalar equivalent often referred to in the literature may lead to mistakes when evaluating the nonlinear refractive index and optimizing designs of new...

  3. Lysozyme contamination facilitates crystallization of a heterotrimeric cortactin–Arg–lysozyme complex

    International Nuclear Information System (INIS)

    Liu, Weizhi; MacGrath, Stacey M.; Koleske, Anthony J.; Boggon, Titus J.

    2012-01-01

    An unusual case of trace amounts of a contaminating protein facilitating formation of a heterotrimeric protein complex. Crystallization of contaminating proteins is a frequently encountered problem for macromolecular crystallographers. In this study, an attempt was made to obtain a binary cocrystal structure of the SH3 domain of cortactin and a 17-residue peptide from the Arg nonreceptor tyrosine kinase encompassing a PxxPxxPxxP (PxxP1) motif. However, cocrystals could only be obtained in the presence of trace amounts of a contaminating protein. A structure solution obtained by molecular replacement followed by ARP/wARP automatic model building allowed a ‘sequence-by-crystallography’ approach to discover that the contaminating protein was lysozyme. This 1.65 Å resolution crystal structure determination of a 1:1:1 heterotrimeric complex of Arg, cortactin and lysozyme thus provides an unusual ‘caveat emptor’ warning of the dangers that underpurified proteins harbor for macromolecular crystallographers

  4. Detecting expression of 5T4 in CTCs and tumor samples from NSCLC patients.

    Directory of Open Access Journals (Sweden)

    Steven R Pirie-Shepherd

    Full Text Available The fetal oncogene 5T4 is a cell surface protein, with overexpression observed in a variety of cancers as compared to normal adult tissue. The ability to select patients with tumors that express high levels of 5T4 may enrich a clinical trial cohort with patients most likely to respond to 5T4 targeted therapy. To that end, we developed assays to measure 5T4 in both tumors and in circulating tumor cells (CTCs. We identified the presence of 5T4 in both adenocarcinoma and squamous cell carcinoma of lung, in all clinical stages and grades of disease. CTCs were identified in peripheral blood from the majority of patients with NSCLC, and 5T4 was detectable in most samples. Although 5T4 was present in both CTCs and tumors in most patients, there was no concordance between relative amount in either sample type. Clinical response rates of patients treated with the therapies directed against 5T4 in early stage clinical trials, as determined by these assays, may provide important insights into the biology of 5T4 in tumors and the mechanisms of action of 5T4-targeting therapy.

  5. Structural analysis of bacteriophage T4 DNA replication: a review in the Virology Journal series on bacteriophage T4 and its relatives

    Directory of Open Access Journals (Sweden)

    Boyer Ryan A

    2010-12-01

    Full Text Available Abstract The bacteriophage T4 encodes 10 proteins, known collectively as the replisome, that are responsible for the replication of the phage genome. The replisomal proteins can be subdivided into three activities; the replicase, responsible for duplicating DNA, the primosomal proteins, responsible for unwinding and Okazaki fragment initiation, and the Okazaki repair proteins. The replicase includes the gp43 DNA polymerase, the gp45 processivity clamp, the gp44/62 clamp loader complex, and the gp32 single-stranded DNA binding protein. The primosomal proteins include the gp41 hexameric helicase, the gp61 primase, and the gp59 helicase loading protein. The RNaseH, a 5' to 3' exonuclease and T4 DNA ligase comprise the activities necessary for Okazaki repair. The T4 provides a model system for DNA replication. As a consequence, significant effort has been put forth to solve the crystallographic structures of these replisomal proteins. In this review, we discuss the structures that are available and provide comparison to related proteins when the T4 structures are unavailable. Three of the ten full-length T4 replisomal proteins have been determined; the gp59 helicase loading protein, the RNase H, and the gp45 processivity clamp. The core of T4 gp32 and two proteins from the T4 related phage RB69, the gp43 polymerase and the gp45 clamp are also solved. The T4 gp44/62 clamp loader has not been crystallized but a comparison to the E. coli gamma complex is provided. The structures of T4 gp41 helicase, gp61 primase, and T4 DNA ligase are unknown, structures from bacteriophage T7 proteins are discussed instead. To better understand the functionality of T4 DNA replication, in depth structural analysis will require complexes between proteins and DNA substrates. A DNA primer template bound by gp43 polymerase, a fork DNA substrate bound by RNase H, gp43 polymerase bound to gp32 protein, and RNase H bound to gp32 have been crystallographically determined. The

  6. Reentrant condensation of lysozyme: Implications for studying dynamics of lysozyme in aqueous solutions of lithium chloride

    Energy Technology Data Exchange (ETDEWEB)

    Mamontov, Eugene [ORNL; O' Neill, Hugh Michael [ORNL

    2014-01-01

    Recent studies have outlined the use of eutectic solution of lithium chloride in water to study microscopic dynamics of lysozyme in an aqueous solvent that is remarkably similar to pure water in many respects, yet allows experiments over a wide temperature range without the solvent crystallization. The eutectic point in (H2O)R(LiCl) system corresponds to R 7.3, and it is of interest to investigate whether less concentrated aqueous solutions of LiCl could be employed in low-temperature studies of a solvated protein. We have investigated a range of concentrations of lysozyme and LiCl in aqueous solutions to identify systems that do not show phase separation and avoid solvent crystallization on cooling down. Compared to the lysozyme concentration in solution, the concentration of LiCl in the aqueous solvent plays the major role in determining systems suitable for low-temperature studies. We have observed interesting and rich phase behavior reminiscent of reentrant condensation of proteins.

  7. Effect of thyroxine on cellular oxygen-consumption and glucose uptake: evidence of an effect of total T4 and not "free T4"

    DEFF Research Database (Denmark)

    Kvetny, J; Matzen, L E

    1990-01-01

    in human mononuclear blood cells. Cells were incubated in protein free medium and in human serum totally depleted of thyroid hormones by resin treatment and fixed amounts of T4 (total T4 = 0-50-100-5000 nmol/l; free T4 = 0-5-11-5600 pmol/l) were added. Thyroxine stimulated glucose uptake and oxygen......Recent studies of cellular T4 and T3 uptake have indicated active transport of the hormones into the cell rather than passive diffusion of the non-protein bound fraction. In order to study the significance of the extracellular environment, oxygen consumption and glucose uptake were examined......K-ATPase by addition of ouabain (9-72 mg/l) did not inhibit T4 stimulation, thus indicating that the ouabain sensitive NaK-ATPase is not a major component of the processes which initiate the intracellular effects of T4. Therefore the stimulation of uptake of oxygen and glucose in human mononuclear blood cells seems...

  8. Bactericidal activity of human lysozyme, muramidase-inactive lysozyme, and cationic polypeptides against Streptococcus sanguis and Streptococcus faecalis: inhibition by chitin oligosaccharides.

    OpenAIRE

    Laible, N J; Germaine, G R

    1985-01-01

    The basis of the bactericidal activity of human lysozyme against Streptococcus sanguis was studied. Experiments were designed to evaluate the role of lysozyme muramidase activity in its bactericidal potency. Inactivation of the muramidase activity of lysozyme was achieved by reduction of essential disulfides with dithiothreitol (DTT) or by incubation with the chitin oligosaccharides chitotriose and chitobiose. Muramidase-inactive lysozyme, prepared by reduction with DTT, was equal in bacteric...

  9. Study on nucleation kinetics of lysozyme crystallization

    Science.gov (United States)

    Lin, Chen; Zhang, Yang; Liu, Jing J.; Wang, Xue Z.

    2017-07-01

    The nucleation kinetics of hen egg-white lysozyme crystallization was investigated using a hot stage cooling crystallizer and a microscope to monitor the solution crystallization process in real time. Images of crystals were continuously recorded under varied precipitant and protein concentrations. The nucleation rate was found to be higher at higher precipitant concentration, and increase monotonically with protein concentration if the precipitant concentration was held constant. Attempt was made to interpret the experimental data using classical nucleation theory. It was found that the model predictions are lower than the experimental values at low supersaturations but agree well with experimental data at high supersaturations. The trends in the experimental data suggest that two nucleation mechanisms might co-exist: heterogeneous nucleation seems to be the dominant at low supersaturation while at higher supersaturation homogeneous nucleation seems to play the major role.

  10. Phylogenetic diversity of T4-like bacteriophages in Lake Baikal, East Siberia.

    Science.gov (United States)

    Butina, Tatyana Vladimirovna; Belykh, Olga I; Maksimenko, Svetlana Yu; Belikov, Sergey I

    2010-08-01

    Among the tailed phages, the myoviruses, those with contractile tails, are widespread and diverse. An important component of the Myoviridae family is the genus 'T4-like viruses'. The present study was aimed at elucidating the molecular diversity of T4-type bacteriophages in Lake Baikal by partial sequencing of g23 genes of T4-type bacteriophages. Our study revealed that the g23 gene sequences investigated were highly diverse and different from those of T4-like bacteriophages and from g23 clones obtained from different environments. Phylogenetic analysis showed that all g23 fragments from Lake Baikal, except for the one sequence, were more closely related to marine T4 cyanophages and to previously described subgroups of uncultured T4 phages from marine and rice field environments.

  11. PROTEOLYTIC REMOVAL OF THE CARBOXYL TERMINUS OF THE T4 GENE 32 HELIX-DESTABILIZING PROTEIN ALTERS THE T4 IN VITRO REPLICATION COMPLEX

    Energy Technology Data Exchange (ETDEWEB)

    Burke, R.L.; Alberts, B.M.; Hosoda, J.

    1980-07-01

    The proteolytic removal of about 60 amino acids from the COOH terminus of the bacteriophage T4 helix-destabilizing protein (gene 32 protein) produces 32*I, a 27,000-dalton fragment which still binds tightly and cooperatively to single-stranded DNA. The substitution of 32*I protein for intact 32 protein in the seven-protein T4 replication complex results in dramatic changes in some of the reactions catalyzed by this in vitro DNA replication system, while leaving others largely unperturbed. (1) Like intact 32 protein, the 32*I protein promotes DNA synthesis by the DNA polymerase when the T4 polymerase accessory proteins (gene 44/62 and 45 proteins) are also present. The host helix-destabilizing protein (Escherichia coli ssb protein) cannot replace the 32*I protein for this synthesis. (2) Unlike intact 32 protein, 32*I protein strongly inhibits DNA synthesis catalyzed by the T4 DNA polymerase alone on a primed single-stranded DNA template. (3) Unlike intact 32 protein, the 32*I protein strongly inhibits RNA primer synthesis catalyzed by the T4 gene 41 and 61 proteins and also reduces the efficiency of RNA primer utilization. As a result, de novo DNA chain starts are blocked completely in the complete T4 replication system, and no lagging strand DNA synthesis occurs. (4) The 32*I protein does not bind to either the T4 DNA polymerase or to the T4 gene 61 protein in the absence of DNA; these associations (detected with intact 32 protein) would therefore appear to be essential for the normal control of 32 protein activity, and to account at least in part for observations 2 and 3, above. We propose that the COOH-terminal domain of intact 32 protein functions to guide its interactions with the T4 DNA polymerase and the T4 gene 61 RNA-priming protein. When this domain is removed, as in 32*I protein, the helix destabilization induced by the protein is controlled inadequately, so that polymerizing enzymes tend to be displaced from the growing 3{prime}-OH end of a

  12. Molecular dynamics simulations of lysozyme in water/sugar solutions

    Energy Technology Data Exchange (ETDEWEB)

    Lerbret, A. [Department of Food Science, Cornell University, 101 Stocking Hall, Ithaca, NY 14853 (United States); Affouard, F. [Laboratoire de Dynamique et Structure des Materiaux Moleculaires, UMR CNRS 8024, Universite Lille I, 59655 Villeneuve d' Ascq Cedex (France)], E-mail: frederic.affouard@univ-lille1.fr; Bordat, P. [Laboratoire de Chimie Theorique et de Physico-Chimie Moleculaire, UMR 5624, Universite de Pau et des Pays de l' Adour, 64000 Pau (France); Hedoux, A.; Guinet, Y.; Descamps, M. [Laboratoire de Dynamique et Structure des Materiaux Moleculaires, UMR CNRS 8024, Universite Lille I, 59655 Villeneuve d' Ascq Cedex (France)

    2008-04-18

    Structural and dynamical properties of the solvent at the protein/solvent interface have been investigated by molecular dynamics simulations of lysozyme in trehalose, maltose and sucrose solutions. Results are discussed in the framework of the bioprotection phenomena. The analysis of the relative concentration of water oxygen atoms around lysozyme suggests that lysozyme is preferentially hydrated. When comparing the three sugars, trehalose is seen more excluded than maltose and sucrose. The preferential exclusion of sugars from the protein surface induces some differences in the behavior of trehalose and maltose, particularly at 50 and 60 wt% concentrations, that are not observed experimentally in binary sugar/mixtures. The dynamical slowing down of the solvent is suggested to mainly arise from the homogeneity of the water/sugar matrices controlled by the percolation of the sugar hydrogen bonds networks. Furthermore, lysozyme strongly increases relaxation times of solvent molecules at the protein/solvent interface.

  13. Studies on isolation and partial purification of lysozyme from egg ...

    African Journals Online (AJOL)

    STORAGESEVER

    2009-01-05

    PAGE. INTRODUCTION. Lysozyme is one of the best-characterized hydrolases, which cleaves β-1,4 ... heteropolymer, which causes lysis of the bacteria con- taining that ... sodium phosphate buffer (pH 7.2). Determining the ...

  14. Laser ablation dynamics and production of thin films of lysozyme

    DEFF Research Database (Denmark)

    Schou, Jørgen; Canulescu, Stela; Matei, Andreea

    Lysozyme is a well-known protein, which is used in food processing because of its bacteriocidal properties. The mass (14307 u) is in the range, in which it easily can be controlled by mass spectrometric methods, for example by MALDI (Matrix assisted laser desorption ionisation). We have recently......, there was a considerable ablation weight loss of lysozyme from each shot. This is the first time the ablation by fs-lasers of a protein has been recorded quantitatively. Films of lysozyme produced by fs-laser irradiation will be analysed by MALDI in order to explore if there also is a significant amount of intact...... these experiments at CNR-SPIN, Napoli, to explore the excitation mechanics by laser impact. Samples of pressed lysozyme prepared in the same manner as in DTU have been irradiated at 523 nm with 300-fs pulses and a fluence of the same order of magnitude as in DYU. Even though the pulse energy was much smaller...

  15. Laser ablation dynamics and production of thin films of lysozyme

    DEFF Research Database (Denmark)

    Canulescu, Stela; Schou, Jørgen; Amoruso, S.

    Lysozyme is a well-known protein, which is used in food processing because of its bactericidal properties. The mass (14307 amu) is in the range in which it easily can be monitored by mass spectrometric methods, for example by MALDI (Matrix assisted laser desorption ionization). We have recently....... This is the first time the ablation by fs-lasers of a protein has been recorded quantitatively. Films of lysozyme produced by fs-laser irradiation were analyzed by MALDI and a significant number of intact molecules in the films with fs-laser deposition was found as well....... impact. Samples of pressed lysozyme prepared in the same manner as in ns-experiments have been irradiated at 527 nm with 300-fs pulses and at at similar fluence as in ns ablation. Even though the pulse energy was much smaller, there was a considerable ablation weight loss of lysozyme from each shot...

  16. Molecular dynamics simulations of lysozyme in water/sugar solutions

    International Nuclear Information System (INIS)

    Lerbret, A.; Affouard, F.; Bordat, P.; Hedoux, A.; Guinet, Y.; Descamps, M.

    2008-01-01

    Structural and dynamical properties of the solvent at the protein/solvent interface have been investigated by molecular dynamics simulations of lysozyme in trehalose, maltose and sucrose solutions. Results are discussed in the framework of the bioprotection phenomena. The analysis of the relative concentration of water oxygen atoms around lysozyme suggests that lysozyme is preferentially hydrated. When comparing the three sugars, trehalose is seen more excluded than maltose and sucrose. The preferential exclusion of sugars from the protein surface induces some differences in the behavior of trehalose and maltose, particularly at 50 and 60 wt% concentrations, that are not observed experimentally in binary sugar/mixtures. The dynamical slowing down of the solvent is suggested to mainly arise from the homogeneity of the water/sugar matrices controlled by the percolation of the sugar hydrogen bonds networks. Furthermore, lysozyme strongly increases relaxation times of solvent molecules at the protein/solvent interface

  17. Destroying activity of magnetoferritin on lysozyme amyloid fibrils

    Energy Technology Data Exchange (ETDEWEB)

    Kopcansky, Peter; Siposova, Katarina [Institute of Experimental Physics, SAS, Watsonova 47, 040 01 Kosice (Slovakia); Melnikova, Lucia, E-mail: melnikova@saske.sk [Institute of Experimental Physics, SAS, Watsonova 47, 040 01 Kosice (Slovakia); Bednarikova, Zuzana [Institute of Experimental Physics, SAS, Watsonova 47, 040 01 Kosice (Slovakia); Institute of Chemical Sciences, Faculty of Sciences, Safarik University, Kosice (Slovakia); Timko, Milan; Mitroova, Zuzana; Antosova, Andrea [Institute of Experimental Physics, SAS, Watsonova 47, 040 01 Kosice (Slovakia); Garamus, Vasil M. [Helmholtz-Zentrum Geesthacht: Centre for Materials and Coastal Research, Max-Planck-Street 1, 21502 Geesthacht (Germany); Petrenko, Viktor I. [Joint Institute for Nuclear Research, Joliot-Curie 6, Dubna, 141980 Moscow Region (Russian Federation); Kyiv Taras Shevchenko National University, Volodymyrska Street 64, Kyiv 01033 (Ukraine); Avdeev, Mikhail V. [Joint Institute for Nuclear Research, Joliot-Curie 6, Dubna, 141980 Moscow Region (Russian Federation); Gazova, Zuzana [Institute of Experimental Physics, SAS, Watsonova 47, 040 01 Kosice (Slovakia); Department of Medical and Clinical Biochemistry and LABMED, Tr. SNP 1, 040 11 Kosice (Slovakia)

    2015-03-01

    Presence of protein amyloid aggregates (oligomers, protofilaments, fibrils) is associated with many diseases as diabetes mellitus or Alzheimer's disease. The interaction between lysozyme amyloid fibrils and magnetoferritin loaded with different amount of iron atoms (168 or 532 atoms) has been investigated by small-angle X-rays scattering and thioflavin T fluorescence measurements. Results suggest that magnetoferritin caused an iron atom-concentration dependent reduction of lysozyme fibril size. - Highlights: • The interaction between lysozyme amyloid fibrils and magnetoferritin loaded with different amount of iron atoms (168 or 532 atoms) has been investigated by small-angle X-rays scattering and thioflavin T fluorescence measurements. • Results suggest that magnetoferritin caused an iron atom-concentration dependent reduction of lysozyme fibril size.

  18. Location of Bromide Ions in Tetragonal Lysozyme Crystals

    Science.gov (United States)

    Lim, Kap; Nadarajah, Arunan; Forsythe, Elizabeth L.; Pusey, Marc L.

    1998-01-01

    Anions have been shown to play a dominant role in the crystallization of chicken egg white lysozyme from salt solutions. Previous studies employing X-ray crystallography had found one chloride ion binding site in the tetragonal crystal form of the protein and four nitrate ion binding sites in the monoclinic form. In this study the anion positions in the tetragonal form were determined from the difference Fourier map obtained from lysozyme crystal grown in bromide and chloride solutions. Five possible anion binding sites were found in this manner. Some of these sites were in pockets containing basic residues while others were near neutral, but polar, residues. The sole chloride ion binding site found in previous studies was confirmed, while four of these sites corresponded to four binding sites found for nitrate ions in monoclinic crystals. The study suggests that most of the anion binding sites in lysozyme remain unchanged, even when different anions and different crystal forms of lysozyme are employed.

  19. The direct piezoelectric effect in the globular protein lysozyme

    Science.gov (United States)

    Stapleton, A.; Noor, M. R.; Sweeney, J.; Casey, V.; Kholkin, A. L.; Silien, C.; Gandhi, A. A.; Soulimane, T.; Tofail, S. A. M.

    2017-10-01

    Here, we present experimental evidence of the direct piezoelectric effect in the globular protein, lysozyme. Piezoelectric materials are employed in many actuating and sensing applications because they can convert mechanical energy into electrical energy and vice versa. Although originally studied in inorganic materials, several biological materials including amino acids and bone, also exhibit piezoelectricity. The exact mechanisms supporting biological piezoelectricity are not known, nor is it known whether biological piezoelectricity conforms strictly to the criteria of classical piezoelectricity. The observation of piezoelectricity in protein crystals presented here links biological piezoelectricity with the classical theory of piezoelectricity. We quantify the direct piezoelectric effect in monoclinic and tetragonal aggregate films of lysozyme using conventional techniques based on the Berlincourt Method. The largest piezoelectric effect measured in a crystalline aggregate film of lysozyme was approximately 6.5 pC N-1. These findings raise fundamental questions as to the possible physiological significance of piezoelectricity in lysozyme and the potential for technical applications.

  20. Dielectric and gravimetric studies of water binding to lysozyme

    International Nuclear Information System (INIS)

    Bone, S.

    1996-01-01

    Time domain dielectric spectroscopy and hydration isotherm measurements as a function of temperature have been applied to hydrated lysozyme powder. Two dielectric dispersions were identified, the first centred at approximately 8 MHz and a second above 1 GHz. The higher dispersion is considered to be the result of rotational relaxation of water molecules bound to the enzyme. In this case the results indicate the existence of a population of 32 water molecules per lysozyme molecule which are irrotationally bound to the lysozyme structure. A larger population of water molecules is relatively free to respond to the electric field and exhibits a dipole moment close to that of vapour phase water molecules. Multi-temperature hydration isotherm measurements are used to calculate enthalpies and entropies associated with the binding of water to lysozyme. Discontinuities both in dielectric and in thermodynamic characteristics in the range 10-14% hydration are interpreted as a re-ordering of the water structure on the enzyme surface

  1. Campylobacter jejuni Group III Phage CP81 Contains Many T4-Like Genes without Belonging to the T4-Type Phage Group: Implications for the Evolution of T4 Phages▿†

    OpenAIRE

    Hammerl, Jens A.; Jäckel, Claudia; Reetz, Jochen; Beck, Sebastian; Alter, Thomas; Lurz, Rudi; Barretto, Caroline; Brüssow, Harald; Hertwig, Stefan

    2011-01-01

    CP81 is a virulent Campylobacter group III phage whose linear genome comprises 132,454 bp. At the nucleotide level, CP81 differs from other phages. However, a number of its structural and replication/recombination proteins revealed a relationship to the group II Campylobacter phages CP220/CPt10 and to T4-type phages. Unlike the T4-related phages, the CP81 genome does not contain conserved replication and virion modules. Instead, the respective genes are scattered throughout the phage genome. ...

  2. L-T4 bioequivalence and hormone replacement studies via feedback control simulations.

    Science.gov (United States)

    Eisenberg, Marisa; Samuels, Mary; DiStefano, Joseph J

    2006-12-01

    FDA Guidance for testing bioequivalence of levothyroxine (L-T(4)) preparations has been challenged by several groups, based on multiple issues. The efficacy of single versus combined hormone therapy also is receiving additional scrutiny. To examine these concerns, we developed a new nonlinear feedback system simulation model of whole-body regulation mechanisms involving dynamics of T(3), T(4), TSH, plasma protein binding, extravascular regulatory enzyme systems, and the hypothalamic-pituitary-thyroid axis, all quantified from human data. To address bioequivalence, we explored how to best account for varying and unmeasured endogenous T(4) following dosing with exogenous oral L-T(4) in euthyroid volunteers in required pharmacokinetic (PK) studies, by simulating various dosing scenarios and developing a new and simple correction method. We computed and assessed dosing error effects and baseline corrections using simulator-predicted endogenous T(4) level variations, due to actual closed-loop effects, and compared these with approximate corrections computed directly from PK data. Predicted dose-responses were quite linear, and for constant baseline, 7-day half-life, and our new formula-correction methods, we established some bounds on bioequivalent dosages. Simulated replacement after thyroidectomy required 141 microg L-T(4) only to normalize T(3) tissue levels and 162 microg L-T(4) to normalize plasma T(3) levels. A combined dose of approximately 103 microg L-T(4) plus approximately 6 microg T(3) ( approximately 18:1 ratio) was needed to normalize both plasma T(3) and T(4) and average tissue T(3) levels. However, simulated average tissue T(3) levels were normalized with standard L-T(4)-only therapy, and plasma T(3) levels were still within the normal range. We suggest a simple and more accurate correction for endogenous T(4) in PK studies. Current standard L-T(4)-only treatment is supported for routine replacement needs.

  3. Multifocal T4 non-small cell lung cancer: a subset with improved prognosis.

    Science.gov (United States)

    Trousse, Delphine; D'Journo, Xavier Benoît; Avaro, Jean-Philippe; Doddoli, Christophe; Giudicelli, Roger; Fuentes, Pierre Antoine; Thomas, Pascal Alexandre

    2008-01-01

    T4-disease for non-small cell lung cancer (NSCLC) includes different conditions: mediastinal invasion, neoplastic pleural cytology, and multifocal disease in the same lobe; regarding the last category, no strict criteria allow to differentiate satellite nodules from synchronous multiple primary tumours. Retrospective study of 56 patients who underwent a complete resection from 1985 to 2006 of a NSCLC graded pT4N0 due to multifocal disease. A small nodule (satellite nodule (pT4sn). Multiple tumours, sized more than 1cm, with an identical histology, located in the same lobe but in different segment were considered as synchronous cancers (pT4sc). There were 44 males and 12 females: 35 patients were graded T4sn and 21 patients T4sc. The median age was 62.5 years. The two groups were similar for sex, age, tobacco consumption, ASA score, NYHA, Charlson's index, spirometric parameters, cardiovascular comorbidity and history of previous extra-thoracic malignancies. All had a complete anatomic resection with mediastinal lymphadenectomy. Thirty-day mortality rate was 3.6%. Overall 5-year and 10-year survival rates were 48.2% and 29.9%, respectively. There was a non-significant trend for a worse survival in T4sn group patients when compared to that of T4sc group patients: 42.9% vs 52.3% at 5 years, and 25% vs 34.9% at 10 years (p=0.62). Multifocal T4 stage IIIB disease is a heterogeneous category where overall prognosis is far better than those of other T4 subgroups. Survival rates associated with pT4sn and pT4sc look roughly similar because of the small size of the subgroups usually submitted to comparison in most series. In the present experience, respective survival figures diverge, suggesting different biological behaviours.

  4. Cloning, characterization, and production of a novel lysozyme by different expression hosts.

    Science.gov (United States)

    Zhang, Haifeng; Fu, Gang; Zhang, Dawei

    2014-10-01

    Lysozyme is a protein found in egg white, tears, saliva, and other secretions. As a marketable natural alternative to preservatives, lysozyme can act as a natural antibiotic. In this study, we have isolated Bacillus licheniformis TIB320 from soil, which contains a lysozyme gene with various features. We have cloned and expressed the lysozyme in E. coli. The antimicrobial activity of the lysozyme showed that it had a broad antimicrobial spectrum against several standard strains. The lysozyme could maintain efficient activities in a pH range between 3 and 9 and from 20°C to 60°C, respectively. The lysozyme was resistant to pepsin and trypsin to some extent at 40°C. Production of the lysozyme was optimized by using various expression strategies in B. subtilis WB800. The lysozyme from B. licheniformis TIB320 will be promising as a food or feed additive.

  5. Investigation of genome sequence and gene expression regulation in T4 related bacteriophages

    OpenAIRE

    Kalinienė, Laura

    2010-01-01

    The complete genome sequence of bacteriophage VR7 has been determined. The genome sequence is 169,285 nt, with an overall G+C content of 40,3%, compared with 35.3 % of T4. VR7 encodes 293 putative protein-encoding open reading frames (ORFs) and tRNAMet. In total, 211 of the 293 VR7 open reading frames encode putative proteins that share 30% ‒ 97% amino acid sequence identity with those found in T4; 46 ORFs resemble genes from other T4-like phages, 9 show similarities to genes of non T4 type p...

  6. Isolation and purification of lysozyme from the hen egg white

    OpenAIRE

    DEKINA S.S.; ROMANOVSKA I.I.; OVSEPYAN A.M.; BODYUL M.G.; TOPTIKOV V.A.

    2015-01-01

    The aim of the research was the development of the method of lysozyme isolation from hen egg proteins. Lysozyme was isolated by differential heat denaturation of proteins with changing of the medium pH value, followed by neutralization, dialysis and additional purification by gel chromatography on Sephadex G-50. Activity was determined by bacteriolytic method (with Micrococcus lysodeikticus 4698 as a substrate). The enzyme purity and molecular mass were determined using SDS-electrophoresis an...

  7. Third party annotation gene data set of eutherian lysozyme genes

    OpenAIRE

    Premzl, Marko

    2014-01-01

    The eutherian comparative genomic analysis protocol annotated most comprehensive eutherian lysozyme gene data set. Among 209 potential coding sequences, the third party annotation gene data set of eutherian lysozyme genes included 116 complete coding sequences that first described seven major gene clusters. As one new framework of future experiments, the present integrated gene annotations, phylogenetic analysis and protein molecular evolution analysis proposed new classification and nomencla...

  8. Modelling of microstructured waveguides using a finite-element-based vectorial mode solver with transparent boundary condition

    NARCIS (Netherlands)

    Uranus, H.P.; Hoekstra, Hugo; van Groesen, Embrecht W.C.; Bienstman, P.; Vanholme, L.

    2004-01-01

    Finite element vectorial optical mode solver is used to analyze microstructured waveguides in a relatively small computational domain. The presentation will consider the computational method, as well as the applications of it on a number of waveguides with 2-D cross section where microstructures are

  9. Propagation-invariant vectorial Bessel beams by use of sub wavelength quantized Pancharatnam-Berry phase optics

    International Nuclear Information System (INIS)

    Niv, A.; Biener, G.; Kleiner, V.; Hasman, E.

    2004-01-01

    Full Text:Propagation-invariant scalar fields have been extensively studied both theoretically and experimentally, since they were proposed by Durnin et al. These fields were employed in applications such as optical tweezers and for transport and guiding of microspheres. Although there has recently been considerable theoretical interest in propagation-invariant vectorial beams, experimental studies of such beams have remained somewhat limited. One of the most interesting types of propagation-invariant vectorial beam is the linearly polarized axially symmetric beam (LPASB) [l]. Recently, we introduced and experimentally demonstrated propagation-invariant vectorial Bessel beams with linearly polarized axial symmetry based on quantized Pancharatnam-Berry phase optical elements (QPBOEs) [21 and an axicon. QP-BOEs utilize the geometric phase that accompanies space-variant polarization manipulations to achieve a desired phase modification [31. To test our approach we formed QPBOEs with different polarization orders as computer-generated space-variant sub wavelength gratings upon GaAs wafers for use with 10.6 micron laser radiation. The resultant beams were also transmitted through a polarizer that produced a unique propagation-invariant scalar beam. This beam has a propeller-shaped intensity pattern that can be rotated by simple rotation of the polarizer. We therefore have demonstrated the formation of a vectorial Bessel beam by using simple, lightweight thin elements and exploited that beam to perform a controlled rotation of a propeller-shaped intensity pattern that can be suitable for optical tweezers

  10. Penetration and fusion of phospholipid vesicles by lysozyme

    International Nuclear Information System (INIS)

    Kim, J.; Kim, H.

    1989-01-01

    The lysozyme-induced fusion of phosphatidylserine/phosphatidylethanolamine vesicles as studied at a wide range of pH is found to correlate well with the binding of this protein to the vesicles. An identical 6000 molecular weight segment of lysozyme at the N-terminal region is found to be protected from tryptic digestion when initially incubated with vesicles at several pH values. Only this segment is labeled by dansyl chloride, which is partitioned into the bilayer. These results suggest the penetration of one segment of lysozyme into the bilayer. Photoactivated labeling of the membrane-penetrating segment of lysozyme with 3-(trifluoromethyl)-3-([ 125 I]iodophenyl)diazirine ([ 125 I]TID) and subsequent identification of the labeled residues by Edman degradation and gamma-ray counting indicate that four amino acids from the N-terminal are located outside the hydrophobic core of the bilayer. Although treatment of the membrane-embedded segment with aminopeptidase failed to cleave any amino acids from the N-terminal, it appears that a loop of lysozyme segment near the N-terminal penetrates into the bilayer at acidic pH. A helical wheel diagram shows that the labeling is done mainly on one surface of the alpha-helix. The penetration kinetics as studied by time-dependent [ 125 I]TID labeling coincide with the fusion kinetics, strongly suggesting that the penetration of the lysozyme segment into the vesicles is the cause of the fusion

  11. Strong and Selective Adsorption of Lysozyme on Graphene Oxide

    Science.gov (United States)

    2015-01-01

    Biosensing methods and devices using graphene oxide (GO) have recently been explored for detection and quantification of specific biomolecules from body fluid samples, such as saliva, milk, urine, and serum. For a practical diagnostics application, any sensing system must show an absence of nonselective detection of abundant proteins in the fluid matrix. Because lysozyme is an abundant protein in these body fluids (e.g., around 21.4 and 7 μg/mL of lysozyme is found in human milk and saliva from healthy individuals, and more than 15 or even 100 μg/mL in patients suffering from leukemia, renal disease, and sarcoidosis), it may interfere with detections and quantification if it has strong interaction with GO. Therefore, one fundamental question that needs to be addressed before any development of GO based diagnostics method is how GO interacts with lysozyme. In this study, GO has demonstrated a strong interaction with lysozyme. This interaction is so strong that we are able to subsequently eliminate and separate lysozyme from aqueous solution onto the surface of GO. Furthermore, the strong electrostatic interaction also renders the selective adsorption of lysozyme on GO from a mixture of binary and ternary proteins. This selectivity is confirmed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), fluorescence spectroscopy, and UV–vis absorption spectroscopy. PMID:24684375

  12. Study of ethanol-lysozyme interactions using neutron diffraction

    International Nuclear Information System (INIS)

    Lehmann, M.S.; Mason, S.A.; McIntyre, G.J.

    1985-01-01

    Single-crystal neutron diffraction has been used to observe the interactions between deuterated ethanol (CD3CD2OH) and lysozyme in triclinic crystals of hen egg white lysozyme soaked in 25% (v/v) ethanol solutions. A total of 6047 observed reflections to a resolution of 2 A were used, and 13 possible ethanol sites were identified. The three highest occupied sites are close to locations for bromoethanol found in an earlier study by Yonath et al. [Yonath, A., Podjarny, A., Honig, B., Traub, W., Sielecki, A., Herzberg, O., and Moult, J. (1978) Biophys. Struct. Mech. 4, 27-36]. Structure refinements including a model for the flat solvent lead to a final crystallographic agreement factor of 0.097. Comparison with earlier neutron studies on triclinic lysozyme showed that neither the molecular structure nor the thermal motions were affected significantly by the ethanol. A detailed analysis of the ethanol-lysozyme contacts showed 61% of these to be with hydrophobic sites, in agreement with the dominant hydrophobic nature of ethanol. This, together with the fact that the molecular structure of lysozyme is not perturbed, suggests a model for denaturation of lysozyme by alcohol, which proceeds via a dehydration of the protein at high alcohol concentration

  13. Lysozyme M deficiency leads to an increased susceptibility to Streptococcus pneumoniae-induced otitis media

    Directory of Open Access Journals (Sweden)

    Woo Jeong-Im

    2008-10-01

    Full Text Available Abstract Background Lysozyme is an antimicrobial innate immune molecule degrading peptidoglycan of the bacterial cell wall. Lysozyme shows the ubiquitous expression in wide varieties of species and tissues including the tubotympanum of mammals. We aim to investigate the effects of lysozyme depletion on pneumococcal clearance from the middle ear cavity. Methods Immunohistochemistry was performed to localize lysozyme in the Eustachian tube. Lysozyme expression was compared between the wild type and the lysozyme M-/- mice using real time quantitative RT-PCR and western blotting. Muramidase activity and bactericidal activity of lysozyme was measured using a lysoplate radial diffusion assay and a liquid broth assay, respectively. To determine if depletion of lysozyme M increases a susceptibility to pneumococal otitis media, 50 CFU of S. pneumoniae 6B were transtympanically inoculated to the middle ear and viable bacteria were counted at day 3 and 7 with clinical grading of middle ear inflammation. Results Immunolabeling revealed that localization of lysozyme M and lysozyme P is specific to some/particular cell types of the Eustachian tube. Lysozyme P of lysozyme M-/- mice was mainly expressed in the submucosal gland but not in the tubal epithelium. Although lysozyme M-/- mice showed compensatory up-regulation of lysozyme P, lysozyme M depletion resulted in a decrease in both muramidase and antimicrobial activities. Deficiency in lysozyme M led to an increased susceptibility to middle ear infection with S. pneumoniae 6B and resulted in severe middle ear inflammation, compared to wild type mice. Conclusion The results suggest that lysozyme M plays an important role in protecting the middle ear from invading pathogens, particularly in the early phase. We suggest a possibility of the exogenous lysozyme as an adjuvant therapeutic agent for otitis media, but further studies are necessary.

  14. Campylobacter jejuni group III phage CP81 contains many T4-like genes without belonging to the T4-type phage group: implications for the evolution of T4 phages.

    Science.gov (United States)

    Hammerl, Jens A; Jäckel, Claudia; Reetz, Jochen; Beck, Sebastian; Alter, Thomas; Lurz, Rudi; Barretto, Caroline; Brüssow, Harald; Hertwig, Stefan

    2011-09-01

    CP81 is a virulent Campylobacter group III phage whose linear genome comprises 132,454 bp. At the nucleotide level, CP81 differs from other phages. However, a number of its structural and replication/recombination proteins revealed a relationship to the group II Campylobacter phages CP220/CPt10 and to T4-type phages. Unlike the T4-related phages, the CP81 genome does not contain conserved replication and virion modules. Instead, the respective genes are scattered throughout the phage genome. Moreover, most genes for metabolic enzymes of CP220/CPt10 are lacking in CP81. On the other hand, the CP81 genome contains nine similar genes for homing endonucleases which may be involved in the attrition of the conserved gene order for the virion core genes of T4-type phages. The phage apparently possesses an unusual modification of C or G bases. Efficient cleavage of its DNA was only achieved with restriction enzymes recognizing pure A/T sites. Uncommonly, phenol extraction leads to a significant loss of CP81 DNA from the aqueous layer, a property not yet described for other phages belonging to the T4 superfamily.

  15. Campylobacter jejuni Group III Phage CP81 Contains Many T4-Like Genes without Belonging to the T4-Type Phage Group: Implications for the Evolution of T4 Phages▿†

    Science.gov (United States)

    Hammerl, Jens A.; Jäckel, Claudia; Reetz, Jochen; Beck, Sebastian; Alter, Thomas; Lurz, Rudi; Barretto, Caroline; Brüssow, Harald; Hertwig, Stefan

    2011-01-01

    CP81 is a virulent Campylobacter group III phage whose linear genome comprises 132,454 bp. At the nucleotide level, CP81 differs from other phages. However, a number of its structural and replication/recombination proteins revealed a relationship to the group II Campylobacter phages CP220/CPt10 and to T4-type phages. Unlike the T4-related phages, the CP81 genome does not contain conserved replication and virion modules. Instead, the respective genes are scattered throughout the phage genome. Moreover, most genes for metabolic enzymes of CP220/CPt10 are lacking in CP81. On the other hand, the CP81 genome contains nine similar genes for homing endonucleases which may be involved in the attrition of the conserved gene order for the virion core genes of T4-type phages. The phage apparently possesses an unusual modification of C or G bases. Efficient cleavage of its DNA was only achieved with restriction enzymes recognizing pure A/T sites. Uncommonly, phenol extraction leads to a significant loss of CP81 DNA from the aqueous layer, a property not yet described for other phages belonging to the T4 superfamily. PMID:21697478

  16. CXCR4 mediated chemotaxis is regulated by 5T4 oncofetal glycoprotein in mouse embryonic cells.

    Directory of Open Access Journals (Sweden)

    Thomas D Southgate

    2010-04-01

    Full Text Available 5T4 oncofetal molecules are highly expressed during development and upregulated in cancer while showing only low levels in some adult tissues. Upregulation of 5T4 expression is a marker of loss of pluripotency in the early differentiation of embryonic stem (ES cells and forms an integrated component of an epithelial-mesenchymal transition, a process important during embryonic development and metastatic spread of epithelial tumors. Investigation of the transcriptional changes in early ES differentiation showed upregulation of CXCL12 and down-regulation of a cell surface protease, CD26, which cleaves this chemokine. CXCL12 binds to the widely expressed CXCR4 and regulates key aspects of development, stem cell motility and tumour metastasis to tissues with high levels of CXCL12. We show that the 5T4 glycoprotein is required for optimal functional cell surface expression of the chemokine receptor CXCR4 and CXCL12 mediated chemotaxis in differentiating murine embryonic stem cells and embryo fibroblasts (MEF. Cell surface expression of 5T4 and CXCR4 molecules is co-localized in differentiating ES cells and MEF. By contrast, differentiating ES and MEF derived from 5T4 knockout (KO mice show only intracellular CXCR4 expression but infection with adenovirus encoding mouse 5T4 restores CXCL12 chemotaxis and surface co-localization with 5T4 molecules. A series of chimeric constructs with interchanged domains of 5T4 and the glycoprotein CD44 were used to map the 5T4 sequences relevant for CXCR4 membrane expression and function in 5T4KO MEF. These data identified the 5T4 transmembrane domain as sufficient and necessary to enable CXCR4 cell surface expression and chemotaxis. Furthermore, some monoclonal antibodies against m5T4 can inhibit CXCL12 chemotaxis of differentiating ES cells and MEF which is not mediated by simple antigenic modulation. Collectively, these data support a molecular interaction of 5T4 and CXCR4 occurring at the cell surface which

  17. Three-dimensional polarization marked multiple-QR code encryption by optimizing a single vectorial beam

    Science.gov (United States)

    Lin, Chao; Shen, Xueju; Hua, Binbin; Wang, Zhisong

    2015-10-01

    We demonstrate the feasibility of three dimensional (3D) polarization multiplexing by optimizing a single vectorial beam using a multiple-signal window multiple-plane (MSW-MP) phase retrieval algorithm. Original messages represented with multiple quick response (QR) codes are first partitioned into a series of subblocks. Then, each subblock is marked with a specific polarization state and randomly distributed in 3D space with both longitudinal and transversal adjustable freedoms. A generalized 3D polarization mapping protocol is established to generate a 3D polarization key. Finally, multiple-QR code is encrypted into one phase only mask and one polarization only mask based on the modified Gerchberg-Saxton (GS) algorithm. We take the polarization mask as the cyphertext and the phase only mask as additional dimension of key. Only when both the phase key and 3D polarization key are correct, original messages can be recovered. We verify our proposal with both simulation and experiment evidences.

  18. Thermodynamics of electron transfer and its coupling to vectorial processes in biological membranes.

    Science.gov (United States)

    Arata, H; Nishimura, M

    1980-11-01

    A method is developed to express the flux of an electron transfer reaction as a function of the conjugate force, the redox potential difference, throughout the nonlinear region. The flux can be expressed by a product of the hyperbolic sine of the force, a factor ("redox-poising parameter") determined by the redox potentials of subsystem (in certain cases by local pH's and pK's of subsystems), and some constants. This is analogous to the expression of the flux of a diffusion process by the product of its force and the concentration of the diffusing species. The redox-poising parameter corresponds to the concentration term. The expression is applied to redox chains in which electron transfers are coupled to vectorial processes such as proton translocation or electric current.

  19. Enhanced activity of lysozyme-AgNP conjugate with synergic antibacterial effect without damaging the catalytic site of lysozyme.

    Science.gov (United States)

    Ernest, Vinita; Gajalakshmi, Sekar; Mukherjee, Amitava; Chandrasekaran, Natarajan

    2014-10-01

    The conjugation of silver nanoparticles (AgNPs) with biomolecules such as oligosaccharides, DNA, proteins has attracted great attention of scientists recently. In this study, lysozyme-AgNP conjugates were evaluated for its synergic antimicrobial effect. AgNPs were synthesized and characterized using UV-Visible, X-ray diffraction analysis (XRD) and atomic force microscopy (AFM). AgNP (0-1 mM) was interacted with lysozyme for multi-spectrophotometric studies. Lysozyme was immobilized on AgNP at different ratios and the resulting nano-bio-conjugate was tested against Escherichia coli for potent synergic antibacterial effects. A surface plasmon peak at 420 nm confirmed the presence of AgNPs and spherical to oval-shaped AgNPs were observed by AFM. The particle size was calculated to be 25 nm by XRD analysis. The maximum immobilization efficiency (98%) was achieved at 0.01:1 ratio of enzyme:AgNP. UV-Visible and fluorescence spectral studies revealed the binding of AgNPs to lysozyme by the formation of ground-state complex and the binding parameters were calculated. Circular dichroism studies confirmed decrease by 11% in the α-helical and 29.32% in β-sheets of lysozyme upon AgNP interaction. FTIR spectra revealed the binding of AgNP through thiol (-SH) linkages of lysozyme. Our results showed that the antimicrobial activity of lysozyme-AgNP conjugate was enhanced up to 86% decrease in the cell growth. In summary, the immobilization of lysozyme on AgNP has yielded a nano-bio-conjugate with synergistic antibacterial properties.

  20. Stoichiometry of vectorial H+ movements coupled to electron transport and to ATP synthesis in mitochondria

    Science.gov (United States)

    Alexandre, Adolfo; Reynafarje, Baltazar; Lehninger, Albert L.

    1978-01-01

    In order to verify more directly our earlier measurements showing that, on the average, close to four vectorial H+ are rejected per pair of electrons passing each of the three energy-conserving sites of the mitochondrial electron transport chain, direct tests of the H+/2e- ratio for sites 2 and 3 were carried out in the presence of permeant charge-compensating cations. Site 2 was examined by utilizing succinate as electron donor and ferricyanide as electron acceptor from mitochondrial cytochrome c; the directly measured H+/2e- ratio was close to 4. Energy-conserving site 3 was isolated for study with ferrocyanide or ascorbate plus tetramethylphenylenediamine as electron donors to cytochrome c and with oxygen as electron acceptor. The directly measured H+/2e- ratio for site 3 was close to 4. The H+/ATP ratio (number of vectorial H+ ejected per ATP hydrolyzed) was determined with a new method in which the steady-state rates of both H+ ejection and ATP hydrolysis were measured in the presence of K+ + valinomycin. The H+/ATP ratio was found to approach 3.0. A proton cycle for oxidative phosphorylation is proposed, in which four electrochemical H+ equivalents are ejected per pair of electrons passing each energy-conserving site; three of the H+ equivalents pass inward to derive ATP synthesis from ADP and phosphate and the fourth H+ is used to bring about the energy-requiring electrogenic expulsion of ATP4- in exchange for extramitochondrial ADP3-, via the H+/H2PO4- symporter. PMID:31621

  1. Structure-sweetness relationship in egg white lysozyme: role of lysine and arginine residues on the elicitation of lysozyme sweetness.

    Science.gov (United States)

    Masuda, Tetsuya; Ide, Nobuyuki; Kitabatake, Naofumi

    2005-10-01

    Lysozyme is one of the sweet-tasting proteins. To clarify the structure-sweetness relationship and the basicity-sweetness relationship in lysozyme, we have generated lysozyme mutants with Pichia systems. Alanine substitution of lysine residues demonstrated that two out of six lysine residues, Lys13 and Lys96, are required for lysozyme sweetness, while the remaining four lysine residues do not play a significant role in the perception of sweetness. Arginine substitution of lysine residues revealed that the basicity, but not the shape, of the side chain plays a significant role in sweetness. Single alanine substitutions of arginine residues showed that three arginine residues, Arg14, Arg21, and Arg73, play significant roles in lysozyme sweetness, whereas Arg45, Arg68, Arg125 and chemical modification by 1,2-cyclohexanedione did not affect sweetness. From investigation of the charge-specific mutations, we found that the basicity of a broad surface region formed by five positively charged residues, Lys13, Lys96, Arg14, Arg21, and Arg73, is required for lysozyme sweetness. Differences in the threshold values among sweet-tasting proteins might be caused by the broadness and/or the density of charged residues on the protein surface.

  2. Modelling T4 cell count as a marker of HIV progression in the ...

    African Journals Online (AJOL)

    Modelling T4 cell count as a marker of HIV progression in the absence of any defense mechanism. VSM Yadavalli, MMO Labeodan, S Udayabaskaran, N Forche. Abstract. The T4 cell count, which is considered one of the markers of disease progression in an HIV infected individual, is modelled in this paper. The World ...

  3. Specific action of T4 endonuclease V on damaged DNA in xeroderma pigmentosum cells in vivo

    International Nuclear Information System (INIS)

    Tanaka, K.; Hayakawa, H.; Sekiguchi, M.; Okada, Y.

    1977-01-01

    The specific action of T4 endonuclease V on damaged DNA in xeroderma pigmentosum cells was examined using an in vivo assay system with hemagglutinating virus of Japan (Sendai virus) inactivated by uv light. A clear dose response was observed between the level of uv-induced unscheduled DNA synthesis of xeroderma pigmentosum cells and the amount of T4 endonuclease V activity added. The T4 enzyme was unstable in human cells, and its half-life was 3 hr. Fractions derived from an extract of Escherichia coli infected with T4v 1 , a mutant defective in the endonuclease V gene, showed no ability to restore the uv-induced unscheduled DNA synthesis of xeroderma pigmentosum cells. However, fractions derived from an extract of T4D-infected E. coli with endonuclease V activity were effective. The T4 enzyme was effective in xeroderma pigmentosum cells on DNA damaged by uv light but not in cells damaged by 4-nitroquinoline 1-oxide. The results of these experiments show that the T4 enzyme has a specific action on human cell DNA in vivo. Treatment with the T4 enzyme increased the survival of group A xeroderma pigmentosum cells after uv irradiation

  4. Results of surgical treatment of T4 non-small cell lung cancer

    NARCIS (Netherlands)

    Pitz, CCM; de la Riviere, AB; van Swieten, HA; Westermann, CJJ; Lammers, JWJ; van den Bosch, JMM

    2003-01-01

    Objective: Because of location and invasion of surrounding structures, the role of surgical treatment for T4 tumors remains unclear. Extended resections carry a high mortality and should be restricted for selected patients. This study clarifies the selection process in non-small cell T4 tumors with

  5. Assembly and dynamics of the bacteriophage T4 homologous recombination machinery

    Directory of Open Access Journals (Sweden)

    Morrical Scott W

    2010-12-01

    Full Text Available Abstract Homologous recombination (HR, a process involving the physical exchange of strands between homologous or nearly homologous DNA molecules, is critical for maintaining the genetic diversity and genome stability of species. Bacteriophage T4 is one of the classic systems for studies of homologous recombination. T4 uses HR for high-frequency genetic exchanges, for homology-directed DNA repair (HDR processes including DNA double-strand break repair, and for the initiation of DNA replication (RDR. T4 recombination proteins are expressed at high levels during T4 infection in E. coli, and share strong sequence, structural, and/or functional conservation with their counterparts in cellular organisms. Biochemical studies of T4 recombination have provided key insights on DNA strand exchange mechanisms, on the structure and function of recombination proteins, and on the coordination of recombination and DNA synthesis activities during RDR and HDR. Recent years have seen the development of detailed biochemical models for the assembly and dynamics of presynaptic filaments in the T4 recombination system, for the atomic structure of T4 UvsX recombinase, and for the roles of DNA helicases in T4 recombination. The goal of this chapter is to review these recent advances and their implications for HR and HDR mechanisms in all organisms.

  6. AtlasT4SS: A curated database for type IV secretion systems

    Directory of Open Access Journals (Sweden)

    Souza Rangel C

    2012-08-01

    Full Text Available Abstract Background The type IV secretion system (T4SS can be classified as a large family of macromolecule transporter systems, divided into three recognized sub-families, according to the well-known functions. The major sub-family is the conjugation system, which allows transfer of genetic material, such as a nucleoprotein, via cell contact among bacteria. Also, the conjugation system can transfer genetic material from bacteria to eukaryotic cells; such is the case with the T-DNA transfer of Agrobacterium tumefaciens to host plant cells. The system of effector protein transport constitutes the second sub-family, and the third one corresponds to the DNA uptake/release system. Genome analyses have revealed numerous T4SS in Bacteria and Archaea. The purpose of this work was to organize, classify, and integrate the T4SS data into a single database, called AtlasT4SS - the first public database devoted exclusively to this prokaryotic secretion system. Description The AtlasT4SS is a manual curated database that describes a large number of proteins related to the type IV secretion system reported so far in Gram-negative and Gram-positive bacteria, as well as in Archaea. The database was created using the RDBMS MySQL and the Catalyst Framework based in the Perl programming language and using the Model-View-Controller (MVC design pattern for Web. The current version holds a comprehensive collection of 1,617 T4SS proteins from 58 Bacteria (49 Gram-negative and 9 Gram-Positive, one Archaea and 11 plasmids. By applying the bi-directional best hit (BBH relationship in pairwise genome comparison, it was possible to obtain a core set of 134 clusters of orthologous genes encoding T4SS proteins. Conclusions In our database we present one way of classifying orthologous groups of T4SSs in a hierarchical classification scheme with three levels. The first level comprises four classes that are based on the organization of genetic determinants, shared homologies, and

  7. Encapsulation and controlled release of lysozyme from electrospun poly(epsilon-caprolactone)/poly(ethylene glycol) non-woven membranes by formation of lysozyme-oleate complexes.

    Science.gov (United States)

    Li, Yan; Jiang, Hongliang; Zhu, Kangjie

    2008-02-01

    In this study, the concept of hydrophobic ion pairing was adopted for incorporating lysozyme into electrospun poly(epsilon-caprolactone) (PCL)/poly(ethylene glycol) (PEG) non-woven membranes. The solubility of lysozyme in organic solvent was enhanced through the formation of lysozyme-oleate complexes, which could be directly loaded into PCL/PEG membranes using electrospinning technique. The resultant PCL/PEG nanofibers have a compact structure with an average diameter ranged from about 0.4 microm to 0.9 microm. The addition of PEG into the PCL nanofibers not only improved the hydrophilicity of the membrane, but also played an important role on in vitro lysozyme release rate. It was found that the release rate of lysozyme was enhanced with the increase of PEG content. In addition, the increase of salt concentration in the release medium accelerated lysozyme release. It has also been shown that the released lysozyme retained most of its enzymatic activity.

  8. Identification of chicken lysozyme g2 and its expression in the intestine.

    Science.gov (United States)

    Nile, C J; Townes, C L; Michailidis, G; Hirst, B H; Hall, J

    2004-11-01

    Lysozyme is an important component of the innate immune system, protecting the gastrointestinal tract from infection. The aim of the present study was to determine if lysozyme is expressed in the chicken ( Gallus gallus) intestine and to characterise the molecular forms expressed. Immunohistochemical staining localised lysozyme to epithelial cells of the villous epithelium along the length of the small intestine. There was no evidence for lysozyme expression in crypt epithelium and no evidence for Paneth cells. Immunoblots of chicken intestinal protein revealed three proteins: a 14-kDa band consistent with lysozyme c, and two additional bands of approximately 21 and 23 kDa, the latter consistent with lysozyme g. RT-PCR analyses confirmed that lysozyme c mRNA is expressed in 4-day, but not older chicken intestine and lysozyme g in 4- to 35-day chicken intestine. A novel chicken lysozyme g2 gene was identified by in silico analyses and mRNA for this lysozyme g2 was identified in the intestine from chickens of all ages. Chicken lysozyme g2 shows similarity with fish lysozyme g, including the absence of a signal peptide and cysteines involved in disulphide bond formation of the mammalian and bird lysozyme g proteins. Analyses using SecretomeP predict that chicken lysozyme g2 may be secreted by the non-classical secretory pathway. We conclude that lysozyme is expressed in the chicken small intestine by villous enterocytes. Lysozyme c, lysozyme g and g2 may fulfil complimentary roles in protecting the intestine.

  9. Genomic characteristics and environmental distributions of the uncultivated Far-T4 phages

    Directory of Open Access Journals (Sweden)

    Simon eRoux

    2015-03-01

    Full Text Available Viral metagenomics (viromics is a tremendous tool to reveal viral diversity and ecosystem functional roles across ecosystems ranging from the human gut to the world's oceans. As with microbes however, there appear vast swaths of 'dark matter' yet to be documented for viruses, even among relatively well-studied viral types. Here, we use viromics to explore the 'Far-T4 phages' sequence space, a neighbor clade from the well-studied T4-like phages that was first detected through PCR study in seawater and subsequently identified in freshwater lakes through 454-sequenced viromes. To advance the description of these viruses beyond this single marker gene, we explore Far-T4 genome fragments assembled from 2 deeply-sequenced freshwater viromes. Single gene phylogenetic trees confirm that the Far-T4 phages are divergent from the T4-like phages, genome fragments reveal largely collinear genome organization, and both data led to the delineation of 5 Far-T4 clades. Three-dimensional models of major capsid proteins are consistent with a T4-like structure, and highlight highly conserved core flanked by variable insertions. Finally, we contextualize these now better characterized Far-T4 phages by re-analyzing 196 previously published viromes. These suggest that Far-T4 are common in freshwater and seawater as only 4 of 82 aquatic viromes lacked Far-T4-like sequences. Variability in representation across the 5 newly identified clades suggests clade-specific niche differentiation may be occurring across the different biomes, though the underlying mechanism remains unidentified. While complete genome assembly from complex communities and the lack of host linkage information still bottleneck virus discovery through viromes, these findings exemplify the power of metagenomics approaches to assess the diversity, evolutionary history, and genomic characteristics of novel uncultivated phages.

  10. Genomic characteristics and environmental distributions of the uncultivated Far-T4 phages.

    Science.gov (United States)

    Roux, Simon; Enault, François; Ravet, Viviane; Pereira, Olivier; Sullivan, Matthew B

    2015-01-01

    Viral metagenomics (viromics) is a tremendous tool to reveal viral taxonomic and functional diversity across ecosystems ranging from the human gut to the world's oceans. As with microbes however, there appear vast swaths of "dark matter" yet to be documented for viruses, even among relatively well-studied viral types. Here, we use viromics to explore the "Far-T4 phages" sequence space, a neighbor clade from the well-studied T4-like phages that was first detected through PCR study in seawater and subsequently identified in freshwater lakes through 454-sequenced viromes. To advance the description of these viruses beyond this single marker gene, we explore Far-T4 genome fragments assembled from two deeply-sequenced freshwater viromes. Single gene phylogenetic trees confirm that the Far-T4 phages are divergent from the T4-like phages, genome fragments reveal largely collinear genome organizations, and both data led to the delineation of five Far-T4 clades. Three-dimensional models of major capsid proteins are consistent with a T4-like structure, and highlight a highly conserved core flanked by variable insertions. Finally, we contextualize these now better characterized Far-T4 phages by re-analyzing 196 previously published viromes. These suggest that Far-T4 are common in freshwater and seawater as only four of 82 aquatic viromes lacked Far-T4-like sequences. Variability in representation across the five newly identified clades suggests clade-specific niche differentiation may be occurring across the different biomes, though the underlying mechanism remains unidentified. While complete genome assembly from complex communities and the lack of host linkage information still bottleneck virus discovery through viromes, these findings exemplify the power of metagenomics approaches to assess the diversity, evolutionary history, and genomic characteristics of novel uncultivated phages.

  11. Lysozyme association with circulating RNA, extracellular vesicles, and chronic stress.

    Science.gov (United States)

    Abey, Sarah K; Yuana, Yuana; Joseph, Paule V; Kenea, Natnael D; Fourie, Nicolaas H; Sherwin, LeeAnne B; Gonye, Gregory E; Smyser, Paul A; Stempinski, Erin S; Boulineaux, Christina M; Weaver, Kristen R; Bleck, Christopher K E; Henderson, Wendy A

    2017-06-01

    Stress has demonstrated effects on inflammation though underlying cell-cell communication mechanisms remain unclear. We hypothesize that circulating RNAs and extracellular vesicles (EVs) in patients with chronic stress contain signals with functional roles in cell repair. Blood transcriptome from patients with Irritable Bowel Syndrome versus controls were compared to identify signaling pathways and effectors. Plasma EVs were isolated (size-exclusion chromatography) and characterized for effectors' presence (immunogold labelling-electron microscopy). Based on transcriptome pathways and EV-labelling, lysozyme's effects on cell migration were tested in human colon epithelial CRL-1790 cells and compared to the effects of CXCL12, a migration inducer (wound assay). The effect of lysozyme on immune-linked mRNA and protein levels in cells which survived following serum starvation and scratch wound were investigated (NanoString). Blood transcriptomes revealed pyridoxal 5'phosphate salvage, pyrimidine ribonucleotides salvage pathways, atherosclerosis, and cell movement signaling with membrane CD9 and extracellular lysozyme as effectors. Plasma EVs showed labelling with CD9, mucins, and lysozyme. This is the first identification of lysozyme on plasma EVs. In CRL-1790 cells, lysozyme induced migration and repaired scratch wound as well as CXCL12. Immune mRNA and protein expressions were altered in cells which survived following serum starvation and scratch wound, with or without lysozyme in serum-free media post-wounding: CD9, IL8, IL6 mRNAs and CD9, NT5E, PD-L1 proteins. Repair and inflammatory signals are identified in plasma EVs and circulating RNAs in chronic stress. Registered clinicaltrials.gov #NCT00824941. This study highlights the role of circulating RNAs and EVs in stress.

  12. A new lysozyme from the eastern oyster, Crassostrea virginica, and a possible evolutionary pathway for i-type lysozymes in bivalves from host defense to digestion

    Science.gov (United States)

    2010-01-01

    Background Lysozymes are enzymes that lyse bacterial cell walls, an activity widely used for host defense but also modified in some instances for digestion. The biochemical and evolutionary changes between these different functional forms has been well-studied in the c-type lysozymes of vertebrates, but less so in the i-type lysozymes prevalent in most invertebrate animals. Some bivalve molluscs possess both defensive and digestive lysozymes. Results We report a third lysozyme from the oyster Crassostrea virginica, cv-lysozyme 3. The chemical properties of cv-lysozyme 3 (including molecular weight, isoelectric point, basic amino acid residue number, and predicted protease cutting sites) suggest it represents a transitional form between lysozymes used for digestion and immunity. The cv-lysozyme 3 protein inhibited the growth of bacteria (consistent with a defensive function), but semi-quantitative RT-PCR suggested the gene was expressed mainly in digestive glands. Purified cv-lysozyme 3 expressed maximum muramidase activity within a range of pH (7.0 and 8.0) and ionic strength (I = 0.005-0.01) unfavorable for either cv-lysozyme 1 or cv-lysozyme 2 activities. The topology of a phylogenetic analysis of cv-lysozyme 3 cDNA (full length 663 bp, encoding an open reading frame of 187 amino acids) is also consistent with a transitional condition, as cv-lysozyme 3 falls at the base of a monophyletic clade of bivalve lysozymes identified from digestive glands. Rates of nonsynonymous substitution are significantly high at the base of this clade, consistent with an episode of positive selection associated with the functional transition from defense to digestion. Conclusion The pattern of molecular evolution accompanying the shift from defensive to digestive function in the i-type lysozymes of bivalves parallels those seen for c-type lysozymes in mammals and suggests that the lysozyme paralogs that enhance the range of physiological conditions for lysozyme activity may provide

  13. A new lysozyme from the eastern oyster, Crassostrea virginica, and a possible evolutionary pathway for i-type lysozymes in bivalves from host defense to digestion

    Directory of Open Access Journals (Sweden)

    Itoh Naoki

    2010-07-01

    Full Text Available Abstract Background Lysozymes are enzymes that lyse bacterial cell walls, an activity widely used for host defense but also modified in some instances for digestion. The biochemical and evolutionary changes between these different functional forms has been well-studied in the c-type lysozymes of vertebrates, but less so in the i-type lysozymes prevalent in most invertebrate animals. Some bivalve molluscs possess both defensive and digestive lysozymes. Results We report a third lysozyme from the oyster Crassostrea virginica, cv-lysozyme 3. The chemical properties of cv-lysozyme 3 (including molecular weight, isoelectric point, basic amino acid residue number, and predicted protease cutting sites suggest it represents a transitional form between lysozymes used for digestion and immunity. The cv-lysozyme 3 protein inhibited the growth of bacteria (consistent with a defensive function, but semi-quantitative RT-PCR suggested the gene was expressed mainly in digestive glands. Purified cv-lysozyme 3 expressed maximum muramidase activity within a range of pH (7.0 and 8.0 and ionic strength (I = 0.005-0.01 unfavorable for either cv-lysozyme 1 or cv-lysozyme 2 activities. The topology of a phylogenetic analysis of cv-lysozyme 3 cDNA (full length 663 bp, encoding an open reading frame of 187 amino acids is also consistent with a transitional condition, as cv-lysozyme 3 falls at the base of a monophyletic clade of bivalve lysozymes identified from digestive glands. Rates of nonsynonymous substitution are significantly high at the base of this clade, consistent with an episode of positive selection associated with the functional transition from defense to digestion. Conclusion The pattern of molecular evolution accompanying the shift from defensive to digestive function in the i-type lysozymes of bivalves parallels those seen for c-type lysozymes in mammals and suggests that the lysozyme paralogs that enhance the range of physiological conditions for

  14. Salt-specific effects in lysozyme solutions

    Directory of Open Access Journals (Sweden)

    T. Janc

    2016-03-01

    Full Text Available The effects of additions of low-molecular-mass salts on the properties of aqueous lysozyme solutions are examined by using the cloud-point temperature, T_{cloud}, measurements. Mixtures of protein, buffer, and simple salt in water are studied at pH=6.8 (phosphate buffer and pH=4.6 (acetate buffer. We show that an addition of buffer in the amount above I_{buffer} = 0.6 mol dm^{-3} does not affect the T_{cloud} values. However, by replacing a certain amount of the buffer electrolyte by another salt, keeping the total ionic strength constant, we can significantly change the cloud-point temperature. All the salts de-stabilize the solution and the magnitude of the effect depends on the nature of the salt. Experimental results are analyzed within the framework of the one-component model, which treats the protein-protein interaction as highly directional and of short-range. We use this approach to predict the second virial coefficients, and liquid-liquid phase diagrams under conditions, where T_{cloud} is determined experimentally.

  15. Terahertz absorption of lysozyme in solution

    Science.gov (United States)

    Martin, Daniel R.; Matyushov, Dmitry V.

    2017-08-01

    Absorption of radiation by solution is described by its frequency-dependent dielectric function and can be viewed as a specific application of the dielectric theory of solutions. For ideal solutions, the dielectric boundary-value problem separates the polar response into the polarization of the void in the liquid, created by the solute, and the response of the solute dipole. In the case of a protein as a solute, protein nuclear dynamics do not project on significant fluctuations of the dipole moment in the terahertz domain of frequencies and the protein dipole can be viewed as dynamically frozen. Absorption of radiation then reflects the interfacial polarization. Here we apply an analytical theory and computer simulations to absorption of radiation by an ideal solution of lysozyme. Comparison with the experiment shows that Maxwell electrostatics fails to describe the polarization of the protein-water interface and the "Lorentz void," which does not anticipate polarization of the interface by the external field (no surface charges), better represents the data. An analytical theory for the slope of the solution absorption against the volume fraction of the solute is formulated in terms of the cavity field response function. It is calculated from molecular dynamics simulations in good agreement with the experiment. The protein hydration shell emerges as a separate sub-ensemble, which, collectively, is not described by the standard electrostatics of dielectrics.

  16. Genetic control of the humoral immune response to avian egg white lysozymes in the chicken

    International Nuclear Information System (INIS)

    Flanagan, M.P.

    1987-01-01

    Chickens from two closely related sublines, GHs-B6 and GHs-B13, differing serologically at the major histocompatibility complex, were significantly different in their humoral response to three avian egg white lysozymes. Specific antisera levels were measured by radioimmunoassay using 125 I-labeled lysozymes. Antibodies elicited in response to these lysozymes are assumed to be directed against sites on these lysozymes where their amino acid sequence differs from that of the recipient G. domesticus egg white lysozyme (HEL). GHs-B6 birds produced a high level of antibody in response to immunization of turkey (TEL), pheasant (PhL) and guinea hen (GHL) lysozymes. GHs-B13 birds produced no detectable antibody to TEL, were intermediate in their response to PhL and equaled the antibody production of GHs-B6 birds in response to GHL. Antisera to each lysozyme were examined for crossreactivity with all other lysozymes by use of a competitive binding assay

  17. Genome of low-temperature T4-related bacteriophage vB_EcoM-VR7.

    Science.gov (United States)

    Kaliniene, Laura; Klausa, Vytautas; Zajančkauskaite, Aurelija; Nivinskas, Rimas; Truncaite, Lidija

    2011-10-01

    The complete genome sequence of the T4-related low-temperature Escherichia coli bacteriophage vB_EcoM-VR7 was determined. The genome sequence is 169,285 bp long, with a G+C content of 40.3%. Overall, 95% of the phage genome is coding. It encodes 293 putative protein-encoding open reading frames (ORFs) and tRNA(Met). More than half (59%) of the genomic DNA lacks significant homology with the DNA of T4, but once translated, 72% of the vB_EcoM-VR7 genome (211 ORFs) encodes protein homologues of T4 genes. Overall, 46 vB_EcoM-VR7 ORFs have no homologues in T4 but are derived from other T4-related phages, nine ORFs show similarities to bacterial or non-T4-related phage genes, and 27 ORFs are unique to vB_EcoM-VR7. This phage lacks several T4 enzymes involved in host DNA degradation; however, there is extensive representation of the DNA replication, recombination and repair enzymes as well as the viral capsid and tail structural genes.

  18. Folding Behaviors of Protein (Lysozyme) Confined in Polyelectrolyte Complex Micelle.

    Science.gov (United States)

    Wu, Fu-Gen; Jiang, Yao-Wen; Chen, Zhan; Yu, Zhi-Wu

    2016-04-19

    The folding/unfolding behavior of proteins (enzymes) in confined space is important for their properties and functions, but such a behavior remains largely unexplored. In this article, we reported our finding that lysozyme and a double hydrophilic block copolymer, methoxypoly(ethylene glycol)5K-block-poly(l-aspartic acid sodium salt)10 (mPEG(5K)-b-PLD10), can form a polyelectrolyte complex micelle with a particle size of ∼30 nm, as verified by dynamic light scattering and transmission electron microscopy. The unfolding and refolding behaviors of lysozyme molecules in the presence of the copolymer were studied by microcalorimetry and circular dichroism spectroscopy. Upon complex formation with mPEG(5K)-b-PLD10, lysozyme changed from its initial native state to a new partially unfolded state. Compared with its native state, this copolymer-complexed new folding state of lysozyme has different secondary and tertiary structures, a decreased thermostability, and significantly altered unfolding/refolding behaviors. It was found that the native lysozyme exhibited reversible unfolding and refolding upon heating and subsequent cooling, while lysozyme in the new folding state (complexed with the oppositely charged PLD segments of the polymer) could unfold upon heating but could not refold upon subsequent cooling. By employing the heating-cooling-reheating procedure, the prevention of complex formation between lysozyme and polymer due to the salt screening effect was observed, and the resulting uncomplexed lysozyme regained its proper unfolding and refolding abilities upon heating and subsequent cooling. Besides, we also pointed out the important role the length of the PLD segment played during the formation of micelles and the monodispersity of the formed micelles. Furthermore, the lysozyme-mPEG(5K)-b-PLD10 mixtures prepared in this work were all transparent, without the formation of large aggregates or precipitates in solution as frequently observed in other protein

  19. Plasticity of the gene functions for DNA replication in the T4-like phages.

    Science.gov (United States)

    Petrov, Vasiliy M; Nolan, James M; Bertrand, Claire; Levy, Dawn; Desplats, Carine; Krisch, H M; Karam, Jim D

    2006-08-04

    We have completely sequenced and annotated the genomes of several relatives of the bacteriophage T4, including three coliphages (RB43, RB49 and RB69), three Aeromonas salmonicida phages (44RR2.8t, 25 and 31) and one Aeromonas hydrophila phage (Aeh1). In addition, we have partially sequenced and annotated the T4-like genomes of coliphage RB16 (a close relative of RB43), A. salmonicida phage 65, Acinetobacter johnsonii phage 133 and Vibrio natriegens phage nt-1. Each of these phage genomes exhibited a unique sequence that distinguished it from its relatives, although there were examples of genomes that are very similar to each other. As a group the phages compared here diverge from one another by several criteria, including (a) host range, (b) genome size in the range between approximately 160 kb and approximately 250 kb, (c) content and genetic organization of their T4-like genes for DNA metabolism, (d) mutational drift of the predicted T4-like gene products and their regulatory sites and (e) content of open-reading frames that have no counterparts in T4 or other known organisms (novel ORFs). We have observed a number of DNA rearrangements of the T4 genome type, some exhibiting proximity to putative homing endonuclease genes. Also, we cite and discuss examples of sequence divergence in the predicted sites for protein-protein and protein-nucleic acid interactions of homologues of the T4 DNA replication proteins, with emphasis on the diversity in sequence, molecular form and regulation of the phage-encoded DNA polymerase, gp43. Five of the sequenced phage genomes are predicted to encode split forms of this polymerase. Our studies suggest that the modular construction and plasticity of the T4 genome type and several of its replication proteins may offer resilience to mutation, including DNA rearrangements, and facilitate the adaptation of T4-like phages to different bacterial hosts in nature.

  20. Effects of Purification on the Crystallization of Lysozyme

    Science.gov (United States)

    Ewing, Felecia L.; Forsythe, Elizabeth L.; Van Der Woerd, Mark; Pusey, Marc L.

    1996-01-01

    We have additionally purified a commercial lysozyme preparation by cation exchange chromatography, followed by recrystallization. This material is 99.96% pure with respect to macromolecular impurities. At basic pH, the purified lysozyme gave only tetragonal crystals at 20 C. Protein used directly from the bottle, prepared by dialysis against distilled water, or which did not bind to the cation exchange column had considerably altered crystallization behavior. Lysozyme which did not bind to the cation exchange column was subsequently purified by size exclusion chromatography. This material gave predominately bundles of rod-shaped crystals with some small tetragonal crystals at lower pHs. The origin of the bundled rod habit was postulated to be a thermally dependent tetragonal- orthorhombic change in the protein structure. This was subsequently ruled out on the basis of crystallization behavior and growth rate experiments. This suggests that heterogeneous forms of lysozyme may be responsible. These results demonstrate three classes of impurities: (1) small molecules, which may be removed by dialysis; (2) macromolecules, which are removable by chromatographic techniques; and (3) heterogeneous forms of the protein, which can be removed in this case by cation exchange chromatography. Of these, heterogeneous forms of the lysozyme apparently have the greatest affect on its crystallization behavior.

  1. ISOLATION AND PURIFICATION OF LYSOZYME FROM THE HEN EGG WHITE

    Directory of Open Access Journals (Sweden)

    S. S.

    2015-12-01

    Full Text Available The aim of the research was the development of the method of lysozyme isolation from hen egg proteins. Lysozyme was isolated by differential heat denaturation of proteins with changing of the medium pH value, followed by neutralization, dialysis and additional purification by gel chromatography on Sephadex G-50. Activity was determined by bacteriolytic method (with Micrococcus lysodeikticus 4698 as a substrate. The enzyme purity and molecular mass were determined using SDS-electrophoresis and massspectrometry. The method of lysozyme isolation from hen egg proteins with the enzyme yield of 3.2 ± 0.2% and bacteriolytic activity of 22 025 ± 1 500 U/mg is modified. According to electrophoresis data, the isolated enzyme is characterized by high degree of purity (~95–98% and is comparable with lysozyme of AppliChem company by main physical and chemical characteristics. The obtaining product is stored in a crystalline form at low temperature (–24 оC for 9 months. The proposed method allows obtaining active and stable lysozyme with high purity from hen egg protein in laboratory conditions for the usage in biotechnology.

  2. MUCOADHESIVE GEL WITH IMMOBILIZED LYSOZYME: PREPARATION AND PROPERTIES

    Directory of Open Access Journals (Sweden)

    Dekina S. S.

    2015-08-01

    Full Text Available The study of non-covalent immobilized lysozyme, as well as physico-chemical and biochemical properties of obtained mucoadhesive gel was the aim of the research. Lysozyme activity was determined by bacteriolytic method (Micrococcus lysodeikticus cells acetone powder was a substrate. Lysozyme immobilization was conducted by the method of entrapment in gel. Enzyme carrier interaction was studied by viscometric, spectrophotometric and spectrofluorimetric methods. Mucoadhesive gel with immobilized lysozyme, possessing antiinflammatory and antimicrobial activities, was prepared. Due to immobilization, protein-polymer complex with the original enzymatic activity was formed. The product is characterized by high mucoadhesive properties, quantitative retaining of protein and bacteriolytic activity, prolonged release of the enzyme, improved biochemical characteristics (extended pH-activity profile, stability in acidic medium and during storage for 2 years, and it is perspective for further studies. The proposed method for lysozyme immobilization in the carboxymethyl cellulose sodium salt gel allows to obtain a stable, highly efficient product, with high adhesive properties for attachment to the mucous membranes, that is promising for use in biomedicine.

  3. Evidence for a third type of UV repair in bacteriophage T4

    International Nuclear Information System (INIS)

    Minderhout, L.A.G. van; Grimbergen, J.

    1976-01-01

    Two classes of UV-sensitive mutants of bacteriophage T4 have been found so far: excision-repair mutants and replication-repair mutants. A third type of UV repair in bacteria was described by Bridges and named ''reinitiation recovery.'' This repair is sensitive to acriflavine and is responsible for the shoulder in some UV-inactivation curves. T4 UV-sensitive mutants which are defective in this repair have not been described. In this paper, evidence is presented for the third UV-repair pathway in phage T4. Recently new UV-sensitive mutants induced in strain uvs5 were isolated

  4. First experiences with the AMERLEX-MAB FREE T4 assay

    International Nuclear Information System (INIS)

    Nijhof, W.A.; Penders, T.J.

    1989-01-01

    The new Amerlex-MAB FT 4 is a quick direct free T 4 assay with good reproducability. The correlation between the Amerlex-MAB FT 4 and the free T 4 of Byk is good. In the non-thyreoidal illness patient group no deviation for the values were found. Amerlex-MAB FT 4 is cheaper, because no total T4 has to be measured. More research has to be done for special patient sera. Disturbing influences as free fatty acids, heparin and auto-antibodies have to be checked. (R.B.). 3 refs.; 3 figs.; 4 tabs

  5. Effects of 18 months of L-T4 replacement in women with subclinical hypothyroidism.

    LENUS (Irish Health Repository)

    Adrees, M

    2012-02-01

    CONTEXT: Some of the cardiovascular and renal abnormalities seen in overt hypothyroidism have also been reported in subclinical hypothyroidism (SCH). Short-term L-T4 replacement in SCH improves cardiovascular risk markers and reduces carotid intima-media thickness (CIMT), a surrogate marker of atherosclerosis. The haemodynamic and renal effects of L-T4 replacement in SCH are poorly understood. OBJECTIVES: To compare cardiovascular risk factors and renal variables in women with SCH and normal women. To study the effects of L-T4 replacement in SCH subjects on these variables and on structural and functional changes in common carotid and brachial arteries. DESIGN: Fifty-six women with SCH before and after L-T4 replacement for 18 months and 56 normal women of similar age distribution were studied. Blood Pressure (BP), plasma lipids and homocysteine were measured and renal function evaluated [estimation of glomerular filtration rate (eGFR) using standard equations and measurement of serum Cystatin-C] in women with SCH before and after 18 months of l-T4, and in healthy women. CIMT and endothelial function (using brachial artery ultrasound) were studied before and after L-T4 in a subgroup of women with SCH. RESULTS: Systolic and diastolic BP, total cholesterol, triglyceride, LDL-cholesterol, lipoprotein(a) and homocysteine were greater in SCH (P < 0.05), and following L-T4 replacement decreased (P < 0.05) to levels that no longer differed from normal subjects. Estimated GFR was reduced and serum Cystatin-C increased (P < 0.05) in SCH. These variables also normalized following L-T4. Following L-T4 replacement the carotid artery baseline diameter increased by 7.1% and CIMT decreased by a mean value of 13%, while brachial artery diameter increased basally by 12.5% and following endothelium-dependent vasodilatation by 17.5% (P < 0.05). However, the increment following reactive hyperaemia did not differ before or following L-T4 replacement. CONCLUSION: Normalization of

  6. Isolation and partial purification of lysozyme from saliva of Bali Cattle ...

    African Journals Online (AJOL)

    SND

    2012-01-26

    Jan 26, 2012 ... lysozyme from saliva of Bali cattle is capable of isolation using aqueous mixtures of PEG 4000/Na2SO4. The lysozyme has a unique potential as an anti-bacterial. Moreover, the natural role of this lysozyme in saliva or in the oral cavity and how the prospect of its application remains to be further investigated ...

  7. Bactericidal activity of human lysozyme, muramidase-inactive lysozyme, and cationic polypeptides against Streptococcus sanguis and Streptococcus faecalis: inhibition by chitin oligosaccharides.

    Science.gov (United States)

    Laible, N J; Germaine, G R

    1985-01-01

    The basis of the bactericidal activity of human lysozyme against Streptococcus sanguis was studied. Experiments were designed to evaluate the role of lysozyme muramidase activity in its bactericidal potency. Inactivation of the muramidase activity of lysozyme was achieved by reduction of essential disulfides with dithiothreitol (DTT) or by incubation with the chitin oligosaccharides chitotriose and chitobiose. Muramidase-inactive lysozyme, prepared by reduction with DTT, was equal in bactericidal potency to native lysozyme. Solutions of native chicken egg white lysozyme and human lysozyme exhibited equal bactericidal potency yet differed ca. fourfold with respect to lytic (muramidase) activity. The above results suggested that the bactericidal activity of lysozyme is not dependent upon muramidase activity. Chitotriose and chitobiose were found to inhibit both lytic and bactericidal activities of lysozyme. The bactericidal activity of muramidase-inactive lysozyme (reduction with DTT) was also inhibited by chitotriose and chitobiose. Further investigations demonstrated that chitotriose and chitobiose were also potent inhibitors of the bactericidal activity of the cationic homopolypeptides poly-L-arginine and poly-D-lysine. These latter results suggested that the essential bactericidal property of lysozyme was its extreme cationic nature and that some bacterial endogenous activities, inhibitable by chitotriose and chitobiose, were essential for expression of the bactericidal activity of either native or muramidase-inactive lysozyme or of the cationic homopolypeptides. Experiments with Streptococcus faecalis whole cells, cell walls, and crude autolysin preparations implicated endogenous autolytic muramidases as the bacterial targets of chitotriose and chitobiose. The essentially identical responses of S. sanguis and S. faecalis to chitotriose in bactericidal assays with muramidase-inactive lysozyme and polylysine suggested that muramidase-like enzymes exist in S

  8. Effect on catalysis by replacement of catalytic residue from hen egg white lysozyme to Venerupis philippinarum lysozyme*

    Science.gov (United States)

    Abe, Yoshito; Kubota, Mitsuru; Takazaki, Shinya; Ito, Yuji; Yamamoto, Hiromi; Kang, Dongchon

    2016-01-01

    Abstract Asn46Asp/Asp52Ser or Asn46Glu/Asp52Ser hen egg white lysozyme (HEL) mutant was designed by introducing the substituted catalytic residue Asp46 or Glu46, respectively, based on Venerupis philippinarum (Vp) lysozyme structure as a representative of invertebrate‐type (i‐type) lyzozyme. These mutations restored the bell‐shaped pH‐dependency of the enzyme activity from the sigmoidal pH‐dependency observed for the Asp52Ser mutant. Furthermore both lysozyme mutants possessed retaining mechanisms like Vp lysozyme and HEL. The Asn46Glu/Asp52Ser mutant, which has a shorter distance between two catalytic residues, formed a glycosyl adduct in the reaction with the N‐acetylglucosamine oligomer. Furthermore, we found the accelerated turnover through its glycosyl adduct formation and decomposition. The turnover rate estimated from the glycosyl formation and decomposition rates was only 20% of the observed hydrolysis rate of the substrate. Based on these results, we discussed the catalytic mechanism of lysozymes. PMID:27291073

  9. Dynamics and Predictors of Serum TSH and fT4 Reference Limits in Early Pregnancy

    DEFF Research Database (Denmark)

    Laurberg, Peter; Andersen, Stine Linding; Hindersson, Peter

    2016-01-01

    CONTEXT: Thyroid hormones are important developmental factors and levels should be adequate both in the pregnant woman and in the fetus. However, there is no consensus on maternal thyroid test reference limits in early pregnancy. OBJECTIVE: Estimation of week-to-week changes in and predictors...... of TSH and free T4 (fT4) reference limits in the first trimester of pregnancy. DESIGN: Measurement of TSH and fT4 in biobank sera collected in pregnancy weeks 5-19 from a random sample of the Danish National Birth Cohort that enrolled 101 032 pregnant in 1996-2002. SETTING: National cohort of pregnant...... women. PARTICIPANTS: Healthy participants (n = 6671) were identified and individual characteristics retrieved using interview data and data from Danish national health registers. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Reference limits for TSH and fT4 in each first trimester pregnancy week...

  10. Adsorption of T4 bacteriophages on planar indium tin oxide surface via controlled surface tailoring.

    Science.gov (United States)

    Liana, Ayu Ekajayanthi; Chia, Ed Win; Marquis, Christopher P; Gunawan, Cindy; Gooding, J Justin; Amal, Rose

    2016-04-15

    The work investigates the influence of surface physicochemical properties of planar indium tin oxide (ITO) as a model substrate on T4 bacteriophage adsorption. A comparative T4 bacteriophage adsorption study shows a significant difference in bacteriophage adsorption observed on chemically modified planar ITO when compared to similarly modified particulate ITO, which infers that trends observed in virus-particle interaction studies are not necessarily transferrable to predict virus-planar surface adsorption behaviour. We also found that ITO surfaces modified with methyl groups, (resulting in increased surface roughness and hydrophobicity) remained capable of adsorbing T4 bacteriophage. The adsorption of T4 onto bare, amine and carboxylic functionalised planar ITO suggests the presence of a unique binding behaviour involving specific functional groups on planar ITO surface beyond the non-specific electrostatic interactions that dominate phage to particle interactions. The paper demonstrates the significance of physicochemical properties of surfaces on bacteriophage-surface interactions. Copyright © 2016 Elsevier Inc. All rights reserved.

  11. Full-vectorial propagation model and modified effective mode area of four-wave mixing in straight waveguides.

    Science.gov (United States)

    Guo, Kai; Friis, Søren M M; Christensen, Jesper B; Christensen, Erik N; Shi, Xiaodong; Ding, Yunhong; Ou, Haiyan; Rottwitt, Karsten

    2017-09-15

    We derive from Maxwell's equations full-vectorial nonlinear propagation equations of four-wave mixing valid in straight semiconductor-on-insulator waveguides. Special attention is given to the resulting effective mode area, which takes a convenient form known from studies in photonic crystal fibers, but has not been introduced in the context of integrated waveguides. We show that the difference between our full-vectorial effective mode area and the scalar equivalent often referred to in the literature may lead to mistakes when evaluating the nonlinear refractive index and optimizing designs of new waveguides. We verify the results of our derivation by comparing it to experimental measurements in a silicon-on-insulator waveguide, taking tolerances on fabrication parameters into account.

  12. Vectorial magnetic field sensing with a dual-core photonic-crystal fiber: Toward fiber-optic stereomagnetometry

    Science.gov (United States)

    Blakley, S.; Becker, J.; Fedotov, I. V.; Kilin, S. Ya.; Sakoda, K.; Zheltikov, A. M.

    2018-02-01

    A monolithic, ultra-compact fiber probe integrating a nitrogen-vacancy diamond microcrystal, a dual-core photonic-crystal fiber (PCF), and a microwave transmission line is shown to enable a highly sensitive dual-channel vectorial magnetic field measurements by means of simultaneous optical detection of magnetic resonances through the two fiber cores. A reliable detection of microscale magnetic-field variations as low as ˜10 µT/(Hz)1/2 is demonstrated using a dual-core PCF with a core-to-core separation of 6 µm. Such a fiber probe is shown to provide a unique tool for a vectorial three-dimensional mapping of weak magnetic fields induced by microscale objects.

  13. Excision repair and patch size in UV-irradiated bacteriophage T4

    International Nuclear Information System (INIS)

    Yarosh, D.B.; Rosenstein, B.S.; Setlow, R.B.

    1981-01-01

    We determined the average size of excision repair patches in repair of UV lesions in bacteriophage T4 by measuring the photolysis of bromodeoxyuridine incorporated during repair. The average patch was small, approximately four nucleotides long. In control experiments with the denV1 excision-deficient mutant, we encountered an artifact, a protein(s) which remained bound to phenol-extracted DNA and prevented nicking by the UV-specific endonucleases of Micrococcus luteus and bacteriophage T4

  14. Survival and prognostic factors of surgically resected T4 non-small cell lung cancer.

    Science.gov (United States)

    Osaki, Toshihiro; Sugio, Kenji; Hanagiri, Takeshi; Takenoyama, Mitsuhiro; Yamashita, Toshihiro; Sugaya, Masakazu; Yasuda, Manabu; Yasumoto, Kosei

    2003-06-01

    Category T4 nonsmall cell lung cancer (NSCLC) encompasses heterogenous subgroups. We retrospectively analyzed the survival of patients with surgically resected T4 NSCLC to evaluate the evidence for prognostic implications according to the subgroups of T4 category, nodal status, and resection completeness. Seventy-six patients with T4N0-2M0 NSCLC were divided into three subgroups within the T4 category: 24 patients with the tumor invading the mediastinal organs (mediastinal group), 16 with a malignant pleural effusion or dissemination (pleural group), and 36 with satellite tumor nodules within the ipsilateral primary tumor lobe (satellite group). Complete resection was possible in 47 patients (61.8%). The pathologic N statuses were N0 in 28, N1 in 13, and N2 in 35 patients. The overall survival of the 76 patients was 19.1% at 5 years. The overall 5-year survivals according to the three subgroups of the T4 category were as follows: mediastinal group, 18.2%; pleural group, 0%; and satellite group, 26.7% (mediastinal/satellite versus pleural, p = 0.037). Factors significantly influencing the overall 5-year survival were the pathologic N status (N2 versus N0-1, p = 0.022) and the completeness of resection (complete versus incomplete, p = 0.0001). A multivariate survival analysis demonstrated that the pathologic N status and the completeness of resection were significant independent predictors of a poorer prognosis even after adjusting for the subgroup of the T4 category. Resectable T4N0-1 NSCLC that is not due to pleural disease deserves consideration of aggressive surgical resection with expected 5-year survival of about 20%.

  15. Sensitive radioimmunoassay of total thyroxine (T4) in horses using a simple extraction method.

    Science.gov (United States)

    Tangyuenyong, Siriwan; Nambo, Yasuo; Nagaoka, Kentaro; Tanaka, Tomomi; Watanabe, Gen

    2017-07-28

    Most thyroid hormone determinations in animals are based on immunoassays adapted from those used to test human samples, which may not reflect the actual values of thyroid hormone in horses because of the presence of binding proteins. The aims of the present study were i) to establish a novel radioimmunoassay (RIA) using a more simple and convenient method to separate binding proteins for the measurement of total thyroxine (T4) in horses and ii) to validate the assay by comparing total T4 concentrations in yearling horses raised in different climates. Blood samples were collected from trained yearlings in Hokkaido (temperate climate) and Miyazaki (subtropical climate) in Japan and from adult horses in estrus and diestrus. T4 was extracted from both serum and plasma using modified acid ethanol cryo-precipitation and sodium acetate ethanol methods. Circulating total T4 concentrations were determined by RIA. T4 concentration by sodium acetate ethanol was appropriately detectable rather than sodium salicylate method and was the same as for acid ethanol method. Furthermore, this sodium acetate ethanol method required fewer extraction steps than the other methods. Circulating T4 concentrations in yearlings were 225.98 ± 20.89 ng/ml, which was higher than the previous reference values. With respect to climate, T4 levels in Hokkaido yearlings tended to be higher than those in Miyazaki yearlings throughout the study period. These results indicated that this RIA protocol using a modified sodium acetate ethanol separation technique might be an appropriate tool for specific measurement of total T4 in horses.

  16. Modeling the growth rates of tetragonal lysozyme crystals

    Science.gov (United States)

    Li, Meirong; Nadarajah, Arunan; Pusey, Marc L.

    1995-11-01

    Although the faceted growth of tetragonal lysozyme crystals is known to occur by 2D nucleation and dislocation-led growth, the measured growth rates do not follow model predictions based on these mechanisms. One possible reason for this deviation is that these models ignore the highly aggregated state of lysozyme in supersaturated solutions. In this study a growth mechanism for tetragonal lysozyme crystals involving aggregation reactions leading to the formation of the growth unit, mass transport of the growth unit to the crystal interface and faceted crystal growth by growth unit addition, is proposed. The distribution of aggregates in lysozyme nutrient solutions were determined from the equilibrium aggregation reactions and comparisons were made with growth rates calculated from the model based on the proposed mechanism and the measured growth rate data. The results indicated than an octamer corresponding to the tetragonal crystal unit cell was the most likely growth unit for the process. Remarkably good fits were obtained with this model to the measured growth rate data for three sets of pH and salt concentrations, suggesting the validity of the proposed mechanism. The values of the kinetic coefficient for the step velocity was in the range for small molecule crystal growth and the heats of reaction compared well with that obtained from lysozyme solubility data. The results presented here suggest that the inorganic and protein crystal growth processes are quite similar in many ways. Lysozyme crystal growth differs primarily due to growth by an aggregate growth unit and in the effect of nutrient solution conditions on the protein aggregation process.

  17. Effects of virus and host genes on recombination among ultraviolet-irradiated bacteriophage T4

    International Nuclear Information System (INIS)

    Priemer, M.M.; Chan, V.L.

    1978-01-01

    The influence of the polA, uvrA, and recA genes of Escherichia coli on recombination among ultraviolet-irradiated T4 bacteriophages was determined with respect to recombination between rII markers and phage yield. The polA and uvrA gene products have no effect on these two aspects of phage DNA metabolism. A recA mutation does not significantly alter rII recombination frequencies in irradiated phage crosses, nor does it greatly change the yield of infectious particles in wild-type phage crosses or crosses in which the phage strains possess the v mutation. However, the same cross experiment performed with a pair of T4x mutants in a recA host demonstrates an 84% reduction in the phage yield in an unirradiated control cross. Furthermore, with increasing doses of uv irradiation, phage productivity of the T4x mutant declines at an accelerated rate compared to T4x + strains crossed in recA cells. Multiplicity reactivation experiments in which wild-type or recombination-deficient (x or y) T4 phages infect wild-type or recombination-deficient (recA) host cells show that irradiated phages can only be reactivated in recA + hosts, regardless of the bacteriophage genotype. These results indicate the involvement of the E. coli recA gene product in normal T4 replication and multiplicity reactivation

  18. Activity-based in vitro selection of T4 DNA ligase

    International Nuclear Information System (INIS)

    Takahashi, Fumio; Funabashi, Hisakage; Mie, Masayasu; Endo, Yaeta; Sawasaki, Tatsuya; Aizawa, Masuo; Kobatake, Eiry

    2005-01-01

    Recent in vitro methodologies for selection and directed evolution of proteins have concentrated not only on proteins with affinity such as single-chain antibody but also on enzymes. We developed a display technology for selection of T4 DNA ligase on ribosome because an in vitro selection method for DNA ligase had never been developed. The 3' end of mRNA encoding the gene of active or inactive T4 DNA ligase-spacer peptide fusion protein was hybridized to dsDNA fragments with cohesive ends, the substrate of T4 DNA ligase. After in vitro translation of the mRNA-dsDNA complex in a rabbit reticulocyte system, a mRNA-dsDNA-ribosome-ligase complex was produced. T4 DNA ligase enzyme displayed on a ribosome, through addition of a spacer peptide, is able to react with dsDNA in the complex. The complex expressing active ligase was biotinylated by ligation with another biotinylated dsDNA probe and selected with streptavidin-coated magnetic beads. We effectively selected active T4 DNA ligase from a small amount of protein. The gene of the active T4 DNA ligase was enriched 40 times from a mixture of active and inactive genes using this selection strategy. This ribosomal display strategy may have high potential to be useful for selection of other enzymes associated with DNA

  19. A new type of UV-sensitive mutant of phage T4D

    International Nuclear Information System (INIS)

    Minderhout, L. van; Grimbergen, J.

    1975-01-01

    Within a group of more than 20 UV-sensitive mutants of T4D, 4 UV-sensitive mutants with the same sensitivity as T4 x were isolated independently of each other. They were uvs9, uvs21, uvs35, and uvs52. The double mutants with x and y 10 were constructed: they are slightly more UV sensitive than T4v 1 . The double mutant with uvs5 was not found. The mutations of uvs9, 21, 35, and 52 are closely linked with v 1 . The photoreactivable sector is 0.4. One of the mutants, uvs52, has the same sensitivity for methyl methanesulphonate as T4 + , shows a stronger multiplicity reactivation than the wild type, shows the same sensitivity relative to T4 + and T4v 1 in Luria-Latarjet tests and in monocomplex UV inactivation, and raises the recombinant frequency in crosses with irradiated phage. The uvs52 + function has the same sensitivity to UV as the v + function. Complementation between uvs52 and v 1 , if present, is difficult to demonstrate owing to an appreciable multiplicity reactivation contribution to increased survival. The possibility that uvs52 is an allele of v 1 is discussed. The observations fit the assumption that uvs52 is an excision-repair mutant with a low excision rate

  20. Spectrophotometric studies on the interaction between (-)-epigallocatechin gallate and lysozyme

    Science.gov (United States)

    Ghosh, Kalyan Sundar; Sahoo, Bijaya Ketan; Dasgupta, Swagata

    2008-02-01

    Various reported antibacterial activities of (-)-epigallocatechin-3-gallate (EGCG), the major polyphenol of green tea prompted us to study its binding with lysozyme. This has been investigated by fluorescence, circular dichroism (CD) and protein-ligand docking. The binding parameters were determined using a modified Stern-Volmer equation. The thermodynamic parameters are indicative of an initial hydrophobic association. The complex is, however, held together predominantly by van der Waals interactions and hydrogen bonding. CD studies do not indicate any significant changes in the secondary structure of lysozyme. Docking studies revealed that specific interactions are observed with residues Trp 62 and Trp 63.

  1. The solubility of hen egg-white lysozyme

    Science.gov (United States)

    Howard, Sandra B.; Twigg, Pamela J.; Baird, James K.; Meehan, Edward J.

    1988-01-01

    The equilibrium solubility of chicken egg-white lysozyme in the presence of crystalline solid state was determined as a function of NaCl concentration, pH, and temperature. The solubility curves obtained represent a region of the lysozyme phase diagram. This diagram makes it possible to determine the supersaturation of a given set of conditions or to achieve identical supersaturations by different combinations of parameters. The temperature dependence of the solubility permits the evaluation of Delta-H of crystallization. The data indicate a negative heat of crystallization for the tetragonal crystal form but a positive heat of crystallization for the high-temperature orthorhombic form.

  2. Identification of some OH radical-induced products of lysozyme

    International Nuclear Information System (INIS)

    Dizdaroglu, M.; Gajewski, E.; Simic, M.G.

    1983-01-01

    OH radical reactions with lysozyme in #betta#-irradiated N 2 O saturated aqueous solutions caused formation of allo-threonine, α-amino-n-butyric acid, o- and m-tyrosines, and 2- and 3-hydroxytyrosines. These identified radiolytic products were characterized by capillary gas chromatography-mass spectrometry as their trimethylsilyl derivatives after HCl-hydrolysis of irradiated lysozyme. Their initial G-values were also determined using gas chromatography. The possible use of these radiolytic products as monitors of radiation-induced damage to proteins and the sites of attack are also discussed. (author)

  3. THE ASSESSMENT OF INSECTICIDAL IMPACT ON THE MALARIA MOSQUITO'S VECTORIAL CAPACITY, FROM DATA ON THE PROPORTION OF PAROUS FEMALES.

    Science.gov (United States)

    GARRETT-JONES, C; GRAB, B

    1964-01-01

    In malaria eradication the residual insecticide exerts upon the mosquito's vectorial capacity a direct insecticidal impact, the order of which may be measured by observing the decrease in the proportion of parous females. The impact is expressed as the product of the degree of reduction of the expectation of infective life (termed the longevity factor of impact) and that of the expectation of life (the density factor). To compute the factors from the proportion parous it is necessary to know also the mean difference in age between the nulliparous and the youngest parous females in the sample, and the sporogonic period of the parasite.Graphs are presented to enable the field worker, who has observed these parameters, to read off from his data the proportion surviving one day, the expectation of infective life and the expectation of life. Examples from the field are used to illustrate the manner of computing the direct insecticidal impact with the aid of the graphs. It is emphasized that this method can only measure the relative impact on vectorial capacity, and will not show whether the actual level of vectorial capacity is such that a malaria reproduction rate of <1 is indicated.

  4. New Vectorial Propulsion System and Trajectory Control Designs for Improved AUV Mission Autonomy.

    Science.gov (United States)

    Masmitja, Ivan; Gonzalez, Julian; Galarza, Cesar; Gomariz, Spartacus; Aguzzi, Jacopo; Del Rio, Joaquin

    2018-04-17

    Autonomous Underwater Vehicles (AUV) are proving to be a promising platform design for multidisciplinary autonomous operability with a wide range of applications in marine ecology and geoscience. Here, two novel contributions towards increasing the autonomous navigation capability of a new AUV prototype (the Guanay II) as a mix between a propelled vehicle and a glider are presented. Firstly, a vectorial propulsion system has been designed to provide full vehicle maneuverability in both horizontal and vertical planes. Furthermore, two controllers have been designed, based on fuzzy controls, to provide the vehicle with autonomous navigation capabilities. Due to the decoupled system propriety, the controllers in the horizontal plane have been designed separately from the vertical plane. This class of non-linear controllers has been used to interpret linguistic laws into different zones of functionality. This method provided good performance, used as interpolation between different rules or linear controls. Both improvements have been validated through simulations and field tests, displaying good performance results. Finally, the conclusion of this work is that the Guanay II AUV has a solid controller to perform autonomous navigation and carry out vertical immersions.

  5. Vectorial Command of Induction Motor Pumping System Supplied by a Photovoltaic Generator

    Science.gov (United States)

    Makhlouf, Messaoud; Messai, Feyrouz; Benalla, Hocine

    2011-01-01

    With the continuous decrease of the cost of solar cells, there is an increasing interest and needs in photovoltaic (PV) system applications following standard of living improvements. Water pumping system powered by solar-cell generators are one of the most important applications. The fluctuation of solar energy on one hand, and the necessity to optimise available solar energy on the other, it is useful to develop new efficient and flexible modes to control motors that entrain the pump. A vectorial control of an asynchronous motor fed by a photovoltaic system is proposed. This paper investigates a photovoltaic-electro mechanic chain, composed of a PV generator, DC-AC converter, a vector controlled induction motor and centrifugal pump. The PV generator is forced to operate at its maximum power point by using an appropriate search algorithm integrated in the vector control. The optimization is realized without need to adding a DC-DC converter to the chain. The motor supply is also ensured in all insolation conditions. Simulation results show the effectiveness and feasibility of such an approach.

  6. A Fast Alternating Minimization Algorithm for Nonlocal Vectorial Total Variational Multichannel Image Denoising

    Directory of Open Access Journals (Sweden)

    Rubing Xi

    2014-01-01

    Full Text Available The variational models with nonlocal regularization offer superior image restoration quality over traditional method. But the processing speed remains a bottleneck due to the calculation quantity brought by the recent iterative algorithms. In this paper, a fast algorithm is proposed to restore the multichannel image in the presence of additive Gaussian noise by minimizing an energy function consisting of an l2-norm fidelity term and a nonlocal vectorial total variational regularization term. This algorithm is based on the variable splitting and penalty techniques in optimization. Following our previous work on the proof of the existence and the uniqueness of the solution of the model, we establish and prove the convergence properties of this algorithm, which are the finite convergence for some variables and the q-linear convergence for the rest. Experiments show that this model has a fabulous texture-preserving property in restoring color images. Both the theoretical derivation of the computation complexity analysis and the experimental results show that the proposed algorithm performs favorably in comparison to the widely used fixed point algorithm.

  7. Invasiveness of Aedes aegypti and Aedes albopictus and Vectorial Capacity for Chikungunya Virus.

    Science.gov (United States)

    Lounibos, Leon Philip; Kramer, Laura D

    2016-12-15

    In this review, we highlight biological characteristics of Aedes aegypti and Aedes albopictus, 2 invasive mosquito species and primary vectors of chikungunya virus (CHIKV), that set the tone of these species' invasiveness, vector competence, and vectorial capacity (VC). The invasiveness of both species, as well as their public health threats as vectors, is enhanced by preference for human blood. Vector competence, characterized by the efficiency of an ingested arbovirus to replicate and become infectious in the mosquito, depends largely on vector and virus genetics, and most A. aegypti and A. albopictus populations thus far tested confer vector competence for CHIKV. VC, an entomological analog of the pathogen's basic reproductive rate (R 0 ), is epidemiologically more important than vector competence but less frequently measured, owing to challenges in obtaining valid estimates of parameters such as vector survivorship and host feeding rates. Understanding the complexities of these factors will be pivotal in curbing CHIKV transmission. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.

  8. Goose-Type Lysozyme Inhibitor (PliG) Enhances Survival of Escherichia coli in Goose Egg Albumen ▿

    OpenAIRE

    Vanderkelen, Lise; Van Herreweghe, Joris M.; Callewaert, Lien; Michiels, Chris W.

    2011-01-01

    The goose-type lysozyme inhibitor PliG enhances the survival of Escherichia coli in goose but not in chicken egg white, which contains goose- and chicken-type lysozymes, respectively. These results indicate that both the type of host lysozyme and the type of bacterial lysozyme inhibitor may affect bacterium-host interactions.

  9. Neutron scattering study on the interaction between polyethylene glycol and lysozyme

    International Nuclear Information System (INIS)

    Magazu, S.; Migliardo, F.; Barreca, D.; Bellocco, E.; Lagana, G.

    2008-01-01

    Recently, new opportunities at the interface of polymer chemistry and biology have been explored. It has been found that polyethylene glycol (PEG) has bioprotectant effects in interacting with protein molecules. In the present paper, small angle neutron scattering (SANS) findings on lysozyme/D 2 O and lysozyme/D 2 O/PEG solutions for temperature values of T=310 and 333 K are shown. We are able to characterise by SANS the dimensions of the protein in solution and emphasise the effects of PEG on lysozyme

  10. Design and creation of a Ca2+ binding site in human lysozyme to enhance structural stability.

    OpenAIRE

    Kuroki, R; Taniyama, Y; Seko, C; Nakamura, H; Kikuchi, M; Ikehara, M

    1989-01-01

    A Ca2+ binding site like an EF-hand motif was designed and created in human lysozyme by replacing both Gln-86 and Ala-92 with aspartic acids by site-directed mutagenesis. The mutant human lysozyme (D86/92-lysozyme) was expressed and secreted by yeast. One Ca2+ was found to bind one molecule of the purified protein with the binding constant 5.0 x 10(6) M-1. The enzymatic activity of holo-D86/92-lysozyme against glycol chitin at 40 degrees C was 2-fold higher than that of the native lysozyme. M...

  11. Functional Characterization of a c-type Lysozyme from Indian Shrimp Fenneropenaeus indicus.

    Science.gov (United States)

    Karthik, Viswanathan; Kamalakannan, Vijayan; Thomas, Ancy; Sudheer, Naduvilamuriparambu Saidumuhammed; Singh, Issac S Bright; Narayanan, Rangarajan Badri

    2014-06-01

    Lysozyme gene from Fenneropenaeus indicus was cloned, expressed in Escherichia coli and characterized. The cDNA consists of 477 base pairs and encodes amino acid sequence of 159 residues. F. indicus lysozyme had high identity (98%) with Fenneropenaeus merguiensis and Fenneropenaeus chinensis and exhibits low to moderate identities with lysozymes of other invertebrates and vertebrates. This lysozyme is presumed to be chicken types as it possesses two catalytic and eight cysteine residues that are conserved across c-type lysozymes and a c-terminal extension, which is a characteristic of lysozymes from marine invertebrates. Further, the antimicrobial properties of the recombinant lysozyme from F. indicus were determined in comparison with recombinant hen egg white lysozyme. This exhibited high activity against a Gram-negative pathogenic bacterium Salmonella typhimurium and two fungal strains Pichia pastoris and Saccharomyces cerevisiae in turbidimetric assay. Distribution of lysozyme gene and protein in tissues of shrimps infected with white spot syndrome virus revealed that the high levels of lysozyme are correlated with low and high viral load in abdominal muscle and tail, respectively. In conclusion, lysozyme from F. indicus has a broad spectrum of antimicrobial properties, which once again emphasizes its role in shrimp innate immune response.

  12. Clinicopathological outcomes of preoperative chemoradiotherapy using S-1 plus Irinotecan for T4 lower rectal cancer.

    Science.gov (United States)

    Beppu, Naohito; Yoshie, Hidenori; Kimura, Fumihiko; Aihara, Tsukasa; Doi, Hiroshi; Kamikonya, Norihiko; Matsubara, Nagahide; Tomita, Naohiro; Yanagi, Hidenori; Yamanaka, Naoki

    2016-07-01

    To investigate the clinicopathological outcomes of patients with T4 lower rectal cancer treated using preoperative chemoradiotherapy with S-1 plus Irinotecan. Between 2005 and 2011, 35 patients with T4M0 lower rectal cancer, diagnosed initially as T4a in 12 and as T4b in 23, were treated with 45 Gy of radiotherapy concomitantly with S-1 plus Irinotecan. The median follow-up period was 50.6 months (range 2-123 months). A total of 32 patients (91.4 %) completed the radiotherapy and 26 (74.3 %) completed the full chemotherapy regimen. Radical surgery was then performed in 33 (94.3 %) of the 35 patients after the exclusion of two patients, who had macroscopic residual disease. The pathological diagnosis was downstaged from T4a to ypT0-3 in all 12 of those patients (100 %) and from T4b to ypT0-4a in 20 of those 23 patients (87.0 %). The tumor regression grade of 1a/1b/2/3 (complete response) was 10/8/15/2, respectively. In terms of long-term survival, the 5-year local relapse-free survival rate was 74.8 % and the recurrence-free survival rate was 52.0 %. This regimen may result in favorable downstaging. Moreover, in this series, pathological evidence of involvement of adjacent organs was rare following preoperative chemoradiotherapy, in the patients with disease diagnosed as T4b at the initial staging.

  13. Antimicrobial activity of T4 bacteriophage conjugated indium tin oxide surfaces.

    Science.gov (United States)

    Liana, Ayu E; Marquis, Christopher P; Gunawan, Cindy; Justin Gooding, J; Amal, Rose

    2018-03-15

    We report the antimicrobial activity of bare and surface functionalized indium tin oxide (ITO) conjugated with T4 bacteriophage towards E. coli. A ∼ 10 3 -fold reduction (99.9%) in the bacterial concentration was achieved within 2 h exposure of E. coli to the bare as well as the amine, carboxylic and methyl functionalized ITO/T4 surfaces. Despite the known differences in bacteriophage loading of these ITO/T4 systems, the almost identical extent of antimicrobial activity of all of the ITO/T4 systems resulted from the release of a comparable amount of infective T4 from the systems. As anticipated, a single dose of immobilized bacteriophage was sufficient to eliminate further surge of bacterial population. Upon the 2 h eradication of the '1st batch' of E. coli population, all of the ITO/T4 systems, each system with 10 2 -fold more suspended bacteriophage (due to propagation of the phage at the expense of the '1st batch' E. coli death), reduced the '2nd batch' of E. coli concentration by ∼10 4 -fold in just 30 min, suggesting the potential of immobilized bacteriophage systems as solution to the issues of antimicrobial agent depletion. All of the ITO/T4 systems maintained their antimicrobial activity in the presence of model food components. The antimicrobial activity was however, affected by pH; at pH 5 whereby the bacteria's growth was physiologically inhibited, generally no reduction in E. coli concentration was detected. The present work provides an understanding of the mode of antimicrobial activity exhibited by an immobilized bacteriophage based substrate and demonstrates efficacy in the presence of food components. Copyright © 2017 Elsevier Inc. All rights reserved.

  14. Effect of adjuvant lithium on thyroxine (T4) concentration after radioactive iodine therapy

    Energy Technology Data Exchange (ETDEWEB)

    Hammond, Emmanuel NiiBoye; Vangu, Mboyo-Di-Tamba Heben Willy [University of the Witwatersrand, Division of Nuclear Medicine and Molecular Imaging, Department of Radiation Sciences, Johannesburg (South Africa)

    2016-10-15

    To study the effect of adjuvant lithium on serum thyroxine (T4) concentrations in patients treated with radioactive iodine (RAI) therapy in our environment. This was a prospective simple randomized comparative, experimental cohort study of patients with hyperthyroidism referred for RAI ablation therapy in the two main academic hospitals in Johannesburg between February 2014 and September 2015. Amongst the 163 participants in the final analysis, 75 received RAI alone and 88 received RAI with lithium. The difference in mean T4 concentrations at 3 months between the RAI-only group (17.67 pmol/l) and the RAI with lithium group (11.55 pmol/l) was significant with a small effect size (U = 2328.5, Z = -2.700, p = 0.007, r = 0.01). Significant decreases in T4 concentrations were observed as early as 1 month after RAI (p = 0.0001) in the RAI with lithium group, but in the RAI-only group, significant decreases in T4 concentrations were observed only at 3 months after RAI therapy (p = 0.000). Women and patients with Graves' disease who received RAI with adjuvant lithium also showed significant decreases in T4 concentrations at 1 month (p = 0.002 and p = 0.003, respectively). Adjuvant lithium leads to an earlier and better response to RAI therapy with lower T4 concentrations that are achieved earlier. This earlier response and decrease in T4 concentrations were noted in patients with Graves' disease and nodular goitre, and in women with hyperthyroidism who received adjuvant lithium therapy. (orig.)

  15. Prognostic factors in resected satellite-nodule T4 non-small cell lung cancer.

    Science.gov (United States)

    Rao, Jagan; Sayeed, Rana A; Tomaszek, Sandra; Fischer, Stefan; Keshavjee, Shaf; Darling, Gail E

    2007-09-01

    The 1997 non-small cell lung cancer staging revisions assigned a T4 descriptor to satellite nodules in the primary tumor lobe. We reviewed our experience of satellite-nodule T4 non-small cell lung cancer following these revisions and evaluated prognostic factors for this group. All patients who underwent resection of non-small cell lung cancer between April 1997 and June 2005 with satellite nodule(s) confirmed at pathologic examination were identified from our institutional Lung Tumor Registry. Case notes and pathology reports were reviewed and data collected on possible prognostic factors. Survival was modeled using the Kaplan-Meier method, and survival differences between groups were analyzed using the log-rank test. From 1,276 non-small cell lung cancer patients who underwent resection, 137 were staged pT4, and 35 were T4-satellite nodules. Median follow-up was 25 months (range, 1 to 102 months). Median main tumor size was 3.0 cm (range, 1 to 9.8 cm). Adenocarcinoma or bronchioloalveolar carcinoma was the predominant histologic diagnosis (n = 28; 80%). One-, 3- and 5-year survival was 86%, 69%, and 57%, respectively; median survival was 68 months. During the same period, 137 patients undergoing resection for all T4 lesions had a 1-, 3-, and 5-year survival of 68%, 53%, and 18%, respectively. Adenocarcinoma or bronchioloalveolar carcinoma histologic diagnosis (adenocarcinoma or bronchioloalveolar carcinoma versus squamous, 75% versus 67% 3-year survival; p = 0.0026), female gender (66% versus 49% for males, 5-year survival; p = 0.041), and absence of vascular invasion (no invasion versus vascular invasion, 74% versus 20% 5-year survival; p = 0.0101) were significant predictors of better survival. Survival for resected T4 non-small cell lung cancer with satellite nodule(s) in the primary lobe is better than for other T4 lesions, and the T4 descriptor may unduly upstage these cases. The current T4 descriptor represents a heterogeneous population.

  16. Genomes of the T4-related bacteriophages as windows on microbial genome evolution.

    Science.gov (United States)

    Petrov, Vasiliy M; Ratnayaka, Swarnamala; Nolan, James M; Miller, Eric S; Karam, Jim D

    2010-10-28

    The T4-related bacteriophages are a group of bacterial viruses that share morphological similarities and genetic homologies with the well-studied Escherichia coli phage T4, but that diverge from T4 and each other by a number of genetically determined characteristics including the bacterial hosts they infect, the sizes of their linear double-stranded (ds) DNA genomes and the predicted compositions of their proteomes. The genomes of about 40 of these phages have been sequenced and annotated over the last several years and are compared here in the context of the factors that have determined their diversity and the diversity of other microbial genomes in evolution. The genomes of the T4 relatives analyzed so far range in size between ~160,000 and ~250,000 base pairs (bp) and are mosaics of one another, consisting of clusters of homology between them that are interspersed with segments that vary considerably in genetic composition between the different phage lineages. Based on the known biological and biochemical properties of phage T4 and the proteins encoded by the T4 genome, the T4 relatives reviewed here are predicted to share a genetic core, or "Core Genome" that determines the structural design of their dsDNA chromosomes, their distinctive morphology and the process of their assembly into infectious agents (phage morphogenesis). The Core Genome appears to be the most ancient genetic component of this phage group and constitutes a mere 12-15% of the total protein encoding potential of the typical T4-related phage genome. The high degree of genetic heterogeneity that exists outside of this shared core suggests that horizontal DNA transfer involving many genetic sources has played a major role in diversification of the T4-related phages and their spread to a wide spectrum of bacterial species domains in evolution. We discuss some of the factors and pathways that might have shaped the evolution of these phages and point out several parallels between their diversity

  17. Genomes of the T4-related bacteriophages as windows on microbial genome evolution

    Directory of Open Access Journals (Sweden)

    Miller Eric S

    2010-10-01

    Full Text Available Abstract The T4-related bacteriophages are a group of bacterial viruses that share morphological similarities and genetic homologies with the well-studied Escherichia coli phage T4, but that diverge from T4 and each other by a number of genetically determined characteristics including the bacterial hosts they infect, the sizes of their linear double-stranded (ds DNA genomes and the predicted compositions of their proteomes. The genomes of about 40 of these phages have been sequenced and annotated over the last several years and are compared here in the context of the factors that have determined their diversity and the diversity of other microbial genomes in evolution. The genomes of the T4 relatives analyzed so far range in size between ~160,000 and ~250,000 base pairs (bp and are mosaics of one another, consisting of clusters of homology between them that are interspersed with segments that vary considerably in genetic composition between the different phage lineages. Based on the known biological and biochemical properties of phage T4 and the proteins encoded by the T4 genome, the T4 relatives reviewed here are predicted to share a genetic core, or "Core Genome" that determines the structural design of their dsDNA chromosomes, their distinctive morphology and the process of their assembly into infectious agents (phage morphogenesis. The Core Genome appears to be the most ancient genetic component of this phage group and constitutes a mere 12-15% of the total protein encoding potential of the typical T4-related phage genome. The high degree of genetic heterogeneity that exists outside of this shared core suggests that horizontal DNA transfer involving many genetic sources has played a major role in diversification of the T4-related phages and their spread to a wide spectrum of bacterial species domains in evolution. We discuss some of the factors and pathways that might have shaped the evolution of these phages and point out several parallels

  18. Role of lysozyme inhibitors in the virulence of avian pathogenic Escherichia coli.

    Science.gov (United States)

    Vanderkelen, Lise; Ons, Ellen; Van Herreweghe, Joris M; Callewaert, Lien; Goddeeris, Bruno M; Michiels, Chris W

    2012-01-01

    Lysozymes are key effectors of the animal innate immunity system that kill bacteria by hydrolyzing peptidoglycan, their major cell wall constituent. Recently, specific inhibitors of the three major lysozyme families occuring in the animal kingdom (c-, g- and i-type) have been discovered in Gram-negative bacteria, and it has been proposed that these may help bacteria to evade lysozyme mediated lysis during interaction with an animal host. Escherichia coli produces two inhibitors that are specific for c-type lysozyme (Ivy, Inhibitor of vertebrate lysozyme; MliC, membrane bound lysozyme inhibitor of c-type lysozyme), and one specific for g-type lysozyme (PliG, periplasmic lysozyme inhibitor of g-type lysozyme). Here, we investigated the role of these lysozyme inhibitors in virulence of Avian Pathogenic E. coli (APEC) using a serum resistance test and a subcutaneous chicken infection model. Knock-out of mliC caused a strong reduction in serum resistance and in in vivo virulence that could be fully restored by genetic complementation, whereas ivy and pliG could be knocked out without effect on serum resistance and virulence. This is the first in vivo evidence for the involvement of lysozyme inhibitors in bacterial virulence. Remarkably, the virulence of a ivy mliC double knock-out strain was restored to almost wild-type level, and this strain also had a substantial residual periplasmic lysozyme inhibitory activity that was higher than that of the single knock-out strains. This suggests the existence of an additional periplasmic lysozyme inhibitor in this strain, and indicates a regulatory interaction in the expression of the different inhibitors.

  19. Role of lysozyme inhibitors in the virulence of avian pathogenic Escherichia coli.

    Directory of Open Access Journals (Sweden)

    Lise Vanderkelen

    Full Text Available Lysozymes are key effectors of the animal innate immunity system that kill bacteria by hydrolyzing peptidoglycan, their major cell wall constituent. Recently, specific inhibitors of the three major lysozyme families occuring in the animal kingdom (c-, g- and i-type have been discovered in Gram-negative bacteria, and it has been proposed that these may help bacteria to evade lysozyme mediated lysis during interaction with an animal host. Escherichia coli produces two inhibitors that are specific for c-type lysozyme (Ivy, Inhibitor of vertebrate lysozyme; MliC, membrane bound lysozyme inhibitor of c-type lysozyme, and one specific for g-type lysozyme (PliG, periplasmic lysozyme inhibitor of g-type lysozyme. Here, we investigated the role of these lysozyme inhibitors in virulence of Avian Pathogenic E. coli (APEC using a serum resistance test and a subcutaneous chicken infection model. Knock-out of mliC caused a strong reduction in serum resistance and in in vivo virulence that could be fully restored by genetic complementation, whereas ivy and pliG could be knocked out without effect on serum resistance and virulence. This is the first in vivo evidence for the involvement of lysozyme inhibitors in bacterial virulence. Remarkably, the virulence of a ivy mliC double knock-out strain was restored to almost wild-type level, and this strain also had a substantial residual periplasmic lysozyme inhibitory activity that was higher than that of the single knock-out strains. This suggests the existence of an additional periplasmic lysozyme inhibitor in this strain, and indicates a regulatory interaction in the expression of the different inhibitors.

  20. 21 CFR 862.1490 - Lysozyme (muramidase) test system.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Lysozyme (muramidase) test system. 862.1490 Section 862.1490 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test...

  1. Lysozyme Catalyzes the Formation of Antimicrobial Silver Nanoparticles (POSTPRINT)

    Science.gov (United States)

    2009-04-01

    molecules on nanopar- ticle surface. Consistent with its observed colloidal in- stability, the measured surface potential of AglysoSDS nanoparticles was...globular state, the protein would take on a new cationic and amphiphilic form. In this uniquely amphipathic state, the lysozyme coating on the particle

  2. Lysozyme Catalyzes the Formation of Antimicrobial Silver Nanoparticles

    Science.gov (United States)

    2009-04-03

    adsorption of anionic SDS molecules on nanopar- ticle surface. Consistent with its observed colloidal in- stability, the measured surface potential of Ag...ionic protein and, in a globular state, the protein would take on a new cationic and amphiphilic form. In this uniquely amphipathic state, the lysozyme

  3. Complex coacervate core micelles with a lysozyme-modified corona

    NARCIS (Netherlands)

    Danial, M.; Klok, H.A.; Norde, W.; Cohen Stuart, M.A.

    2007-01-01

    This paper describes the preparation, characterization, and enzymatic activity of complex coacervate core micelles (C3Ms) composed of poly(acrylic acid) (PAA) and poly(N-methyl-2-vinyl pyridinium iodide)-b-poly(ethylene oxide) (PQ2VP-PEO) to which the antibacterial enzyme lysozyme is end-attached.

  4. Adsorption of lysozyme unto silica and polystyrene surfaces in ...

    African Journals Online (AJOL)

    user

    2011-04-11

    Apr 11, 2011 ... polystyrene interfaces was studied at varying lysozyme concentrations and ionic strength. The studies revealed an ... adsorption densities of 1.34 x10-6 mol g-1 and 1.57 x10-6 mol g-1 obtained for silica and polystyrene respectively at the ..... Differential scanning calometry and FTIR analysis. J. Colloid.

  5. Binding of Lysozyme to Spherical Poly(styrenesulfonate Gels

    Directory of Open Access Journals (Sweden)

    Martin Andersson

    2018-01-01

    Full Text Available Polyelectrolyte gels are useful as carriers of proteins and other biomacromolecules in, e.g., drug delivery. The rational design of such systems requires knowledge about how the binding and release are affected by electrostatic and hydrophobic interactions between the components. To this end we have investigated the uptake of lysozyme by weakly crosslinked spherical poly(styrenesulfonate (PSS microgels and macrogels by means of micromanipulator assisted light microscopy and small angle X-ray scattering (SAXS in an aqueous environment. The results show that the binding process is an order of magnitude slower than for cytochrome c and for lysozyme binding to sodium polyacrylate gels under the same conditions. This is attributed to the formation of very dense protein-rich shells in the outer layers of the microgels with low permeability to the protein. The shells in macrogels contain 60 wt % water and nearly charge stoichiometric amounts of lysozyme and PSS in the form of dense complexes of radius 8 nm comprising 30–60 lysozyme molecules. With support from kinetic modelling results we propose that the rate of protein binding and the relaxation rate of the microgel are controlled by the protein mass transport through the shell, which is strongly affected by hydrophobic and electrostatic interactions. The mechanism explains, in turn, an observed dependence of the diffusion rate on the apparent degree of crosslinking of the networks.

  6. Effect of molecular anisotropy on the nucleation of lysozyme

    NARCIS (Netherlands)

    Drenth, J; Dijkstra, K; Haas, C; Leppert, J; Ohlenschlager, O

    2003-01-01

    The growth of protein crystals is preceded by an induction period during which the protein solution prepares itself for crystallization. We have measured the length of the induction period for lysozyme as a function of the temperature for a solution of 16.9 mg/mL at pH 4.5 with 5% NaCl as the

  7. ASSOCIATION OF LYSOZYME TO PHOSPHOLIPID SURFACES AND VESICLE FUSION

    NARCIS (Netherlands)

    ARNOLD, K; HOEKSTRA, D; OHKI, S

    1992-01-01

    Lysozyme-induced fusion of phosphatidylserine (PS) vesicles was studied as a function of pH. Fusion, monitored by lipid-mixing, was measured by following the dilution of pyrene-labelled phosphatidylcholine, incorporated in PS vesicles, into unlabelled bilayers. It is demonstrated that

  8. Monosize magnetic hydrophobic beads for lysozyme purification under magnetic field

    Energy Technology Data Exchange (ETDEWEB)

    Altintas, Evrim Banu [Department of Chemistry, Hacettepe University, Beytepe, Ankara (Turkey); Denizli, Adil, E-mail: denizli@hacettepe.edu.tr [Department of Chemistry, Hacettepe University, Beytepe, Ankara (Turkey)

    2009-06-01

    Monosize and magnetic poly(glycidyl methacrylate-N-methacryloyl-(L)-tryptophan) [mPGMATrp] beads (1.6 {mu}m in diameter) were used for hydrophobic affinity capture of lysozyme from chicken egg-white. N-methacryloyl-(L)-tryptophan (MATrp), which gives hydrophobicity to the resulting polymer, was synthesized by reacting methacryloyl chloride and L-tryptophan methyl ester then characterized by Nuclear Magnetic Resonance (NMR). mPGMATrp beads were produced by dispersion polymerization in the presence of magnetite nano-powder. mPGMATrp beads were characterized by means of swelling studies, elemental analysis, Fourier Transform Infrared Spectroscopy (FTIR) and Scanning Electron Microscope (SEM). Lysozyme adsorption experiments were performed under different experimental conditions (i.e., lysozyme concentration, temperature, and ionic strength) in magnetically stabilized fluidized bed system, (MSFB). Maximum adsorption capacity was 263.9 mg/g. It was observed that mPGMATrp beads can be used without significant loss in lysozyme adsorption capacity after 25 adsorption-elution cycle.

  9. Locations of Bromide Ions in Tetragonal Lysozyme Crystals

    Science.gov (United States)

    Lim, Kap; Nadarajah, Arunan; Forsythe, Elizabeth L.; Pusey, Marc L.

    1998-01-01

    Anions have been shown to play a dominant role in the crystallization of chicken egg-white lysozyme from salt solutions. Previous studies employing X-ray crystallography have found one chloride ion binding site in the tetragonal crystal form of the protein and four nitrate ion binding sites in the monoclinic form. In this study the anion positions in the tetragonal form were determined from the difference Fourier map obtained from lysozyme crystals grown in bromide and chloride solutions. Five possible anion-binding sites were found in this manner. Some of these sites were in pockets containing basic residues while others were near neutral, but polar, residues. The sole chloride ion binding site found in previous studies was confirmed, while four further sites were found which corresponded to the four binding sites found for nitrate ions in monoclinic crystals. The study suggests that most of the anion-binding sites in lysozyme remain unchanged even when different anions and different crystal forms of lysozyme are employed.

  10. Monosize magnetic hydrophobic beads for lysozyme purification under magnetic field

    International Nuclear Information System (INIS)

    Altintas, Evrim Banu; Denizli, Adil

    2009-01-01

    Monosize and magnetic poly(glycidyl methacrylate-N-methacryloyl-(L)-tryptophan) [mPGMATrp] beads (1.6 μm in diameter) were used for hydrophobic affinity capture of lysozyme from chicken egg-white. N-methacryloyl-(L)-tryptophan (MATrp), which gives hydrophobicity to the resulting polymer, was synthesized by reacting methacryloyl chloride and L-tryptophan methyl ester then characterized by Nuclear Magnetic Resonance (NMR). mPGMATrp beads were produced by dispersion polymerization in the presence of magnetite nano-powder. mPGMATrp beads were characterized by means of swelling studies, elemental analysis, Fourier Transform Infrared Spectroscopy (FTIR) and Scanning Electron Microscope (SEM). Lysozyme adsorption experiments were performed under different experimental conditions (i.e., lysozyme concentration, temperature, and ionic strength) in magnetically stabilized fluidized bed system, (MSFB). Maximum adsorption capacity was 263.9 mg/g. It was observed that mPGMATrp beads can be used without significant loss in lysozyme adsorption capacity after 25 adsorption-elution cycle.

  11. Antimicrobial activity of lysozyme with special relevance to milk

    African Journals Online (AJOL)

    STORAGESEVER

    2008-12-29

    Dec 29, 2008 ... and secretions of mammals including milk, saliva, tears, urine, respiratory and cervical secretions ..... normally resistant to native lysozyme (Pellegrini et al.,. 1992; Ibrahim et al., 1993; 1996; 1996a; Düring .... ability to bind calcium ions, stability to heat and/or pH, the isoelectric point (Table 2). In addition, the ...

  12. Quantitative tear lysozyme assay: a new technique for transporting specimens.

    OpenAIRE

    Seal, D V; Mackie, I A; Coakes, R L; Farooqi, B

    1980-01-01

    We have developed a method for assaying the concentration of tear lysozyme using eluates of tear fluid collected on filter paper discs. Specimens can be stored and transported to remote laboratories for assay. We have shown that the 'indirect' or eluate method gives statistically comparable results to the 'direct' method using fresh, neat tear fluid.

  13. Quantitative tear lysozyme assay: a new technique for transporting specimens.

    Science.gov (United States)

    Seal, D V; Mackie, I A; Coakes, R L; Farooqi, B

    1980-09-01

    We have developed a method for assaying the concentration of tear lysozyme using eluates of tear fluid collected on filter paper discs. Specimens can be stored and transported to remote laboratories for assay. We have shown that the 'indirect' or eluate method gives statistically comparable results to the 'direct' method using fresh, neat tear fluid.

  14. Performance Evaluation of the MyT4 Technology for Determining ART Eligibility.

    Science.gov (United States)

    Sitoe, Nádia; Macamo, Rosa; Meggi, Bindiya; Tobaiwa, Ocean; Loquiha, Osvaldo; Bollinger, Timothy; Vojnov, Lara; Jani, Ilesh

    2016-01-01

    In resource-limited countries, CD4 T-cell (CD4) testing continues to be used for determining antiretroviral therapy (ART) initiation eligibility and opportunistic infection monitoring. To support expanded access to CD4 testing, simple and robust technologies are necessary. We conducted this study to evaluate the performance of a new Point-of-Care (POC) CD4 technology, the MyT4, compared to conventional laboratory CD4 testing. EDTA venous blood from 200 HIV-positive patients was tested in the laboratory using the MyT4 and BD FACSCalibur™. The MyT4 had an r2 of 0.82 and a mean bias of 12.3 cells/μl. The MyT4 had total misclassifications of 14.7% and 8.8% when analyzed using ART eligibility thresholds of 350 and 500 cells/μl, respectively. We conclude that the MyT4 performed well in classifying patients using the current ART initiation eligibility thresholds in Mozambique when compared to the conventional CD4 technology.

  15. The sequences and activities of RegB endoribonucleases of T4-related bacteriophages.

    Science.gov (United States)

    Piesiniene, Lina; Truncaite, Lidija; Zajanckauskaite, Aurelija; Nivinskas, Rimas

    2004-01-01

    The RegB endoribonuclease encoded by bacteriophage T4 is a unique sequence-specific nuclease that cleaves in the middle of GGAG or, in a few cases, GGAU tetranucleotides, preferentially those found in the Shine-Dalgarno regions of early phage mRNAs. In this study, we examined the primary structures and functional properties of RegB ribonucleases encoded by T4-related bacteriophages. We show that all but one of 36 phages tested harbor the regB gene homologues and the similar signals for transcriptional and post-transcriptional autogenous regulation of regB expression. Phage RB49 in addition to gpRegB utilizes Escherichia coli endoribonuclease E for the degradation of its transcripts for gene regB. The deduced primary structure of RegB proteins of 32 phages studied is almost identical to that of T4, while the sequences of RegB encoded by phages RB69, TuIa and RB49 show substantial divergence from their T4 counterpart. Functional studies using plasmid-phage systems indicate that RegB nucleases of phages T4, RB69, TuIa and RB49 exhibit different activity towards GGAG and GGAU motifs in the specific locations. We expect that the availability of the different phylogenetic variants of RegB may help to localize the amino acid determinants that contribute to the specificity and cleavage efficiency of this processing enzyme.

  16. Analysis of T4SS-induced signaling by H. pylori using quantitative phosphoproteomics

    Directory of Open Access Journals (Sweden)

    Frithjof eGlowinski

    2014-07-01

    Full Text Available Helicobacter pylori is a Gram-negative bacterial pathogen colonizing the human stomach. Infection with H. pylori causes chronic inflammation of the gastric mucosa and may lead to peptic ulceration and/or gastric cancer. A major virulence determinant of H. pylori is the type IV secretion system (T4SS, which is used to inject the virulence factor CagA into the host cell, triggering a wide range of cellular signaling events. Here, we used a phosphoproteomic approach to investigate tyrosine signaling in response to host-pathogen interaction, using stable isotope labeling in cell culture (SILAC of AGS cells to obtain a differential picture between multiple infection conditions. Cells were infected with wild type H. pylori P12, a P12ΔCagA deletion mutant, and a P12ΔT4SS deletion mutant to compare signaling changes over time and in the absence of CagA or the T4SS. Tryptic peptides were enriched for tyrosine (Tyr phosphopeptides and analysed by nano-LC-Orbitrap MS. In total, 58 different phosphosites were found to be regulated following infection. The majority of phosphosites identified were kinases of the MAPK familiy. CagA and the T4SS were found to be key regulators of Tyr phosphosites. Our findings indicate that CagA primarily induces activation of ERK1 and integrin linked factors, whereas the T4SS primarily modulates JNK and p38 activation.

  17. Does Warming a Lysozyme Solution Cook Ones Data?

    Science.gov (United States)

    Pusey, Marc; Burke, Michael; Judge, Russell

    2000-01-01

    Chicken egg white lysozyme has a well characterized thermally driven phase transition. Between pH 4.0 and 5.2, the transition temperature, as defined by the point where the tetragonal and orthorhombic solubility are equal, is a function of the pH, salt (precipitant) type and concentration, and most likely of the buffer concentration as well. This phase transition can be carried out with protein solution alone, prior to the initiation of the crystallization process. We have now measured the kinetics of this process and investigated its reversibility. An aliquot of a stock protein solution is held at a given temperature, and at periodic intervals used to set up batch crystallization experiments. The batch solutions were incubated at 20 C until macroscopic crystals were obtained, at which point the number of crystals in each well were counted. The transition effects increased with temperature, slowly falling off at 30 C with a half time (time to approx. 1/2 the t = 0 number of crystals) of approx. 5 hours, and an estimated half time of approx. 0.5 hours at 43 C. Further, the process was not reversible by simple cooling. After holding a lysozyme solution at 37 C (prior to addition of precipitant) for 16 hours, then cooling and holding it at 4 C, no return to the pre-warmed nucleation kinetics are observed after at least 4 weeks. Thus every thermal excursion above the phase transition point results in a further decrease in the nucleation rate of that solution, the extent being a function of the time and temperature. Orthorhombic lysozyme crystals apparently do not undergo the flow-induced growth cessation of tetragonal lysozyme crystals. We have previously shown that putting the protein in the orthorhombic form does not affect the averaged face growth kinetics, only nucleation, for tetragonal crystals. We may be able to use this differential behavior to elucidate how flow affects tile lysozyme crystal growth process.

  18. T4 RIA precision profiles automatically computed using a small programmable pocket calculator HP-41 CV

    International Nuclear Information System (INIS)

    Ditu, M.; Waud, J.M.; Hamilton, R.G.; Wagner, H.N.

    1984-01-01

    Precision profiles as useful tool for assurance of assay quality in 10 independent T4 RIAs have been automatically obtained using a small programmable pocket calculator HP-41 CV. For each T4 assay batch, the within-assay coefficient of variation varied from 7 to 11% in the hormone concentration range of 2 to 20μg/dl. The difference in coefficient of variation for all the 10 successive assay batches of T4 never exceeded 1% in the same hormone concentrations regions. All the above findings demonstrate that the precision profile can be used as a powerful tool for assessing the assay quality and consistency of overall random error between successive assay batches [fr

  19. Zinc(II) and the single-stranded DNA binding protein of bacteriophage T4

    International Nuclear Information System (INIS)

    Gauss, P.; Krassa, K.B.; McPheeters, D.S.; Nelson, M.A.; Gold, L.

    1987-01-01

    The DNA binding domain of the gene 32 protein of the bacteriophage T4 contains a single zinc-finger sequence. The gene 32 protein is an extensively studied member of a class of proteins that bind relatively nonspecifically to single-stranded DNA. The authors have sequenced and characterized mutations in gene 32 whose defective proteins are activated by increasing the Zn(II) concentration in the growth medium. The results identify a role for the gene 32 protein in activation of T4 late transcription. Several eukaryotic proteins with zinc fingers participate in activation of transcription, and the gene 32 protein of T4 should provide a simple, well-characterized system in which genetics can be utilized to study the role of a zinc finger in nucleic acid binding and gene expression

  20. T4 syndrome - A distinct theoretical concept or elusive clinical entity? A case report.

    Science.gov (United States)

    Hirai, Patricia Miyuki; Thomson, Oliver P

    2016-10-01

    T4 syndrome has existed as a clinical concept for more than three decades and it has been identified as a source of upper extremity (UE) symptoms. This case report explores the clinical reasoning in the diagnoses and management of a patient with symptoms consistent with T4-type syndrome and critically discusses the concept of T4 syndrome using recent research to help explain the clinical presentation. Manual therapy treatment focused on stimulation of the sympathetic ganglia, decreasing local upper thoracic pain and UE referral pattern noted during passive examination. The successful outcomes included immediate and lasting symptom relief after upper thoracic spinal manipulation. Although treatment has been based on the theory that mechanical thoracic dysfunction can produce sympathetic nervous system (SNS) referred pain, the role the sympathetic reflexes potentially plays on the referral symptoms to the UE presently remains unclear. Copyright © 2016 Elsevier Ltd. All rights reserved.

  1. Time course and vectorial nature of albumin metabolism in isolated perfused rabbit PCT

    International Nuclear Information System (INIS)

    Park, C.H.

    1988-01-01

    The time course and vectorial nature of renal metabolism of albumin (Alb) were studied. The tubular absorption, accumulation, and hydrolysis of Alb and the release of the hydrolysis products were determined in the isolated rabbit proximal convoluted tubule (PCT) perfused with tritiated Alb ([ 3 H3C]Alb) at 36.4 micrograms/ml. The Alb absorption across the apical membrane was constant (99.9 +/- 4.9 x 10(-3) ng.min-1.mm-1). In contrast, the accumulation and hydrolysis of Alb in the cells increased nonlinearly with time. The bulk of the tritium that accumulated in the cells was associated with intact [ 3 H3C]Alb. Only the final hydrolysis products were released from the cells and these first appeared in the peritubular bath 6-7 min after the start of perfusion of the tubule with [ 3 H3C]Alb. The hydrolysis product was not detectable in the tubule lumen. The proteolytic activity correlated linearly with the protein load to the cells, characteristic of first-order kinetics and a high-capacity system. The results suggest that the renal tubular handling of proteins proceeds from the apical to the basolateral aspect of the cell. The transcellular processing of Alb is rapid and can occur in 6-7 min. The accumulation of intact protein in the cell and the first-order kinetics of hydrolysis of the absorbed protein suggest that the rate-limiting step in proximal tubular handling of proteins may include the initial hydrolysis of protein or reside in steps that precede the hydrolysis

  2. Ukraine’s Multi-Vectorial Foreign Policy: Looking West while not Overlooking its Eastern Neighbour

    Directory of Open Access Journals (Sweden)

    María Raquel Freire

    2009-05-01

    Full Text Available La Ucrania post-soviética ha perseguido un curso independiente en su política exterior desde que lograse la independencia en 1991. Sin embargo su posición geográfica condicionó mucho el contorno de su política exterior. De hecho, mientras que un acercamiento al Oeste ha sido valorado grandemente, las relaciones con Rusia han seguido siendo el pilar central de Ucrania, intentándose por tanto, mirar hacia el Oeste sin perder de vista a su vecino oriental. En tal contexto, ¿cómo podrían conciliar las autoridades ucranianas los diferentes vectores de la política exterior de su país?¿En qué medida podrían pervivir los principios de la Revolución Naranja entre un contexto interior tan difícil y un contexto exterior complejo?¿En qué medida pueden afectar a las opciones en política exterior de Kiev la influencia occidental y las interferencias rusas? Intentando encontrar respuestas a tales preguntas, este artículo tiene como objetivo la deconstrucción de los varios círculos en la política exterior multi-vectorial de Kiev que se encargan de atender a la política exterior de una manera integrada y donde las formulaciones interiores se ven condicionadas por las opciones externas, echando luz de esta manera sobre cómo las interrelaciones e interconexiones se desarrollan en el proceso de toma de decisiones y en el curso elegido.

  3. Time course and vectorial nature of albumin metabolism in isolated perfused rabbit PCT.

    Science.gov (United States)

    Park, C H

    1988-09-01

    The time course and vectorial nature of renal metabolism of albumin (Alb) were studied. The tubular absorption, accumulation, and hydrolysis of Alb and the release of the hydrolysis products were determined in the isolated rabbit proximal convoluted tubule (PCT) perfused with tritiated Alb ([3H3C]Alb) at 36.4 micrograms/ml. The Alb absorption across the apical membrane was constant (99.9 +/- 4.9 x 10(-3) ng.min-1.mm-1). In contrast, the accumulation and hydrolysis of Alb in the cells increased nonlinearly with time. The bulk of the tritium that accumulated in the cells was associated with intact [3H3C]Alb. Only the final hydrolysis products were released from the cells and these first appeared in the peritubular bath 6-7 min after the start of perfusion of the tubule with [3H3C]Alb. The hydrolysis product was not detectable in the tubule lumen. The proteolytic activity correlated linearly with the protein load to the cells, characteristic of first-order kinetics and a high-capacity system. The results suggest that the renal tubular handling of proteins proceeds from the apical to the basolateral aspect of the cell. The transcellular processing of Alb is rapid and can occur in 6-7 min. The accumulation of intact protein in the cell and the first-order kinetics of hydrolysis of the absorbed protein suggest that the rate-limiting step in proximal tubular handling of proteins may include the initial hydrolysis of protein or reside in steps that precede the hydrolysis.

  4. DDiT4L promotes autophagy and inhibits pathological cardiac hypertrophy in response to stress.

    Science.gov (United States)

    Simonson, Bridget; Subramanya, Vinita; Chan, Mun Chun; Zhang, Aifeng; Franchino, Hannabeth; Ottaviano, Filomena; Mishra, Manoj K; Knight, Ashley C; Hunt, Danielle; Ghiran, Ionita; Khurana, Tejvir S; Kontaridis, Maria I; Rosenzweig, Anthony; Das, Saumya

    2017-02-28

    Physiological cardiac hypertrophy, in response to stimuli such as exercise, is considered adaptive and beneficial. In contrast, pathological cardiac hypertrophy that arises in response to pathological stimuli such as unrestrained high blood pressure and oxidative or metabolic stress is maladaptive and may precede heart failure. We found that the transcript encoding DNA damage-inducible transcript 4-like (DDiT4L) was expressed in murine models of pathological cardiac hypertrophy but not in those of physiological cardiac hypertrophy. In cardiomyocytes, DDiT4L localized to early endosomes and promoted stress-induced autophagy through a process involving mechanistic target of rapamycin complex 1 (mTORC1). Exposing cardiomyocytes to various types of pathological stress increased the abundance of DDiT4L , which inhibited mTORC1 but activated mTORC2 signaling. Mice with conditional cardiac-specific overexpression of DDiT4L had mild systolic dysfunction, increased baseline autophagy, reduced mTORC1 activity, and increased mTORC2 activity, all of which were reversed by suppression of transgene expression. Genetic suppression of autophagy also reversed cardiac dysfunction in these mice. Our data showed that DDiT4L may be an important transducer of pathological stress to autophagy through mTOR signaling in the heart and that DDiT4L could be therapeutically targeted in cardiovascular diseases in which autophagy and mTOR signaling play a major role. Copyright © 2017, American Association for the Advancement of Science.

  5. Immunogenicity studies of proteins forming the T4 phage head surface.

    Science.gov (United States)

    Dąbrowska, Krystyna; Miernikiewicz, Paulina; Piotrowicz, Agnieszka; Hodyra, Katarzyna; Owczarek, Barbara; Lecion, Dorota; Kaźmierczak, Zuzanna; Letarov, Andrey; Górski, Andrzej

    2014-11-01

    Advances in phage therapy and novel applications of phages in biotechnology encourage interest in phage impact on human and animal immunity. Here we present comparative studies of immunogenic properties of T4 phage head surface proteins gp23*, gp24*, Hoc, and Soc, both as elements of the phage capsid and as isolated agents. Studies comprise evaluation of specific antibodies in the human population, analysis of the proteins' impact on the primary and secondary responses in mice, and the effect of specific antibodies on phage antibacterial activity in vitro and in vivo in mice. In humans, natural antibodies specific to T4-like phages were abundant (81% of investigated sera). Among those, significantly elevated levels of IgG antibodies only against major head protein (gp23*) were found, which probably reflected cross-reactions of T4 with antibodies induced by other T4-like phages. Both IgM and IgG antibodies were induced mostly by gp23* and Hoc, while weak (gp24*) and very weak (Soc) reactivities of other head proteins were noticed. Thus, T4 head proteins that markedly contribute to immunological memory to the phage are highly antigenic outer capsid protein (Hoc) and major capsid protein (gp23*). Specific anti-gp23* and anti-Hoc antibodies substantially decreased T4 phage activity in vitro and to some extent in vivo. Cooperating with antibodies, the immune complement system also contributed to annihilating phages. Current descriptions of phage immunogenicity and its biological consequences are still vague and incomplete; thus, the central problem of this work is timely and may have strong practical implications. Here is presented the very first description of the contribution of bacteriophage proteins to immunological memory of the phage. Understanding of interactions between phages and mammalian immunology may help in biotechnological adaptations of phages for therapeutic requirements as well as for better appreciation of phage ecology and their role in the biosphere

  6. Covalent Modification of Bacteriophage T4 DNA Inhibits CRISPR-Cas9.

    Science.gov (United States)

    Bryson, Alexandra L; Hwang, Young; Sherrill-Mix, Scott; Wu, Gary D; Lewis, James D; Black, Lindsay; Clark, Tyson A; Bushman, Frederic D

    2015-06-16

    The genomic DNAs of tailed bacteriophages are commonly modified by the attachment of chemical groups. Some forms of DNA modification are known to protect phage DNA from cleavage by restriction enzymes, but others are of unknown function. Recently, the CRISPR-Cas nuclease complexes were shown to mediate bacterial adaptive immunity by RNA-guided target recognition, raising the question of whether phage DNA modifications may also block attack by CRISPR-Cas9. We investigated phage T4 as a model system, where cytosine is replaced with glucosyl-hydroxymethylcytosine (glc-HMC). We first quantified the extent and distribution of covalent modifications in T4 DNA by single-molecule DNA sequencing and enzymatic probing. We then designed CRISPR spacer sequences targeting T4 and found that wild-type T4 containing glc-HMC was insensitive to attack by CRISPR-Cas9 but mutants with unmodified cytosine were sensitive. Phage with HMC showed only intermediate sensitivity. While this work was in progress, another group reported examples of heavily engineered CRISRP-Cas9 complexes that could, in fact, overcome the effects of T4 DNA modification, indicating that modifications can inhibit but do not always fully block attack. Bacteria were recently found to have a form of adaptive immunity, the CRISPR-Cas systems, which use nucleic acid pairing to recognize and cleave genomic DNA of invaders such as bacteriophage. Historic work with tailed phages has shown that phage DNA is often modified by covalent attachment of large chemical groups. Here we demonstrate that DNA modification in phage T4 inhibits attack by the CRISPR-Cas9 system. This finding provides insight into mechanisms of host-virus competition and also a new set of tools that may be useful in modulating the activity of CRISPR-Cas9 in genome engineering applications. Copyright © 2015 Bryson et al.

  7. Prognostic Factors Affecting Survival After Multivisceral Resection in Patients with Clinical T4b Gastric Cancer.

    Science.gov (United States)

    Mita, Kazuhito; Ito, Hideto; Katsube, Toshio; Tsuboi, Ayaka; Yamazaki, Nobuyoshi; Asakawa, Hideki; Hayashi, Takashi; Fujino, Keiichi

    2017-12-01

    The prognosis and survival of patients with advanced gastric cancer is poor. Although completeness of resection (R0) is one of the most important factors affecting survival, multivisceral resection (MVR) for locally advanced (clinical T4b, cT4b) gastric cancer remains controversial. The aim of this study was to evaluate the factors affecting prognosis and survival after MVR in patients with cT4b gastric cancer. Between 2005 and 2015, we retrospectively reviewed the medical records of 103 patients who underwent MVR for cT4b gastric cancer with suspected direct invasion to adjacent organs. Patient characteristics, related complications, long-term survival, and prognostic factors of cT4b gastric cancer were analyzed. Postoperative mortality and morbidity rates of patients after MVR were 1.0 and 37.9%, respectively. R0 resection was achieved in 82.5% patients, all of whom had a significantly improved survival rate. Overall survival rates at 1 and 3 years were 78.3 and 47.7% for R0 resection and 46.6 and 14.3% for R1 resection, respectively (R0 vs. R1, P < 0.002). Multivariate analysis revealed that completeness of resection (R0) was an independent prognostic factor associated with longer survival. In patients with cT4b gastric cancer, gastrectomy with MVR to achieve an R0 resection can be performed with acceptable postoperative morbidity and mortality rates and can have a positive impact on long-term survival.

  8. Genomic characteristics and environmental distributions of the uncultivated Far-T4 phages

    OpenAIRE

    Roux, Simon; Enault, François; Ravet, Viviane; Pereira, Olivier; Sullivan, Matthew B.

    2015-01-01

    Viral metagenomics (viromics) is a tremendous tool to reveal viral taxonomic and functional diversity across ecosystems ranging from the human gut to the world's oceans. As with microbes however, there appear vast swaths of ?dark matter? yet to be documented for viruses, even among relatively well-studied viral types. Here, we use viromics to explore the ?Far-T4 phages? sequence space, a neighbor clade from the well-studied T4-like phages that was first detected through PCR study in seawater ...

  9. Preparation of quality control samples for thyroid hormones T3 and T4 in radioimmunoassay techniques

    International Nuclear Information System (INIS)

    Ahmed, F.O.A.

    2006-03-01

    Today, the radioimmunoassay becomes one of the best techniques for quantitative analysis of very low concentration of different substances. RIA is being widely used in medical and research laboratories. To maintain high specificity and accuracy in RIA and other related techniques the quality controls must be introduced. In this dissertation quality control samples for thyroid hormones (Triiodothyronine T3 and Thyroxin T4), using RIA techniques. Ready made chinese T4, T3 RIA kits were used. IAEA statistical package were selected.(Author)

  10. De evaluatie van Corning "MAGIC T4-RIA" en de vergelijking met Corning "IMMOPHASE FT4-RIA"

    NARCIS (Netherlands)

    Elvers LH; Smit PJ; Loeber JG

    1985-01-01

    Beschreven wordt de evaluatie van "MAGIC T4-RIA" (MAGIC) en de vergelijking met de totaal T4 resultaten van "IMMOPHASE FT4-RIA" (IMMO). MAGIC is in de practische uitvoerbaarheid eenvoudig en weinig arbeidsintensief. Voor 3 rattesera met 44, 95 en 167 nmol T4/l bedroeg de

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2012-07-11

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

  12. Generation of recombinant destabilase-lysozyme from medicinal leeches in three different expression systems.

    Science.gov (United States)

    Manuvera, Valentin A; Kurdyumov, Alexey S; Filonova, Kseniya A; Lazarev, Vassili N

    2015-12-01

    Destabilase-lysozyme (mlDL) is an enzyme secreted by the salivary gland cells of medicinal leeches. Destabilase-lysozyme possesses lysozyme and isopeptidase activities. We generated recombinant destabilase-lysozyme isoform 2 in three expression systems, i.e., in the bacteria Escherichia coli, in the yeast Pichia pastoris, and in the human cell line Expi293F. In E. coli, we generated both polypeptide in inclusion bodies that was later undergone to the refolding and soluble protein that had been fused with the chaperone SlyD. The chaperone was later cleaved by a specific TEV-protease. In cultures of the yeast P. pastoris and the human cell line Expi293F, the soluble form of destabilase-lysozyme was accumulated in the culture media. For the generated enzymes, we determined the lysozyme, isopeptidase and fibrinolytic activities and tested their general antimicrobial effects. The comparisons of the enzymes generated in the different expression systems revealed that all of the destabilase-lysozymes obtained in the soluble forms possessed equal levels of lysozyme, isopeptidase and fibrinolytic activities that exceeded several to ten times the levels of the same activities of the destabilase-lysozyme renaturated from the inclusion bodies. A similar pattern of the differences in the levels of the general antimicrobial effects was observed for the destabilase-lysozymes generated in the soluble form and as inclusion bodies. Copyright © 2015 Elsevier Inc. All rights reserved.

  13. Measuring Plasticity with Orientation Contrast Microscopy in Aluminium 6061-T4

    NARCIS (Netherlands)

    Ghodrat, S.; Riemslag, A.C.; Kestens, L.A.I.

    2017-01-01

    Orientation contrast microscopy (i.e., electron backscattered diffraction, EBSD) was employed to monitor the plastic strain in loaded tensile samples of aluminium alloy Al6061 in T4 condition. The kernel average misorientation (KAM) is known to be an appropriate parameter in orientation contrast

  14. Radioimmunoassay of serum T3, T4 and TSH during anesthesia and operation

    International Nuclear Information System (INIS)

    Gosheva-Antonova, Ts.; Zakharieva, B.; Kurtev, I.

    1987-01-01

    The serum concentrations of thyroxine (T 3 ), triiodothyronine (T 4 ) and thyroid-stimulating hormone (TSH) were determined in 31 partients before and during urologic operations on the 30th and 60th minute since the onset of the operation, performed under endotracheal halotane or neuroleptanesthesia (NLA) in assisted breathing and intravenous drip anesthesia with ketalar-diazepam in spontaneous breathing. There was statistically significant rise in T 4 level, decrease in T 3 and negligible changes in TSH level, in patients operated under halotane anesthesia. In those operated under NLA, T 4 tended initially to be elevated, with subseguent fall to starting level, with a tendency toward rise in TSH and stable unchanged T 3 level. Ketalar-diazepam anesthesia was applied only to patients subjected to transurethral resections. T 4 in them tended to be decreased, while T 3 and TSH showed negligible changes. Since the operations of patients anesthesized with halotane and NLA had similar localizations and severity, the differences in the thyroid hormone reactions could be associated with the type of anesthesia. The negligible changes in TSH are highly suggestive that this hormone is not influenced by the operation stress and anesthetics, and does hot exert regulating effect upon the thyroid status under these conditions. The milder reactions in patients operated under ketalar-diazepam anestesia may largely be associated with the milder operation stress in transurethal resection

  15. The isolation and characterization of Stenotrophomonas maltophilia T4-like bacteriophage DLP6.

    Directory of Open Access Journals (Sweden)

    Danielle L Peters

    Full Text Available Increasing isolation of the extremely antibiotic resistant bacterium Stenotrophomonas maltophilia has caused alarm worldwide due to the limited treatment options available. A potential treatment option for fighting this bacterium is 'phage therapy', the clinical application of bacteriophages to selectively kill bacteria. Bacteriophage DLP6 (vB_SmoM-DLP6 was isolated from a soil sample using clinical isolate S. maltophilia strain D1571 as host. Host range analysis of phage DLP6 against 27 clinical S. maltophilia isolates shows successful infection and lysis in 13 of the 27 isolates tested. Transmission electron microscopy of DLP6 indicates that it is a member of the Myoviridae family. Complete genome sequencing and analysis of DLP6 reveals its richly recombined evolutionary history, featuring a core of both T4-like and cyanophage genes, which suggests that it is a member of the T4-superfamily. Unlike other T4-superfamily phages however, DLP6 features a transposase and ends with 229 bp direct terminal repeats. The isolation of this bacteriophage is an exciting discovery due to the divergent nature of DLP6 in relation to the T4-superfamily of phages.

  16. Structural studies on metal-containing enzymes: T4 endonuclease VII and D. gigas formate dehydrogenase

    NARCIS (Netherlands)

    Raaijmakers, H.C.A.

    2001-01-01

    Many biological processes require metal ions, and many of these metal-ion functions involve metalloproteins. The metal ions in metalloproteins are often critical to the protein's function, structure, or stability. This thesis focuses on two of these proteins, bacteriophage T4 endonuclease

  17. A unique intracellular compartment formed during the oligotrophic growth of Rhodococcus erythropolis N9T-4.

    Science.gov (United States)

    Yoshida, Nobuyuki; Yano, Takanori; Kedo, Kaori; Fujiyoshi, Takuya; Nagai, Rina; Iwano, Megumi; Taguchi, Eiji; Nishida, Tomoki; Takagi, Hiroshi

    2017-01-01

    Rhodococcus erythropolis N9T-4, isolated from stored crude oil, shows extremely oligotrophic features and can grow on a basal medium without any additional carbon, nitrogen, sulfur, and energy sources, but requires CO 2 for its oligotrophic growth. Transmission electron microscopic observation showed that a relatively large and spherical compartment was observed in a N9T-4 cell grown under oligotrophic conditions. In most cases, only one compartment was observed per cell, but in some cases, it was localized at each pole of the cell, suggesting that it divides at cell division. We termed this unique bacterial compartment an oligobody. The oligobody was not observed or very rarely observed in small sizes under nutrient rich conditions, whereas additional carbon sources did not affect oligobody formation. Energy dispersive X-ray spectroscopy analysis revealed remarkable peaks corresponding to phosphorus and potassium in the oligobody. The oligobodies in N9T-4 cells could be stained by Toluidine blue, suggesting that the oligobody is composed of inorganic polyphosphate and is a type of acidocalcisome. Two genes-encoding polyphosphate kinases, ppk1 and ppk2, were found in the N9T-4 genome: ppk1 disruption caused a negative effect on the formation of the oligobody. Although it was suggested that the oligobody plays an important role for the oligotrophic growth, both ppk-deleted mutants showed the same level of oligotrophic growth as the wild-type strain.

  18. Intracellular accumulation of trehalose and glycogen in an extreme oligotroph, Rhodococcus erythropolis N9T-4.

    Science.gov (United States)

    Yano, Takanori; Funamizu, Yuhei; Yoshida, Nobuyuki

    2016-01-01

    An extreme oligotroph, Rhodococcus erythropolis N9T-4, showed intracellular accumulation of trehalose and glycogen under oligotrophic conditions. No trehalose accumulation was observed in cells grown on the rich medium. Deletion of the polyphosphate kinase genes enhanced the trehalose accumulation and decreases the intracellular glycogen contents, suggesting an oligotrophic relationship between among the metabolic pathways of trehalose, glycogen, and inorganic polyphosphate biosyntheses.

  19. Phylogenetic Diversity of T4-Type Phages in Sediments from the Subtropical Pearl River Estuary.

    Science.gov (United States)

    He, Maoqiu; Cai, Lanlan; Zhang, Chuanlun; Jiao, Nianzhi; Zhang, Rui

    2017-01-01

    Viruses are an abundant and active component of marine sediments and play a significant role in microbial ecology and biogeochemical cycling at local and global scales. To obtain a better understanding of the ecological characteristics of the viriobenthos, the abundance and morphology of viruses and the diversity and community structure of T4-type phages were systematically investigated in the surface sediments of the subtropical Pearl River Estuary (PRE). Viral abundances ranged from 4.49 × 10 8 to 11.7 × 10 8 viruses/g and prokaryotic abundances ranged from 2.63 × 10 8 to 9.55 × 10 8 cells/g, and both decreased from freshwater to saltwater. Diverse viral morphotypes, including tailed, spherical, filamentous, and rod-shaped viruses, were observed using transmission electron microscopy. Analysis of the major capsid gene ( g23 ) indicated that the sediment T4-type phages were highly diverse and, similar to the trend in viral abundances, their diversity decreased as the salinity increased. Phylogenetic analysis suggested that most of the g23 operational taxonomic units were affiliated with marine, paddy soil, and lake groups. The T4-type phage communities in freshwater and saltwater sediments showed obvious differences, which were related to changes in the Pearl River discharge. The results of this study demonstrated both allochthonous and autochthonous sources of the viral community in the PRE sediments and the movement of certain T4-type viral groups between the freshwater and saline water biomes.

  20. Phylogenetic Diversity of T4-Type Phages in Sediments from the Subtropical Pearl River Estuary

    Directory of Open Access Journals (Sweden)

    Maoqiu He

    2017-05-01

    Full Text Available Viruses are an abundant and active component of marine sediments and play a significant role in microbial ecology and biogeochemical cycling at local and global scales. To obtain a better understanding of the ecological characteristics of the viriobenthos, the abundance and morphology of viruses and the diversity and community structure of T4-type phages were systematically investigated in the surface sediments of the subtropical Pearl River Estuary (PRE. Viral abundances ranged from 4.49 × 108 to 11.7 × 108 viruses/g and prokaryotic abundances ranged from 2.63 × 108 to 9.55 × 108 cells/g, and both decreased from freshwater to saltwater. Diverse viral morphotypes, including tailed, spherical, filamentous, and rod-shaped viruses, were observed using transmission electron microscopy. Analysis of the major capsid gene (g23 indicated that the sediment T4-type phages were highly diverse and, similar to the trend in viral abundances, their diversity decreased as the salinity increased. Phylogenetic analysis suggested that most of the g23 operational taxonomic units were affiliated with marine, paddy soil, and lake groups. The T4-type phage communities in freshwater and saltwater sediments showed obvious differences, which were related to changes in the Pearl River discharge. The results of this study demonstrated both allochthonous and autochthonous sources of the viral community in the PRE sediments and the movement of certain T4-type viral groups between the freshwater and saline water biomes.

  1. Platinum(II) complexes block the entry of T4 phage DNA into the host cells

    DEFF Research Database (Denmark)

    Kerszman, Gustaw; Josephsen, Jens; Fernholm, Bo

    1979-01-01

    The efficiency of multiplicity reactivation of T4 particles inactivated by platinum(II) complexes is very low. The same is true for marker rescue and functional survival of genes. This can be at least partly explained by the inability of most inactivated virus particles to introduce their DNA...

  2. The isolation and characterization of Stenotrophomonas maltophilia T4-like bacteriophage DLP6.

    Science.gov (United States)

    Peters, Danielle L; Stothard, Paul; Dennis, Jonathan J

    2017-01-01

    Increasing isolation of the extremely antibiotic resistant bacterium Stenotrophomonas maltophilia has caused alarm worldwide due to the limited treatment options available. A potential treatment option for fighting this bacterium is 'phage therapy', the clinical application of bacteriophages to selectively kill bacteria. Bacteriophage DLP6 (vB_SmoM-DLP6) was isolated from a soil sample using clinical isolate S. maltophilia strain D1571 as host. Host range analysis of phage DLP6 against 27 clinical S. maltophilia isolates shows successful infection and lysis in 13 of the 27 isolates tested. Transmission electron microscopy of DLP6 indicates that it is a member of the Myoviridae family. Complete genome sequencing and analysis of DLP6 reveals its richly recombined evolutionary history, featuring a core of both T4-like and cyanophage genes, which suggests that it is a member of the T4-superfamily. Unlike other T4-superfamily phages however, DLP6 features a transposase and ends with 229 bp direct terminal repeats. The isolation of this bacteriophage is an exciting discovery due to the divergent nature of DLP6 in relation to the T4-superfamily of phages.

  3. UV irradiation impairs in vivo encapsidation of bacteriophage T4 DNA

    International Nuclear Information System (INIS)

    Zachary, A.; Black, L.W.

    1984-01-01

    T4 DNA structural requirements for encapsidation in vivo were investigated, using thin-section electron microscopy to quantitate the kinetics and yields of head intermediates after synchronous DNA packaging into accumulated processed proheads. UV irradiation (254 nm) of T4-infected bacteria just before initiation of encapsidation resulted in a reduction in the rate of DNA packaged measured by electron microscopy and in the yield of viable phage progeny. In UV-irradiated infections with excision-deficient mutants (denV-), the extent of packaging decline was proportional to the UV dose and phage yields were lower than expected based on the packaging levels observed by microscopy. Rescue analysis of progeny from such infections revealed elevated levels of nonviable virions. Pyrimidine dimers were encapsidated in denV- infections, but in excision-competent infections (denV+) dimers were not packaged. A UV-independent, 15 to 20% packaging arrest was also observed when denV endonuclease was inactive during encapsidation, indicating a denV requirement to achieve normal T4 packaging levels. Pyrimidine dimers apparently represent or induce transient blockage of DNA encapsidation or both, causing a decline in the rate. This is in contrast to other DNA structural blocks to packaging induced by mutations in T4 genes 30 and 49, which appear to arrest the process

  4. The complete AdS_3 x S^3 x T^4 worldsheet S-matrix

    NARCIS (Netherlands)

    Borsato, Riccardo|info:eu-repo/dai/nl/370952170; Sax, Olof Ohlsson; Sfondrini, Alessandro|info:eu-repo/dai/nl/330983083; jr, Bogdan Stefanski

    2014-01-01

    We derive the non-perturbative worldsheet S matrix for fundamental excitations of Type IIB superstring theory on AdS_3 x S^3 x T^4 with Ramond-Ramond flux. To this end, we study the off-shell symmetry algebra of the theory and its representations. We use these to determine the S matrix up to scalar

  5. Ethnic differences in TSH but not in free T4 concentrations or TPO antibodies during pregnancy

    NARCIS (Netherlands)

    Benhadi, N.; Wiersinga, W. M.; Reitsma, J. B.; Vrijkotte, T. G. M.; van der Wal, M. F.; Bonsel, G. J.

    2007-01-01

    OBJECTIVE: To describe the TSH, free T4 and thyroid peroxidase antibody (TPO-Ab) concentrations during pregnancy among four ethnic groups and to determine reference values for these parameters during normal pregnancy. METHODS: Cross-sectional study of a cohort of 3270 pregnant women living in the

  6. In vitro selection of optimal DNA substrates for T4 RNA ligase

    Science.gov (United States)

    Harada, Kazuo; Orgel, Leslie E.

    1993-01-01

    We have used in vitro selection techniques to characterize DNA sequences that are ligated efficiently by T4 RNA ligase. We find that the ensemble of selected sequences ligated about 10 times as efficiently as the random mixture of sequences used as the input for selection. Surprisingly, the majority of the selected sequences approximated a well-defined consensus sequence.

  7. Potentially pathogenic Acanthamoeba genotype T4 isolated from dental units and emergency combination showers.

    Science.gov (United States)

    Castro-Artavia, Esteban; Retana-Moreira, Lissette; Lorenzo-Morales, Jacob; Abrahams-Sandí, Elizabeth

    2017-12-01

    Acanthamoeba is the genus of free-living amoebae that is most frequently isolated in nature. To date, 20 Acanthamoeba genotypes have been described. Genotype T4 is responsible for approximately 90% of encephalitis and keratitis cases. Due to the ubiquitous presence of amoebae, isolation from environmental sources is not uncommon; to determine the clinical importance of an isolation, it is necessary to have evidence of the pathogenic potential of amoebae. The aim of this study was to physiologically characterise 8 Acanthamoeba T4 isolates obtained from dental units and emergency combination showers and to determine their pathogenic potential by employing different laboratory techniques. Eight axenic cultures of Acanthamoeba genotype T4 were used in pathogenic potential assays. Osmotolerance, thermotolerance, determination and characterisation of extracellular proteases and evaluation of cytopathic effects in MDCK cells were performed. All of the isolates were osmotolerant, thermotolerant and had serine proteases from 44-122 kDa. Two isolates had cytopathic effects on the MDCK cell monolayer. The presence of Acanthamoeba T4 with pathogenic potential in areas such as those tested in this study reaffirms the need for adequate cleaning and maintenance protocols to reduce the possibility of infection with free-living amoebae.

  8. Potentially pathogenic Acanthamoeba genotype T4 isolated from dental units and emergency combination showers

    Directory of Open Access Journals (Sweden)

    Esteban Castro-Artavia

    Full Text Available BACKGROUND Acanthamoeba is the genus of free-living amoebae that is most frequently isolated in nature. To date, 20 Acanthamoeba genotypes have been described. Genotype T4 is responsible for approximately 90% of encephalitis and keratitis cases. Due to the ubiquitous presence of amoebae, isolation from environmental sources is not uncommon; to determine the clinical importance of an isolation, it is necessary to have evidence of the pathogenic potential of amoebae. OBJECTIVE The aim of this study was to physiologically characterise 8 Acanthamoeba T4 isolates obtained from dental units and emergency combination showers and to determine their pathogenic potential by employing different laboratory techniques. METHODS Eight axenic cultures of Acanthamoeba genotype T4 were used in pathogenic potential assays. Osmotolerance, thermotolerance, determination and characterisation of extracellular proteases and evaluation of cytopathic effects in MDCK cells were performed. FINDINGS All of the isolates were osmotolerant, thermotolerant and had serine proteases from 44-122 kDa. Two isolates had cytopathic effects on the MDCK cell monolayer. MAIN CONCLUSION The presence of Acanthamoeba T4 with pathogenic potential in areas such as those tested in this study reaffirms the need for adequate cleaning and maintenance protocols to reduce the possibility of infection with free-living amoebae.

  9. New Insights into DNA Polymerase Function Revealed by Phosphonoacetic Acid-Sensitive T4 DNA Polymerases.

    Science.gov (United States)

    Zhang, Likui

    2017-11-20

    The bacteriophage T4 DNA polymerase (pol) and the closely related RB69 DNA pol have been developed into model enzymes to study family B DNA pols. While all family B DNA pols have similar structures and share conserved protein motifs, the molecular mechanism underlying natural drug resistance of nonherpes family B DNA pols and drug sensitivity of herpes DNA pols remains unknown. In the present study, we constructed T4 phages containing G466S, Y460F, G466S/Y460F, P469S, and V475W mutations in DNA pol. These amino acid substitutions replace the residues in drug-resistant T4 DNA pol with residues found in drug-sensitive herpes family DNA pols. We investigated whether the T4 phages expressing the engineered mutant DNA pols were sensitive to the antiviral drug phosphonoacetic acid (PAA) and characterized the in vivo replication fidelity of the phage DNA pols. We found that G466S substitution marginally increased PAA sensitivity, whereas Y460F substitution conferred resistance. The phage expressing a double mutant G466S/Y460F DNA pol was more PAA-sensitive. V475W T4 DNA pol was highly sensitive to PAA, as was the case with V478W RB69 DNA pol. However, DNA replication was severely compromised, which resulted in the selection of phages expressing more robust DNA pols that have strong ability to replicate DNA and contain additional amino acid substitutions that suppress PAA sensitivity. Reduced replication fidelity was observed in all mutant phages expressing PAA-sensitive DNA pols. These observations indicate that PAA sensitivity and fidelity are balanced in DNA pols that can replicate DNA in different environments.

  10. Engineering of Bacteriophage T4 Genome Using CRISPR-Cas9.

    Science.gov (United States)

    Tao, Pan; Wu, Xiaorong; Tang, Wei-Chun; Zhu, Jingen; Rao, Venigalla

    2017-10-20

    Bacteriophages likely constitute the largest biomass on Earth. However, very few phage genomes have been well-characterized, the tailed phage T4 genome being one of them. Even in T4, much of the genome remained uncharacterized. The classical genetic strategies are tedious, compounded by genome modifications such as cytosine hydroxylmethylation and glucosylation which makes T4 DNA resistant to most restriction endonucleases. Here, using the type-II CRISPR-Cas9 system, we report the editing of both modified (ghm-Cytosine) and unmodified (Cytosine) T4 genomes. The modified genome, however, is less susceptible to Cas9 nuclease attack when compared to the unmodified genome. The efficiency of restriction of modified phage infection varied greatly in a spacer-dependent manner, which explains some of the previous contradictory results. We developed a genome editing strategy by codelivering into E. coli a CRISPR-Cas9 plasmid and a donor plasmid containing the desired mutation(s). Single and multiple point mutations, insertions and deletions were introduced into both modified and unmodified genomes. As short as 50-bp homologous flanking arms were sufficient to generate recombinants that can be selected under the pressure of CRISPR-Cas9 nuclease. A 294-bp deletion in RNA ligase gene rnlB produced viable plaques, demonstrating the usefulness of this editing strategy to determine the essentiality of a given gene. These results provide the first demonstration of phage T4 genome editing that might be extended to other phage genomes in nature to create useful recombinants for phage therapy applications.

  11. Marine T4-type bacteriophages, a ubiquitous component of the dark matter of the biosphere

    Science.gov (United States)

    Filée, Jonathan; Tétart, Françoise; Suttle, Curtis A.; Krisch, H. M.

    2005-08-01

    Tailed bacteriophages are the most abundant biological entities in marine environments. However, most of these marine phages are uncharacterized because few of their hosts have been cultivated. To learn more about such phages, we designed a set of degenerate PCR primers for phage T4 g23, which encodes the major capsid protein in all of the T4-type phages, an important family of the tailed phage. These primers were used to amplify g23-related sequences from diverse marine environments (fjords and bays of British Columbia, the eastern Gulf of Mexico, and the western Arctic Ocean) revealing a remarkable level of molecular diversity, which in some cases was correlated with morphological variation of the virions. Phylogenetic analysis showed that although some of these sequences were closely related to well studied subgroups of the T4-type phage, such as the T-evens, the majority of them belong to five previously uncharacterized subgroups. These data indicate that the host range of T4-type phages is much broader than previously imagined and that the laboratory isolate T4 belongs to a phage family that is extraordinarily widespread and diverse in the biosphere. Author contributions: J.F., F.T., and H.M.K. designed research; J.F. and F.T. performed research; C.A.S. contributed new reagents/analytic tools; J.F., C.A.S., and H.M.K. analyzed data; and J.F., C.A.S., and H.M.K. wrote the paper.This paper was submitted directly (Track II) to the PNAS office.Data deposition: The nucleotide sequences of g23 reported in this paper have been deposited in the GenBank database (accession nos. DQ105858-DQ105942).

  12. Prognostic Value of Subclassification Using MRI in the T4 Classification Nasopharyngeal Carcinoma Intensity-Modulated Radiotherapy Treatment

    Energy Technology Data Exchange (ETDEWEB)

    Chen Lei [State Key Laboratory of Oncology in South China, Department of Radiation Oncology, Sun Yat-sen University Cancer Center, Guangzhou (China); Liu Lizhi [State Key Laboratory of Oncology in South China, Imaging Diagnosis and Interventional Center, Sun Yat-sen University Cancer Center, Guangzhou (China); Chen Mo; Li Wenfei; Yin Wenjing [State Key Laboratory of Oncology in South China, Department of Radiation Oncology, Sun Yat-sen University Cancer Center, Guangzhou (China); Lin Aihua [Department of Medical Statistics and Epidemiology, School of Public Health, Sun Yat-sen University, Guangzhou (China); Sun Ying [State Key Laboratory of Oncology in South China, Department of Radiation Oncology, Sun Yat-sen University Cancer Center, Guangzhou (China); Li Li [State Key Laboratory of Oncology in South China, Imaging Diagnosis and Interventional Center, Sun Yat-sen University Cancer Center, Guangzhou (China); Ma Jun, E-mail: majun2@mail.sysu.edu.cn [State Key Laboratory of Oncology in South China, Department of Radiation Oncology, Sun Yat-sen University Cancer Center, Guangzhou (China)

    2012-09-01

    Purpose: To subclassify patients with the T4 classification nasopharyngeal carcinoma (NPC), according to the seventh edition of the American Joint Committee on Cancer staging system, using magnetic resonance imaging (MRI), and to evaluate the prognostic value of subclassification after intensity-modulated radiotherapy (IMRT). Methods and Materials: A total of 140 patients who underwent MRI and were subsequently histologically diagnosed with nondisseminated classification T4 NPC received IMRT as their primary treatment and were included in this retrospective study. T4 patients were subclassified into two grades: T4a was defined as a primary nasopharyngeal tumor with involvement of the masticator space only; and T4b was defined as involvement of the intracranial region, cranial nerves, and/or orbit. Results: The 5-year overall survival (OS) rate and distant metastasis-free survival (DMFS) rate for T4a patients (82.5% and 87.0%, respectively), were significantly higher than for T4b patients (62.6% and 66.8%; p = 0.033 and p = 0.036, respectively). The T4a/b subclassification was an independent prognostic factor for OS (hazard ratio = 2.331, p = 0.032) and DMFS (hazard ratio = 2.602, p = 0.034), and had no significant effect on local relapse-free survival. Conclusions: Subclassification of T4 patients, as T4a or T4b, using MRI according to the site of invasion, has prognostic value for the outcomes of IMRT treatment in NPC.

  13. 2012 ETA Guidelines: The Use of L-T4 + L-T3 in the Treatment of Hypothyroidism

    DEFF Research Database (Denmark)

    Nygaard, Birte

    2012-01-01

    Background: Data suggest symptoms of hypothyroidism persist in 5–10% of levothyroxine (L-T4)-treated hypothyroid patients with normal serum thyrotrophin (TSH). The use of L-T4 + liothyronine (L-T3) combination therapy in such patients is controversial. The ETA nominated a task force to review....... There is insufficient evidence that L-T4 + L-T3 combination therapy is better than L-T4 monotherapy, and it is recommended that L-T4 monotherapy remains the standard treatment of hypothyroidism. L-T4 + L-T3 combination therapy might be considered as an experimental approach in compliant L-T4-treated hypothyroid...

  14. Interaction of anthraquinone dyes with lysozyme: Evidences from spectroscopic and docking studies

    Energy Technology Data Exchange (ETDEWEB)

    Paramaguru, G.; Kathiravan, A.; Selvaraj, S.; Venuvanalingam, P. [School of Chemistry, Bharathidasan University, Tiruchirappalli 620024, Tamil Nadu (India); Renganathan, R., E-mail: rrengas@gmail.com [School of Chemistry, Bharathidasan University, Tiruchirappalli 620024, Tamil Nadu (India)

    2010-03-15

    The interaction between lysozyme and anthraquinone dyes such as Alizarin Red S, Acid blue 129 and Uniblue was studied using steady state, time resolved fluorescence measurements and docking studies. Addition of anthraquinone dyes effectively quenched the intrinsic fluorescence of lysozyme. Fluorescence quenching of lysozyme by dyes has revealed the formation of complex. The number of binding sites (n) and binding constant (K) for all the three dyes was calculated by relevant fluorescence quenching data. Based on Foerster's non-radiative energy transfer theory, distance (r{sub 0}) between the donor (lysozyme) and acceptor (dyes) as well as the critical energy transfer distance (R{sub 0}) has also been calculated. The interaction between dyes and lysozyme occurs through static quenching mechanism as confirmed by time resolved spectroscopy. The conformational change of lysozyme has been analyzed using synchronous fluorescence measurement. Finally, docking studies revealed that specific interactions were observed with the residue of Trp 62.

  15. Interaction of anthraquinone dyes with lysozyme: Evidences from spectroscopic and docking studies

    International Nuclear Information System (INIS)

    Paramaguru, G.; Kathiravan, A.; Selvaraj, S.; Venuvanalingam, P.; Renganathan, R.

    2010-01-01

    The interaction between lysozyme and anthraquinone dyes such as Alizarin Red S, Acid blue 129 and Uniblue was studied using steady state, time resolved fluorescence measurements and docking studies. Addition of anthraquinone dyes effectively quenched the intrinsic fluorescence of lysozyme. Fluorescence quenching of lysozyme by dyes has revealed the formation of complex. The number of binding sites (n) and binding constant (K) for all the three dyes was calculated by relevant fluorescence quenching data. Based on Foerster's non-radiative energy transfer theory, distance (r 0 ) between the donor (lysozyme) and acceptor (dyes) as well as the critical energy transfer distance (R 0 ) has also been calculated. The interaction between dyes and lysozyme occurs through static quenching mechanism as confirmed by time resolved spectroscopy. The conformational change of lysozyme has been analyzed using synchronous fluorescence measurement. Finally, docking studies revealed that specific interactions were observed with the residue of Trp 62.

  16. [Survival and prognostic factors in resected satellite-nodule T4 non-small cell lung cancer].

    Science.gov (United States)

    Ma, Kai; Wang, Tian-you; He, Bao-liang; Chang, Dong; Hu, Xiao-dan; Yin, Zhi-yi; Jiang, Hua; Cui, Yong; Gao, Zhi; Gong, Min

    2009-01-15

    To study the survival and prognostic implication in surgically resected satellite-nodule T4 (T4 satellite) non-small cell lung cancer (NSCLC). From January 1995 to March 2005, the complete resection was performed to 42 patients with NSCLC who were postoperatively identified as pathologic-stage T4 satellite. Survival and associations between clinicopathological parameters and prognosis were analyzed. Thirty-two patients with pathologic stage local-invasion T4 (T4 invasion) NSCLC who underwent resection at the same time were also analyzed. The 1-, 3- and 5-year survival was 76.2%, 57.1% and 46.0% for patients with T4 satellite, while 62.3%, 31.5% and 20.0% for patients with T4 invasion. There was a significant higher survival in T4 satellite group when compared to that in T4 invasion group (P satellite N0M0 got a better survival than those with T4 satellite N1-2M0, T4 invasion N0M0 and T4 invasion N1 -2M0 (P satellite, univariate analysis showed that histology, main tumor size, lymph node status and adjuvant chemotherapy were linked with survival, while main tumor size, lymph node status and adjuvant chemotherapy served as the independent prognostic factors with multivariate analysis. Patients with completely resected T4 satellite NSCLC have a better prognosis than those with T4 invasion. Main tumor size over 3 cm, lymph node metastasis or no adjuvant chemotherapy means an unfavorable prognosis.

  17. [Characteristics of thermal inactivation of lysozyme in solution].

    Science.gov (United States)

    Tarun, E I; Eremin, A N; Metelitsa, D I

    1986-01-01

    In the buffer solution (pH 6,2) at 20-80 degrees, the lysozyme thermoinactivation was studied by monitoring of its activity decrease in the lysis of M. lysodeicticus cells. Protein inactivation was characterized by effective pseudofirst order rate constants which depend on enzyme concentration and are described by equation k = k0 . exp [-alpha 0 (1-gamma/T) [E]0], where k0 is inactivation rate constant at "infinite" enzyme dilution, [E0] is an initial lysozyme concentration, alpha 0 and gamma are the coefficients independent on [E0]. By extrapolation of the "k" dependencies on [E]0 the constants k0 were determined. In the range 40-70 degrees C, the rate constant k0 is equal 4,0 X 10(11) . exp (-24 200/RT) sec-1.

  18. A Lysozyme with Antifungal Activity from Pithecellobium dulce Seeds

    Directory of Open Access Journals (Sweden)

    Ploypat Niyomploy

    2011-01-01

    Full Text Available A protein of an apparent molecular mass of 14.4 kDa with antifungal activity was isolated from the seeds of Pithecellobium dulce using extraction with 100 mM Tris-HCl buffer (pH=8.0, precipitation with 80 % ammonium sulfate, and bioassay purification via Resource Q anion exchange chromatography and Superdex 200 gel filtration chromatography. The purified protein was putatively identified by tandem mass spectrometry with Mascot database searching, with the partial amino acid sequences showing a high degree of similarity to chicken egg white lysozyme. This putative plant lysozyme expressed antifungal activity with a rather high thermal stability of up to 80 °C for 15 min (at pH=8.0. It exerted an antifungal action towards Macrophomina phaseolina but displayed no antifungal activity against two other isolates, Phymatotrichopsis omnivora and Fusarium avenaceum.

  19. Protein crystal growth - Growth kinetics for tetragonal lysozyme crystals

    Science.gov (United States)

    Pusey, M. L.; Snyder, R. S.; Naumann, R.

    1986-01-01

    Results are reported from theoretical and experimental studies of the growth rate of lysozyme as a function of diffusion in earth-gravity conditions. The investigations were carried out to form a comparison database for future studies of protein crystal growth in the microgravity environment of space. A diffusion-convection model is presented for predicting crystal growth rates in the presence of solutal concentration gradients. Techniques used to grow and monitor the growth of hen egg white lysozyme are detailed. The model calculations and experiment data are employed to discuss the effects of transport and interfacial kinetics in the growth of the crystals, which gradually diminished the free energy in the growth solution. Density gradient-driven convection, caused by presence of the gravity field, was a limiting factor in the growth rate.

  20. Tetragonal Chicken Egg White Lysozyme Solubility in Sodium Chloride Solutions

    Science.gov (United States)

    Forsythe, Elizabeth L.; Judge, Russell A.; Pusey, Marc L.

    1998-01-01

    The solubility of chicken egg white lysozyme, crystallized in the tetragonal form was measured in sodium chloride solutions from 1.6 to 30.7 C, using a miniature column solubility apparatus. Sodium chloride solution concentrations ranged from 1 to 7% (w/v). The solutions were buffered with 0.1 M sodium acetate buffer with the solubility being measured at pH values in 0.2 pH unit increments in the range pH 4.0 to 5.4, with data also included at pH 4.5. Lysozyme solubility was found to increase with increases in temperature and decreasing salt concentration. Solution pH has a varied and unpredictable effect on solubility.

  1. The solubility of hen egg-white lysozyme

    Science.gov (United States)

    Howard, Sandra B.; Twigg, Pamela J.; Baird, James K.; Meehan, Edward J.

    1988-07-01

    The equilibrium solubility of chicken egg-white lysozyme in the presence of crystalline solid state was determined as a function of NaCl concentration, pH, and temperature. The solubility curves obtained represent a region of the lysozyme phase diagram. This diagram makes it possible to determine the supersaturation of a given set of conditions or to achieve identical supersaturations by different combinations of parameters. The temperature dependence of the solubility permits the evaluation of ΔH of crystallization and we present a method for its calculation. The data indicates a negative heat of crystallization for the tetragonal crystal form but a positive heat of crystallization for the high temperature orthorhombic form.

  2. Analysis of two lysozyme genes and antimicrobial functions of their recombinant proteins in Asian seabass.

    Directory of Open Access Journals (Sweden)

    Gui Hong Fu

    Full Text Available Lysozymes are important proteins of the innate immune system for the defense against bacterial infection. We cloned and analyzed chicken-type (c-type and goose-type (g-type lysozymes from Asian seabass (Lates calcarifer. The deduced amino acid sequence of the c-type lysozyme contained 144 residues and possessed typical structure residues, conserved catalytic residues (Glu(50 and Asp(67 and a "GSTDYGIFQINS" motif. The deduced g-type lysozyme contained 187 residues and possessed a goose egg white lysozyme (GEWL domain containing three conserved catalytic residues (Glu(71, Asp(84, Asp(95 essential for catalytic activity. Real time quantitative PCR (qRT-PCR revealed that the two lysozyme genes were constitutively expressed in all the examined tissues. The c-type lysozyme was most abundant in liver, while the g-type lysozyme was predominantly expressed in intestine and weakly expressed in muscle. The c-type and g-type transcripts were up-regulated in the kidney, spleen and liver in response to a challenge with Vibrio harveyi. The up-regulation of the c-type lysozyme was much stronger than that of the g-type lysozyme in kidney and spleen. The recombinant proteins of the c-type and g-type lysozymes showed lytic activities against the bacterial pathogens Vibrio harveyi and Photobacterium damselae in a dosage-dependent manner. We identified single nucleotide polymorphisms (SNPs in the two lysozyme genes. There were significant associations of these polymorphisms with resistance to the big belly disease. These results suggest that the c- and g-type genes play an important role in resistance to bacterial pathogens in fish. The SNP markers in the two genes associated with the resistance to bacterial pathogens may facilitate the selection of Asian seabass resistant to bacterial diseases.

  3. Lysozyme amyloidosis – a case report and review of the literature

    OpenAIRE

    Pleyer, Christopher; Flesche, Jan; Saeed, Fahad

    2015-01-01

    Lysozyme amyloidosis is an exceedingly rare hereditary autosomal dominant amyloidosis, which is characterized by the precipitation of lysozyme protein within the body, leading to multi-organ dysfunction. Herein, we present the case of a U.S. family affected by lysozyme amyloidosis. In particular, we report pericardial disease involvement leading to recurrent pericardial effusion, which to our knowledge has not been described yet. To our knowledge, we have also for the first time identified th...

  4. Analysis of two lysozyme genes and antimicrobial functions of their recombinant proteins in Asian seabass.

    Science.gov (United States)

    Fu, Gui Hong; Bai, Zhi Yi; Xia, Jun Hong; Liu, Feng; Liu, Peng; Yue, Gen Hua

    2013-01-01

    Lysozymes are important proteins of the innate immune system for the defense against bacterial infection. We cloned and analyzed chicken-type (c-type) and goose-type (g-type) lysozymes from Asian seabass (Lates calcarifer). The deduced amino acid sequence of the c-type lysozyme contained 144 residues and possessed typical structure residues, conserved catalytic residues (Glu(50) and Asp(67)) and a "GSTDYGIFQINS" motif. The deduced g-type lysozyme contained 187 residues and possessed a goose egg white lysozyme (GEWL) domain containing three conserved catalytic residues (Glu(71), Asp(84), Asp(95)) essential for catalytic activity. Real time quantitative PCR (qRT-PCR) revealed that the two lysozyme genes were constitutively expressed in all the examined tissues. The c-type lysozyme was most abundant in liver, while the g-type lysozyme was predominantly expressed in intestine and weakly expressed in muscle. The c-type and g-type transcripts were up-regulated in the kidney, spleen and liver in response to a challenge with Vibrio harveyi. The up-regulation of the c-type lysozyme was much stronger than that of the g-type lysozyme in kidney and spleen. The recombinant proteins of the c-type and g-type lysozymes showed lytic activities against the bacterial pathogens Vibrio harveyi and Photobacterium damselae in a dosage-dependent manner. We identified single nucleotide polymorphisms (SNPs) in the two lysozyme genes. There were significant associations of these polymorphisms with resistance to the big belly disease. These results suggest that the c- and g-type genes play an important role in resistance to bacterial pathogens in fish. The SNP markers in the two genes associated with the resistance to bacterial pathogens may facilitate the selection of Asian seabass resistant to bacterial diseases.

  5. Analysis of Two Lysozyme Genes and Antimicrobial Functions of Their Recombinant Proteins in Asian Seabass

    Science.gov (United States)

    Fu, Gui Hong; Bai, Zhi Yi; Xia, Jun Hong; Liu, Feng; Liu, Peng; Yue, Gen Hua

    2013-01-01

    Lysozymes are important proteins of the innate immune system for the defense against bacterial infection. We cloned and analyzed chicken-type (c-type) and goose-type (g-type) lysozymes from Asian seabass (Lates calcarifer). The deduced amino acid sequence of the c-type lysozyme contained 144 residues and possessed typical structure residues, conserved catalytic residues (Glu50 and Asp67) and a “GSTDYGIFQINS” motif. The deduced g-type lysozyme contained 187 residues and possessed a goose egg white lysozyme (GEWL) domain containing three conserved catalytic residues (Glu71, Asp84, Asp95) essential for catalytic activity. Real time quantitative PCR (qRT-PCR) revealed that the two lysozyme genes were constitutively expressed in all the examined tissues. The c-type lysozyme was most abundant in liver, while the g-type lysozyme was predominantly expressed in intestine and weakly expressed in muscle. The c-type and g-type transcripts were up-regulated in the kidney, spleen and liver in response to a challenge with Vibrio harveyi. The up-regulation of the c-type lysozyme was much stronger than that of the g-type lysozyme in kidney and spleen. The recombinant proteins of the c-type and g-type lysozymes showed lytic activities against the bacterial pathogens Vibrio harveyi and Photobacterium damselae in a dosage-dependent manner. We identified single nucleotide polymorphisms (SNPs) in the two lysozyme genes. There were significant associations of these polymorphisms with resistance to the big belly disease. These results suggest that the c- and g-type genes play an important role in resistance to bacterial pathogens in fish. The SNP markers in the two genes associated with the resistance to bacterial pathogens may facilitate the selection of Asian seabass resistant to bacterial diseases. PMID:24244553

  6. Structural Basis of Protein Oxidation Resistance: A Lysozyme Study

    OpenAIRE

    Girod, Marion; Enjalbert, Quentin; Brunet, Claire; Antoine, Rodolphe; Lemoine, Jérôme; Lukac, Iva; Radman, Miroslav; Krisko, Anita; Dugourd, Philippe

    2014-01-01

    Accumulation of oxidative damage in proteins correlates with aging since it can cause irreversible and progressive degeneration of almost all cellular functions. Apparently, native protein structures have evolved intrinsic resistance to oxidation since perfectly folded proteins are, by large most robust. Here we explore the structural basis of protein resistance to radiation-induced oxidation using chicken egg white lysozyme in the native and misfolded form. We study the differential resistan...

  7. Lysozyme as diffusion tracer for measuring aqueous solution viscosity.

    Science.gov (United States)

    Parmar, Avanish S; Muschol, Martin

    2009-11-01

    Measuring tracer diffusion provides a convenient approach for monitoring local changes in solution viscosity or for determining viscosity changes in response to multiple solution parameters including pH, temperature, salt concentrations or salt types. One common limitation of tracer diffusion in biologically relevant saline solutions is the loss of colloidal stability and aggregation of the tracer particles with increasing ionic strength. Using dynamic light scattering to measure tracer diffusion, we compared the performance of two different types of tracer particles, polystyrene nanobeads vs. the small protein lysozyme, for viscosity measurements of saline solutions. Polystyrene beads provide reliable values for water viscosity, but begin flocculating at ionic strengths exceeding about 100mM. Using lysozyme, in contrast, we could map out viscosity changes of saline solutions for a variety of different salts, for salt concentrations up to 1M, over a wide range of pH values, and over the temperature range most relevant for biological systems (5-40 degrees C). Due to its inherently high structural and colloidal stability, lysozyme provides a convenient and reliable tracer particle for all these measurements, and its use can be readily extended to other optical approaches towards localized measurements of tracer diffusion such as fluorescence correlation spectroscopy.

  8. Influence of surface charge on lysozyme adsorption to ceria nanoparticles

    International Nuclear Information System (INIS)

    Wang Binghui; Wu Peng; Yokel, Robert A.; Grulke, Eric A.

    2012-01-01

    Understanding mechanisms for forming protein coronas on nanomaterial surfaces is essential to designing drug delivery systems and designing and interpreting the results of nanomaterial toxicity tests. The study reports the adsorption behavior of a positively charged protein, lysozyme, on cerium dioxide (ceria) nanoparticles with three different surface charges. Adsorption isotherms were modeled with the Toth and Sips equations. Isotherm loading levels were compared to monolayer coverage estimate for ‘side-on’ and ‘end-on’ lysozyme orientations as well as random packing (jamming) and maximum packing limits. Evaluation of adsorption site energy distributions (generated using the model coefficients) suggested that the negatively charged ceria surface had a very broad site energy distribution and that its surface heterogeneity controls the adsorption process. By contrast, the adsorption of lysozyme on the positively charged nanoparticles appears to be influenced by lateral effects from adsorbed protein species. The results illustrate the importance of nanoparticle surface chemistry to protein adsorption. The modeling and site energy distribution evaluations may be useful for interpreting the formation of protein coronas on nanoparticles.

  9. Structural basis of protein oxidation resistance: a lysozyme study.

    Science.gov (United States)

    Girod, Marion; Enjalbert, Quentin; Brunet, Claire; Antoine, Rodolphe; Lemoine, Jérôme; Lukac, Iva; Radman, Miroslav; Krisko, Anita; Dugourd, Philippe

    2014-01-01

    Accumulation of oxidative damage in proteins correlates with aging since it can cause irreversible and progressive degeneration of almost all cellular functions. Apparently, native protein structures have evolved intrinsic resistance to oxidation since perfectly folded proteins are, by large most robust. Here we explore the structural basis of protein resistance to radiation-induced oxidation using chicken egg white lysozyme in the native and misfolded form. We study the differential resistance to oxidative damage of six different parts of native and misfolded lysozyme by a targeted tandem/mass spectrometry approach of its tryptic fragments. The decay of the amount of each lysozyme fragment with increasing radiation dose is found to be a two steps process, characterized by a double exponential evolution of their amounts: the first one can be largely attributed to oxidation of specific amino acids, while the second one corresponds to further degradation of the protein. By correlating these results to the structural parameters computed from molecular dynamics (MD) simulations, we find the protein parts with increased root-mean-square deviation (RMSD) to be more susceptible to modifications. In addition, involvement of amino acid side-chains in hydrogen bonds has a protective effect against oxidation Increased exposure to solvent of individual amino acid side chains correlates with high susceptibility to oxidative and other modifications like side chain fragmentation. Generally, while none of the structural parameters alone can account for the fate of peptides during radiation, together they provide an insight into the relationship between protein structure and susceptibility to oxidation.

  10. Comparative insight into surfactants mediated amyloidogenesis of lysozyme.

    Science.gov (United States)

    Chaturvedi, Sumit K; Khan, Javed M; Siddiqi, Mohammad K; Alam, Parvez; Khan, Rizwan H

    2016-02-01

    Electrostatic and hydrophobic interactions have an important role in the protein aggregation. In this study, we have investigated the effect of charge and hydrophobicity of oppositely charged surfactants i.e., anionic (AOT and SDS) and cationic (CTAB and DTAB) on hen egg white lysozyme at pH 9.0 and 13.0, respectively. We have employed various methods such as turbidity measurements, Rayleigh light scattering, ThT, Congo red and ANS dye binding assays, far-UV CD, atomic force microscopy, transmission electron and fluorescence microscopy. At lower molar ratio, both anionic and cationic surfactants promote amyloid fibril formation in lysozyme at pH 9.0 and 13.0, respectively. The aggregation was proportionally increased with respect to protein concentration and hydrophobicity of surfactant. The morphology of aggregates at both the pH was fibrillar in structure, as visualized by dye binding and microscopic imaging techniques. Initially, the interaction between surfactants and lysozyme was electrostatic and then hydrophobic as investigated by ITC. This study demonstrates the crucial role of charge and hydrophobicity during amyloid fibril formation. Copyright © 2015 Elsevier B.V. All rights reserved.

  11. Antimicrobial peptide lysozyme has the potential to promote mouse hair follicle growth in vitro.

    Science.gov (United States)

    Su, Yongsheng; Liu, Hui; Wang, Jin; Lin, Bojie; Miao, Yong; Hu, Zhiqi

    2015-10-01

    Lysozyme is a well-known antimicrobial peptide that exists widely in mammalian skin and it is also expressed by pilosebaceous units. However, the exact location of lysozyme in hair follicles and whether it exerts any direct effects on hair follicle growth are unclear. To determine whether lysozyme affected hair growth in vitro, micro-dissected mouse vibrissae follicles (VFs) were treated in serum-free organ culture for 3 days with lysozyme (1-10μg/ml). After that, the effects of lysozyme on dermal papilla (DP) cells were also investigated. Lysozyme was mainly identified in DP and dermal sheath regions of VF by immunochemistry. In addition, 5-10μg/ml lysozyme had a promoting effect on shaft production. It was also associated with significant proliferation of matrix keratinocytes by immunofluorescence observation. Furthermore, lysozyme promoted hair growth by increasing the levels of alkaline phosphatase and lymphoid enhancer factor 1 in DP, as determined by Western blotting. These results indicate that lysozyme is a promoter of VF growth via enhancing the hair-inductive capacity of DP cells during organ culture. Copyright © 2015 Elsevier GmbH. All rights reserved.

  12. Parallel tempering Monte Carlo simulations of lysozyme orientation on charged surfaces

    Science.gov (United States)

    Xie, Yun; Zhou, Jian; Jiang, Shaoyi

    2010-02-01

    In this work, the parallel tempering Monte Carlo (PTMC) algorithm is applied to accurately and efficiently identify the global-minimum-energy orientation of a protein adsorbed on a surface in a single simulation. When applying the PTMC method to simulate lysozyme orientation on charged surfaces, it is found that lysozyme could easily be adsorbed on negatively charged surfaces with "side-on" and "back-on" orientations. When driven by dominant electrostatic interactions, lysozyme tends to be adsorbed on negatively charged surfaces with the side-on orientation for which the active site of lysozyme faces sideways. The side-on orientation agrees well with the experimental results where the adsorbed orientation of lysozyme is determined by electrostatic interactions. As the contribution from van der Waals interactions gradually dominates, the back-on orientation becomes the preferred one. For this orientation, the active site of lysozyme faces outward, which conforms to the experimental results where the orientation of adsorbed lysozyme is co-determined by electrostatic interactions and van der Waals interactions. It is also found that despite of its net positive charge, lysozyme could be adsorbed on positively charged surfaces with both "end-on" and back-on orientations owing to the nonuniform charge distribution over lysozyme surface and the screening effect from ions in solution. The PTMC simulation method provides a way to determine the preferred orientation of proteins on surfaces for biosensor and biomaterial applications.

  13. Engineering Escherichia coli for Soluble Expression and Single Step Purification of Active Human Lysozyme

    Science.gov (United States)

    Lamppa, John W.; Tanyos, Sam A.; Griswold, Karl E.

    2012-01-01

    Genetically engineered variants of human lysozyme represent promising leads in the battle against drug-resistant bacterial pathogens, but early stage development and testing of novel lysozyme variants is constrained by the lack of a robust, scalable and facile expression system. While wild type human lysozyme is reportedly produced at 50 – 80 kg per hectare of land in recombinant rice, this plant-based system is not readily scaled down to bench top production, and it is therefore not suitable for development and characterization of novel lysozyme variants. Here, we describe a novel and efficient expression system capable of producing folded, soluble and functional human lysozyme in E. coli cells. To achieve this goal, we simultaneously co-express multiple protein folding chaperones as well as harness the lysozyme inhibitory protein, Ivy. Our strategy exploits E. coli’s ease of culture, short doubling time, and facile genetics to yield upwards of 30 mg/L of soluble lysozyme in a bioreactor system, a 3000-fold improvement over prior efforts in E. coli. Additionally, molecular interactions between lysozyme and a his-tagged Ivy allows for one-step purification by IMAC chromatography, yielding as much as 21 mg/L of purified enzyme. We anticipate that our expression and purification platform will facilitate further development of engineered lysozymes having utility in disease treatment and other practical applications. PMID:23220215

  14. The interaction of lysozyme with caffeine, theophylline and theobromine in solution.

    Science.gov (United States)

    Zhang, Hong-Mei; Tang, Bo-Ping; Wang, Yan-Qing

    2010-10-01

    The interactions of lysozyme with caffeine (Caf), theophylline (Tph) and theobromine (Tbr) were investigated using UV-Vis absorption, fluorescence, synchronous fluorescence, and three-dimensional fluorescence spectra techniques. The results revealed that Caf (Tph or Tbr) caused the fluorescence quenching of lysozyme by the formation of Caf (Tph or Tbr)-lysozyme complex. The binding constants (K(A)) and thermodynamic parameters (ΔG°, ΔH°, ΔS°) at two different temperatures, the binding locality, and the binding power were obtained. The results showed that the process of binding Caf (Tph or Tbr) to lysozyme was a spontaneous molecular interaction procedure and the hydrophobic and electrostatic interactions play a major role in stabilizing the complex; The distance r between donor (lysozyme) and acceptor (Caf, Tph or Tbr) was obtained according to fluorescence resonance energy transfer. The effect of Caf (Tph or Tbr) on the conformation of lysozyme was analyzed using synchronous fluorescence and three-dimensional fluorescence spectra techniques. The results showed that the binding of Caf (Tph or Tbr) to lysozyme induced some micro-environmental and conformational changes in lysozyme and disturbed the environment of the polypeptide of lysozyme.

  15. Protective properties of lysozyme on β-amyloid pathology: implications for Alzheimer disease.

    Science.gov (United States)

    Helmfors, Linda; Boman, Andrea; Civitelli, Livia; Nath, Sangeeta; Sandin, Linnea; Janefjord, Camilla; McCann, Heather; Zetterberg, Henrik; Blennow, Kaj; Halliday, Glenda; Brorsson, Ann-Christin; Kågedal, Katarina

    2015-11-01

    The hallmarks of Alzheimer disease are amyloid-β plaques and neurofibrillary tangles accompanied by signs of neuroinflammation. Lysozyme is a major player in the innate immune system and has recently been shown to prevent the aggregation of amyloid-β1-40 in vitro. In this study we found that patients with Alzheimer disease have increased lysozyme levels in the cerebrospinal fluid and lysozyme co-localized with amyloid-β in plaques. In Drosophila neuronal co-expression of lysozyme and amyloid-β1-42 reduced the formation of soluble and insoluble amyloid-β species, prolonged survival and improved the activity of amyloid-β1-42 transgenic flies. This suggests that lysozyme levels rise in Alzheimer disease as a compensatory response to amyloid-β increases and aggregation. In support of this, in vitro aggregation assays revealed that lysozyme associates with amyloid-β1-42 and alters its aggregation pathway to counteract the formation of toxic amyloid-β species. Overall, these studies establish a protective role for lysozyme against amyloid-β associated toxicities and identify increased lysozyme in patients with Alzheimer disease. Therefore, lysozyme has potential as a new biomarker as well as a therapeutic target for Alzheimer disease. Copyright © 2015. Published by Elsevier Inc.

  16. Characterization of expression, activity and role in antibacterial immunity of Anopheles gambiae lysozyme c-1

    OpenAIRE

    Kajla, Mayur K.; Andreeva, Olga; Gilbreath, Thomas M.; Paskewitz, Susan M.

    2010-01-01

    There are eight lysozyme genes in the Anopheles gambiae genome. Transcripts of one of these genes, LYSC-1, increased in Anopheles gambiae cell line 4a3B by 24 h after exposure to heat-killed Micrococcus luteus. Lysozyme activity was also identified in conditioned media from the cell line from which the protein was purified to homogeneity using ion exchange and gel filtration. Mass spectrometric analysis of the purified protein showed 100% identity to lysozyme c-1. Purified lysozyme c-1 was te...

  17. Engineering Escherichia coli for soluble expression and single step purification of active human lysozyme.

    Science.gov (United States)

    Lamppa, John W; Tanyos, Sam A; Griswold, Karl E

    2013-03-10

    Genetically engineered variants of human lysozyme represent promising leads in the battle against drug-resistant bacterial pathogens, but early stage development and testing of novel lysozyme variants is constrained by the lack of a robust, scalable and facile expression system. While wild type human lysozyme is reportedly produced at 50–80 kg per hectare of land in recombinant rice, this plant-based system is not readily scaled down to bench top production, and it is therefore not suitable for development and characterization of novel lysozyme variants. Here, we describe a novel and efficient expression system capable of producing folded, soluble and functional human lysozyme in Escherichia coli cells. To achieve this goal, we simultaneously co-express multiple protein folding chaperones as well as harness the lysozyme inhibitory protein, Ivy. Our strategy exploits E. coli's ease of culture, short doubling time, and facile genetics to yield upwards of 30 mg/l of soluble lysozyme in a bioreactor system, a 3000-fold improvement over prior efforts in E. coli. Additionally, molecular interactions between lysozyme and a his-tagged Ivy allows for one-step purification by IMAC, yielding as much as 21 mg/l of purified enzyme. We anticipate that our expression and purification platform will facilitate further development of engineered lysozymes having utility in disease treatment and other practical applications. Copyright © 2012 Elsevier B.V. All rights reserved.

  18. Chronopotentiometric sensing of specific interactions between lysozyme and the DNA aptamer.

    Science.gov (United States)

    Ostatná, Veronika; Kasalová-Vargová, Veronika; Kékedy-Nagy, László; Černocká, Hana; Ferapontova, Elena E

    2017-04-01

    Specific DNA-protein interactions are vital for cellular life maintenance processes, such as transcriptional regulation, chromosome maintenance, replication and DNA repair, and their monitoring gives valuable information on molecular-level organization of those processes. Here, we propose a new method of label-free electrochemical sensing of sequence specific binding between the lysozyme protein and a single stranded DNA aptamer specific for lysozyme (DNA apta ) that exploits the constant current chronopotentiometric stripping (CPS) analysis at modified mercury electrodes. Specific lysozyme-DNA apta binding was distinguished from nonspecific lysozyme-DNA interactions at thioglycolic acid-modified mercury electrodes, but not at the dithiothreitol-modified or bare mercury electrodes. Stability of the surface-attached lysozyme-DNA apta layer depended on the stripping current (I str ) intensity, suggesting that the integrity of the layer critically depends on the time of its exposure to negative potentials. Stabilities of different lysozyme-DNA complexes at the negatively polarized electrode surface were tested, and it was shown that structural transitions of the specific lysozyme-DNA apta complexes occur in the I str ranges different from those observed for assemblies of lysozyme with DNA sequences capable of only nonspecific lysozyme-DNA interactions. Thus, the CPS allows distinct discrimination between specific and non-specific protein-DNA binding and provides valuable information on stability of the nucleic acid-protein interactions at the polarized interfaces. Copyright © 2016 Elsevier B.V. All rights reserved.

  19. [Highly active fractions of the medicinal leech recombinant destabilase-lysozyme].

    Science.gov (United States)

    Fadeeva, Iu I; Antipova, N V; Baskova, I P; Zavalova, L L

    2014-01-01

    From the highly purified but lowly active recombinant protein Destabilas-Lysozyme (Dest-Lys) by use cation-exchange column TSK CM 3-SW chromatography, it was separated non-active fraction IV, contained 90% of protein. Fractions I, II and III, represented proteins with lysozyme and isopeptidase activities. Their lysozyme activity correlates with the activity of natural Des-Lys. The ratio of the activities in fractions I - III is such, that maximal lysozyme activity is concentrated in fraction III, isopeptidase - in fraction I. It is discussed the possibility of Dest-Lys different functions regulation is depended on the formation of protein complex forms.

  20. O-specific polysaccharide confers lysozyme resistance to extraintestinal pathogenic Escherichia coli.

    Science.gov (United States)

    Bao, Yinli; Zhang, Haobo; Huang, Xinxin; Ma, Jiale; Logue, Catherine M; Nolan, Lisa K; Li, Ganwu

    2018-12-31

    Extraintestinal pathogenic Escherichia coli (ExPEC) is the leading cause of bloodstream and other extraintestinal infections in human and animals. The greatest challenge encountered by ExPEC during an infection is posed by the host defense mechanisms, including lysozyme. ExPEC have developed diverse strategies to overcome this challenge. The aim of this study was to characterize the molecular mechanism of ExPEC resistance to lysozyme. For this, 15,000 transposon mutants of a lysozyme-resistant ExPEC strain NMEC38 were screened; 20 genes were identified as involved in ExPEC resistance to lysozyme-of which five were located in the gene cluster between galF and gnd, and were further confirmed to be involved in O-specific polysaccharide biosynthesis. The O-specific polysaccharide was able to inhibit the hydrolytic activity of lysozyme; it was also required by the complete lipopolysaccharide (LPS)-mediated protection of ExPEC against the bactericidal activity of lysozyme. The O-specific polysaccharide was further shown to be able to directly interact with lysozyme. Furthermore, LPS from ExPEC strains of different O serotypes was also able to inhibit the hydrolytic activity of lysozyme. Because of their cell surface localization and wide distribution in Gram-negative bacteria, O-specific polysaccharides appear to play a long-overlooked role in protecting bacteria against exogenous lysozyme.

  1. The interaction of the protein lysozyme with bacteria E. coli observed using nanodiamond labelling

    Energy Technology Data Exchange (ETDEWEB)

    Perevedentseva, Elena; Cheng, C-Y; Chung, P-H; Tu, J-S; Hsieh, Y-H; Cheng, C-L [Department of Physics, National Dong Hwa University, Hualien 97401, Taiwan (China)

    2007-08-08

    The application of a nanometre-sized diamond in Raman-detectable biolabelling is demonstrated in this study. The interaction of a lysozyme-nanodiamond complex with bacteria E. coli was observed via Raman mapping using the diamond Raman signal as the labelling marker. The results are compared with scanning electron microscope observations, and the adsorbed lysozyme's functionality is analysed. High antibacterial activity of lysozyme-nanodiamond complex was observed, equivalent to active lysozyme in solution. The results suggest that nanodiamond labelling can be effective and that it can be applied in ambient conditions without complicated sample pre-treatments.

  2. The interaction of the protein lysozyme with bacteria E. coli observed using nanodiamond labelling

    International Nuclear Information System (INIS)

    Perevedentseva, Elena; Cheng, C-Y; Chung, P-H; Tu, J-S; Hsieh, Y-H; Cheng, C-L

    2007-01-01

    The application of a nanometre-sized diamond in Raman-detectable biolabelling is demonstrated in this study. The interaction of a lysozyme-nanodiamond complex with bacteria E. coli was observed via Raman mapping using the diamond Raman signal as the labelling marker. The results are compared with scanning electron microscope observations, and the adsorbed lysozyme's functionality is analysed. High antibacterial activity of lysozyme-nanodiamond complex was observed, equivalent to active lysozyme in solution. The results suggest that nanodiamond labelling can be effective and that it can be applied in ambient conditions without complicated sample pre-treatments

  3. Serum TBG and T4 concentration in non-thyroidal diseases

    International Nuclear Information System (INIS)

    Sasaki, Y.; Tobari, C.; Sekita, N.; Onodera, Y.; Asazu, M.; Someya, K.

    1983-01-01

    Routinely available radioassay kits have recently enabled the measurement of serum concentrations of thyroxine binding globulin (TBG) and thyroxine (T 4 ), both total (TT 4 ) and free (FT 4 ) in various disease conditions. Serum TBG and T 4 level were measured in variety of non-thyroidal diseases, of which significance was evaluated in comparison with that in thyroidal diseases. Abnormal serum TBG concentrations in various non-thyroidal diseases and pregnancy result in abnormal serum TT 4 levels, which may cause difficulty in differentiation of these conditions from hyper- or hypothyroidal states. Serum FT 4 levels give better indicator than TT 4 , though the difference among RIA kits are considerably large. However, measurement of serum FT 4 levels alone is not sufficient to distinguish non-thyroidal disease from thyroidal diseases with abnormal thyroidal function. The differentiation has to be based on the combination of clinical findings and results of multiple thyroidal function tests

  4. Characterization of a bacteriophage T4 mutant lacking DNA-dependent ATPase

    International Nuclear Information System (INIS)

    Behme, M.T.; Ebisuzaki, K.

    1975-01-01

    A DNA-dependent ATPase has previously been purified from bacteriophage T4-infected Escherichia coli. A mutant phage strain lacking this enzyme has been isolated and characterized. Although the mutant strain produced no detectable DNA-dependent ATPase, growth properties were not affected. Burst sizes were similar for the mutant phage and T4D in polAl, recB, recC, uvrA, uvrB, uvrC, and various DNA-negative E. coli. UV sensitivity and genetic recombination were normal in a variety of E. coli hosts. Mapping data indicate that the genetic locus controlling the mutant occurs near gene 56. The nonessential nature of this gene is discussed

  5. Coupling DNA unwinding activity with primer synthesis in the bacteriophage T4 primosome

    Science.gov (United States)

    Manosas, Maria; Spiering, Michelle M.; Zhuang, Zhihao; Benkovic, Stephen J.; Croquette, Vincent

    2009-01-01

    The unwinding and priming activities of the bacteriophage T4 primosome, which consists of a hexameric helicase (gp41) translocating 5′ to 3′ and an oligomeric primase (gp61) synthesizing primers 5′ to 3′, has been investigated on DNA hairpins manipulated by a magnetic trap. We find that the T4 primosome continuously unwinds the DNA duplex while allowing for primer synthesis through a primosome disassembly mechanism or a novel DNA looping mechanism. A fused gp61-gp41 primosome unwinds and primes DNA exclusively via the DNA looping mechanism. Other proteins within the replisome control the partitioning of these two mechanisms disfavoring primosome disassembly thereby increasing primase processivity. In contrast priming in bacteriophage T7 involves discrete pausing of the primosome and in Escherichia coli appears to be associated primarily with dissociation of the primase from the helicase. Thus nature appears to use several strategies to couple the disparate helicase and primase activities within primosomes. PMID:19838204

  6. Lysozyme-immobilized electrospun PAMA/PVA and PSSA-MA/PVA ion-exchange nanofiber for wound healing.

    Science.gov (United States)

    Tonglairoum, Prasopchai; Ngawhirunpat, Tanasait; Rojanarata, Theerasak; Opanasopit, Praneet

    2014-08-27

    Abstract This research was aimed to develop the lysozyme immobilized ion-exchange nanofiber mats for wound healing. To promote the healing process, the PSSA-MA/PVA and PAMA ion-exchange nanofiber mats were fabricated to mimic the extracellular matrix structure using electrospinning process followed by thermally crosslinked. Lysozyme was immobilized on the ion-exchane nanofibers by an adsorption method. The ion-exchange nanofibers were investigated using SEM, FTIR and XRPD. Moreover, the lysozyme-immobilized ion-exchange nanofibers were further investigated for lysozyme content and activity, lysozyme release and wound healing activity. The fiber diameters of the mats were in the nanometer range. Lysozyme was gradually absorbed into the PSSA-MA/PVA nanofiber with higher extend than that is absorbed on the PAMA/PVA nanofiber and exhibited higher activity than lysozyme-immobilized PAMA/PVA nanofiber. The total contents of lysozyme on the PSSA-MA/PVA and PAMA/PVA nanofiber were 648 and 166 µg/g, respectively. FTIR and lysozyme activity results confirmed the presence of lysozyme on the nanofiber mats. The lysozyme was released from the PSSA-MA/PVA and PAMA/PVA nanofiber in the same manner. The lysozyme-immobilized PSSA-MA/PVA nanofiber mats and lysozyme-immobilized PAMA/PVA nanofiber mats exhibited significantly faster healing rate than gauze and similar to the commercial antibacterial gauze dressing. These results suggest that these nanofiber mats could provide the promising candidate for wound healing application.

  7. Pulsating strings from two-dimensional CFT on (T4N/S(N

    Directory of Open Access Journals (Sweden)

    Carlos Cardona

    2015-04-01

    Full Text Available We propose a state from the two-dimensional conformal field theory on the orbifold (T4N/S(N as a dual description for a pulsating string moving in AdS3. We show that, up to first order in the deforming parameter, the energy in both descriptions has the same dependence on the mode number, but with a non-trivial function of the coupling.

  8. Expression of the bacteriophage T4 denV structural gene in Escherichia coli

    International Nuclear Information System (INIS)

    Recinos, A. III; Augustine, M.L.; Higgins, K.M.; Lloyd, R.S.

    1986-01-01

    The expression of the T4 denV gene, which previously had been cloned in plasmid constructs downstream of the bacteriophage lambda hybrid promoter-operator oLpR, was analyzed under a variety of growth parameters. Expression of the denV gene product, endonuclease V, was confirmed in DNA repair-deficient Escherichia coli (uvrA recA) by Western blot analyses and by enhancements of resistance to UV irradiation

  9. T4 phage and its head surface proteins do not stimulate inflammatory mediator production.

    Directory of Open Access Journals (Sweden)

    Paulina Miernikiewicz

    Full Text Available Viruses are potent activators of the signal pathways leading to increased cytokine or ROS production. The effects exerted on the immune system are usually mediated by viral proteins. Complementary to the progress in phage therapy practice, advancement of knowledge about the influence of bacteriophages on mammalian immunity is necessary. Particularly, the potential ability of phage proteins to act like other viral stimulators of the immune system may have strong practical implications for the safety and efficacy of bacteriophage therapy. Here we present studies on the effect of T4 phage and its head proteins on production of inflammatory mediators and inflammation-related factors: IL-1α, IL-1β, IL-2, IL-6, IL-10, IL-12 p40/p70, IFN-γ, TNF-α, MCP-1, MIG, RANTES, GCSF, GM-CSF and reactive oxygen species (ROS. Plasma cytokine profiles in an in vivo mouse model and in human blood cells treated with gp23*, gp24*, Hoc and Soc were evaluated by cytokine antibody arrays. Cytokine production and expression of CD40, CD80, CD86 and MHC class II molecules were also investigated in mouse bone marrow-derived dendritic cells treated with whole T4 phage particle or the same capsid proteins. The influence of T4 and gp23*, gp24*, Hoc and Soc on reactive oxygen species generation was examined in blood cells using luminol-dependent chemiluminescence assay. In all performed assays, the T4 bacteriophage and its capsid proteins gp23*, gp24*, Hoc and Soc did not affect production of inflammatory-related cytokines or ROS. These observations are of importance for any medical or veterinary application of bacteriophages.

  10. Diversity of structure and function of DNA polymerase (gp43) of T4-related bacteriophages.

    Science.gov (United States)

    Petrov, V M; Karam, J D

    2004-11-01

    The replication DNA polymerase (gp43) of the bacteriophage T4 is a member of the pol B family of DNA polymerases, which are found in all divisions of life in the biosphere. The enzyme is a modularly organized protein that has several activities in one polypeptide chain (approximately 900 amino acid residues). These include two catalytic functions, POL (polymerase) and EXO (3 -exonuclease), and specific binding activities to DNA, the mRNA for gp43, deoxyribonucleotides (dNTPs), and other T4 replication proteins. The gene for this multifunctional enzyme (gene 43) has been preserved in evolution of the diverse group of T4-like phages in nature, but has diverged in sequence, organization, and specificity of the binding functions of the gene product. We describe here examples of T4-like phages where DNA rearrangements have created split forms of gene 43 consisting of two cistrons instead of one. These gene 43 variants specify separate gp43A (N-terminal) and gp43B (C-terminal) subunits of a split form of gp43. Compared to the monocistronic form, the interruption in contiguity of the gene 43 reading frame maps in a highly diverged sequence separating the code for essential components of two major modules of this pol B enzyme, the FINGERS and PALM domains, which contain the dNTP binding pocket and POL catalytic residues of the enzyme. We discuss the biological implications of these gp43 splits and compare them to other types of pol B splits in nature. Our studies suggest that DNA mobile elements may allow genetic information for pol B modules to be exchanged between organisms.

  11. Three magnetic particles solid phase radioimmunoassay for T4: Comparison of their results with established methods

    International Nuclear Information System (INIS)

    Bashir, T.

    1996-01-01

    The introduction of solid phase separation techniques is an important improvement in radioimmunoassays and immunoradiometric assays. Magnetic particle solid phase method has additional advantages over others, as the separation is rapid and centrifugation is not required. Three types of magnetic particles have been studied in T 4 RIA and the results have been compared with commercial kits and other established methods. (author). 4 refs, 9 figs, 2 tabs

  12. Tensile behavior of friction stir welded AA 6061-T4 aluminum alloy joints

    International Nuclear Information System (INIS)

    Heidarzadeh, A.; Khodaverdizadeh, H.; Mahmoudi, A.; Nazari, E.

    2012-01-01

    Highlights: ► Range of parameters for defect-free friction stir welded AA 6061-T4 was reached. ► A model was developed for predicting UTS and EL of friction stir welded AA 6061-T4. ► The maximum values of UTS and EL of joints were estimated by developed model. ► The optimum values of FSW process parameters were determined. -- Abstract: In this investigation response surface methodology based on a central composite rotatable design with three parameters, five levels and 20 runs, was used to develop a mathematical model predicting the tensile properties of friction stir welded AA 6061-T4 aluminum alloy joints at 95% confidence level. The three welding parameters considered were tool rotational speed, welding speed and axial force. Analysis of variance was applied to validate the predicted model. Microstructural characterization and fractography of joints were examined using optical and scanning electron microscopes. Also, the effects of the welding parameters on tensile properties of friction stir welded joints were analyzed in detail. The results showed that the optimum parameters to get a maximum of tensile strength were 920 rev/min, 78 mm/min and 7.2 kN, where the maximum of tensile elongation was obtained at 1300 rev/min, 60 mm/min and 8 kN.

  13. Processive nicking activity of T4 endonuclease V on UV-irradiated chromatin

    International Nuclear Information System (INIS)

    Gruskin, E.A.; Lloyd, R.S.

    1986-01-01

    T4 endonuclease V initiates the excision repair of pyrimidine dimers in UV-irradiated T4 infected E. coli cells. The pyrimidine dimer specific nicking activity of T4 endonuclease V functions by a processive scanning on UV-irradiated DNA. Previously it has been demonstrated that introduction of endonuclease V into repair-deficient human cells causes a restoration of UV survival in these cells. This demonstrates that endonuclease V is competent to incise mammalian DNA at the site of pyrimidine dimers. In order to assess the ability of endonuclease V to act processively on DNA associated as chromatin, minichromosomes were prepared for use as a substrate. Form I DNA was reconstituted with H3, H4 +/- H1 histones by sequential dialysis steps from 2.0 M NaCl to 50 mM NaCl. Time course reactions were performed with minichromosomes containing 10 and 25 dimers per molecule. In each case the rate of disappearance of form I DNA which was associated as chromatin was decreased relative to that of naked form I DNA. Concurrent with that observation, the rate and extent of appearance of form III DNA was increased with the DNA in minichromosomes relative to naked DNA. This is diagnostic of an enhancement of processivity. The inclusion of H1 in the minichromosomes resulted in a slight additional increase in processivity relative to minichromosomes consisting only of H3 and H4

  14. Sequence organization and control of transcription in the bacteriophage T4 tRNA region.

    Science.gov (United States)

    Broida, J; Abelson, J

    1985-10-05

    Bacteriophage T4 contains genes for eight transfer RNAs and two stable RNAs of unknown function. These are found in two clusters at 70 X 10(3) base-pairs on the T4 genetic map. To understand the control of transcription in this region we have completed the sequencing of 5000 base-pairs in this region. The sequence contains a part of gene 3, gene 1, gene 57, internal protein I, the tRNA genes and five open reading frames which most likely code for heretofore unidentified proteins. We have used subclones of the region to investigate the kinetics of transcription in vivo. The results show that transcription in this region consists of overlapping early, middle and late transcripts. Transcription is directed from two early promoters, one or two middle promoters and perhaps two late promoters. This region contains all of the features that are seen in T4 transcription and as such is a good place to study the phenomenon in more detail.

  15. High diversity and potential origins of T4-type bacteriophages on the surface of Arctic glaciers.

    Science.gov (United States)

    Bellas, Christopher M; Anesio, Alexandre M

    2013-09-01

    Tailed bacteriophages are the most abundant viruses in the biosphere. Here we examined the T4-type bacteriophage community inhabiting the surface of two glaciers in Svalbard. We used a molecular approach to target g23, the major capsid protein gene, to demonstrate that in the extreme cryoconite hole habitats the T4-type phages are surprisingly diverse. Phylogenetic analysis revealed that cryoconite hole sediments harbour a mixed phage community spanning multiple T4-type phage subgroups. The majority (71 %) of phage sequences clustered into three novel phylogenetically distinct groups, whilst the remainder clustered with known marine and soil derived phage sequences. The meltwater in cryoconite holes also contained a further distinct phage community which was related to previously detected marine phage variants. The ability of phages to move between marine and glacial habitats was tested in a transplantation experiment. Phages from the nearby marine fjord were found to be capable of initiating infection of supraglacial bacteria, suggesting suitable hosts could be found by non-native phages. Together this evidence suggests that the surface of glaciers contain both novel and cosmopolitan phages, some of which may have arrived in the cryosphere from other biomes.

  16. Quantitation of T-3 (triiodothyronine) and T-4 (thyroxine) in serum and plasma. Draft report

    International Nuclear Information System (INIS)

    Ceglowski, W.; Williams, R.B.

    1981-12-01

    This report summarizes an examination of the published literature concerning the quantitation of thyroxine and triiodothyronine in the clinical laboratory. It therefore details the precision, accuracy, sensitivity, and specificity obtainable in various commercial systems and those devised in the clinical laboratory. The data produced by several of the procedures often indicate that improvements in these parameters would enhance overall assay performance and increase the reliability of the clinical interpretation derivable from assay results. For T-3 and T-4 in vitro assays a very large number of systems exist and are currently being utilized in clinical laboratories in this country. For the sake of brevity some systems, while mentioned, are not reviewed in exhaustive detail. Radioimmunoassay, as the most frequently performed assay for both T-3 and T-4 is extensively reviewed. Also discussed with particular interest are assay systems which will undoubtedly impact on the future course of thyroid hormone assessment in the clinical laboratory, namely enzyme immunoassay and fluorescent immunoassay. The state of the art in T-4 measurements in neonates, because it is such a critical area for application of in vitro thyroid testing, is given detailed review. The quantitation of free thyroxine has been discussed in detail. These assays have been gaining more frequent use in the clinical laboratory and increased commercial system development

  17. Synthesis and magnetic property of T4 virus-supported gold-coated iron ternary nanocomposite

    Energy Technology Data Exchange (ETDEWEB)

    Xu Ziming; Sun Hongjing; Gao Faming, E-mail: fmgao@ysu.edu.cn; Hou Li; Li Na [Yanshan University, Key Laboratory of Applied Chemistry (China)

    2012-12-15

    Herein, we present a novel method based on the use of the symmetrical T4 bacteriophage capsid as a scaffold for preparing the gold-coated iron ternary core/shell nanostructure. Results showed that the thick gold shell was obtained to effectively protect Fe core from oxidation. Magnetic measurements showed that the nanocomposites were superparamagnetic at room temperature with a blocking temperature of about 35 K. At 3 K, its coercivity of 1142.86 Oe was larger than the existing experimental values. The magnetic property of Au/T4 was also tested, demonstrating the source of the magnetic sample arising from the Fe core only. The absorption spectrum of the Fe-Au/T4 complex was measured and compared with gold/virus. Different thickness gold shells were controlled in the synthesis by tuning the Au salt addition. On the basis of results and discussion, we further speculated the general growing mechanism of the template-supported Fe-Au process.

  18. Urea, Uric Acid, Prolactin and fT4 Concentrations in Aqueous Humor of Keratoconus Patients.

    Science.gov (United States)

    Stachon, Tanja; Stachon, Axel; Hartmann, Ulrike; Seitz, Berthold; Langenbucher, Achim; Szentmáry, Nóra

    2017-06-01

    Keratoconus is a noninflammatory disease of the cornea associated with progressive thinning and conical shape. Metabolic alterations in the urea cycle, with changes in collagen fibril stability, oxidative stress, thyroid hormones and prolactin with regulatory effect on biosynthesis and biomechanical stability of corneal stroma, may all play a role in keratoconus etiology. Our purpose was to determine urea, uric acid, prolactin and free thyroxin (fT4) concentrations in human aqueous humor (hAH) of keratoconus and cataract patients. hAH was collected from 100 keratoconus (penetrating keratoplasty) (41.9 ± 14.9 years, 69 males) and 100 cataract patients (cataract surgery) (71.2 ± 12.4 years, 58 males). Urea, uric acid, prolactin and fT4 concentrations were measured by Siemens clinical chemistry or immunoassay system. For statistical analysis, a generalized linear model (GLM) was used. Urea concentration was 11.88 ± 3.03 mg/dl in keratoconus and 16.44 ± 6.40 mg/dl in cataract patients, uric acid 2.04 ± 0.59 mg/dl in keratoconus and 2.18 ± 0.73 mg/dl in cataract groups. Prolactin concentration was 3.18 ± 0.34 ng/ml in keratoconus and 3.33 ± 0.32 ng/ml in cataract patients, fT4 20.57 ± 4.76 pmol/l in KC and 19.06 ± 3.86 pmol/l in cataract group. Urea concentration was effected through gender (p = 0.039), age (p = 0.001) and diagnosis (p = 0.025). Uric acid concentration was not effected through any of the analyzed parameters (p > 0.056). Prolactin and fT4 concentration were effected only through diagnosis (p = 0.009 and p = 0.006). Urea and prolactin concentrations are decreased, fT4 concentration is increased in aqueous humor of keratoconus patients, and uric acid concentration remains unchanged. Urea concentration in aqueous humor is also increased in older and male patients. Therefore, metabolic disorder and hormonal balance may both have an impact on keratoconus development. Further studies are necessary to assess the specific impact.

  19. Alterations in gp37 expand the host range of a T4-like phage.

    Science.gov (United States)

    Chen, Mianmian; Zhang, Lei; Abdelgader, Sheikheldin A; Yu, Li; Xu, Juntian; Yao, Huochun; Lu, Chengping; Zhang, Wei

    2017-09-22

    The use of phages as antibacterial agents is limited by their generally narrow host range. The aim of this study was to make a T4-like phage, WG01, obtain the host range of another T4-like phage, QL01, by replacing its host determinant gene region with that of QL01. This process triggered a direct expansion of the WG01 host range. The offspring of WG01 obtained the host ranges of both QL01 and WG01, as well as the ability to infect eight additional host bacteria in comparison to the wildtype strains. WQD had the widest host range; therefore, the corresponding QD fragments could be used for constructing a homologous sequence library. Moreover, after a sequencing analysis of gene37, we identified two different mechanisms responsible for the expanded host range: 1) the first generation of WG01 formed chimeras without mutations; and 2) the second generation of WG01 mutants formed from the chimeras. The expansion of the host range indicated that regions other than the C-terminal region may indirectly change the receptor specificity by altering the supportive capacity of the binding site. Additionally, we also found that the subsequent generations acquired a novel means of expanding the host range through acquiring a wider temperature range for lysis by exchanging gene37. The method developed in this work offers a quick way to change or expand the host range of a phage. Future clinical applications for screening phages against a given clinical isolate could be achieved after acquiring more suitable homologous sequences. IMPORTANCE T4-like phages have been established as safe in numerous phage therapy applications. The primary drawbacks to the use of phages as therapeutic agents include their highly specific host range. Thus, changing or expanding the host range of T4-like phages is beneficial for selecting phages for phage therapy. In this study, the host range of one T4-like phage WG01 was expanded using genetic manipulation. The WG01 derivatives acquired a novel means

  20. The tertiary structure of an i-type lysozyme isolated from the common orient clam (Meretrix lusoria).

    Science.gov (United States)

    Kuwano, Yuko; Yoneda, Kazunari; Kawaguchi, Yuya; Araki, Tomohiro

    2013-11-01

    To evaluate the structure-function relationships of invertebrate lysozymes, a new invertebrate-type (i-type) lysozyme was isolated from the common orient clam (Meretrix lusoria) and the tertiary structure of this enzyme was determined. Comparison of the tertiary structure of this enzyme with those of chicken and Venerupi philippinarum lysozymes revealed that the location of the side chain of the second catalytic residue, an aspartic acid, and the N-acetylglucosamine trimer bound at subsites A-C were different. Furthermore, the amino acid electrostatically interacting with Asp30 in V. philippinarum lysozyme, Lys108, was substituted by Gly in M. lusoria lysozyme and no other possible amino acid that could contribute to this interaction was found in M. lusoria lysozyme. It therefore seems that the substitutions of the amino acids at the interface of the V. philippinarum lysozyme dimer are likely to change the oligomeric state of the M. lusoria lysozyme.

  1. Development, characterization, and technical applications of a fish lysozyme-specific monoclonal antibody (mAb M24-2)

    OpenAIRE

    Marsh, Marlee B.; Rice, Charles D.

    2009-01-01

    Lysozyme is one of several humoral and cellular factors associated with front line, innate immunity in all vertebrates. Historically, circulating lysozyme has been quantified in teleosts by measuring enzymatic activity against heat-killed Mycococcus lysodieticus using whole serum or plasma at a low pH. However, the amount of serum or plasma required for standard lysozyme activity exceeds that which can be easily acquired from small fish, thus making lysozyme a difficult endpoint to measure in...

  2. Molecular cloning and characterization of a lysozyme cDNA from the mole cricket Gryllotalpa orientalis (Orthoptera: Gryllotalpidae).

    Science.gov (United States)

    Kwon, Hyojung; Bang, Kyeongrin; Lee, Minsup; Cho, Saeyoull

    2014-09-01

    A full-length lysozyme cDNA from Gryllotalpa orientalis was cloned and sequenced. The deduced amino acid sequence of the lysozyme protein was 143 amino acids in length, with a calculated molecular mass of 15.84 kDa and an isoelectric point of 4.74. Sequence motifs, together with alignment and phylogenetic results, confirmed that G. orientalis lysozyme belongs to the C (chicken)-type lysozyme family of proteins. The protein sequence of lysozyme from G. orientalis showed high identity to that of Drosophila melanogaster (51.7 %); however, in contrast to D. melanogaster lysozyme, G. orientalis lysozyme was immune inducible and expressed in a wide range of tissues. Expression of G. orientalis lysozyme mRNA was highest at 8 h post-infection and subsequently decreased with time after bacterial infection. We also expressed G. orientalis lysozyme protein in vitro using the pET expression system. Compared with the negative control, over-expressed G. orientalis lysozyme showed antimicrobial activity against Gram-negative bacteria Escherichia coli and Gram-positive bacteria Bacillus subtilis by radial diffusion assay, with minimal inhibitory concentration values of 30.3 and 7.55 µM, respectively. These results indicate that G. orientalis lysozyme may have stronger antimicrobial activity than other lysozymes against a broad range of microorganisms.

  3. Isolation and partial purification of lysozyme from saliva of Bali cattle ...

    African Journals Online (AJOL)

    About 100 μg/ml lysozyme of 14.2 kDa, determined through sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), per 4800 μg/ml saliva was obtained. Based on turbidity assay of Staphylococcus aureus, it was revealed that inhibition activities of 40 μg/ml isolated lysozyme were comparable to 800 μg/ml ...

  4. Characterization of lysozyme films produced by matrix assisted pulsed laser evaporation (MAPLE)

    DEFF Research Database (Denmark)

    Purice, Andreea; Schou, Jørgen; Kingshott, Peter

    2007-01-01

    Thin lysozyme films of thickness up to more than 100 nm have been produced in a dry environment by MAPLE (matrix assisted pulsed laser evaporation) from a water ice matrix. Analysis of the films demonstrates that a significant part of the lysozyme molecules is transferred to the substrate without...

  5. Surface morphology of thin lysozyme films produced by matrix-assisted pulsed laser evaporation (MAPLE)

    DEFF Research Database (Denmark)

    Purice, Andreea; Schou, Jørgen; Pryds, Nini

    2007-01-01

    Thin films of the protein, lysozyme, have been deposited by the matrix-assisted pulsed laser evaporation (MAPLE) technique. Frozen targets of 0.3-1.0 wt.% lysozyme dissolved in ultrapure water were irradiated by laser light at 355 mn with a fluence of 2 J/cm(2). The surface quality of the thin...

  6. Salt-Induced Disintegration of Lysozyme-Containing Polyelectrolyte Complex Micelles

    NARCIS (Netherlands)

    Lindhoud, Saskia; Voorhaar, Lenny; de Vries, Renko; Schweins, Ralf; Stuart, Martien A. Cohen; Norde, Willem

    2009-01-01

    The salt-induced disintegration of lysozyme-filled polyelectrolyte complex micelles, consisting of positively charged homopolymers (PDMAEMA(150)), negatively charged diblock copolymers (PAA(42)-PAAm(417)) and lysozyme, has been Studied with dynamic light scattering (DL) and small-angle neutron

  7. Salt-Induced Disintegration of Lysozyme-Containing Polyelectrolyte Complex Micelles

    NARCIS (Netherlands)

    Lindhoud, S.; Cohen Stuart, M.A.; Norde, W.; Vries, de R.J.; Schweins, R.; Voorhaar, L.

    2009-01-01

    The salt-induced disintegration of lysozyme-filled polyelectrolyte complex micelles, consisting of positively charged homopolymers (PDMAEMA150), negatively charged diblock copolymers (PAA42-PAAm417), and lysozyme, has been studied with dynamic light scattering (DLS) and small-angle neutron

  8. Isothermal Titration Calorimetry and Macromolecular Visualization for the Interaction of Lysozyme and Its Inhibitors

    Science.gov (United States)

    Wei, Chin-Chuan; Jensen, Drake; Boyle, Tiffany; O'Brien, Leah C.; De Meo, Cristina; Shabestary, Nahid; Eder, Douglas J.

    2015-01-01

    To provide a research-like experience to upper-division undergraduate students in a biochemistry teaching laboratory, isothermal titration calorimetry (ITC) is employed to determine the binding constants of lysozyme and its inhibitors, N-acetyl glucosamine trimer (NAG[subscript 3]) and monomer (NAG). The extremely weak binding of lysozyme/NAG is…

  9. Drug delivery to the kidneys and the bladder with the low molecular weight protein lysozyme

    NARCIS (Netherlands)

    Kok, Robbert J.; Haas, Marijke; Moolenaar, Frits; de Zeeuw, Dick; Meijer, Dirk K.F.

    1998-01-01

    The low molecular weight protein (LMWP) lysozyme is a suitable drug carrier for renal drug targeting. When the tubular reabsorption of a can be prevented, the protein will be excreted in the urine. In this way, lysozyme (LZM) conjugates might also be used as carriers for targeting to the urinary

  10. Thermodynamic Exploration of Eosin-Lysozyme Binding: A Physical Chemistry and Biochemistry Laboratory Experiment

    Science.gov (United States)

    Huisman, Andrew J.; Hartsell, Lydia R.; Krueger, Brent P.; Pikaart, Michael J.

    2010-01-01

    We developed a modular pair of experiments for use in the undergraduate physical chemistry and biochemistry laboratories. Both experiments examine the thermodynamics of the binding of a small molecule, eosin Y, to the protein lysozyme. The assay for binding is the quenching of lysozyme fluorescence by eosin through resonant energy transfer. In…

  11. Stability of lysozyme in aqueous extremolyte solutions during heat shock and accelerated thermal conditions

    NARCIS (Netherlands)

    Avanti, Christina; Saluja, Vinay; Van Streun, Erwin L. P.; Frijlink, Henderik W.; Hinrichs, Wouter L. J.

    2014-01-01

    The purpose of this study was to investigate the stability of lysozyme in aqueous solutions in the presence of various extremolytes (betaine, hydroxyectoine, trehalose, ectoine, and firoin) under different stress conditions. The stability of lysozyme was determined by Nile red Fluorescence

  12. Drug delivery to the kidneys and the bladder with the low molecular weight protein lysozyme

    NARCIS (Netherlands)

    Kok, RJ; Haas, M; Moolenaar, F; de Zeeuw, D; Meijer, DKF

    1998-01-01

    The low molecular weight protein (LMWP) lysozyme is a suitable drug carrier for renal drug targeting. When the tubular reabsorption of a LMWP can be prevented, the protein will be excreted in the urine. In this way lysozyme (LZM) conjugates might also be used as carriers for targeting to the urinary

  13. Genetic control of the humoral immune response to avian egg white lysozymes in the chicken

    Energy Technology Data Exchange (ETDEWEB)

    Flanagan, M.P.

    1987-01-01

    Chickens from two closely related sublines, GHs-B6 and GHs-B13, differing serologically at the major histocompatibility complex, were significantly different in their humoral response to three avian egg white lysozymes. Specific antisera levels were measured by radioimmunoassay using /sup 125/I-labeled lysozymes. Antibodies elicited in response to these lysozymes are assumed to be directed against sites on these lysozymes where their amino acid sequence differs from that of the recipient G. domesticus egg white lysozyme (HEL). GHs-B6 birds produced a high level of antibody in response to immunization of turkey (TEL), pheasant (PhL) and guinea hen (GHL) lysozymes. GHs-B13 birds produced no detectable antibody to TEL, were intermediate in their response to PhL and equaled the antibody production of GHs-B6 birds in response to GHL. Antisera to each lysozyme were examined for crossreactivity with all other lysozymes by use of a competitive binding assay.

  14. Suppressive effects of lysozyme on polyphosphate-mediated vascular inflammatory responses

    Energy Technology Data Exchange (ETDEWEB)

    Chung, Jiwoo [College of Pharmacy, CMRI, Research Institute of Pharmaceutical Sciences, BK21 Plus KNU Multi-Omics Based Creative Drug Research Team, Kyungpook National University, Daegu 41566 (Korea, Republic of); Ku, Sae-Kwang [Department of Anatomy and Histology, College of Korean Medicine, Daegu Haany University, Gyeongsan 38610 (Korea, Republic of); Lee, Suyeon [College of Pharmacy, CMRI, Research Institute of Pharmaceutical Sciences, BK21 Plus KNU Multi-Omics Based Creative Drug Research Team, Kyungpook National University, Daegu 41566 (Korea, Republic of); Bae, Jong-Sup, E-mail: baejs@knu.ac.kr [College of Pharmacy, CMRI, Research Institute of Pharmaceutical Sciences, BK21 Plus KNU Multi-Omics Based Creative Drug Research Team, Kyungpook National University, Daegu 41566 (Korea, Republic of)

    2016-06-10

    Lysozyme, found in relatively high concentration in blood, saliva, tears, and milk, protects us from the ever-present danger of bacterial infection. Previous studies have reported proinflammatory responses of endothelial cells to the release of polyphosphate(PolyP). In this study, we examined the anti-inflammatory responses and mechanisms of lysozyme and its effects on PolyP-induced septic activities in human umbilical vein endothelial cells (HUVECs) and mice. The survival rates, septic biomarker levels, behavior of human neutrophils, and vascular permeability were determined in PolyP-activated HUVECs and mice. Lysozyme suppressed the PolyP-mediated vascular barrier permeability, upregulation of inflammatory biomarkers, adhesion/migration of leukocytes, and activation and/or production of nuclear factor-κB, tumor necrosis factor-α, and interleukin-6. Furthermore, lysozyme demonstrated protective effects on PolyP-mediated lethal death and the levels of the related septic biomarkers. Therefore, these results indicated the therapeutic potential of lysozyme on various systemic inflammatory diseases, such as sepsis or septic shock. -- Highlights: •PolyP is shown to be an important mediator of vascular inflammation. •Lysozyme inhibited PolyP-mediated hyperpermeability. •Lysozyme inhibited PolyP-mediated septic response. •Lysozyme reduced PolyP-induced septic mortality.

  15. The effects of xylitol and sorbitol on lysozyme- and peroxidase-related enzymatic and candidacidal activities.

    Science.gov (United States)

    Kim, Bum-Soo; Chang, Ji-Youn; Kim, Yoon-Young; Kho, Hong-Seop

    2015-07-01

    To investigate whether xylitol and sorbitol affect enzymatic and candidacidal activities of lysozyme, the peroxidase system, and the glucose oxidase-mediated peroxidase system. Xylitol and sorbitol were added to hen egg-white lysozyme, bovine lactoperoxidase, glucose oxidase-mediated peroxidase, and whole saliva in solution and on hydroxyapatite surfaces. The enzymatic activities of lysozyme, peroxidase, and glucose oxidase-mediated peroxidase were determined by the turbidimetric method, the NbsSCN assay, and production of oxidized o-dianisidine, respectively. Candidacidal activities were determined by comparing colony forming units using Candida albicans ATCC strains 10231, 11006, and 18804. While xylitol and sorbitol did not affect the enzymatic activity of hen egg-white lysozyme both in solution and on hydroxyapatite surfaces, they did inhibit the enzymatic activity of salivary lysozyme significantly in solution, but not on the surfaces. Xylitol and sorbitol enhanced the enzymatic activities of both bovine lactoperoxidase and salivary peroxidase significantly in a dose-dependent manner in solution, but not on the surfaces. Sorbitol, but not xylitol, inhibited the enzymatic activity of glucose oxidase-mediated peroxidase significantly. Both xylitol and sorbitol did not affect candidacidal activities of hen egg-white lysozyme, the bovine lactoperoxidase system, or the glucose oxidase-mediated bovine lactoperoxidase system. Xylitol and sorbitol inhibited salivary lysozyme activity, but enhanced both bovine lactoperoxidase and salivary peroxidase activities significantly in solution. Xylitol and sorbitol did not augment lysozyme- and peroxidase-related candidacidal activities. Copyright © 2015 Elsevier Ltd. All rights reserved.

  16. Suppressive effects of lysozyme on polyphosphate-mediated vascular inflammatory responses

    International Nuclear Information System (INIS)

    Chung, Jiwoo; Ku, Sae-Kwang; Lee, Suyeon; Bae, Jong-Sup

    2016-01-01

    Lysozyme, found in relatively high concentration in blood, saliva, tears, and milk, protects us from the ever-present danger of bacterial infection. Previous studies have reported proinflammatory responses of endothelial cells to the release of polyphosphate(PolyP). In this study, we examined the anti-inflammatory responses and mechanisms of lysozyme and its effects on PolyP-induced septic activities in human umbilical vein endothelial cells (HUVECs) and mice. The survival rates, septic biomarker levels, behavior of human neutrophils, and vascular permeability were determined in PolyP-activated HUVECs and mice. Lysozyme suppressed the PolyP-mediated vascular barrier permeability, upregulation of inflammatory biomarkers, adhesion/migration of leukocytes, and activation and/or production of nuclear factor-κB, tumor necrosis factor-α, and interleukin-6. Furthermore, lysozyme demonstrated protective effects on PolyP-mediated lethal death and the levels of the related septic biomarkers. Therefore, these results indicated the therapeutic potential of lysozyme on various systemic inflammatory diseases, such as sepsis or septic shock. -- Highlights: •PolyP is shown to be an important mediator of vascular inflammation. •Lysozyme inhibited PolyP-mediated hyperpermeability. •Lysozyme inhibited PolyP-mediated septic response. •Lysozyme reduced PolyP-induced septic mortality.

  17. Production of active lysozyme films by matrix assisted pulsed laser evaporation at 355 nm

    DEFF Research Database (Denmark)

    Purice, Andreea; Schou, Jørgen; Kingshott, P.

    2007-01-01

    Thin lysozyme films have been produced in a dry environment by MAPLE (matrix assisted pulsed laser evaporation) from a water ice matrix irradiated by laser light at 355 nm above the absorption threshold of the protein. A significant part of the lysozyme molecules are transferred to the film without...

  18. Pulsed laser deposition of lysozyme: the dependence on shot numbers and the angular distribution

    DEFF Research Database (Denmark)

    Constantinescu, C.; Matei, A.; Schou, Jørgen

    2013-01-01

    . This was verified by matrix-assisted laser desorption ionization (MALDI) spectrometry of thin films deposited on silicon substrates. The deposition rate of lysozyme was found to decrease with the number of shots and was correlated with increasing thermal damage of the lysozyme. This was monitored by measurements...

  19. Concurrent chemotherapy for T4 classification nasopharyngeal carcinoma in the era of intensity-modulated radiotherapy.

    Directory of Open Access Journals (Sweden)

    Cai-neng Cao

    Full Text Available To evaluate concurrent chemotherapy for T4 classification nasopharyngeal carcinoma (NPC treated by intensity-modulated radiotherapy (IMRT.From July 2004 to June 2011, 180 non-metastatic T4 classification NPC patients were retrospectively analyzed. Of these patients, 117 patients were treated by concurrent chemoradiotherapy (CCRT using IMRT and 63 cases were treated by IMRT alone.The median follow-up time was 58.97 months (range, 2.79-114.92 months. For all the patients, the 1, 3 and 5-year local failure-free survival (LFFS rates were 97.7%, 89.2% and 85.9%, regional failure free survival (RFFS rates were 98.9%, 94.4% and 94.4%, distant failure-free survival (DFFS rates were 89.7%, 79.9% and 76.2%, and overall survival (OS rates were 92.7%, 78.9% and 65.3%, respectively. No statistically significant difference was observed in LFFS, RFFS, DFFS and OS between the CCRT group and the IMRT alone group. No statistically significant difference was observed in acute toxicity except leukopenia (p = 0.000 during IMRT between the CCRT group and the IMRT alone group.IMRT alone for T4 classification NPC achieved similar treatment outcomes in terms of disease local control and overall survival as compared to concurrent chemotherapy plus IMRT. However, this is a retrospective study with a limited number of patients, such results need further investigation in a prospective randomized clinical trial.

  20. Intensity-modulated radiation therapy for T4 nasopharyngeal carcinoma. Treatment results and locoregional

    Energy Technology Data Exchange (ETDEWEB)

    Chen, J.L.Y.; Tsai, C.L.; Chen, W.Y.; Wang, C.W. [National Taiwan Univ. Hospital, Taipei (China). Div. of Radiation Oncology; Huang, Y.S.; Chen, Y.F. [National Taiwan Univ. Hospital, Taipei (China). Dept. of Medical Imaging; Kuo, S.H. [National Taiwan Univ. Hospital, Taipei (China). Div. of Radiation Oncology; National Taiwan Univ. College of Medicine, Taipei (China). Graduate Inst. of Clinical Medicine; Hong, R.L. [National Taiwan Univ. Hospital, Taipei (China). Div. of Medical Oncology; Ko, J.Y.; Lou, P.J. [National Taiwan Univ. Hospital, Taipei (China). Dept. of Otolaryngology

    2013-12-15

    Purpose: The purpose of this work was to examine outcomes in patients with T4 nasopharyngeal carcinoma (NPC) treated with intensity-modulated radiation therapy (IMRT). Methods and materials: Between 2007 and 2010, 154 patients with nonmetastatic T4 NPC were treated with IMRT to a total dose of 70 Gy in 33-35 fractions. In addition, 97 % of patients received concurrent platinum-based chemotherapy. The median follow-up time was 52.8 months. Results: The rates of 5-year actuarial locoregional control, distant metastasis-free survival, progression free-survival, and overall survival (OS) were 81.2, 72.2, 61.9, and 78.1 %, respectively. A total of 27 patients had locoregional recurrence: 85.2 % in-field failures, 11.1 % marginal failures, and 3.7 % out-of-field failures. Fourteen patients with locoregional recurrence received aggressive treatments, including nasopharyngectomy, neck dissection, or re-irradiation, and the 5-year OS rate tended to be better (61.9 %) compared to those receiving conservative treatment (32.0 %, p = 0.051). In patients treated with 1 course of radiotherapy, grade {>=} 3 toxicities of ototoxicity, neck fibrosis, xerostomia, epistaxis, and radiographic temporal lobe necrosis occurred in 18.2, 9.8, 6.3, 2.1, and 5.6 % of patients, respectively. Increased ototoxicity, osteonecrosis, severe nasal bleeding, and temporal necrosis were observed in patients treated by re-irradiation. Conclusion: IMRT offers good locoregional control in patients with T4 NPC. For patients with locoregional recurrence after definitive radiotherapy, aggressive local treatment may be considered for a better outcome. (orig.)

  1. Specificity of interactions among the DNA-packaging machine components of T4-related bacteriophages.

    Science.gov (United States)

    Gao, Song; Rao, Venigalla B

    2011-02-04

    Tailed bacteriophages use powerful molecular motors to package the viral genome into a preformed capsid. Packaging at a rate of up to ∼2000 bp/s and generating a power density twice that of an automobile engine, the phage T4 motor is the fastest and most powerful reported to date. Central to DNA packaging are dynamic interactions among the packaging components, capsid (gp23), portal (gp20), motor (gp17, large "terminase"), and regulator (gp16, small terminase), leading to precise orchestration of the packaging process, but the mechanisms are poorly understood. Here we analyzed the interactions between small and large terminases of T4-related phages. Our results show that the gp17 packaging ATPase is maximally stimulated by homologous, but not heterologous, gp16. Multiple interaction sites are identified in both gp16 and gp17. The specificity determinants in gp16 are clustered in the diverged N- and C-terminal domains (regions I-III). Swapping of diverged region(s), such as replacing C-terminal RB49 region III with that of T4, switched ATPase stimulation specificity. Two specificity regions, amino acids 37-52 and 290-315, are identified in or near the gp17-ATPase "transmission" subdomain II. gp16 binding at these sites might cause a conformational change positioning the ATPase-coupling residues into the catalytic pocket, triggering ATP hydrolysis. These results lead to a model in which multiple weak interactions between motor and regulator allow dynamic assembly and disassembly of various packaging complexes, depending on the functional state of the packaging machine. This might be a general mechanism for regulation of the phage packaging machine and other complex molecular machines.

  2. Purification of phage display-modified bacteriophage T4 by affinity chromatography

    Directory of Open Access Journals (Sweden)

    Figura Grzegorz

    2011-05-01

    Full Text Available Abstract Background Affinity chromatography is one of the most efficient protein purification strategies. This technique comprises a one-step procedure with a purification level in the order of several thousand-fold, adaptable for various proteins, differentiated in their size, shape, charge, and other properties. The aim of this work was to verify the possibility of applying affinity chromatography in bacteriophage purification, with the perspective of therapeutic purposes. T4 is a large, icosahedral phage that may serve as an efficient display platform for foreign peptides or proteins. Here we propose a new method of T4 phage purification by affinity chromatography after its modification with affinity tags (GST and Histag by in vivo phage display. As any permanent introduction of extraneous DNA into a phage genome is strongly unfavourable for medical purposes, integration of foreign motifs with the phage genome was not applied. The phage was propagated in bacteria expressing fusions of the phage protein Hoc with affinity tags from bacterial plasmids, independently from the phage expression system. Results Elution profiles of phages modified with the specific affinity motifs (compared to non-specific phages document their binding to the affinity resins and effective elution with standard competitive agents. Non-specific binding was also observed, but was 102-105 times weaker than the specific one. GST-modified bacteriophages were also effectively released from glutathione Sepharose by proteolytic cleavage. The possibility of proteolytic release was designed at the stage of expression vector construction. Decrease in LPS content in phage preparations was dependent on the washing intensity; intensive washing resulted in preparations of 11-40 EU/ml. Conclusions Affinity tags can be successfully incorporated into the T4 phage capsid by the in vivo phage display technique and they strongly elevate bacteriophage affinity to a specific resin. Affinity

  3. Evidence that the heterogeneity of a T4 population is the result of heritable traits.

    Directory of Open Access Journals (Sweden)

    Zachary J Storms

    Full Text Available Many bacteriophage populations display heterogeneity in their adsorption characteristics; a portion of the phage population remains free in solution throughout adsorption experiments (residual fraction. This residual fraction generally constitutes a minority of phages that exhibit significantly slower adsorption kinetics than the main phage stock (main fraction. While this phenomenon is likely the result of evolutionary driving forces, the present study demonstrates that the residual fraction is not always the result of phenotypic variations within a single genotype, as is generally thought. Experiments with phage T4 showed that two subgroups with distinct adsorption traits that were passed on to their progeny could be isolated from the original phage stock. Sequencing of genes involved in adsorption revealed two point mutations in gene 37 of residual fraction isolates, which resulted in modifications to the long tail-fiber, the organelle of attachment and host cell recognition. Adsorption studies consistently showed that T4 phage stocks amplified from residual fraction isolates had significantly lower adsorption efficiencies than those amplified from main fractions. The conducted experiments provide convincing evidence that the observed heterogeneity in T4 adsorption behavior is the result of conserved mutations to the phage genome and is not exclusively the result of phenotypic variations within the population. While it is believed high mutation rates exist to hasten phage adaptation, this study shows that this bet hedging strategy can also, in the short term, inadvertently handicap the phage's adsorption capabilities to a given host under normal infection conditions, resulting in the residual fraction observed in adsorption experiments.

  4. Oncological outcomes following radical prostatectomy for patients with pT4 prostate cancer

    Directory of Open Access Journals (Sweden)

    Dharam Kaushik

    Full Text Available ABSTRACT Objectives: Radical prostatectomy (RP for locally advanced prostate cancer may reduce the risk of metastasis and cancer-specific death. Herein, we evaluated the outcomes for patients with pT4 disease treated with RP. Materials and methods: Among 19,800 men treated with RP at Mayo Clinic from 1987 to 2010, 87 were found to have pT4 tumors. Biochemical recurrence (BCR-free survival, systemic progression (SP free survival and overall survival (OS were estimated using the Kaplan-Meier method and compared with the log-rank test. Cox proportional hazards regression models were used to assess the association of clinic-pathological features with outcome. Results: Median follow-up was 9.8 years (IQR 3.6, 13.4. Of the 87 patients, 50 (57.5% were diagnosed with BCR, 30 (34.5% developed SP, and 38 (43.7% died, with 11 (12.6% dying of prostate cancer. Adjuvant androgen deprivation therapy was administered to 77 men, while 32 received adjuvant external beam radiation therapy. Ten-year BCR-free survival, SP-free survival, and OS was 37%, 64%, and 70% respectively. On multivariate analysis, the presence of positive lymph nodes was marginally significantly associated with patients' risk of BCR (HR: 1.94; p=0.05, while both positive lymph nodes (HR 2.96; p=0.02 and high pathologic Gleason score (HR 1.95; p=0.03 were associated with SP. Conclusions: Patients with pT4 disease may experience long-term survival following RP, and as such, when technically feasible, surgical resection should be considered in the multimodal treatment approach to these men.

  5. Intensity-modulated radiation therapy for T4 nasopharyngeal carcinoma. Treatment results and locoregional

    International Nuclear Information System (INIS)

    Chen, J.L.Y.; Tsai, C.L.; Chen, W.Y.; Wang, C.W.; Huang, Y.S.; Chen, Y.F.; Kuo, S.H.; National Taiwan Univ. College of Medicine, Taipei; Hong, R.L.; Ko, J.Y.; Lou, P.J.

    2013-01-01

    Purpose: The purpose of this work was to examine outcomes in patients with T4 nasopharyngeal carcinoma (NPC) treated with intensity-modulated radiation therapy (IMRT). Methods and materials: Between 2007 and 2010, 154 patients with nonmetastatic T4 NPC were treated with IMRT to a total dose of 70 Gy in 33-35 fractions. In addition, 97 % of patients received concurrent platinum-based chemotherapy. The median follow-up time was 52.8 months. Results: The rates of 5-year actuarial locoregional control, distant metastasis-free survival, progression free-survival, and overall survival (OS) were 81.2, 72.2, 61.9, and 78.1 %, respectively. A total of 27 patients had locoregional recurrence: 85.2 % in-field failures, 11.1 % marginal failures, and 3.7 % out-of-field failures. Fourteen patients with locoregional recurrence received aggressive treatments, including nasopharyngectomy, neck dissection, or re-irradiation, and the 5-year OS rate tended to be better (61.9 %) compared to those receiving conservative treatment (32.0 %, p = 0.051). In patients treated with 1 course of radiotherapy, grade ≥ 3 toxicities of ototoxicity, neck fibrosis, xerostomia, epistaxis, and radiographic temporal lobe necrosis occurred in 18.2, 9.8, 6.3, 2.1, and 5.6 % of patients, respectively. Increased ototoxicity, osteonecrosis, severe nasal bleeding, and temporal necrosis were observed in patients treated by re-irradiation. Conclusion: IMRT offers good locoregional control in patients with T4 NPC. For patients with locoregional recurrence after definitive radiotherapy, aggressive local treatment may be considered for a better outcome. (orig.)

  6. Immunological response in egg-sensitive adults challenged with cheese containing or not containing lysozyme.

    Science.gov (United States)

    Rossi, Filippo; Iaconelli, Amerigo; Fiorentini, Lucia; Zito, Francesco; Donati, Maria Benedett; De Cristofaro, Maria Laura; Piva, Gianfranco; Mingrone, Geltrude

    2012-12-01

    Lysozyme is an enzyme that hydrolyzes bacterial peptidoglicans. For this reason, it is used in cheese manufacturing in order to prevent a defect of long-ripened hard cheese called "late blowing" due to the outgrowth of spores of Clostridium tyrobutyricum and Clostridium butyricum. Moreover, germination of Listeria monocytogenes spores into vegetative cells is also sensitive to lysozyme. The enzyme can be an allergenic molecule, and for this reason there are concerns about its use in food industry. The immunological and clinical response of consumption of lysozyme-containing cheese has been evaluated in 25 egg-sensitive subjects with or without lysozyme sensitization. A total of 25 egg-sensitive subjects were enrolled in this study. All the subjects were already treated for egg-sensitization and presented a positive skin prick test. All the subjects had a body mass index ≤ 25 kg/m(2) and were in the age range of 20-50 years. Each subject was studied twice and received randomly 30 g of Grana Padano (containing lysozyme) or TrentinGrana cheese (lysozyme-free) of two different aging periods: 16 or 24 months. A washout period of 1 week between each cheese intake was adopted. Blood samples were taken in fasting conditions and 1 hour after cheese intake and IgA, total IgE, and lysozyme-, ovomucoid-, and ovalbumin-specific IgE were measured. No adverse reactions were observed in both groups of patients after cheese samples were given. Lysozyme did not determine any variation of specific IgE compared with basal level. In lysozyme-sensitive patients a significant relationship between IgA and lysozyme-specific IgE was observed when lysozyme-containing cheese was given, confirming that lysozyme can pass the gut barrier. Neither adverse events nor immunological responses were observed after ingestion of cheese containing lysozyme. However, the immunological properties of peptides deriving from cheese protein hydrolysis need to be clarified, as does the effect of lysozyme on

  7. Absent endometrium due to balanced translocation [t(4;20] presenting as primary amenorrhea

    Directory of Open Access Journals (Sweden)

    Aruna Nigam

    2014-01-01

    Full Text Available Primary amenorrhea is defined as the absence of menarche by 16-18 years of age in the presence of well-developed secondary sexual characters. An incidence of 1-3% has been reported in women of reproductive age group. The etiology varies with anatomical, genetic and hormonal factors implicated in the causation of primary amenorrhea. We present a case of absent endometrium due to balanced reciprocal translocation (RCPTR, 46 XX t (4;20(q12;q13.1 as primary amenorrhea.

  8. Reconstrucción "Módulo D" aparcamiento Madrid Barajas T-4

    OpenAIRE

    Corres Peiretti, Hugo; Romero Rey, Eduardo

    2008-01-01

    En este artículo se describen los daños producidos a la estructura del aparcamiento de la terminal T4 de Barajas, tras el atentado del 2006. Se describen las zonas y elementos que se vieron afectados, y el tipo de daños que se detectarón. Tras analizar esa información se explica como se produzco el derrumbamiento y los modos de fallos que se observarón en la estructura. Inmediatamente despues de le explosión empiezan las labores de desemcombro y de demolición de los elementos afectados. Depen...

  9. Human Volunteers Receiving Escherichia coli Phage T4 Orally: a Safety Test of Phage Therapy

    OpenAIRE

    Bruttin, Anne; Brüssow, Harald

    2005-01-01

    Fifteen healthy adult volunteers received in their drinking water a lower Escherichia coli phage T4 dose (103 PFU/ml), a higher phage dose (105 PFU/ml), and placebo. Fecal coliphage was detected in a dose-dependent way in volunteers orally exposed to phage. All volunteers receiving the higher phage dose showed fecal phage 1 day after exposure; this prevalence was only 50% in subjects receiving the lower phage dose. No fecal phage was detectable a week after a 2-day course of oral phage applic...

  10. T-4 handbook of material properties data bases. Volume 1c. Equations of state

    International Nuclear Information System (INIS)

    Holian, K.S.

    1984-11-01

    This manual is a compilation of descriptions of the equations of state (EOS) in the T-4 computerized library of material properties tables. The introduction gives a brief descriptions of the library and of the physics theories and models which were used to calculate the equations of state. Then each EOS is described in detail. First, various physical parameters of each theoretical EOS are tabulated and compared with experiments when available. Then the method of generating the EOS is briefly described. Finally, the tabels are plotted in terms of pressure and energy vs density along line of constant temperature

  11. Cloning and expression of the genes coding for tube associated proteins of bacteriophage T4

    International Nuclear Information System (INIS)

    Nieradko, J.

    1995-01-01

    Genes 29, 48 and 54 of bacteriophage T4, coding for specific tube associated proteins, were cloned to the expression vector pT7-5. The molecular mass of the products of these genes was estimated to be 64, 39 and 36 kDa, respectively. The examined genes are co-transcribed with genes 51, 27 and 28 from the same DNA strand and a common late promoter sequence located downstream of gene 51. (author). 15 refs, 3 figs, 1 tab

  12. Fatigue crack growth in 2017A-T4 alloy subjected to proportional bending with torsion

    Directory of Open Access Journals (Sweden)

    D. Rozumek

    2017-10-01

    Full Text Available The paper presents the results of tests on the fatigue crack growth for a constant moment amplitude under combined bending with torsion in the aluminium alloy AW-2017A-T4. The tests were performed under different values of the load ratio R. Plane specimens with stress concentrators in form of the external one-sided sharp notch were tested. A non-uniform fatigue cracks growth on both lateral surfaces of specimens was observed during experimental tests. Fatigue cracks were developing in the specimens in two stages; quarter-elliptic edge cracks were observed at the beginning, then evolving into through cracks

  13. The Anti-sigma Factor RsiV Is a Bacterial Receptor for Lysozyme: Co-crystal Structure Determination and Demonstration That Binding of Lysozyme to RsiV Is Required for σV Activation.

    Directory of Open Access Journals (Sweden)

    Jessica L Hastie

    2016-09-01

    Full Text Available σ factors provide RNA polymerase with promoter specificity in bacteria. Some σ factors require activation in order to interact with RNA polymerase and transcribe target genes. The Extra-Cytoplasmic Function (ECF σ factor, σV, is encoded by several Gram-positive bacteria and is specifically activated by lysozyme. This activation requires the proteolytic destruction of the anti-σ factor RsiV via a process of regulated intramembrane proteolysis (RIP. In many cases proteases that cleave at site-1 are thought to directly sense a signal and initiate the RIP process. We previously suggested binding of lysozyme to RsiV initiated the proteolytic destruction of RsiV and activation of σV. Here we determined the X-ray crystal structure of the RsiV-lysozyme complex at 2.3 Å which revealed that RsiV and lysozyme make extensive contacts. We constructed RsiV mutants with altered abilities to bind lysozyme. We find that mutants that are unable to bind lysozyme block site-1 cleavage of RsiV and σV activation in response to lysozyme. Taken together these data demonstrate that RsiV is a receptor for lysozyme and binding of RsiV to lysozyme is required for σV activation. In addition, the co-structure revealed that RsiV binds to the lysozyme active site pocket. We provide evidence that in addition to acting as a sensor for the presence of lysozyme, RsiV also inhibits lysozyme activity. Thus we have demonstrated that RsiV is a protein with multiple functions. RsiV inhibits σV activity in the absence of lysozyme, RsiV binds lysozyme triggering σV activation and RsiV inhibits the enzymatic activity of lysozyme.

  14. The Anti-sigma Factor RsiV Is a Bacterial Receptor for Lysozyme: Co-crystal Structure Determination and Demonstration That Binding of Lysozyme to RsiV Is Required for σV Activation.

    Science.gov (United States)

    Hastie, Jessica L; Williams, Kyle B; Bohr, Lindsey L; Houtman, Jon C; Gakhar, Lokesh; Ellermeier, Craig D

    2016-09-01

    σ factors provide RNA polymerase with promoter specificity in bacteria. Some σ factors require activation in order to interact with RNA polymerase and transcribe target genes. The Extra-Cytoplasmic Function (ECF) σ factor, σV, is encoded by several Gram-positive bacteria and is specifically activated by lysozyme. This activation requires the proteolytic destruction of the anti-σ factor RsiV via a process of regulated intramembrane proteolysis (RIP). In many cases proteases that cleave at site-1 are thought to directly sense a signal and initiate the RIP process. We previously suggested binding of lysozyme to RsiV initiated the proteolytic destruction of RsiV and activation of σV. Here we determined the X-ray crystal structure of the RsiV-lysozyme complex at 2.3 Å which revealed that RsiV and lysozyme make extensive contacts. We constructed RsiV mutants with altered abilities to bind lysozyme. We find that mutants that are unable to bind lysozyme block site-1 cleavage of RsiV and σV activation in response to lysozyme. Taken together these data demonstrate that RsiV is a receptor for lysozyme and binding of RsiV to lysozyme is required for σV activation. In addition, the co-structure revealed that RsiV binds to the lysozyme active site pocket. We provide evidence that in addition to acting as a sensor for the presence of lysozyme, RsiV also inhibits lysozyme activity. Thus we have demonstrated that RsiV is a protein with multiple functions. RsiV inhibits σV activity in the absence of lysozyme, RsiV binds lysozyme triggering σV activation and RsiV inhibits the enzymatic activity of lysozyme.

  15. Characterization of lysozyme films produced by matrix assisted pulsed laser evaporation (MAPLE)

    International Nuclear Information System (INIS)

    Purice, Andreea; Schou, Jorgen; Kingshott, Peter; Pryds, Nini; Dinescu, Maria

    2007-01-01

    Thin lysozyme films of thickness up to more than 100 nm have been produced in a dry environment by MAPLE (matrix assisted pulsed laser evaporation) from a water ice matrix. Analysis of the films demonstrates that a significant part of the lysozyme molecules is transferred to the substrate without decomposition and that the protein activity is preserved. The film deposition rate for 1 wt% lysozyme has a maximum at 2 J/cm 2 of about 1 ng/cm 2 per laser shot. During the film production the deposition rate is constant without any sign of depletion or accumulation effects in the water ice target or in the growing film. Scanning electron microscopy (SEM) images demonstrate that the silicon substrate is completely covered by lysozyme films thicker than 100 nm. Deposition was also made from a target with pressed (100%) solid lysozyme, but the deposition was difficult to handle and with a much slower rate than that from a water ice matrix

  16. Structure of an antibody-lysozyme complex unexpected effect of conservative mutation.

    Science.gov (United States)

    Chacko, S; Silverton, E; Kam-Morgan, L; Smith-Gill, S; Cohen, G; Davies, D

    1995-01-20

    The structure of the complex between the Fab HyHEL-5 and chicken lysozyme revealed a large interface region containing 23 lysozyme and 28 Fab residues. Arg68 of the lysozyme is centrally placed in this interface and theoretical studies together with binding assays of this Fab to different avian lysozymes have previously shown that this arginine residue is an important contributor to the binding. The Arg68-->Lys mutant binds 10(3) times less well to the HyHEL-5 Fab. We have examined the refined crystal structure of the complex of this mutant lysozyme with the Fab. No global changes occur, but there is an introduction of a new water molecule into the interface that mediates the hydrogen bonding interactions between the lysine and residues on the Fab. These data are compared with the effects of similar changes on the inhibition of serine proteases such as trypsin where the energetic effects of this substitution are small.

  17. Laser ablation of lysozyme with UV, visible and infrared femto- and nanosecond pulses

    DEFF Research Database (Denmark)

    Schou, Jørgen; Canulescu, Stela; Matei, Andreea

    Lysozyme is an interesting molecule for laser ablation of organic materials, because the ablation has been comprehensively studied, it is a medium heavy molecule with a mass of 14305 Da, which can be detected by standard techniques, and because it is used as a bactericidal protein in the food...... industry. Lysozyme molecules do not absorb energy for wavelengths above 310 nm, but nevertheless there is a strong mass loss by ablation for laser irradiation in the visible regime. The total ablation yield of lysozyme at 355 nm and at 2 J/cm2 is about 155 µg/pulse, possibly one of the highest ablation...... yields ever measured. The mass loss is mainly caused by fragmentation of the lysozyme into simple gases, such as H2S, H2O and CO2 , which are rapidly pumped away in the vacuum chamber. We have investigated the mass loss by ablation of lysozyme in all regimes to see whether a similar mechanism governs...

  18. Pulsed laser deposition of lysozyme: the dependence on shot numbers and the angular distribution

    Science.gov (United States)

    Constantinescu, C.; Matei, A.; Schou, J.; Canulescu, S.; Dinescu, M.

    2013-12-01

    The ejection of molecules from a pressed solid target of lysozyme induced by laser ablation in the UV-regime at a wavelength of 355 nm was investigated. The ablation studies were carried out in vacuum at a laser fluence of 2 J/cm2 for which a significant fraction of proteins remains intact. This was verified by matrix-assisted laser desorption ionization (MALDI) spectrometry of thin films deposited on silicon substrates. The deposition rate of lysozyme was found to decrease with the number of shots and was correlated with increasing thermal damage of the lysozyme. This was monitored by measurements of the optical reflectivity of dry lysozyme. The angular distribution of the mass deposition can be fitted well by Anisimov's hydrodynamic model. The total deposited yield over the entire hemisphere from direct laser ablation of lysozyme was estimated from this model and found to be three orders of magnitude less than the ablated mass.

  19. Dynamics of hydration in hen egg white lysozyme.

    Science.gov (United States)

    Sterpone, F; Ceccarelli, M; Marchi, M

    2001-08-10

    We investigate the hydration dynamics of a small globular protein, hen egg-white lysozyme. Extensive simulations (two trajectories of 9 ns each) were carried out to identify the time-scales and mechanism of water attachment to this protein. The location of the surface and integral water molecules in lysozyme was also investigated. Three peculiar temporal scales of the hydration dynamics can be discerned: two among these, with sub-nanosecond mean residence time, tau(w), are characteristic of surface hydration water; the slower time-scale (tau(w) approximately 2/3 ns) is associated with buried water molecules in hydrophilic pores and in superficial clefts. The computed tau(w) values in the two independent runs fall in a similar range and are consistent with each other, thus adding extra weight to our result. The tau(w) of surface water obtained from the two independent trajectories is 20 and 24 ps. In both simulations only three water molecules are bound to lysozyme for the entire length of the trajectories, in agreement with nuclear magnetic relaxation dispersion estimates. Locations other than those identified in the protein crystal are found to be possible for these long-residing water molecules. The dynamics of the hydration water molecules observed in our simulations implies that each water molecule visits a multitude of residues during the lifetime of its bound with the protein. The number of residues seen by a single water molecule increases with the time-scale of its residence time and, on average, is equal to one only for the water molecules with shorter residence time. Thus, tau(w) values obtained from inelastic neutron scattering and based on jump-diffusion models are likely not to account for the contribution of water molecules with longer residence time. Copyright 2001 Academic Press.

  20. Structural basis of protein oxidation resistance: a lysozyme study.

    Directory of Open Access Journals (Sweden)

    Marion Girod

    Full Text Available Accumulation of oxidative damage in proteins correlates with aging since it can cause irreversible and progressive degeneration of almost all cellular functions. Apparently, native protein structures have evolved intrinsic resistance to oxidation since perfectly folded proteins are, by large most robust. Here we explore the structural basis of protein resistance to radiation-induced oxidation using chicken egg white lysozyme in the native and misfolded form. We study the differential resistance to oxidative damage of six different parts of native and misfolded lysozyme by a targeted tandem/mass spectrometry approach of its tryptic fragments. The decay of the amount of each lysozyme fragment with increasing radiation dose is found to be a two steps process, characterized by a double exponential evolution of their amounts: the first one can be largely attributed to oxidation of specific amino acids, while the second one corresponds to further degradation of the protein. By correlating these results to the structural parameters computed from molecular dynamics (MD simulations, we find the protein parts with increased root-mean-square deviation (RMSD to be more susceptible to modifications. In addition, involvement of amino acid side-chains in hydrogen bonds has a protective effect against oxidation Increased exposure to solvent of individual amino acid side chains correlates with high susceptibility to oxidative and other modifications like side chain fragmentation. Generally, while none of the structural parameters alone can account for the fate of peptides during radiation, together they provide an insight into the relationship between protein structure and susceptibility to oxidation.

  1. Surface Immobilized His-tagged Azurin as a Model Interface for the Investigation of Vectorial Electron Transfer in Biological Systems

    International Nuclear Information System (INIS)

    Casalini, Stefano; Berto, Marcello; Kovtun, Alessandro; Operamolla, Alessandra; Di Rocco, Giulia; Facci, Paolo; Liscio, Andrea; Farinola, Gianluca M.; Borsari, Marco; Bortolotti, Carlo A.

    2015-01-01

    A model system for the electrochemical investigation of vectorial electron transfer in biological systems was designed, assembled and characterized. Gold electrodes, functionalized with a -OCH 3 terminated, aromatic self-assembled monolayer, were used as a substrate for the adsorption of variants of copper-containing, redox metalloprotein azurin. The engineered azurin bears a polyhistidine tag at its C-terminus. Thanks to the presence of the solvent exposed tag, which chelates Cu 2+ ions in solution, we introduced an exogenous redox centre. The different reduction potentials of the two redox centres and their positioning with respect to the surface are such that electron transfer from the exogenous copper centre and the electrode is mediated by the native azurin active site, closely paralleling electron transfer processes in naturally occurring multicentre metalloproteins.

  2. DENV gene of bacteriophage T4 codes for both pyrimidine dimer-DNA glycosylase and apyrimidinic endonuclease activities

    International Nuclear Information System (INIS)

    McMillan, S.; Edenberg, H.J.; Radany, E.H.; Friedberg, R.C.; Friedberg, E.C.

    1981-01-01

    Recent studies have shown that purified preparations of phage T4 UV DNA-incising activity (T4 UV endonuclease or endonuclease V of phase T4) contain a pyrimidine dimer-DNA glycosylase activity that catalyzes hydrolysis of the 5' glycosyl bond of dimerized pyrimidines in UV-irradiated DNA. Such enzyme preparations have also been shown to catalyze the hydrolysis of phosphodiester bonds in UV-irradiated DNA at a neutral pH, presumably reflecting the action of an apurinic/apyrimidinic endonuclease at the apyrimidinic sites created by the pyrimidine dimer-DNA glycosylase. In this study we found that preparations of T4 UV DNA-incising activity contained apurinic/apyrimidinic endonuclease activity that nicked depurinated form I simian virus 40 DNA. Apurinic/apyrimidinic endonuclease activity was also found in extracts of Escherichia coli infected with T4 denV + phage. Extracts of cells infected with T4 denV mutants contained significantly lower levels of apurinic/apyrimidinic endonuclease activity; these levels were no greater than the levels present in extracts of uninfected cells. Furthermore, the addition of DNA containing UV-irradiated DNA and T4 enzyme resulted in competition for pyrimidine dimer-DNA glycosylase activity against the UV-irradiated DNA. On the basis of these results, we concluded that apurinic/apyrimidinic endonuclease activity is encoded by the denV gene of phage T4, the same gene that codes for pyrimidine dimer-DNA glycosylase activity

  3. Genome analysis of phage JS98 defines a fourth major subgroup of T4-like phages in Escherichia coli.

    Science.gov (United States)

    Zuber, Sophie; Ngom-Bru, Catherine; Barretto, Caroline; Bruttin, Anne; Brüssow, Harald; Denou, Emmanuel

    2007-11-01

    Numerous T4-like Escherichia coli phages were isolated from human stool and environmental wastewater samples in Bangladesh and Switzerland. The sequences of the major head gene (g23) revealed that these coliphages could be placed into four subgroups, represented by the phages T4, RB69, RB49, and JS98. Thus, JS98 defines a new major subgroup of E. coli T4-like phages. We conducted an analysis of the 169-kb JS98 genome sequence. Overall, 198 of the 266 JS98 open reading frames (ORFs) shared amino acid sequence identity with the reference T4 phage, 41 shared identity with other T4-like phages, and 27 ORFs lacked any database matches. Genes on the plus strand encoded virion proteins, which showed moderate to high sequence identity with T4 proteins. The right genome half of JS98 showed a higher degree of sequence conservation with T4 and RB69, even for the nonstructural genes, than did the left genome half, containing exclusively nonstructural genes. Most of the JS98-specific genes were found in the left genome half. Two came as a hypervariability cluster, but most represented isolated genes, suggesting that they were acquired separately in multiple acquisition events. No evidence for DNA exchange between JS98 phage and the E. coli host genome or coliphages other than T4 was observed. No undesired genes which could compromise its medical use were detected in the JS98 genome sequence.

  4. Control of solvent evaporation in hen egg white lysozyme crystallization

    Science.gov (United States)

    Wilson, L. J.; Suddath, F. L.

    1992-01-01

    An investigation of the role of solvent evaporation in tetragonal lysozyme crystallization was preformed with a device that employs N2(g) to control the evaporation of solvent from a micro-volume crystallization hanging drop. The number of crystals was found to vary with the rate at which the final supersaturation level was achieved. It was found that the more rapid the approach to supersaturation the larger the number of crystals. Accordingly, the crystals reached a smaller terminal size. Elongation of the (110) face parallel to the four-fold axis was observed with the slower evaporation rates.

  5. Observance of polymorphic behaviour during dissolution of insulin and lysozyme

    Directory of Open Access Journals (Sweden)

    A. Bernardo

    2005-09-01

    Full Text Available Although protein crystallization is a unit operation with potentially high separation factors, it has not been widely used in industry. Protein crystallization studies and practices have hitherto been largely limited to crystallography protocols. Knowledge of the behaviour of protein in solution would help to overcome empiric limitations in protein crystallisation. Thus, dissolution of porcine insulin and hen egg white lysozyme was studied and an unusual variation in solute concentration, with a concentration peak for short dissolution times, was verified. Polymorphic behaviour of protein in solution was observed, which altered physical properties such as solubility.

  6. Fourier transform infrared studies in solid egg white lysozyme

    International Nuclear Information System (INIS)

    Rivzi, T.Z.

    1994-12-01

    Fourier Transform Infrared (FTIR) Spectroscopy is the most recent addition to the arsenal of bioanalytical techniques capable of providing information about the secondary structure of proteins in a variety of environments. FTIR spectra have been obtained in solid egg white lysozyme. The spectra display the usual amide I, II and III bands. Secondary structural information obtained from the spectra after applying resolution enhancement techniques to the amide I band has been found consistent with the x-ray crystallographic data of the protein and also to the spectroscopic data of the protein in aqueous solution. (author). 17 refs, 6 figs, 2 tabs

  7. Identification and characterization of a highly thermostable bacteriophage lysozyme

    OpenAIRE

    Lavigne, R; Briers, Y; Hertveldt, K; ROBBEN, Johan; Volckaert, G

    2004-01-01

    Pseudomonas aeruginosa bacteriophage phiKMV is a T7-like lytic phage. Liquid chromatography-mass spectrometry of the structural proteins revealed gene product 36 (gp36) as part of the phiKMV phage particle. The presence of a lysozyme domain in the C terminal of this protein (gp36C) was verified by turbidimetric assays on chloroform-treated P. aeruginosa PAO1 and Escherichia coli WK6 cells. The molecular mass (20,884 Da) and pI (6.4) of recombinant gp36C were determined, as were the optimal en...

  8. The effect of mineral substrates on the crystallization of lysozyme

    Science.gov (United States)

    Kimble, W. L.; Paxton, T. E.; Rousseau, R. W.; Sambanis, A.

    1998-05-01

    The effects of exogenous mineral substrates on the induction time for nucleation, and on the number, morphology and purity of protein crystals were investigated in a series of experiments using chicken egg-white lysozyme (CEWL) as a model protein. CEWL was crystallized using the vapor-diffusion technique in the absence of substrates (control) and in the presence of mineral substrates exhibiting various degrees of crystalline lattice match to CEWL (experiments). Results indicate that mineral substrates with a close lattice match to CEWL had a greater influence on the induction time for nucleation and crystal properties.

  9. Beneficial effects of increased lysozyme levels in Alzheimer's disease modelled in Drosophila melanogaster.

    Science.gov (United States)

    Sandin, Linnea; Bergkvist, Liza; Nath, Sangeeta; Kielkopf, Claudia; Janefjord, Camilla; Helmfors, Linda; Zetterberg, Henrik; Blennow, Kaj; Li, Hongyun; Nilsberth, Camilla; Garner, Brett; Brorsson, Ann-Christin; Kågedal, Katarina

    2016-10-01

    Genetic polymorphisms of immune genes that associate with higher risk to develop Alzheimer's disease (AD) have led to an increased research interest on the involvement of the immune system in AD pathogenesis. A link between amyloid pathology and immune gene expression was suggested in a genome-wide gene expression study of transgenic amyloid mouse models. In this study, the gene expression of lysozyme, a major player in the innate immune system, was found to be increased in a comparable pattern as the amyloid pathology developed in transgenic mouse models of AD. A similar pattern was seen at protein levels of lysozyme in human AD brain and CSF, but this lysozyme pattern was not seen in a tau transgenic mouse model. Lysozyme was demonstrated to be beneficial for different Drosophila melanogaster models of AD. In flies that expressed Aβ 1-42 or AβPP together with BACE1 in the eyes, the rough eye phenotype indicative of toxicity was completely rescued by coexpression of lysozyme. In Drosophila flies bearing the Aβ 1-42 variant with the Arctic gene mutation, lysozyme increased the fly survival and decreased locomotor dysfunction dose dependently. An interaction between lysozyme and Aβ 1-42 in the Drosophila eye was discovered. We propose that the increased levels of lysozyme, seen in mouse models of AD and in human AD cases, were triggered by Aβ 1-42 and caused a beneficial effect by binding of lysozyme to toxic species of Aβ 1-42 , which prevented these from exerting their toxic effects. These results emphasize the possibility of lysozyme as biomarker and therapeutic target for AD. © 2016 The Authors. The FEBS Journal published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.

  10. Lysozyme in water-acetonitrile mixtures: Preferential solvation at the inner edge of excess hydration

    Science.gov (United States)

    Sirotkin, Vladimir A.; Kuchierskaya, Alexandra A.

    2017-06-01

    Preferential solvation/hydration is an effective way for regulating the mechanism of the protein destabilization/stabilization. Organic solvent/water sorption and residual enzyme activity measurements were performed to monitor the preferential solvation/hydration of hen egg-white lysozyme at high and low water content in acetonitrile at 25 °C. The obtained results show that the protein destabilization/stabilization depends essentially on the initial hydration level of lysozyme and the water content in acetonitrile. There are three composition regimes for the dried lysozyme. At high water content, the lysozyme has a higher affinity for water than for acetonitrile. The residual enzyme activity values are close to 100%. At the intermediate water content, the dehydrated lysozyme has a higher affinity for acetonitrile than for water. A minimum on the residual enzyme activity curve was observed in this concentration range. At the lowest water content, the organic solvent molecules are preferentially excluded from the dried lysozyme, resulting in the preferential hydration. The residual catalytic activity is ˜80%, compared with that observed after incubation in pure water. Two distinct schemes are operative for the hydrated lysozyme. At high and intermediate water content, lysozyme is preferentially hydrated. However, in contrast to the dried protein, at the intermediate water content, the initially hydrated lysozyme has the increased preferential hydration parameters. At low water content, the preferential binding of the acetonitrile molecules to the initially hydrated lysozyme was detected. No residual enzyme activity was observed in the water-poor acetonitrile. Our data clearly show that the initial hydration level of the protein macromolecules is one of the key factors that govern the stability of the protein-water-organic solvent systems.

  11. Hormonal evaluation of T4 and T3 through radioimmunoassay in younglings of rats with hypothyroidism

    International Nuclear Information System (INIS)

    Silveira, M.F.G.; Silva, I.M.S.; Pereira, S.S.L.; Souza, G.M.L.; Carvalho, E.F.M.B.; Cavalcante, C.V.G.; Lima Filho, G.L.; Catanho, M.T.J.A.

    2000-01-01

    The onset of fetal thyroid function occurs about 17-18 days after conception in the rat. The maternal hypothyroidism which occurs during gestation provokes alteration in the rat after birth. Due to this alteration, we decided to analyze the hormonal modification in the newborn rats. The hypothyroidism was induced in normal dams, which were being treated for 7 days with MMI (in the concentration of 0,03% in drinking water) before mating. Another dam group which was submitted to an induction of hypothyroidism maintained the treatment with MMI for 13 days during gestation. The hormones were assessed by radioimmunoassay technique. It was seen that the rats which were born from hypothyroid dams suffered alterations on its T 4 and T 3 hormone levels concerning to 10, 30 and 60 days after birth. There was also modifications on their weight and size. The growth is affected throughout post-natal life by thyroid hormones, which has a facilitator influence on growth hormone economy, as opposed to the inhibitory effects on TSH economy. The administration of MMI bars the fetal thyroid gland function, causing a decrease of both T 4 and T 3 levels, even after the birth, indicating that the maternal hypothyroidism influences on the post-natal life of the rat. (author)

  12. Acanthamoeba T4, T5 and T11 isolated from mineral water bottles in southern Brazil.

    Science.gov (United States)

    Maschio, Vinicius José; Chies, Fernanda; Carlesso, Ana Maris; Carvalho, Amanda; Rosa, Sayonara Peixoto; Van Der Sand, Sueli Teresinha; Rott, Marilise Brittes

    2015-01-01

    Acanthamoeba is a protist potential pathogen, capable of causing a blinding keratitis in contact lens wearers and disseminated infection, leading to granulomatous amebic encephalitis in immunocompromised individuals. This amoeba is a ubiquitous organism that has been isolated from various domestic water systems, such as cooling towers and hospital water networks. The objective of this work was to investigate the presence of Acanthamoeba in mineral water bottles marketed in Porto Alegre, southern Brazil. Positive samples were further classified at the genotype level after sequencing the ASA.S1 region of 18S rDNA gene. Six of the eight isolates belonged to T5 genotype, one to T4 genotype, and one was T11. Several genotypes have been reported worldwide as causative of pathologies in humans, including genotypes T4, T5 and T11. Overall, the widespread distribution of potentially pathogenic Acanthamoeba strains in the studied source demands more awareness within the public and health professionals, because this pathogen is emerging as a risk for human health worldwide.

  13. Isolation and genetic properties of a bacteriophage T4 uvs X mutant

    International Nuclear Information System (INIS)

    Childs, J.D.

    1980-01-01

    A technique for isolating UV-sensitive mutants of bacteriophage T4 and a relatively simple method for localizing the UV-sensitive mutations on the genetic map are described. Of 13 mutants isolated 5 were shown to be den V mutants and 1 was shown to be a uvs X mutant. Of the remainder, 4 are probably uvs X mutants (based on their map location) while the indentity of the remaining 3 has not been determined. The uvs X mutant (uvsX102) had enhanced UV and γ-ray sensitivity compared to the only other well characterized mutant in this gene, uvsX1 (T4x). Both uvsX1 and uvsX102 were more UV-sensitivity when plated on the su - E. coli B hosts, B and S/6, compared to the su + K12 host CR63, and both reduced the frequency of recombination to the same extent (2-3 fold). The uvsX gene is located between gene 41 and βgt. (orig.)

  14. The effect of bacteriophages T4 and HAP1 on in vitro melanoma migration

    Directory of Open Access Journals (Sweden)

    Boratyński Janusz

    2009-01-01

    Full Text Available Abstract Background The antibacterial activity of bacteriophages has been described rather well. However, knowledge about the direct interactions of bacteriophages with mammalian organisms and their other, i.e. non-antibacterial, activities in mammalian systems is quite scarce. It must be emphasised that bacteriophages are natural parasites of bacteria, which in turn are parasites or symbionts of mammals (including humans. Bacteriophages are constantly present in mammalian bodies and the environment in great amounts. On the other hand, the perspective of the possible use of bacteriophage preparations for antibacterial therapies in cancer patients generates a substantial need to investigate the effects of phages on cancer processes. Results In these studies the migration of human and mouse melanoma on fibronectin was inhibited by purified T4 and HAP1 bacteriophage preparations. The migration of human melanoma was also inhibited by the HAP1 phage preparation on matrigel. No response of either melanoma cell line to lipopolysaccharide was observed. Therefore the effect of the phage preparations cannot be attributed to lipopolysaccharide. No differences in the effects of T4 and HAP1 on melanoma migration were observed. Conclusion We believe that these observations are of importance for any further attempts to use bacteriophage preparations in antibacterial treatment. The risk of antibiotic-resistant hospital infections strongly affects cancer patients and these results suggest the possibility of beneficial phage treatment. We also believe that they will contribute to the general understanding of bacteriophage biology, as bacteriophages, extremely ubiquitous entities, are in permanent contact with human organisms.

  15. The effect of bacteriophages T4 and HAP1 on in vitro melanoma migration.

    Science.gov (United States)

    Dabrowska, Krystyna; Skaradziński, Grzegorz; Jończyk, Paulina; Kurzepa, Aneta; Wietrzyk, Joanna; Owczarek, Barbara; Zaczek, Maciej; Switała-Jeleń, Kinga; Boratyński, Janusz; Poźniak, Gryzelda; Maciejewska, Magdalena; Górski, Andrzej

    2009-01-20

    The antibacterial activity of bacteriophages has been described rather well. However, knowledge about the direct interactions of bacteriophages with mammalian organisms and their other, i.e. non-antibacterial, activities in mammalian systems is quite scarce. It must be emphasised that bacteriophages are natural parasites of bacteria, which in turn are parasites or symbionts of mammals (including humans). Bacteriophages are constantly present in mammalian bodies and the environment in great amounts. On the other hand, the perspective of the possible use of bacteriophage preparations for antibacterial therapies in cancer patients generates a substantial need to investigate the effects of phages on cancer processes. In these studies the migration of human and mouse melanoma on fibronectin was inhibited by purified T4 and HAP1 bacteriophage preparations. The migration of human melanoma was also inhibited by the HAP1 phage preparation on matrigel. No response of either melanoma cell line to lipopolysaccharide was observed. Therefore the effect of the phage preparations cannot be attributed to lipopolysaccharide. No differences in the effects of T4 and HAP1 on melanoma migration were observed. We believe that these observations are of importance for any further attempts to use bacteriophage preparations in antibacterial treatment. The risk of antibiotic-resistant hospital infections strongly affects cancer patients and these results suggest the possibility of beneficial phage treatment. We also believe that they will contribute to the general understanding of bacteriophage biology, as bacteriophages, extremely ubiquitous entities, are in permanent contact with human organisms.

  16. Mini Transsternal Approach to the Anterior High Thoracic Spine (T1–T4 Vertebrae

    Directory of Open Access Journals (Sweden)

    Christian Brogna

    2016-01-01

    Full Text Available Purpose. The anterior high thoracic spine is one of the most complex segments to be accessed surgically due to anatomical constraints and transitional characteristics. We describe in detail the mini transsternal approach to metastatic, infective, traumatic, and degenerative pathologies of T1 to T4 vertebral bodies. We analyse our surgical series, indications, and outcomes. Methods. Over a 5-year period 18 consecutive patients with thoracic myelopathy due to metastatic, infective, traumatic, and degenerative pathologies with T1 to T4 vertebral bodies involvement received a mini transsternal approach with intraoperative monitoring. Frankel scoring system was used to grade the neurological status. Results. Mean follow-up was 40 months. 78% patients improved in Frankel grade after surgery and 22% patients remained unchanged. Average operation time was 210 minutes. There were no intraoperative complications. One patient developed postoperative pneumonia successfully treated with antibiotics. Conclusion. The mini transsternal is a safe approach for infective, metastatic, traumatic, and degenerative lesions affecting the anterior high thoracic spine and the only one allowing an early and direct visualisation of the anterior theca. This approach overcomes the anatomical constraints of this region and provides adequate room for optimal reconstruction and preservation of spinal alignment in the cervicothoracic transition zone with good functional patient outcomes.

  17. Efficacy and safety of treating T4 oral cavity tumors with primary chemoradiotherapy.

    Science.gov (United States)

    Cohen, Ezra E W; Baru, Joshua; Huo, Dezheng; Haraf, Daniel J; Crowley, Maureen; Witt, Mary Ellyn; Blair, Elizabeth A; Weichselbaum, Ralph R; Rosen, Fred; Vokes, Everett E; Stenson, Kerstin

    2009-08-01

    Patients with T4 oral cavity (OC) tumors are often treated with surgery followed by adjuvant chemoradiotherapy (CRT). We performed a retrospective review of 4 multi-institutional phase II studies estimating long-term toxicity, locoregional control (LC), progression-free survival (PFS), and overall survival (OS) of primary CRT. Thirty-nine subjects were identified; 16 (42%) with bony involvement. Median radiotherapy dose delivered to primary tumor was 74 Gy. Five-year OS, PFS, and LC rates were 56%, 51%, and 75%, respectively. Sixty-nine percent of subjects with bony involvement never relapsed. Seven subjects developed osteoradionecrosis. Bone involvement with primary tumor did not appear to be associated with increased risk of death, relapse, or long-term complication. These data suggest that primary CRT is an effective treatment approach in patients with T4 OC tumors including those with bony involvement producing LC, survival, and complication rates comparable to historical series. Prospective clinical trials should evaluate primary surgical versus CRT treatment in these patients. Copyright 2009 Wiley Periodicals, Inc.

  18. Acanthamoeba genotypes T3 and T4 as causative agents of amoebic keratitis in Mexico.

    Science.gov (United States)

    Omaña-Molina, Maritza; Vanzzini-Zago, Virginia; Hernandez-Martinez, Dolores; Gonzalez-Robles, Arturo; Salazar-Villatoro, Lizbeth; Ramirez-Flores, Elizabeth; Oregon-Miranda, Eric; Lorenzo-Morales, Jacob; Martinez-Palomo, Adolfo

    2016-02-01

    Free-living amoebae (FLA) are widely distributed worldwide. Some genera included in this group act as opportunistic pathogens causing fatal encephalitis and Acanthamoeba keratitis (AK), a sight-threatening infection of the cornea associated with the use of soft contact lenses that could even end in blindness if an early diagnosis and treatment are not achieved. Furthermore, the numbers of AK cases keep rising worldwide mainly due to an increase of contact lens wearers and lack of hygiene in the maintenance of lenses and their cases. In Mexico, no cases of AK have been described so far although the isolation of other pathogenic FLA such as Naegleria fowleri and Balamuthia mandrillaris from both clinical and environmental sources has been reported. The present study reports two cases of Acanthamoeba keratitis diagnosed in two patients admitted to the Hospital "Luis Sánchez Bulnes" for Blindness Prevention in Mexico City, Mexico. Corneal scrapes and contact lenses were checked for the presence of Acanthamoeba strains in both patients. Strains were axenized after initial isolation to classify at the genotype level. After sequencing the diagnostic fragment 3 (DF3) region located on the 18S ribosomal DNA (rDNA) gene of Acanthamoeba, genotype T3 and genotype T4 were identified in clinical case 1 and 2, respectively. To our knowledge, these are the first reported cases of AK in Mexico in the literature and the first description of Acanthamoeba genotypes T3 and T4 as causative agents of amoebic infection.

  19. Lysozyme's lectin-like characteristics facilitates its immune defense function

    KAUST Repository

    Zhang, Ruiyan

    2017-06-06

    Interactions between human lysozyme (HL) and the lipopolysaccharide (LPS) of Klebsiella pneumoniae O1, a causative agent of lung infection, were identified by surface plasmon resonance. To characterize the molecular mechanism of this interaction, HL binding to synthetic disaccharides and tetrasaccharides representing one and two repeating units, respectively, of the O-chain of this LPS were studied. pH-dependent structural rearrangements of HL after interaction with the disaccharide were observed through nuclear magnetic resonance. The crystal structure of the HL-tetrasaccharide complex revealed carbohydrate chain packing into the A, B, C, and D binding sites of HL, which primarily occurred through residue-specific, direct or water-mediated hydrogen bonds and hydrophobic contacts. Overall, these results support a crucial role of the Glu35/Asp53/Trp63/Asp102 residues in HL binding to the tetrasaccharide. These observations suggest an unknown glycan-guided mechanism that underlies recognition of the bacterial cell wall by lysozyme and may complement the HL immune defense function.

  20. A new membrane-based crystallization technique: tests on lysozyme

    Science.gov (United States)

    Curcio, Efrem; Profio, Gianluca Di; Drioli, Enrico

    2003-01-01

    The great importance of protein science both in industrial and scientific fields, in conjunction with the intrinsic difficulty to grow macromolecular crystals, stimulates the development of new observations and ideas that can be useful in initiating more systematic studies using novel approaches. In this regard, an innovative technique, based on the employment of microporous hydrophobic membranes in order to promote the formation of lysozyme crystals from supersaturated solutions, is introduced in this work. Operational principles and possible advantages, both in terms of controlled extraction of solvent by acting on the concentration of the stripping solution and reduced induction times, are outlined. Theoretical developments and experimental results concerning the mass transfer, in vapour phase, through the membrane are presented, as well as the results from X-ray diffraction to 1.7 Å resolution of obtained lysozyme crystals using NaCl as the crystallizing agent and sodium acetate as the buffer. Crystals were found to be tetragonal with unit cell dimensions of a= b=79.1 Å and c=37.9 Å; the overall Rmerge on intensities in the resolution range from 25 to 1.7 Å was, in the best case, 4.4%.

  1. Terahertz mechanical vibrations in lysozyme: Raman spectroscopy vs modal analysis

    Science.gov (United States)

    Carpinteri, Alberto; Lacidogna, Giuseppe; Piana, Gianfranco; Bassani, Andrea

    2017-07-01

    The mechanical behaviour of proteins is receiving an increasing attention from the scientific community. Recently it has been suggested that mechanical vibrations play a crucial role in controlling structural configuration changes (folding) which govern proteins biological function. The mechanism behind protein folding is still not completely understood, and many efforts are being made to investigate this phenomenon. Complex molecular dynamics simulations and sophisticated experimental measurements are conducted to investigate protein dynamics and to perform protein structure predictions; however, these are two related, although quite distinct, approaches. Here we investigate mechanical vibrations of lysozyme by Raman spectroscopy and linear normal mode calculations (modal analysis). The input mechanical parameters to the numerical computations are taken from the literature. We first give an estimate of the order of magnitude of protein vibration frequencies by considering both classical wave mechanics and structural dynamics formulas. Afterwards, we perform modal analyses of some relevant chemical groups and of the full lysozyme protein. The numerical results are compared to experimental data, obtained from both in-house and literature Raman measurements. In particular, the attention is focused on a large peak at 0.84 THz (29.3 cm-1) in the Raman spectrum obtained analyzing a lyophilized powder sample.

  2. Genetically enhanced lysozyme evades a pathogen derived inhibitory protein.

    Science.gov (United States)

    Dostal, Sarah M; Fang, Yongliang; Guerrette, Jonathan C; Scanlon, Thomas C; Griswold, Karl E

    2015-04-17

    The accelerating spread of drug-resistant bacteria is creating demand for novel antibiotics. Bactericidal enzymes, such as human lysozyme (hLYZ), are interesting drug candidates due to their inherent catalytic nature and lack of susceptibility to the resistance mechanisms typically directed toward chemotherapeutics. However, natural antibacterial enzymes have their own limitations. For example, hLYZ is susceptible to pathogen derived inhibitory proteins, such as Escherichia coli Ivy. Here, we describe proof of concept studies demonstrating that hLYZ can be effectively redesigned to evade this potent lysozyme inhibitor. Large combinatorial libraries of hLYZ were analyzed using an innovative screening platform based on microbial coculture in hydrogel microdroplets. Isolated hLYZ variants were orders of magnitude less susceptible to E. coli Ivy yet retained high catalytic proficiency and inherent antibacterial activity. Interestingly, the engineered escape variants showed a disadvantageous increase in susceptibility to the related Ivy ortholog from Pseudomonas aeruginosa as well as an unrelated E. coli inhibitory protein, MliC. Thus, while we have achieved our original objective with respect to escaping E. coli Ivy, engineering hLYZ for broad-spectrum evasion of proteinaceous inhibitors will require consideration of the complex and varied determinants that underlie molecular recognition by these emerging virulence factors.

  3. The Effect of Protein Impurities on Lysozyme Crystal Growth

    Science.gov (United States)

    Judge, Russell A.; Forsythe, Elizabeth L.; Pusey, Marc L.

    1998-01-01

    While bulk crystallization from impure solutions is used industrially as a purification step for a wide variety of materials, it is a technique that has rarely been used for proteins. Proteins have a reputation for being difficult to crystallize and high purity of the initial crystallization solution is considered paramount for success in the crystallization. Although little is written on the purifying capability of protein crystallization or of the effect of impurities on the various aspects of the crystallization process, recent published reports show that crystallization shows promise and feasibility as a purification technique for proteins. In order to further examine the issue of purity in macromolecule crystallization this study investigates the effect of the protein impurities, avidin, ovalbumin and conalbumin, at concentrations up to 50%, on the solubility, crystal face growth rates and crystal purity, of the protein lysozyme. Solubility was measured in batch experiments while a computer controlled video microscope system was used to measure the f {101} and {101} lysozyme crystal face growth rates. While little effect was observed on solubility and high crystal purity was obtained (>99.99%), the effect of the impurities on the face growth rates varied from no effect to a significant face specific effect leading to growth cessation, a phenomenon that is frequently observed in protein crystal growth. The results shed interesting light on the effect of protein impurities on protein crystal growth and strengthen the feasibility of using crystallization as a unit operation for protein purification.

  4. Bacteriophage T4 Infection of Stationary Phase E. coli: Life after Log from a Phage Perspective.

    Science.gov (United States)

    Bryan, Daniel; El-Shibiny, Ayman; Hobbs, Zack; Porter, Jillian; Kutter, Elizabeth M

    2016-01-01

    Virtually all studies of phage infections investigate bacteria growing exponentially in rich media. In nature, however, phages largely encounter non-growing cells. Bacteria entering stationary phase often activate well-studied stress defense mechanisms that drastically alter the cell, facilitating its long-term survival. An understanding of phage-host interactions in such conditions is of major importance from both an ecological and therapeutic standpoint. Here, we show that bacteriophage T4 can efficiently bind to, infect and kill E. coli in stationary phase, both in the presence and absence of a functional stationary-phase sigma factor, and explore the response of T4-infected stationary phase cells to the addition of fresh nutrients 5 or 24 h after that infection. An unexpected new mode of response has been identified. "Hibernation" mode is a persistent but reversible dormant state in which the infected cells make at least some phage enzymes, but halt phage development until appropriate nutrients become available before producing phage particles. Our evidence indicates that the block in hibernation mode occurs after the middle-mode stage of phage development; host DNA breakdown and the incorporation of the released nucleotides into phage DNA indicate that the enzymes of the nucleotide synthesizing complex, under middle-mode control, have been made and assembled into a functional state. Once fresh glucose and amino acids become available, the standard lytic infection process rapidly resumes and concentrations of up to 10(11) progeny phage (an average of about 40 phage per initially present cell) are produced. All evidence is consistent with the hibernation-mode control point lying between middle mode and late mode T4 gene expression. We have also observed a "scavenger" response, where the infecting phage takes advantage of whatever few nutrients are available to produce small quantities of progeny within 2 to 5 h after infection. The scavenger response seems able

  5. Bacteriophage T4 Infection of Stationary Phase E. coli: Life after Log from a Phage Perspective

    Directory of Open Access Journals (Sweden)

    Elizabeth Martin Kutter

    2016-09-01

    Full Text Available Virtually all studies of phage infections investigate bacteria growing exponentially in rich media. In nature, however, phages largely encounter non-growing cells. Bacteria entering stationary phase often activate well-studied stress defense mechanisms that drastically alter the cell, facilitating its long-term survival. An understanding of phage-host interactions in such conditions is of major importance from both an ecological and therapeutic standpoint. Here, we show that bacteriophage T4 can efficiently bind to, infect and kill E. coli in stationary phase, both in the presence and absence of a functional stationary-phase sigma factor, and explore the response of T4-infected stationary phase cells to the addition of fresh nutrients 5 or 24 hours after that infection. An unexpected new mode of response has been identified. Hibernation mode is a persistent but reversible dormant state in which the infected cells make at least some phage enzymes, but halt phage development until appropriate nutrients become available before producing phage particles. Our evidence indicates that the block in hibernation mode occurs after the middle-mode stage of phage development; host DNA breakdown and the incorporation of the released nucleotides into phage DNA indicate that the enzymes of the nucleotide synthesizing complex, under middle-mode control, have been made and assembled into a functional state. Once fresh glucose and amino acids become available, the standard lytic infection process rapidly resumes and concentrations of up to 1011 progeny phage (an average of about 40 phage per initially-present cell are produced. All evidence is consistent with the hibernation-mode control point lying between middle mode and late mode T4 gene expression. We have also observed a scavenger response, where the infecting phage takes advantage of whatever few nutrients are available to produce small quantities of progeny within 2 to 5 hours after infection. The scavenger

  6. Modelling T4 cell count as a marker of HIV progression in the absence of any defence mechanism

    Directory of Open Access Journals (Sweden)

    VSS Yadavalli

    2010-12-01

    Full Text Available The T4 cell count, which is considered one of the markers of disease progression in an HIV infected individual, is modelled in this paper. The World Health Organisation has recently advocated that countries encourage HIV infected individuals to commence antiretroviral treatments once their T4 cell count drops below 350 cells per ml of blood (this threshold was formerly 200 cells per ml of blood. This recommendation is made because when the T4 cell count is low, the T4 cells are unable to mount an effective immune response against antigens and any such foreign matters in the body, and consequently the individual becomes susceptible to opportunistic infections and lymphomas. A stochastic catastrophe model is developed in this paper to obtain the mean, variance and covariance of the uninfected, infected and lysed T4 cells. The amount of toxin produced in an HIV infected person from the time of infection to a later time may also be obtained from the model. Numerical illustrations of the correlation structures between uninfected and infected T4 cells, and between the infected and lysed T4 cells are also presented.

  7. Virtual unit delay for digital frequency adaptive T/4 delay phase-locked loop system

    DEFF Research Database (Denmark)

    Yang, Yongheng; Zhou, Keliang; Blaabjerg, Frede

    2016-01-01

    /processor with a fixed sampling rate considering the cost and complexity, where the number of unit delays that have been adopted should be an integer. For instance, in conventional digital control systems, a single-phase T/4 Delay Phase-Locked Loop (PLL) system takes 50 unit delays (i.e., in a 50-Hz system...... Delay PLL system should be done in its implementation. This process will result in performance degradation in the digital control system, as the exactly required number of delays is not realized. Hence, in this paper, a Virtual Unit Delay (VUD) has been proposed to address such challenges to the digital......Digital micro-controllers/processors enable the cost-effective control of grid-connected power converter systems in terms of system monitoring, signal processing (e.g., grid synchronization), control (e.g., grid current and voltage control), etc. Normally, the control is implemented in a micro-controller...

  8. The nucleotide sequence of threonine transfer RNA coded by bacteriophage T4

    International Nuclear Information System (INIS)

    Guthrie, C.; Scholla, C.A.; Yesian, H.; Abelson, J.

    1978-01-01

    The nucleotide sequence of a low molecular weight RNA coded by bacteriophage T4 (and previously identified as species α) has been determined. The molecule is of particular biological interest for its associated biosynthetic properties. This RNA is 76 nucleotides in length, contains eight modified bases, and can be arranged in a cloverleaf configuration common to tRNAs. The anticodon sequence is UGU, which corresponds to the threonine-specific codons ACsub(G)sup(A). The nucleotide sequence was determined primarily by nearest-neighbour analysis of RNA synthesized in vitro using [α- 32 P] nucleoside triphosphates. Using the single-strand specific nuclease S1, two in vivo labelled half-molecules were generated and analysed. This information together with restrictions imposed by nearest-neighbour data, provided a unique linear sequence of nucleotides with the features of secondary structure common to tRNA molecules. (author)

  9. Production of antisera and development of radioimmunoassay for serum T3, T4, and ferritin

    International Nuclear Information System (INIS)

    Elhag, Omer Mohamed Abdalla

    1998-05-01

    In this study twelve local rabbits and sixteen New-zealand rabbits were subjected to immunization against T3 and T4 immunogens. Two local sheep (ovis aris) were immunized against human liver ferritin. The T3 and T4 immunogens were prepared by conjugation of the haptens to carrier proteins (bovine serum albumin ''BSA'' and horse serum protein ''HSP''), using water soluble carboiimide as coupling agent. The local and New-zealand rabbits were immunized against these conjugates emulsified in freund's complete adjuvant (FCA) in the first and second injections, and emulsified in freund's incomplete adjuvant (FIA) in the following injections. The blood samples obtained from rabbits after each injection were tested for antibodies as well as for the effect of immunization on rabbits biochemical and haematological parameters. The blood samples obtained from sheep were tested for anti-ferritin antibodies using crude antiserum, then this antiserum was purified using ammonium sulphate. A part of it was adsorbed physically onto polystyrene beads while the other part was linked chemically to magnitisable particles inorder to develop to IRMAs. The purified antiferritin antibody was diluted 200,000 folds before being coated to polystyrene beads, and different dilutions were tried with coupling to magnetic solid phase. Optimization and validation procedures for the two IRMAs ferritin were performed. The results obtained showed poor response of rabbits to immunization against T3 and T4 immunogen conjugates, where the percent bound (B%) of tracer with the antibody ranged from (0.0-22%) for local rabbits using charcoal seperation technique, and (0.0-2.9%) using second antibody precipitation technique. The B% for the antiserum obtained from New-zealand rabbits ranged from (0.0-18.1) using second antibody precipitation technique. Serum T3, T4 and TSH of the immunized rabbits were measured and found to be not significantly different form the controls (p=0.2211, 0.098, 0.35 respectively

  10. Pollution reduction technology program for class T4(JT8D) engines

    Science.gov (United States)

    Roberts, R.; Fiorentino, A. J.; Diehl, L. A.

    1977-01-01

    The technology required to develop commercial gas turbine engines with reduced exhaust emissions was demonstrated. Can-annular combustor systems for the JT8D engine family (EPA class T4) were investigated. The JT8D turbofan engine is an axial-flow, dual-spool, moderate-bypass-ratio design. It has a two-stage fan, a four-stage low-pressure compressor driven by a three-stage low-pressure turbine, and a seven-stage high-pressure compressor driven by a single-stage high-pressure turbine. A cross section of the JT8D-17 showing the mechanical configuration is given. Key specifications for this engine are listed.

  11. Study of the non-covalent interactions of ginsenosides and lysozyme using electrospray ionization mass spectrometry.

    Science.gov (United States)

    Tang, Jun; Fu, Qiang; Cui, Meng; Xing, Junpeng; Liu, Zhiqiang; Liu, Shuying

    2015-11-15

    Ginsenosides are an important class of natural products extracted from ginseng that possess various important biological activities. Studies of interactions of ginsenosides with proteins are essential for comprehensive understanding of the biological activities of ginsenosides. In this study, the interactions of ginsenosides with lysozyme were investigated by electrospray ionization mass spectrometry (ESI-MS). Both protopanaxadiol-type and protopanaxatriol-type ginsenosides were chosen to explore the interactions of ginsenosides towards lysozyme near the physiological conditions by direct ESI-MS, respectively. Comparative experiments were conducted to confirm the interactions were specific. In addition, the dissociation constants of ginsenoside-lysozyme complexes were determined by a ESI-MS titration strategy. The results showed ginsenosides bound to lysozyme at the stoichiometries of 1:1 and 2:1. The association constants of ginsenosides to lysozyme were in the order of Re>Rd>Rf>Rg2 >Rg3 . According to their structures, the binding affinities associated with the type of aglycone and the type and the number of sugar moieties linked on the aglycone. It has been demonstrated that ESI-MS is a powerful tool to probe the non-covalent interactions between lysozyme and ginsenosides. These results provide insights into the interaction of ginsenosides with lysozyme at the molecular level. The developed strategy could be applied to determine the interactions of proteins with other natural products. Copyright © 2015 John Wiley & Sons, Ltd.

  12. Investigating the effects of erythrosine B on amyloid fibril formation derived from lysozyme.

    Science.gov (United States)

    Kuo, Chun-Tien; Chen, Yi-Lin; Hsu, Wei-Tse; How, Su-Chun; Cheng, Yu-Hong; Hsueh, Shu-Shun; Liu, Hwai-Shen; Lin, Ta-Hsien; Wu, Josephine W; Wang, Steven S-S

    2017-05-01

    Formation of amyloid fibrils has been associated with at least 30 different protein aggregation diseases. The 129-residue polypeptide hen lysozyme, which is structurally homologous to human lysozyme, has been demonstrated to exhibit amyloid fibril-forming propensity in vitro. This study is aimed at exploring the influence of erythrosine B on the in vitro amyloid fibril formation of hen lysozyme at pH 2.0 and 55°C using ThT binding assay, transmission electron microscopy, far-UV circular dichroism absorption spectroscopy, 1-anilinonaphthalene-8-sulfonic acid fluorescence spectroscopy, and synchronous fluorescence study. We found that lysozyme fibrillogenesis was dose-dependently suppressed by erythrosine B. In addition, our far-UV CD and ANS fluorescence data showed that, as compared with the untreated lysozyme control, the α-to-ß transition and exposure of hydrophobic clusters in lysozyme were reduced upon treatment with erythrosine B. Moreover, it could be inferred that the binding of erythrosine B occurred in the vicinity of the tryptophan residues. Finally, molecular docking and molecular dynamics simulations were further employed to gain some insights into the possible binding site(s) and interactions between lysozyme and erythrosine B. We believe the results obtained here may contribute to the development of potential strategies/approaches for the suppression of amyloid fibrillogenesis, which is implicated in amyloid pathology. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. Interaction and inhibitory influence of the azo dye carmoisine on lysozyme amyloid fibrillogenesis.

    Science.gov (United States)

    Basu, Anirban; Suresh Kumar, Gopinatha

    2017-07-25

    The binding of the common food colorant carmoisine and its inhibitory effect on amyloid fibrillation in lysozyme have been investigated. Since humans are increasingly exposed to various food colorants like carmoisine, such studies are highly relevant. In the presence of lysozyme, the carmoisine absorption spectrum exhibited hypochromic changes. The intrinsic fluorescence of lysozyme was also quenched on interaction. Time-resolved fluorescence results suggested that the binding mechanism involved ground state complexation. The binding was predominantly dominated by non-polyelectrolytic forces. The molecular distance between the donor (lysozyme) and the acceptor (carmoisine), calculated from FRET theory, was found to be 3.37 nm, indicating that carmoisine binds close to Trp-62/63 residues in the β-domain of the protein. Information on alterations in the microenvironment surrounding the Trp-residues was also obtained from synchronous fluorescence data. Carmoisine binding induced significant loss in the alpha helical organization of lysozyme. The binding, nevertheless, did not influence the thermal stability of lysozyme significantly. The binding reaction was exothermic and driven by large negative enthalpy and small but favourable entropic contributions. Thioflavin T assay, far-UV circular dichroism studies and AFM imaging profiles testified that carmoisine had a significant inhibitory effect on amyloid fibrillogenesis in lysozyme. Carmoisine also had a definitive defibrillating effect on existing fibrils. The results may provide new insights for designing new small molecule inhibitors for amyloid related diseases.

  14. Complex coacervates of hyaluronic acid and lysozyme: effect on protein structure and physical stability.

    Science.gov (United States)

    Water, Jorrit J; Schack, Malthe M; Velazquez-Campoy, Adrian; Maltesen, Morten J; van de Weert, Marco; Jorgensen, Lene

    2014-10-01

    Complex coacervates of hyaluronic acid and lysozyme, a model protein, were formed by ionic interaction using bulk mixing and were characterized in terms of binding stoichiometry and protein structure and stability. The complexes were formed at pH 7.2 at low ionic strength (6mM) and the binding stoichiometry was determined using solution depletion and isothermal titration calorimetry. The binding stoichiometry of lysozyme to hyaluronic acid (870 kDa) determined by solution depletion was found to be 225.9 ± 6.6 mol, or 0.1 bound lysozyme molecules per hyaluronic acid monomer. This corresponded well with that obtained by isothermal titration calorimetry of 0.09 bound lysozyme molecules per hyaluronic acid monomer. The complexation did not alter the secondary structure of lysozyme measured by Fourier-transform infrared spectroscopy overlap analysis and had no significant impact on the Tm of lysozyme determined by differential scanning calorimetry. Furthermore, the protein stability of lysozyme was found to be improved upon complexation during a 12-weeks storage study at room temperature, as shown by a significant increase in recovered protein when complexed (94 ± 2% and 102 ± 5% depending on the polymer-protein weight to weight ratio) compared to 89 ± 2% recovery for uncomplexed protein. This study shows the potential of hyaluronic acid to be used in combination with complex coacervation to increase the physical stability of pharmaceutical protein formulations. Copyright © 2014 Elsevier B.V. All rights reserved.

  15. Influence of graphene oxide and reduced graphene oxide on the activity and conformation of lysozyme.

    Science.gov (United States)

    Bai, Yitong; Ming, Zhu; Cao, Yuye; Feng, Shicheng; Yang, Hua; Chen, Lingyun; Yang, Sheng-Tao

    2017-06-01

    The dramatically different bio-effects of graphene and graphene oxide (GO) have been widely observed in diverse biological systems, which determine the applications and toxicity of graphene materials. To elucidate the mechanism at molecular level, it is urgent to investigate the enzyme-graphene interaction and its consequences. In this study, we comparatively studied the influence of GO and reduced GO (RGO) on the activity and conformation of lysozyme to provide better understandings of their different bio-effects. Both GO and RGO adsorbed large quantities of lysozyme after incubation. GO inhibited lysozyme activity seriously, while RGO nearly had no influence on the enzyme activity. The different inhibitions of enzyme activity could be explained by the lysozyme conformational changes, where GO induced more changes to the protein conformation according to UV-vis absorbance, far-UV circular dichroism spectra, intrinsic fluorescence quenching, and infrared spectra. Based on the spectroscopic changes of lysozyme, GO induced the loss of secondary structure and exposed the active site of lysozyme more to the aqueous environment. In addition, neither GO nor RGO induced the fibrillation of lysozyme after 12d incubation. The results collectively indicated that the oxidation degree significantly impacted the enzyme-graphene interaction. The implications to the designs of enzyme-graphene system for bio-related applications and the toxicological effects of graphene materials are discussed. Copyright © 2017 Elsevier B.V. All rights reserved.

  16. Effects of penicillin and lysozyme on the immunofluorescent and precipitin reactivity of group D streptococci.

    Science.gov (United States)

    Watson, B K; Moellering, R C; Kunz, L J

    1976-07-01

    Damage to the cell wall by growth in the presence of penicillin or by treatment with lysozyme enhanced the immunofluorescent (fluorescent antibody, FA) reactivity to group D streptococci. The optimum concentration and time of treatment with lysozyme varied inversely with the initial FA reactivity of the strain. Speciation of the organisms by a series of biochemical and physiologic tests suggested that the differences in initial FA reactivity were species-related. Thus, S. faecalis strains were the most FA-reactive and most sensitive to lysozyme. S. faecium strains were less FA-reactive and lysozyme-sensitive. S. bovis strains proved to be least FA-sensitive and were most resistant to lysozyme. Treatment with lysozyme was also effective in preparing extracts of group D antigen from all three species for Lancefield grouping by the precipitin test. The lysozyme extracts, moreover, produced much stronger reactions than those made from comparable volumes of cells by the methods of Lancefield or of Rantz and Randall.

  17. High-dose radiotherapy alone for patients with T4-stage laryngeal cancer

    Energy Technology Data Exchange (ETDEWEB)

    Mucha-Malecka, A. [Maria Sklodowska-Curie Memorial Institute, Krakow (Poland). Dept. of Radiation Oncology; Skladowski, K. [Maria Sklodowska-Curie Memorial Institute, Gliwice (Poland). Dept. of Radiation Oncology

    2013-08-15

    Background and purpose: The purpose of this retrospective study was to report on the efficacy of radiotherapy alone in patients with T4-stage laryngeal cancer and to establish the prognostic value of (a) the size and location of the extralaryngeal tumor extensions and (b) of emergency tracheostomy. Patients and methods: A group of 114 patients were treated with definitive radiotherapy between 1990 and 1996. The piriform recess was involved in 37 cases (33 %), the base of the tongue and glosso-epiglottic vallecula in 34 cases (30 %), and the hypopharyngeal wall in 10 cases (9 %). In 16 cases (14 %), emergency tracheostomy was performed before radiotherapy. The mean total dose was 68 Gy (range, 60-77.6 Gy). The mean treatment time was 49 days (range, 42-74 days). Results: Actuarial 3-year local control (LC) was noted in 42 % of patients, disease-free survival (DFS) in 35 %, and overall survival (OS) in 40 %. The best prognosis was for the lesion suspected of cartilage infiltration: 56 % 3-year LC. The worst results were noted in the cases with massive infiltrations spreading from the larynx through the hypopharynx: 13 % 3-year LC. Emergency tracheostomy before radiotherapy was significantly connected with the worst treatment results (p = 0.000): 3-year LC in patients with tracheostomy was 0 % vs. 48 % in patients without tracheostomy. Conclusion: Conventional radiotherapy of T4 laryngeal cancer is a method of treatment with limited effectiveness. The efficacy of radiotherapy is dependent on the location and extent of extralaryngeal infiltrations. Emergency tracheostomy is a prognostic factor connected with the worst prognosis. (orig.)

  18. High-dose radiotherapy alone for patients with T4-stage laryngeal cancer

    International Nuclear Information System (INIS)

    Mucha-Malecka, A.; Skladowski, K.

    2013-01-01

    Background and purpose: The purpose of this retrospective study was to report on the efficacy of radiotherapy alone in patients with T4-stage laryngeal cancer and to establish the prognostic value of (a) the size and location of the extralaryngeal tumor extensions and (b) of emergency tracheostomy. Patients and methods: A group of 114 patients were treated with definitive radiotherapy between 1990 and 1996. The piriform recess was involved in 37 cases (33 %), the base of the tongue and glosso-epiglottic vallecula in 34 cases (30 %), and the hypopharyngeal wall in 10 cases (9 %). In 16 cases (14 %), emergency tracheostomy was performed before radiotherapy. The mean total dose was 68 Gy (range, 60-77.6 Gy). The mean treatment time was 49 days (range, 42-74 days). Results: Actuarial 3-year local control (LC) was noted in 42 % of patients, disease-free survival (DFS) in 35 %, and overall survival (OS) in 40 %. The best prognosis was for the lesion suspected of cartilage infiltration: 56 % 3-year LC. The worst results were noted in the cases with massive infiltrations spreading from the larynx through the hypopharynx: 13 % 3-year LC. Emergency tracheostomy before radiotherapy was significantly connected with the worst treatment results (p = 0.000): 3-year LC in patients with tracheostomy was 0 % vs. 48 % in patients without tracheostomy. Conclusion: Conventional radiotherapy of T4 laryngeal cancer is a method of treatment with limited effectiveness. The efficacy of radiotherapy is dependent on the location and extent of extralaryngeal infiltrations. Emergency tracheostomy is a prognostic factor connected with the worst prognosis. (orig.)

  19. Consequences of tumor planning target volume reduction in treatment of T2-T4 laryngeal cancer.

    Science.gov (United States)

    Vugts, Cornelia A J M; Terhaard, Chris H J; Philippens, Marielle E P; Pameijer, Frank A; Kasperts, Nicolien; Raaijmakers, Cornelis P J

    2014-09-04

    Since lymph nodes volumes are generally four times the volume of the primary PTV, the advantage of using tight margins around the primary PTV is not clear. Therefore treatment margins of T2-T4 laryngeal carcinoma for IMRT are generally chosen in such a way that the PTV is comparable to that in conventional radiotherapy. The aim of this study is to quantify the effect of volume reduction of the primary PTV of T2-T4 laryngeal carcinoma with regard to late toxicity despite elective irradiation of lymph node levels II to IV. Two treatment plans based on conservative (GTV-PTV = 15 mm and 20 mm cranial), and on evidence-based tight margins (GTV-PTV = 8 mm) were calculated for 16 patients. Toxicity effects were estimated based on the dose distributions. Compared to conservative margins, using tight margins resulted in: 1) significant reduction of the normal tissue complication probability (NTCP) for swallowing muscles and submandibular glands, 2) significant reduction of the mean dose in all organs at risk (OAR), 3) a mean dose smaller than 60 Gy for all OARs except for the laryngeal cartilages. When the lymph node levels II to IV were prescribed with an elective dose, an NTCP reduction of 53% for the swallowing muscles and of 23% for the submandibular glands was found by using tight instead of conservative margins. When positive nodes were present, NTCP reduction amounted to 29% and 15%, respectively. There is a potential benefit in realizing evidence-based tight margins for laryngeal cancer patients despite elective irradiation of lymph node levels II to IV.

  20. Structure-Function Analysis of the DNA Translocating Portal of the Bacteriophage T4 Packaging Machine

    Science.gov (United States)

    Padilla-Sanchez, Victor; Gao, Song; Kim, Hyung Rae; Kihara, Daisuke; Sun, Lei; Rossmann, Michael G.; Rao, Venigalla B.

    2013-01-01

    Tailed bacteriophages and herpesviruses consist of a structurally well conserved dodecameric portal at a special five-fold vertex of the capsid. The portal plays critical roles in head assembly, genome packaging, neck/tail attachment, and genome ejection. Although the structures of portals from phages φ29, SPP1 and P22 have been determined, their mechanistic roles have not been well understood. Structural analysis of phage T4 portal (gp20) has been hampered because of its unusual interaction with the E. coli inner membrane. Here, we predict atomic models for the T4 portal monomer and dodecamer, and fit the dodecamer into the cryoEM density of the phage portal vertex. The core structure, like that from other phages, is cone-shaped with the wider end containing the “wing” and “crown” domains inside the phage head. A long “stem” encloses a central channel, and a narrow “stalk” protrudes outside the capsid. A biochemical approach was developed to analyze portal function by incorporating plasmid-expressed portal protein into phage heads and determining the effect of mutations on head assembly, DNA translocation, and virion production. We found that the protruding loops of the stalk domain are involved in assembling the DNA packaging motor. A loop that connects the stalk to the channel might be required for communication between the motor and portal. The “tunnel” loops that project into the channel are essential for sealing the packaged head. These studies established that the portal is required throughout the DNA packaging process, with different domains participating at different stages of genome packaging. PMID:24126213

  1. Structure-function analysis of the DNA translocating portal of the bacteriophage T4 packaging machine.

    Science.gov (United States)

    Padilla-Sanchez, Victor; Gao, Song; Kim, Hyung Rae; Kihara, Daisuke; Sun, Lei; Rossmann, Michael G; Rao, Venigalla B

    2014-03-06

    Tailed bacteriophages and herpesviruses consist of a structurally well conserved dodecameric portal at a special 5-fold vertex of the capsid. The portal plays critical roles in head assembly, genome packaging, neck/tail attachment, and genome ejection. Although the structures of portals from phages φ29, SPP1, and P22 have been determined, their mechanistic roles have not been well understood. Structural analysis of phage T4 portal (gp20) has been hampered because of its unusual interaction with the Escherichia coli inner membrane. Here, we predict atomic models for the T4 portal monomer and dodecamer, and we fit the dodecamer into the cryo-electron microscopy density of the phage portal vertex. The core structure, like that from other phages, is cone shaped with the wider end containing the "wing" and "crown" domains inside the phage head. A long "stem" encloses a central channel, and a narrow "stalk" protrudes outside the capsid. A biochemical approach was developed to analyze portal function by incorporating plasmid-expressed portal protein into phage heads and determining the effect of mutations on head assembly, DNA translocation, and virion production. We found that the protruding loops of the stalk domain are involved in assembling the DNA packaging motor. A loop that connects the stalk to the channel might be required for communication between the motor and the portal. The "tunnel" loops that project into the channel are essential for sealing the packaged head. These studies established that the portal is required throughout the DNA packaging process, with different domains participating at different stages of genome packaging. © 2013.

  2. Nueva Área Terminal (NAT del aeropuerto de Madrid-Barajas T-4

    Directory of Open Access Journals (Sweden)

    Lamela, A.

    2006-03-01

    Full Text Available This paper deals with the New Terminal Area of the Barajas airport. Its first part indicates the participants in the design and construction, as well as some general data. In the second part, organised like a data chart, technical data, such as general information, construction aspects, parking area, data about the building itself and others, are expressed in a concise way. The third part analyzes the construction aspects of the NAT Building, related to the power efficiency. The paper ends with a note that the author of the project has considered of interest about the functionality of the T4. Drawings, details and photographs that show the work in their total magnitude are also included.El articulo que se presenta trata de la Nueva Área Terminal (NAT del aeropuerto de Barajas y está diferenciado en una primera parte, donde se indican los créditos del proyecto y de la construcción, así como algunos datos generales. En el punto 2 y organizado como ficha, aparecen de forma escueta los datos técnicos, tales como: información general, sobre la construcción, sobre los aparcamientos, sobre el edificio terminal y otras. En el punto 3 se analizan los aspectos de la edificación NAT, relacionados con la eficiencia energética y finaliza el artículo con una nota sobre la funcionalidad de la T4, que el autor del proyecto ha considerado de interés. Se incluyen planos detalles y fotografías que muestran la obra en su total magnitud.

  3. Functional Analysis of the Bacteriophage T4 Rad50 Homolog (gp46) Coiled-coil Domain.

    Science.gov (United States)

    Barfoot, Tasida; Herdendorf, Timothy J; Behning, Bryanna R; Stohr, Bradley A; Gao, Yang; Kreuzer, Kenneth N; Nelson, Scott W

    2015-09-25

    Rad50 and Mre11 form a complex involved in the detection and processing of DNA double strand breaks. Rad50 contains an anti-parallel coiled-coil with two absolutely conserved cysteine residues at its apex. These cysteine residues serve as a dimerization domain and bind a Zn(2+) cation in a tetrathiolate coordination complex known as the zinc-hook. Mutation of the zinc-hook in bacteriophage T4 is lethal, indicating the ability to bind Zn(2+) is critical for the functioning of the MR complex. In vitro, we found that complex formation between Rad50 and a peptide corresponding to the C-terminal domain of Mre11 enhances the ATPase activity of Rad50, supporting the hypothesis that the coiled-coil is a major conduit for communication between Mre11 and Rad50. We constructed mutations to perturb this domain in the bacteriophage T4 Rad50 homolog. Deletion of the Rad50 coiled-coil and zinc-hook eliminates Mre11 binding and ATPase activation but does not affect its basal activity. Mutation of the zinc-hook or disruption of the coiled-coil does not affect Mre11 or DNA binding, but their activation of Rad50 ATPase activity is abolished. Although these mutants excise a single nucleotide at a normal rate, they lack processivity and have reduced repetitive exonuclease rates. Restricting the mobility of the coiled-coil eliminates ATPase activation and repetitive exonuclease activity, but the ability to support single nucleotide excision is retained. These results suggest that the coiled-coiled domain adopts at least two conformations throughout the ATPase/nuclease cycle, with one conformation supporting enhanced ATPase activity and processivity and the other supporting nucleotide excision. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  4. Experimental infection of T4 Acanthamoeba genotype determines the pathogenic potential.

    Science.gov (United States)

    Alves, Daniella de Sousa Mendes Moreira; Moraes, Aline Silva; Alves, Luciano Moreira; Gurgel-Gonçalves, Rodrigo; Lino Junior, Ruy de Souza; Cuba-Cuba, César Augusto; Vinaud, Marina Clare

    2016-09-01

    T4 is the Acanthamoeba genotype most related to cases of granulomatous amoebic encephalitis (GAE) in immunocompromised patients and of keratitis in contact lens wearers. The determination of the pathogenic potential of Acanthamoeba clinical and environmental isolates using experimental models is extremely important to elucidate the capacity of free-living organisms to establish and cause disease in hosts. The aim of this study was to compare and evaluate the histopathology and culture between two different routes of experimental infection of T4 Acanthamoeba isolated from environmental and clinical source in mice (intracranial and intraperitoneal). Swiss isogenic healthy mice were inoculated with 10(4) trophozoites by intracranial (IC) and intraperitoneal (IP) routes and observed during 21 days. The brains from animals inoculated by the IC route were collected and from the animals of the IP inoculation group, the brains, livers, kidneys, spleens, and lungs were removed. The organs were prepared and appropriately divided to be evaluated with histopathology and culture. There was no significant difference between the inoculation routes in terms of isolates recovery (χ(2) = 0.09; p = 0.76). In the IC group, isolate recovery rate was significantly higher in histopathology than the one achieved by culture (χ(2) = 6.45; p Acanthamoeba in both routes. This work represents the first in vivo pathogenicity assay of primary isolation source in Central region of Brazil showing in vivo pathogenicity and hematogenous spread capacity of these protozoa, improving the knowledge on free-living amoebae isolates.

  5. Genome Analysis of Phage JS98 Defines a Fourth Major Subgroup of T4-Like Phages in Escherichia coli▿ †

    OpenAIRE

    Zuber, Sophie; Ngom-Bru, Catherine; Barretto, Caroline; Bruttin, Anne; Brüssow, Harald; Denou, Emmanuel

    2007-01-01

    Numerous T4-like Escherichia coli phages were isolated from human stool and environmental wastewater samples in Bangladesh and Switzerland. The sequences of the major head gene (g23) revealed that these coliphages could be placed into four subgroups, represented by the phages T4, RB69, RB49, and JS98. Thus, JS98 defines a new major subgroup of E. coli T4-like phages. We conducted an analysis of the 169-kb JS98 genome sequence. Overall, 198 of the 266 JS98 open reading frames (ORFs) shared ami...

  6. Control of Electrostatic Interactions Between F-Actin And Genetically Modified Lysozyme in Aqueous Media

    International Nuclear Information System (INIS)

    Sanders, L.K.; Xian, W.; Guaqueta, C.; Strohman, M.; Vrasich, C.R.; Luijten, E.; Wong, G.C.L.

    2009-01-01

    The aim for deterministic control of the interactions between macroions in aqueous media has motivated widespread experimental and theoretical work. Although it has been well established that like-charged macromolecules can aggregate under the influence of oppositely charged condensing agents, the specific conditions for the stability of such aggregates can only be determined empirically. We examine these conditions, which involve an interplay of electrostatic and osmotic effects, by using a well defined model system composed of F-actin, an anionic rod-like polyelectrolyte, and lysozyme, a cationic globular protein with a charge that can be genetically modified. The structure and stability of actin-lysozyme complexes for different lysozyme charge mutants and salt concentrations are examined by using synchrotron x-ray scattering and molecular dynamics simulations. We provide evidence that supports a structural transition from columnar arrangements of F-actin held together by arrays of lysozyme at the threefold interstitial sites of the actin sublattice to marginally stable complexes in which lysozyme resides at twofold bridging sites between actin. The reduced stability arises from strongly reduced partitioning of salt between the complex and the surrounding solution. Changes in the stability of actin-lysozyme complexes are of biomedical interest because their formation has been reported to contribute to the persistence of airway infections in cystic fibrosis by sequestering antimicrobials such as lysozyme. We present x-ray microscopy results that argue for the existence of actin-lysozyme complexes in cystic fibrosis sputum and demonstrate that, for a wide range of salt conditions, charge-reduced lysozyme is not sequestered in ordered complexes while retaining its bacterial killing activity.

  7. Control of electrostatic interactions between F-actin and genetically modified lysozyme in aqueous media

    International Nuclear Information System (INIS)

    Sanders, Lori K.; Xian, Wujing; Guaqueta, Camilo; Strohman, Michael J.; Vrasich, Chuck R.; Luijten, Erik; Wong, Gerard C.L.

    2007-01-01

    The aim for deterministic control of the interactions between macroions in aqueous media has motivated widespread experimental and theoretical work. Although it has been well established that like-charged macromolecules can aggregate under the influence of oppositely charged condensing agents, the specific conditions for the stability of such aggregates can only be determined empirically. We examine these conditions, which involve an interplay of electrostatic and osmotic effects, by using a well defined model system composed of F-actin, an anionic rod-like polyelectrolyte, and lysozyme, a cationic globular protein with a charge that can be genetically modified. The structure and stability of actin-lysozyme complexes for different lysozyme charge mutants and salt concentrations are examined by using synchrotron x-ray scattering and molecular dynamics simulations. We provide evidence that supports a structural transition from columnar arrangements of F-actin held together by arrays of lysozyme at the threefold interstitial sites of the actin sublattice to marginally stable complexes in which lysozyme resides at twofold bridging sites between actin. The reduced stability arises from strongly reduced partitioning of salt between the complex and the surrounding solution. Changes in the stability of actin-lysozyme complexes are of biomedical interest because their formation has been reported to contribute to the persistence of airway infections in cystic fibrosis by sequestering antimicrobials such as lysozyme. We present x-ray microscopy results that argue for the existence of actin-lysozyme complexes in cystic fibrosis sputum and demonstrate that, for a wide range of salt conditions, charge-reduced lysozyme is not sequestered in ordered complexes while retaining its bacterial killing activity.

  8. Studies on the role of insect hemolymph polypeptides: Galleria mellonella anionic peptide 2 and lysozyme.

    Science.gov (United States)

    Sowa-Jasiłek, Aneta; Zdybicka-Barabas, Agnieszka; Stączek, Sylwia; Wydrych, Jerzy; Mak, Paweł; Jakubowicz, Teresa; Cytryńska, Małgorzata

    2014-03-01

    The lysozymes are well known antimicrobial polypeptides exhibiting antibacterial and antifungal activities. Their antibacterial potential is related to muramidase activity and non-enzymatic activity resembling the mode of action of cationic defense peptides. However, the mechanisms responsible for fungistatic and/or fungicidal activity of lysozyme are still not clear. In the present study, the anti-Candida albicans activity of Galleria mellonella lysozyme and anionic peptide 2 (AP2), defense factors constitutively present in the hemolymph, was examined. The lysozyme inhibited C. albicans growth in a dose-dependent manner. The decrease in the C. albicans survival rate caused by the lysozyme was accompanied by a considerable reduction of the fungus metabolic activity, as revealed by LIVE/DEAD staining. In contrast, although AP2 reduced C. albicans metabolic activity, it did not influence its survival rate. Our results suggest fungicidal action of G. mellonella lysozyme and fungistatic activity of AP2 toward C. albicans cells. In the presence of AP2, the anti-C. albicans activity of G. mellonella lysozyme increased. Moreover, when the fungus was incubated with both defense factors, true hyphae were observed besides pseudohyphae and yeast-like C. albicans cells. Atomic force microscopy analysis of the cells exposed to the lysozyme and/or AP2 revealed alterations in the cell surface topography and properties in comparison with the control cells. The results indicate synergistic action of G. mellonella AP2 and lysozyme toward C. albicans. The presence of both factors in the hemolymph of naive larvae suggests their important role in the early stages of immune response against fungi in G. mellonella. Copyright © 2014 Elsevier Inc. All rights reserved.

  9. Biomimetic mineralization of calcium carbonate/carboxymethylcellulose microspheres for lysozyme immobilization

    International Nuclear Information System (INIS)

    Lu Zheng; Zhang Juan; Ma Yunzi; Song Siyue; Gu Wei

    2012-01-01

    Porous calcium carbonate/carboxymethylcellulose (CaCO 3 /CMC) microspheres were prepared by the biomimetic mineralization method for lysozyme immobilization via adsorption. The size and morphology of CaCO 3 /CMC microspheres were characterized by transmitted electron microscopy (TEM) and zeta potential measurement. The lysozyme immobilization was verified by Fourier transform infrared (FTIR) spectroscopy. The effects of pHs and temperatures on lysozyme adsorption were investigated as well. It was revealed that CaCO 3 /CMC microspheres could immobilize lysozyme efficiently via electrostatic interactions and a maximum adsorption capacity of 450 mg/g was achieved at pH 9.2 and 25 °C. Moreover, it was found that the adsorption process fitted well with the Langmuir isothermal model. In addition, UV, fluorescence, and circular dichroism (CD) spectroscopic studies showed that lysozyme maintained its original secondary structure during the adsorption/desorption process. Our study therefore demonstrated that CaCO 3 /CMC microsphere can be used as a cost-effective and efficient support for lysozyme immobilization. - Graphical abstract: CaCO 3 /CMC microsphere was prepared by a facile biomimetic mineralization method and can be used as an efficient and cost-effective support for lysozyme immobilization. Highlights: ► CaCO 3 /CMC microspheres were prepared by the biomimetic mineralization method. ► Lysozyme was efficiently immobilized to CaCO 3 /CMC microspheres via adsorption. ► A maximum adsorption capacity of 450 mg/g was obtained at pH 9.2 and 25 °C. ► The original secondary structure of lysozyme was maintained upon immobilization.

  10. The antibacterial protein lysozyme identified as the termite egg recognition pheromone.

    Directory of Open Access Journals (Sweden)

    Kenji Matsuura

    Full Text Available Social insects rely heavily on pheromone communication to maintain their sociality. Egg protection is one of the most fundamental social behaviours in social insects. The recent discovery of the termite-egg mimicking fungus 'termite-ball' and subsequent studies on termite egg protection behaviour have shown that termites can be manipulated by using the termite egg recognition pheromone (TERP, which strongly evokes the egg-carrying and -grooming behaviours of workers. Despite the great scientific and economic importance, TERP has not been identified because of practical difficulties. Herein we identified the antibacterial protein lysozyme as the TERP. We isolated the target protein using ion-exchange and hydrophobic interaction chromatography, and the MALDI-TOF MS analysis showed a molecular size of 14.5 kDa. We found that the TERP provided antibacterial activity against a gram-positive bacterium. Among the currently known antimicrobial proteins, the molecular size of 14.5 kDa limits the target to lysozyme. Termite lysozymes obtained from eggs and salivary glands, and even hen egg lysozyme, showed a strong termite egg recognition activity. Besides eggs themselves, workers also supply lysozyme to eggs through frequent egg-grooming, by which egg surfaces are coated with saliva containing lysozyme. Reverse transcript PCR analysis showed that mRNA of termite lysozyme was expressed in both salivary glands and eggs. Western blot analysis confirmed that lysozyme production begins in immature eggs in queen ovaries. This is the first identification of proteinaceous pheromone in social insects. Researchers have focused almost exclusively on hydrocarbons when searching for recognition pheromones in social insects. The present finding of a proteinaceous pheromone represents a major step forward in, and result in the broadening of, the search for recognition pheromones. This novel function of lysozyme as a termite pheromone illuminates the profound influence

  11. Activity of lysozyme on Lactobacillus hilgardii strains isolated from Port wine.

    Science.gov (United States)

    Dias, Rita; Vilas-Boas, Eduardo; Campos, Francisco M; Hogg, Tim; Couto, José António

    2015-08-01

    This work evaluated the effect of lysozyme on lactobacilli isolated from Port wine. Bacterial growth experiments were conducted in MRS/TJ medium and inactivation studies were performed in phosphate buffer (KH2PO4), distilled water and wine supplemented with different concentrations of lysozyme. The response of bacteria to lysozyme was found to be highly strain dependent. Some strains of Lactobacillus hilgardii together with Lactobacillus collinoides and Lactobacillus fructivorans were found to be resistant to concentrations of lysozyme as high as 2000 mg/L. It was observed that among the L. hilgardii taxon the resistant strains possess an S-layer coat. Apparently, the strains of L. collinoides and L. fructivorans studied are also S-layer producers as suggested by the total protein profile obtained by SDS-PAGE. Thus, the hypothetical protective role of the S-layer against the action of lysozyme was investigated. From the various treatments used to remove the protein from the surface of the cells, the one employing LiCl (5 M) was the most effective. LiCl pre-treated cells exposed to lysozyme (2000 mg/L) in KH2PO4 buffer maintained its resistance. However, when cells were suspended in distilled water an increased sensitivity to lysozyme was observed. Moreover, it was found that the addition of ethanol (20% v/v) to the suspension medium (distilled water) triggered a strong inactivation effect especially on cells previously treated with LiCl (reduction of >6 CFU log cycles). The results suggest that the S-layer exerts a protective effect against lysozyme and that the cell suspension medium influences the bacteriolysis efficiency. It was also noted that ethanol enhances the inactivation effect of lysozyme. Copyright © 2015 Elsevier Ltd. All rights reserved.

  12. Selectivity and localization of lysozyme uptake in contemporary hydrogel contact lens materials.

    Science.gov (United States)

    Heynen, Miriam; Babaei Omali, Negar; Fadli, Zohra; Coles-Brennan, Chantal; Subbaraman, Lakshman N; Jones, Lyndon

    2017-09-01

    The purpose of this study was to investigate the early and selective uptake of lysozyme and the location of deposited lysozyme on contemporary hydrogel contact lens (CL) materials after exposure to an artificial tear solution (ATS) for 16 h. Seven different hydrogel CL materials [polymacon, omafilcon A, nelfilcon A, nesofilcon A, ocufilcon B, etafilcon A (Acuvue Moist), and etafilcon A (Acuvue Define)] were incubated in an ATS for various times. Total protein deposition was determined using a modified Bradford technique. Lysozyme, lactoferrin, and albumin deposition on CLs were determined using 125 I-radiolabeling method. A confocal laser scanning microscopy (CLSM) technique was utilized to map the location of lysozyme uptake in an asymmetric environment. All lens materials had significant amounts of lysozyme after 1 min of exposure to ATS. After 16 h of incubation, higher levels of total protein deposited on the two etafilcon A-based lenses (Moist and Define), followed by ocufilcon B and both were significantly higher than all other CLs tested (p = 0.0001). The two etafilcon A materials (Moist and Define) also deposited the highest amounts of lysozyme (514.8 ± 28.4 and 527.1 ± 14.7 μg/lens respectively) when compared to other test CLs (p = 0.0001). The CLSM technique revealed that the non-ionic CLs tended to have symmetric distribution of lysozyme throughout the lens materials, while the ionic CLs had an asymmetric distribution, with the highest concentration of lysozyme on and near the exposed surface. The quantity and nature of proteins deposited on CLs varies, depending upon the chemical composition of the material. Among the various lenses tested, etafilcon A deposited the highest amount of total protein, most of it represented by lysozyme, which was largely located near the surface of the lens.

  13. Chicken-type lysozyme in channel catfish: expression analysis, lysozyme activity, and efficacy as immunostimulant against Aeromonas hydrophila infection.

    Science.gov (United States)

    Pridgeon, Julia W; Klesius, Phillip H; Dominowski, Paul J; Yancey, Robert J; Kievit, Michele S

    2013-09-01

    To understand whether chicken-type lysozyme (Lys-c) in channel catfish was induced by infection of Aeromonas hydrophila, the transcriptional levels of Lys-c in skin, gut, liver, spleen, posterior kidney, and blood cells in healthy channel catfish was compared to that in channel catfish infected with A. hydrophila by bath immersion. Quantitative PCR revealed that the transcription levels of Lys-c in infected catfish were significantly (P lysozyme-c (pcDNA-Lys-c) was transfected in channel catfish gill cells G1B, the over-expression of pcDNA-Lys-c offered significant (P < 0.05) protection to G1B against A. hydrophila infection. When channel catfish were intraperitoneally injected with QCDCR adjuvant formulated pcDNA-Lys-c and challenged with a highly virulent A. hydrophila strain AL-09-71 at 1-, 2-, 14-, and 28-days post treatment, pcDNA-Lys-c offered 75%, 100%, 60%, and 77% protection to channel catfish, respectively. Macrophages of fish treated with pcDNA-Lys-c produced significantly (P < 0.05) higher amounts of reactive oxygen species and nitric oxide than that of fish treated with pcDNA vector alone. Taken together, our results suggest that pcDNA-Lys-c could be used as a novel immunostimulant to protect channel catfish against A. hydrophila infection. Published by Elsevier Ltd.

  14. Electrostatic assembly of protein lysozyme on DNA visualized by atomic force microscopy

    International Nuclear Information System (INIS)

    Yang Tao; Wei Gang; Li Zhuang

    2007-01-01

    In the present work, atomic force microscopy (AFM) has been used to study the assembly of protein lysozyme on DNA molecule. Based on the electrostatic interaction, the positively charged lysozyme can easily bind onto the negatively charged DNA molecule surface. The protein molecules appear as globular objects on the DNA scaffold, which are distinguishable in the AFM images. At the same time, lysozyme molecules can be assembled onto DNA as dense or sporadic pattern by varying the protein concentration. This work may provide fundamental aspects for building protein nanostructures and studying of DNA-protein interaction

  15. [Method of estimation of lysozyme activity in serum with chitin azure as substrate].

    Science.gov (United States)

    Pisarzhevskií, S A; Globa, A G; Shirshov, O N; Marchuk, A I; Svetukhin, A M; Karelin, A A

    1997-01-01

    The method of estimation of lysozyme activity in serum when chitin azure is used as substrate is described. The basis of incubation medium is 0.1 M acetate buffer, pH 5.0. It was judged about activity of lysozyme by fluorescence of azure in incubation medium at the end of three hour incubation at 37 degrees C after separation on non-reacted chitin azure by short time centrifugation. It was shown that development of wound infection is accompanied by increasing of chitinase activity of lysozyme in serum.

  16. Effect of zinc on lysozyme-like activity of the seastar Marthasterias glacialis (Echinodermata, Asteroidea) mucus.

    Science.gov (United States)

    Stabili, L; Pagliara, P

    2009-03-01

    Lysozyme represents the best characterized enzyme involved in the self-defense from bacteria. In this study we analysed the effects of zinc on the lysozyme-like activity of the seastar Marthasterias glacialis mucus. This activity, detected by measuring the cleared lysis area of dried Micrococcus lysodeikticus cell walls on Petri dishes, was significantly reduced in presence of zinc. The results are discussed in the light of elucidating the possible relationship between environmental contaminants and increased disease susceptibility in seastars due to the decrease of antibacterial protection. The benefits of using the test of lysozyme activity to monitoring environmental pollution are highlighted.

  17. Bacteriolytic Activity Of Human Interleukin-2, Chicken Egg Lysozyme In The Presence Of Potential Effectors

    OpenAIRE

    Levashov, P. A.; Matolygina, D. A.; Ovchinnikova, E. D.; Atroshenko, D. L.; Savin, S. S.; Belogurova, N. G.; Smirnov, S. A.; Tishkov, V. I.; Levashov, A. V.

    2017-01-01

    The bacteriolytic activity of interleukin-2 and chicken egg lysozyme in the presence of various substances has been studied. Glycine and lysine do not affect the activity of interleukin-2 but increase that of lysozyme, showing a bell-shape concentration dependence peaking at 1.5 mM glycine and 18 mM lysine. Arginine and glutamate activate both interleukin-2 and lysozyme with a concentration dependence of the saturation type. Aromatic amino acids have almost no effect on the activity of both i...

  18. Transient expression of the chicken lysozyme gene after transfer into human cells.

    OpenAIRE

    Matthias, P D; Renkawitz, R; Grez, M; Schütz, G

    1982-01-01

    The chicken lysozyme gene was inserted into an SV40-based plasmid vector, and the recombinants were transfected into the human cell lines HeLa and MCF-7. Correct and efficient transient expression directed by the lysozyme promoter was found in both of these cell lines, as determined by S1 nuclease mapping and Northern blot analysis of the RNAs made. SV40 sequences dramatically enhance the expression of the lysozyme gene. This enhancing effect is only acting in cis and is distance/orientation ...

  19. The lectin domain of UDP-N-acetyl-D-galactosamine: polypeptide N-acetylgalactosaminyltransferase-T4 directs its glycopeptide specificities

    DEFF Research Database (Denmark)

    Hassan, H; Reis, C A; Bennett, E P

    2000-01-01

    , most notably two sites in the MUC1 tandem repeat, is entirely dependent on the glycosylation-dependent function of GalNAc-T4. Here we present data that a putative lectin domain found in the C terminus of GalNAc-T4 functions as a GalNAc lectin and confers its glycopeptide specificity. A single amino...... acid substitution in the lectin domain of a secreted form of GalNAc-T4 selectively blocked GalNAc-glycopeptide activity, while the general activity to peptides exerted by this enzyme was unaffected. Furthermore, the GalNAc-glycopeptide activity of wild-type secreted GalNAc-T4 was selectively inhibited......The initiation step of mucin-type O-glycosylation is controlled by a large family of homologous UDP-GalNAc:polypeptide N-acetylgalactosaminyltransferases (GalNAc-transferases). Differences in kinetic properties, substrate specificities, and expression patterns of these isoenzymes provide...

  20. Early determination of the congenital hypothyroidism by means of the cuantification of STH and T4 by radioimmunoassay method

    International Nuclear Information System (INIS)

    De Penados, R.C.

    1986-11-01

    It was investigated the advantages and disadvantages and conditions of working for the diagnosis of the congenital hypothyroidism by RIA, and its future implementation. It was determined concentration of neonatal STH and neonatal T4 in whole blood of 517 new borns, obtained by capilar punction and recollected in filter paper. 261 samples were found with levels of T4 between normal limits and STH between 0 and 12.5 u U/ml correspond to a normal thyroid function. 29 samples with normal T4 and STH above 30 u U/ml, correspond to a possible primary hypothyroidism. 171 samples with normal T4 and STH between 12.5 and 30 u U/ml were found. (Author)

  1. Incorporation of T4 phage DNA into a specific DNA fraction from the higher plant Matthiola incana.

    Science.gov (United States)

    Gradmann-Rebel, W; Hemleben, V

    1976-01-01

    Isolated T4 phage DNA (sigma=1.694 g/ml) is applied to seedlings of the crucifer Matthiola incana (DNA density sigma=1.698 g/ml). The phage DNA can partly be reextracted from the plants in a specific DNA fraction, which is predominantly characterized by its unusual high density (high density complex=HDC; sigma=1.724 g/ml). DNA:DNA hybridization studies show that phage specific DNA sequences are preserved in the HDC. Results of BrdUrd labeling of the plant DNA before and during incubation with T4DNA suggest that the HDC is composed of T4DNA and a plant DNA component of high density. The analysis of ultrasonicated HDC confirms this suggestion. The ability of plant cells to recognize and handle T4 DNA specifically is discussed.

  2. The peripheral conversion of thyroxine (T4) into triiodothyronine (T3) and reverse triiodothyronine (rT3)

    International Nuclear Information System (INIS)

    Wiersinga, W.M.

    1979-01-01

    The aim of this study was to delineate several physiological, pathological and pharmacological factors involved in the peripheral conversion of thyroxine (T 4 ), using radioimmunoassay. The determination of normal values of these tests under basal circumstances and after stimulation with thyrotropin-releasing-hormone is presented, and some physiological factors which may modulate the conversion of T 4 are discussed. Results are presented of the thyroid function tests in patients with thyroid disease and with acute non-thyroidal diseases. (Auth.)

  3. T cell antigen receptor expression by subsets of Ly-2-L3T4- (CD8-CD4-) thymocytes

    DEFF Research Database (Denmark)

    Wilson, A; Ewing, T; Owens, T

    1988-01-01

    The V beta 8-specific mAb F23.1 and KJ16 were used as fluorescent stains to test for TCR expression on the surface of subpopulations of early, CD4-CD8- (L3T4-Ly-2-) thymocytes from adult CBA mice. A surprisingly high proportion (27%) of Ly-2-L3T4- thymocytes were strongly F23.1 and KJ16 positive...

  4. Isozymes of Lysozyme in Leukocytes and Egg White: Evidence for the Species-Specific Control of Egg-White Lysozyme Synthesis

    Science.gov (United States)

    Hindenburg, Alexander; Spitznagel, John; Arnheim, Norman

    1974-01-01

    Two structurally distinct forms of eggwhite lysozyme (EC 3.2.1.17) are known. The egg white of some species contains both of these forms, while the egg white of other species appears to contain only one or the other of them. We have immunological and electrophoretic evidence that the chicken, which has only one lysozyme type in its egg white, contains both types in its polymorphonuclear leukocytes. Experiments on Embden goose bone marrow showed that this tissue also contains both lysozymes, even though the egg white of this species contains only one of them. Our studies suggest that many avian species have the genetic loci that code for both forms of lysozyme, but that a species-specific regulatory mechanism controls whether one or the other or both of them are expressed during egg white production. The fact that two distinct lysozymes are present in chicken leukocytes may be of significance to the antibacterial mechanism of these cells, especially in light of the fact that they lack myeloperoxidase, an important leukocyte enzyme in mammals. Images PMID:4209557

  5. Structure of the small outer capsid protein, Soc: a clamp for stabilizing capsids of T4-like phages.

    Science.gov (United States)

    Qin, Li; Fokine, Andrei; O'Donnell, Erin; Rao, Venigalla B; Rossmann, Michael G

    2010-01-29

    Many viruses need to stabilize their capsid structure against DNA pressure and for survival in hostile environments. The 9-kDa outer capsid protein (Soc) of bacteriophage T4, which stabilizes the virus, attaches to the capsid during the final stage of maturation. There are 870 Soc molecules that act as a "glue" between neighboring hexameric capsomers, forming a "cage" that stabilizes the T4 capsid against extremes of pH and temperature. Here we report a 1.9 A resolution crystal structure of Soc from the bacteriophage RB69, a close relative of T4. The RB69 crystal structure and a homology model of T4 Soc were fitted into the cryoelectron microscopy reconstruction of the T4 capsid. This established the region of Soc that interacts with the major capsid protein and suggested a mechanism, verified by extensive mutational and biochemical studies, for stabilization of the capsid in which the Soc trimers act as clamps between neighboring capsomers. The results demonstrate the factors involved in stabilizing not only the capsids of T4-like bacteriophages but also many other virus capsids.

  6. Intracavitary 'T4 immunotherapy' of malignant mesothelioma using pan-ErbB re-targeted CAR T-cells.

    Science.gov (United States)

    Klampatsa, Astero; Achkova, Daniela Y; Davies, David M; Parente-Pereira, Ana C; Woodman, Natalie; Rosekilly, James; Osborne, Georgina; Thayaparan, Thivyan; Bille, Andrea; Sheaf, Michael; Spicer, James F; King, Juliet; Maher, John

    2017-05-01

    Malignant mesothelioma remains an incurable cancer. We demonstrated that mesotheliomas expressed EGFR (79.2%), ErbB4 (49.0%) and HER2 (6.3%), but lacked ErbB3. At least one ErbB family member was expressed in 88% of tumors. To exploit ErbB dysregulation in this disease, patient T-cells were engineered by retroviral transduction to express a panErbB-targeted chimeric antigen receptor (CAR), co-expressed with a chimeric cytokine receptor that allows interleukin (IL)-4 mediated CAR T-cell proliferation. This combination is referred to as T4 immunotherapy. T-cells from mesothelioma patients were uniformly amenable to T4 genetic modification and expansion/enrichment thereafter using IL-4. Patient-derived T4 + T-cells were activated upon contact with a panel of four mesothelioma cell lines, leading to cytotoxicity and cytokine release in all cases. Adoptive transfer of T4 immunotherapy to SCID Beige mice with an established bioluminescent LO68 mesothelioma xenograft was followed by regression or eradication of disease in all animals. Despite the established ability of T4 immunotherapy to elicit cytokine release syndrome in SCID Beige mice, therapy was very well tolerated. These findings provide a strong rationale for the clinical evaluation of intracavitary T4 immunotherapy to treat mesothelioma. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. Reduction and repopulation of recipient T4+ and T8+ T-lymphocytes in allogeneic bone marrow transplantation

    International Nuclear Information System (INIS)

    Gratama, J.W.; van den Bergh, R.L.; Naipal, A.; D'Amaro, J.; Zwaan, F.E.; Jansen, J.; de Gast, G.C.

    1986-01-01

    In eight recipients of allogeneic bone marrow grafts who had sex-mismatched donors, the reduction and subsequent repopulation of T4+ and T8+ T-lymphocytes of recipient origin were studied. The origin of the donor-recipient T4+ and T8+ T cells was studied using quinacrine staining of Y chromatin combined with T-cell typing for T4 and T8. Following chemoradiotherapy and bone marrow transplantation (BMT), T cells reached their nadir at a median of five (range 1-8) days after BMT. T8+ T cells decreased at a faster rate from the peripheral blood than T4+ T cells. The first T cells that appeared in the circulation at day 12 were predominantly T4+, and a large number of them were of recipient origin. Thereafter, they gradually decreased, and the numbers of T cells of donor origin increased. In the patients who had no or only minor complications, T4+ and T8+ T cells of donor origin repopulated the blood at similar rates. This pattern, however, was modified by severe graft-versus-host disease or by cytomegalovirus infection

  8. Estimation of vectorial capacity of Anopheles minimus Theobald & An. fluviatilis James (Diptera: Culicidae in a malaria endemic area of Odisha State, India

    Directory of Open Access Journals (Sweden)

    K Gunasekaran

    2014-01-01

    Full Text Available Background & objectives: Anopheles minimus and An. fluviatilis were incriminated as the major malaria vectors in Keonjhar district of Odisha State recently. This study was carried out to elucidate the potential role of these two vector species in transmission of malaria during different seasons, and vectorial capacity of these species was also estimated. Methods: Three hilly and forested villages of Keonjhar district were randomly selected. Vectorial capacity (C was calculated using the Macdonald′s formula as modified by Garret-Jones. The human landing density of the vector species was obtained from all night human landing collections (bait protected by bed-net. Man feeding habit was estimated by multiplying the human blood index with feeding frequency, which was obtained on daily basis from the duration of gonotrophic cycle. The probability of survival through the extrinsic incubation cycle was calculated from the probability of survival through one day and duration of sporogonic cycle. Results: The estimated vectorial capacity of An. minimus varied between 0.014 and 1.09 for Plasmodium falciparum (Pf and between 0.1 and 1.46 for P. vivax (Pv. The C of An. minimus for both Pf and Pv was higher during rainy season than the other two seasons. The estimated C of An. fluviatilis varied between 0.04 and 1.28 for Pf and between 0.20 and 1.54 for Pv. Interpretation & conclusions: Based on the estimated values of vectorial capacity of the two vector species, the area could be stratified and such stratification would reflect the difference in the intensity of transmission between different strata and accordingly the appropriate control strategy could be adopted for each stratum.

  9. A Manin-Mumford theorem for the maximal compact subgroup of a universal vectorial extension of a product of elliptic curves

    OpenAIRE

    Jones, Gareth; Schmidt, Harry

    2018-01-01

    We study the intersection of an algebraic variety with the maximal compact subgroup of a universal vectorial extension of a product of elliptic curves. For this intersection we show a Manin-Mumford type statement. This answers some questions posed by Corvaja-Masser-Zannier which arose in connection with their investigation of the intersection of a curve with real analytic subgroups of various algebraic groups. They prove finiteness in the situation of a single elliptic curve. Using Khovanskii...

  10. Crystallization of Chicken Egg White Lysozyme from Assorted Sulfate Salts

    Science.gov (United States)

    Forsythe, Elizabeth L.; Snell, Edward H.; Malone, Christine C.; Pusey, Marc L.

    1999-01-01

    Chicken egg white lysozyme has been found to crystallize from ammonium, sodium, potassium, rubidium, magnesium, and manganese sulfates at acidic and basic pH, with protein concentrations from 60 to 190 mg/ml. Crystals have also been grown at 4 C in the absence of any other added salts using isoionic lysozyme which was titrated to pH 4.6 with dilute sulfuric acid. Four different crystal forms have been obtained, depending upon the temperature, protein concentration, and precipitating salt employed. Crystals grown at 15 C were generally tetragonal, with space group P4(sub 3)2(sub 1)2. Crystallization at 20 C typically resulted in the formation of orthorhombic crystals, space group P2(sub 1)2(sub 1)2(sub 1). The tetragonal reversible reaction orthorhombic transition appeared to be a function of both the temperature and protein concentration, occurring between 15 and 20 C and between 100 and 125 mg/ml protein concentration. Crystallization from 1.2 M magnesium sulfate at pH 7.8 gave a trigonal crystal, space group P3(sub 1)2(sub 1), a = b = 87.4, c = 73.7, gamma = 120 deg, which diffracted to 2.8 A. Crystallization from ammonium sulfate at pH 4.6, generally at lower temperatures, was also found to result in a monoclinic form. space group C2, a = 65.6, b = 95.0, c = 41.2, beta = 119.2 deg. A crystal of approximately 0.2 x 0.2 x 0.5 mm grown from bulk solution diffracted to approximately 3.5 A.

  11. Protein-encapsulated gold cluster aggregates: the case of lysozyme

    Science.gov (United States)

    Baksi, Ananya; Xavier, Paulrajpillai Lourdu; Chaudhari, Kamalesh; Goswami, N.; Pal, S. K.; Pradeep, T.

    2013-02-01

    We report the evolution and confinement of atomically precise and luminescent gold clusters in a small protein, lysozyme (Lyz) using detailed mass spectrometric (MS) and other spectroscopic investigations. A maximum of 12 Au0 species could be bound to a single Lyz molecule irrespective of the molar ratio of Lyz : Au3+ used for cluster growth. The cluster-encapsulated protein also forms aggregates similar to the parent protein. Time dependent studies reveal the emergence of free protein and the redistribution of detached Au atoms, at specific Lyz to Au3+ molar ratios, as a function of incubation time, proposing inter-protein metal ion transfer. The results are in agreement with the studies of inter-protein metal transfer during cluster growth in similar systems. We believe that this study provides new insights into the growth of clusters in smaller proteins.We report the evolution and confinement of atomically precise and luminescent gold clusters in a small protein, lysozyme (Lyz) using detailed mass spectrometric (MS) and other spectroscopic investigations. A maximum of 12 Au0 species could be bound to a single Lyz molecule irrespective of the molar ratio of Lyz : Au3+ used for cluster growth. The cluster-encapsulated protein also forms aggregates similar to the parent protein. Time dependent studies reveal the emergence of free protein and the redistribution of detached Au atoms, at specific Lyz to Au3+ molar ratios, as a function of incubation time, proposing inter-protein metal ion transfer. The results are in agreement with the studies of inter-protein metal transfer during cluster growth in similar systems. We believe that this study provides new insights into the growth of clusters in smaller proteins. Electronic supplementary information (ESI) available. See DOI: 10.1039/c2nr33180b

  12. Crystallization of chicken egg white lysozyme from assorted sulfate salts

    Science.gov (United States)

    Forsythe, Elizabeth L.; Snell, Edward H.; Malone, Christine C.; Pusey, Marc L.

    1999-01-01

    Chicken egg white lysozyme has been found to crystallize from ammonium, sodium, potassium, rubidium, magnesium, and manganese sulfates at acidic and basic pH, with protein concentrations from 60 to 190 mg/ml. Crystals have also been grown at 4°C in the absence of any other added salts using isoionic lysozyme which was titrated to pH 4.6 with dilute sulfuric acid. Four different crystal forms have been obtained, depending upon the temperature, protein concentration, and precipitating salt employed. Crystals grown at 15°C were generally tetragonal, with space group P4 32 12. Crystallization at 20°C typically resulted in the formation of orthorhombic crystals, space group P2 12 12 1. The tetragonal ↔ orthorhombic transition appeared to be a function of both the temperature and protein concentration, occurring between 15 and 20°C and between 100 and 125 mg/ml protein concentration. Crystallization from 1.2 M magnesium sulfate at pH 7.8 gave a trigonal crystal, space group P3 12 1, a= b=87.4, c=73.7, γ=120°, which diffracted to 2.8 Å. Crystallization from ammonium sulfate at pH 4.6, generally at lower temperatures, was also found to result in a monoclinic form, space group C2, a=65.6, b=95.0, c=41.2, β=119.2°. A crystal of ˜0.2×0.2×0.5 mm grown from bulk solution diffracted to ˜3.5 Å.

  13. Crystallization of Chicken Egg White Lysozyme from Sulfate Salts

    Science.gov (United States)

    Forsythe, Elizabeth; Pusey, Marc

    1998-01-01

    It has been "known" that chicken egg white lysozyme does not crystallize from sulfate, particularly ammonium sulfate, salts, but instead gives amorphous precipitates. This has been the basis of several studies using lysozyme comparing macromolecule crystal nucleation and amorphous precipitation. Recently Ries-Kautt et al (Acta Cryst D50, (1994) 366) have shown that purified isoionic CEWL could be crystallized from low concentrations of sulfate at basic pH, and we subsequently showed that in fact CEWL could be purified in both the tetragonal and orthorhombic forms using ammonium sulfate over the pH range 4.0 to 7.8 (Acta Cryst D53, (1997) 795). We have now extended these observations to include a range of common sulfate salts, specifically sodium, potassium, rubidium, magnesium, and manganese sulfates. In all cases but the manganese sulfates both the familiar tetragonal and orthorhombic forms were obtained, with unit cell dimensions close to those known for the "classic" sodium chloride crystallized forms. Manganese sulfate has only yielded orthorhombic crystals to date. All crystallizations were carried out using low (typically less than or equal to 6 M) salt and high (greater than approximately 90 mg/ml) protein concentrations. As with ammonium sulfate, the tetragonal - orthorhombic phase shift appears to be a function of both the temperature and the protein concentration, with higher temperatures and concentrations favoring the orthorhombic and lower the tetragonal form. The phase change range is somewhat reduced for the sulfate salts, depending upon conditions being typically between approximately 15 - 20 C. Both the magnesium and manganese sulfates gave crystals at salt concentrations over 0.6 M as well, with magnesium sulfate giving a very slowly nucleating and growing hexagonal form. A triclinic crystal form, characterized by aggressively small crystals (typically 0.1 mm in size) has been occasionally obtained from ammonium sulfate. Finally, preliminary spot

  14. Neutron crystallography of hen egg-white lysozyme at pH4.9

    International Nuclear Information System (INIS)

    Maeda, Mitsuru; Fujiwara, Satoru; Niimura, Nobuo; Yonezawa, Yasushige

    2001-01-01

    In order to elucidate protein stability and function, it is important to know states of protonation of each amino acid in the protein under different pH. To answer this problem, we have been studying the neutron protein crystallography of hen egg-white (HEW) lysozyme under different pH. Neutron diffraction experiments of single crystals of HEW lysozyme at pH4.9 were carried out with BIX-II at JAERI. The state of protonation of active site in the HEW lysozyme was investigated; there was a hydrogen (deuterium) atom bound to carboxylate oxygen atom of Glu 35 and no hydrogen (deuterium) atom bound to that of Asp 52. The results agreed with the proposed catalytic mechanism of active site in the HEW lysozyme. (author)

  15. SANS study of Lysozyme vs. BSA protein adsorption on silica nanoparticles

    Science.gov (United States)

    Kumar, Sugam; Aswal, V. K.; Kohlbrecher, J.

    2012-06-01

    Lysozyme (M.W. 14.7 kD) and BSA (M.W. 66.7 kD) are two most commonly studied model proteins in literature. Lysozyme (cationic) and BSA (anionic) are oppositely charged at pH 7 and their interaction with anionic silica nanoparticles has been studied using small-angle neutron scattering (SANS). Measurements were carried out on fixed 1 wt% concentration of nanoparticles and varying concentration of protein in the range 0.5 to 2 wt%. It is found that both the proteins adsorb on the nanoparticles where strong interaction of lysozyme leads to the aggregation of nanoparticles but the system remains stable with BSA. Adsorption increases with protein concentration and has been found much larger for lysozyme.

  16. Renaturation of reduced hen egg white lysozyme containing two blocked sulfhydryl groups

    Energy Technology Data Exchange (ETDEWEB)

    Acharya, A.S.; Taniuchi, H.

    1980-10-01

    Formation of native lysozyme from the reduced form involves many pathways in two processes: incorrect pairing of half-cystine residues by oxidation and rearrangement of disulfide (SS) bonds. The energy barrier against suflhydryl (SH)-disulfide interchange of the native or nativelike species thus formed causes accumulation of these species. For example, the enzymatically active isomers containing three presumably native SS bonds and one open SS bond may be thermodynamically favorable over the nonnative isomers and can be formed from reduced lysozyme or lysozyme containing scrambled SS bonds by nonobligatory and flexible pathways. As an extension of these observations formation of nativelike species from reduced lysozyme containing the average of two carboxymethyl (CM)-cysteine was investigated.

  17. Evaluation of immobilized-lysozyme by means of TOF-SIMS

    Science.gov (United States)

    Okada, Keigo; Aoyagi, Satoka; Dohi, Makoto; Kato, Nobuhiko; Kudo, Masahiro; Tozu, Miyako; Miyayama, Takuya; Sanada, Noriaki

    2008-12-01

    Evaluation of immobilized-proteins on bio-devices is important for the development of sophisticated devices. Lysozyme molecules immobilized on substrates were evaluated by means of time-of-flight secondary ion mass spectrometry (TOF-SIMS). Two types of the lysozyme-immobilized samples were prepared by controlling the binding parts, i.e., the amino groups or carboxyl groups, of the protein. The TOF-SIMS spectra of each sample were analyzed with mutual information to select fragment ions specific to each sample. According to the results, differences between the samples being immobilized in the different ways are suggested, and the surface structure of the lysozyme molecule immobilized at amino groups is determined based on three-dimensional structure of lysozyme in the Protein Data Bank.

  18. Investigation of factors affecting the stability of lysozyme spray dried from ethanol-water solutions

    DEFF Research Database (Denmark)

    Ji, Shuying; Thulstrup, Peter Waaben; Mu, Huiling

    2017-01-01

    Formulation composition and processing conditions can be adjusted to enhance the structural integrity as well as the bioactivity of proteins in the spray drying process. In this study, lysozyme was chosen as a model pharmaceutical protein to study these aspects when spray drying from water......-ethanol mixtures. The effect of formulation additives (trehalose, Tween 20 and phosphate-buffered saline) and processing conditions (inlet temperature and storage time of lysozyme in the feed solution before the spray drying process) on the protein bioactivity was investigated. The results showed...... that the bioactivities of spray dried lysozyme with these additives were about 5-10% higher than that without additives. The bioactivity of the spray dried lysozyme was found to increase with a decrease in the inlet temperature from 130°C to 80°C, with similar findings when shortening the storage time of the feed...

  19. Poly(hydroxyethyl methacrylate) based magnetic nanoparticles for lysozyme purification from chicken egg white.

    Science.gov (United States)

    Köse, Kazım; Denizli, Adil

    2013-02-01

    The purpose of this article is to synthesize poly(hydroxyethyl methacrylate-N-methacryloyl-(L)-tryptophan) [mPHEMATrp] magnetic nanoparticles for lysozyme purification from chicken egg white. mPHEMATrp nanoparticles (38 nm in diameter) were synthesized by surfactant-free emulsion polymerization. Specific surface area of the mPHEMATrp nanoparticles was calculated to be 896 m2/g. Elemental analysis of mPHEMATrp nanoparticles was estimated in the range of 5.7-48.3 µmol MATrp/g. The maximum lysozyme adsorption capacity of the mPHEMATrp nanoparticles was 376.1 mg/g. The mPHEMATrp nanoparticles could be used five times without decreasing the lysozyme adsorption capacity significantly. The results indicate that the mPHEMATrp nanoparticles promise high selectivity for lysozyme purification.

  20. Lysozyme crystallization by vapor diffusion: characterization and modeling in the absence and presence of exogenous minerals

    Science.gov (United States)

    Kimble, W. L.; Rousseau, R. W.; Sambanis, A.

    1995-01-01

    A model accounting for water evaporation and crystal growth was synthesized to simulate protein concentration profiles in the crystallization wells of a vapor-diffusion apparatus. The model calculations were compared with experimental results obtained with chicken egg white lysozyme crystallized in the absence and presence of exogenous mineral particles. The model predicted the increase in protein concentration during water evaporation and the decrease during crystal growth. The effects of magnetite, galena and chalcopyrite on the time profile of dissolved lysozyme concentration appeared minimal, except for the occurrence of earlier nucleation in the presence of magnetite. Few of the lysozyme crystals formed were physically associated with these minerals. More protein crystals were associated with topaz, lepidolite and apophyllite, which exhibit a close match of their crystalline lattice to that of lysozyme.

  1. Effect of polyols on the conformational stability and biological activity of a model protein lysozyme.

    Science.gov (United States)

    Singh, Somnath; Singh, Jagdish

    2003-01-01

    The purpose of this study was to investigate the stabilizing action of polyols against various protein degradation mechanisms (eg, aggregation, deamidation, oxidation), using a model protein lysozyme. Differential scanning calorimeter (DSC) was used to measure the thermodynamic parameters, mid point transition temperature and calorimetric enthalpy, in order to evaluate conformational stability. Enzyme activity assay was used to corroborate the DSC results. Mannitol, sucrose, lactose, glycerol, and propylene glycol were used as polyols to stabilize lysozyme against aggregation, deamidation, and oxidation. Mannitol was found to stabilize lysozyme against aggregation, sucrose against deamidation both at neutral pH and at acidic pH, and lactose against oxidation. Stabilizers that provided greater conformational stability of lysozyme against various degradation mechanisms also protected specific enzyme activity to a greater extent. It was concluded that DSC and bioassay could be valuable tools for screening stabilizers in protein formulations.

  2. Improvement on the Fatigue Performance of 2024-T4 Alloy by Synergistic Coating Technology

    Directory of Open Access Journals (Sweden)

    Xi-Shu Wang

    2014-05-01

    Full Text Available In this paper, rotating bending fatigue tests of 2024-T4 Al alloy with different oxide coatings were carried out. Compared to the uncoated and previously reported oxide coatings of aluminum alloys, the fatigue strength is able to be enhanced by using a novel oxide coating with sealing pore technology. These results indicate that the better the coating surface quality is, the more excellent the fatigue performance under rotating bending fatigue loading is. The improvement on the fatigue performance is mainly because the fatigue crack initiation and the early stage of fatigue crack growth at the coating layer can be delayed after PEO coating with pore sealing. Therefore, it is a so-called synergistic coating technology for various uses, including welding thermal cracks and filling micro-pores. The effects of different oxide coatings on surface hardness, compressive residual stress, morphology and fatigue fracture morphology are discussed. A critical compressive residual stress of about 95–100 MPa is proposed.

  3. Single-stranded DNA library preparation from highly degraded DNA using T4 DNA ligase.

    Science.gov (United States)

    Gansauge, Marie-Theres; Gerber, Tobias; Glocke, Isabelle; Korlevic, Petra; Lippik, Laurin; Nagel, Sarah; Riehl, Lara Maria; Schmidt, Anna; Meyer, Matthias

    2017-06-02

    DNA library preparation for high-throughput sequencing of genomic DNA usually involves ligation of adapters to double-stranded DNA fragments. However, for highly degraded DNA, especially ancient DNA, library preparation has been found to be more efficient if each of the two DNA strands are converted into library molecules separately. We present a new method for single-stranded library preparation, ssDNA2.0, which is based on single-stranded DNA ligation with T4 DNA ligase utilizing a splinter oligonucleotide with a stretch of random bases hybridized to a 3΄ biotinylated donor oligonucleotide. A thorough evaluation of this ligation scheme shows that single-stranded DNA can be ligated to adapter oligonucleotides in higher concentration than with CircLigase (an RNA ligase that was previously chosen for end-to-end ligation in single-stranded library preparation) and that biases in ligation can be minimized when choosing splinters with 7 or 8 random nucleotides. We show that ssDNA2.0 tolerates higher quantities of input DNA than CircLigase-based library preparation, is less costly and better compatible with automation. We also provide an in-depth comparison of library preparation methods on degraded DNA from various sources. Most strikingly, we find that single-stranded library preparation increases library yields from tissues stored in formalin for many years by several orders of magnitude. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  4. Oral T4-like phage cocktail application to healthy adult volunteers from Bangladesh

    Energy Technology Data Exchange (ETDEWEB)

    Sarker, Shafiqul Alam, E-mail: sasarker@icddrb.org [International Centre for Diarrhoeal Diseases Research, Bangladesh (icddr,b), 68 Shaheed Tajuddin Ahmed Sharani, Mohakhali, Dhaka 1212 (Bangladesh); McCallin, Shawna; Barretto, Caroline [Nestle Research Centre, Nestec Ltd., Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland); Berger, Bernard, E-mail: bernard.berger@rdls.nestle.com [Nestle Research Centre, Nestec Ltd., Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland); Pittet, Anne-Cecile [Nestle Research Centre, Nestec Ltd., Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland); Sultana, Shamima, E-mail: shamima@icddrb.org [International Centre for Diarrhoeal Diseases Research, Bangladesh (icddr,b), 68 Shaheed Tajuddin Ahmed Sharani, Mohakhali, Dhaka 1212 (Bangladesh); Krause, Lutz, E-mail: ltz.krause@gmail.com [Nestle Research Centre, Nestec Ltd., Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland); Huq, Sayeda, E-mail: sayeeda@mail.icddrb.org [International Centre for Diarrhoeal Diseases Research, Bangladesh (icddr,b), 68 Shaheed Tajuddin Ahmed Sharani, Mohakhali, Dhaka 1212 (Bangladesh); Bibiloni, Rodrigo, E-mail: Rodrigo.Bibiloni@agresearch.co.nz [Nestle Research Centre, Nestec Ltd., Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland); Bruttin, Anne, E-mail: anne.bruttin@rdls.nestle.com [Nestle Research Centre, Nestec Ltd., Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland); Reuteler, Gloria, E-mail: gloria.reuteler@rdls.nestle.com [Nestle Research Centre, Nestec Ltd., Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland); Bruessow, Harald, E-mail: harald.bruessow@rdls.nestle.com [Nestle Research Centre, Nestec Ltd., Vers-chez-les-Blanc, CH-1000 Lausanne 26 (Switzerland)

    2012-12-20

    The genomic diversity of 99 T4-like coliphages was investigated by sequencing an equimolar mixture with Illumina technology and screening them against different databases for horizontal gene transfer and undesired genes. A 9-phage cocktail was given to 15 healthy adults from Bangladesh at a dose of 3 Multiplication-Sign 10{sup 9} and 3 Multiplication-Sign 10{sup 7} plaque-forming units and placebo respectively. Phages were detected in 64% of the stool samples when subjects were treated with higher titer phage, compared to 30% and 28% with lower-titer phage and placebo, respectively. No Escherichia coli was present in initial stool samples, and no amplification of phage was observed. One percent of the administered oral phage was recovered from the feces. No adverse events were observed by self-report, clinical examination, or from laboratory tests for liver, kidney, and hematology function. No impact of oral phage was seen on the fecal microbiota composition with respect to bacterial 16S rRNA from stool.

  5. Expression of lysozyme in the life history of the house fly (Musca domestica l.).

    Science.gov (United States)

    Nayduch, Dana; Joyner, Chester

    2013-07-01

    From egg to adult, all life history stages of house flies associate with septic environments teeming with bacteria. House fly lysozyme was first identified in the larval midgut, where it is used for digestion of microbe-rich meals because of its broad-spectrum activity against gram-positive and gram-negative bacteria as well as fungi. This study aimed to determine the temporal expression of lysozyme in the life history of house flies (from egg through adults) on both the mRNA and protein level, and to determine the tissue-specific expression of lysozyme in adult flies induced by feeding Staphylococcus aureus. From 30-min postoviposition through adulthood, all life history stages of the house fly express lysozyme on the mRNA level. In adult flies, lysozyme is expressed both locally in the alimentary canal and systemically in the fat body. Interestingly, we found that during the normal life history of flies, lysozyme protein was only detected in larval stages and older adults, likely because of ingestion of immune-stimulating levels of bacteria, not experienced during egg, pupa, and teneral adult stages. Constitutive expression on the mRNA level implies that this effector is a primary defense molecule in all stages of the house fly life history, and that a mechanism for posttranscriptional control of mature lysozyme enzyme expression may be present. Lysozyme active enzyme primarily serves both a digestive and defensive function in larval and adult flies, and may be a key player in the ability of Musca domestica L. to thrive in microbe-rich environments.

  6. Tracking lysozyme unfolding during salt-induced precipitation with hydrogen exchange and mass spectrometry.

    Science.gov (United States)

    Tobler, S A; Sherman, N E; Fernandez, E J

    We utilized electrospray ionization mass spectrometry (ESI-MS) and hydrogen-deuterium exchange (HX) to detect unfolding of hen egg white lysozyme during salt-induced precipitation. Deuterated lysozyme was dissolved in protonated buffer at pH 2.16 and precipitated with ammonium sulfate, sodium chloride, and potassium thiocyanate. ESI-MS was used to detect mass differences in lysozyme due to the loss of deuterons for solvent protons, providing insight on the conformational history of the protein during the labeling experiment. Precipitation with ammonium sulfate and sodium chloride did not unfold lysozyme, consistent with the known stabilizing effects of kosmotropic salts. Potassium thiocyanate, an aggressive chaotrope, was an effective precipitant at 0.2 M, but also disrupted lysozyme structure and caused the formation of precipitate fractions that did not readily redissolve into aqueous solution without the use of a chemical denaturant. Precipitation with 1.0 M thiocyanate resulted in faster rates of unfolding and larger amounts of the insoluble precipitate. The unfolding kinetics were biphasic, exhibiting a slow phase after a few hours that presumably reflected a smaller propensity for lysozyme to unfold in the precipitated state. Bimodal mass distributions in the ESI-MS spectra for the thiocyanate precipitates indicate two states for lysozyme in this system, a native and a molten globule-like partially unfolded state. ESI-MS analysis of the insoluble precipitates indicated that they consisted primarily of protein molecules that had unfolded. Investigation of the HX behavior of lysozyme in a KSCN solution at low protein concentrations confirmed the destabilizing effect of the salt on the protein structure, even when there was almost no solid phase present. The HX/ESI-MS results provide insight into the mechanism combining precipitation and denaturation for such a system, both in terms of obtaining quantitative kinetic and stability information and the

  7. Affinity purification of hen egg lysozyme using sephadex G75 | Islam ...

    African Journals Online (AJOL)

    We found lysozyme binds Sephadex G75, a dextran-based matrix routinely used for Gel-filtration chromatography, in a pH dependent manner. The binding is rapid and specific in a buffer containing 25 mM NaCl at pH 8.0, and requires only 0.1 ml of swollen Sephadex G75 suspension per mg of lysozyme. The bound ...

  8. Influence of different restorative materials on lysozyme and amylase activity of the salivary pellicle in situ.

    Science.gov (United States)

    Hannig, Christian; Wasser, Mathias; Becker, Klaus; Hannig, Matthias; Huber, Karin; Attin, Thomas

    2006-09-15

    Lysozyme and amylase are the most abundant enzymatic components in the salivary pellicle. The purpose of the present study was to determine the influence of different substrata on amylase and lysozyme activity in salivary pellicles formed in situ. Slabs (5 mm diameter) of bovine dentine and enamel, of titanium, gold alloy, resin composite, PMMA, amalgam, and feldspar ceramic were fixed on the buccal sites of individual splints worn by six subjects for 30 min to allow pellicle formation. Thereafter, slabs were removed from the trays and rinsed with running water. Lysozyme activity was determined via lysis of Micrococcus lysodeicticus. Amylase activity was measured with a photometric method using 2-chloro-4-nitrophenyl-4-O-beta-D-galactopyranosylmaltotriosid (GalG2CNP) as substrate. Both pellicle enzymes were evaluated in the immobilized as well as in the desorbed state. Salivary enzyme activities were also measured. All investigated pellicles exhibited lysozyme and amylase activity. Great intraindividual and interindividual differences were observed. Over all samples, immobilized amylase activity amounted to 0.65 +/- 0.64 mU/cm2. Immobilized lysozyme activity was 5.04 +/- 1.55 U/cm2. There were no major effects of the substratum on pellicle-bound amylase and lysozyme activity. Immobilized and desorbed enzyme activities revealed a strong correlation (lysozyme: r = 0.700; amylase: r = 0.990). Salivary enzyme activities had only little impact on pellicle-bound enzyme activities. Amylase and lysozyme are incorporated in the acquired in situ pellicle on different solid surfaces in an active conformation. Dental material and enzyme activity in the saliva have only little impact on enzymatic activity in the pellicle in situ.

  9. Effect of lysozyme on "flor" velum yeasts in the biological aging of sherry wines.

    Science.gov (United States)

    Roldán, Ana; Lasanta, Cristina; Caro, Ildefonso; Palacios, Víctor

    2012-05-01

    Biological aging is a key step in the production of Sherry wine classified as "fine". During this stage, a film of yeast referred to as "flor velum" covers the surface of the wine and substantially alters its characteristics. Other microorganisms may coexist with flor yeasts, such as lactic acid bacteria and non-Saccharomyces yeasts, whose growth may be favored under certain conditions, causing organoleptic deviations and deterioration of the wine. To prevent the development of lactic bacteria, lysozyme usage has been introduced. Lysozyme is a hydrolytic enzyme with muramidase activity that can lyse gram-positive bacteria; its use in winemaking was approved by the OIV in 1997 (resolution OENO 10/97). Thus far, the use of lysozyme during the production of Sherry wines is not widespread despite its effectiveness in controlling lactic acid bacteria. However, there have been no studies on the effect of lysozyme on flor velum. The aim of this study was to determine the influence of lysozyme on yeast growth and the formation, development and metabolism of flor velum during the biological aging process of Sherry wine. The results indicate that lysozyme does not affect the flor yeast during the fermentative stage or biofilm stage. However, if yeast inoculation is carried out under submerged culture conditions during biological aging, low doses of lysozyme (≥12.5 g/hL) affect cell multiplication and the membrane hydrophobicity of the yeast, inhibiting their aggregation and flotation and the subsequent development of flor velum. Thus, the yeast inoculation protocol and the methodology used for the addition of lysozyme influence velum development, its metabolism and the wine characteristics. Copyright © 2011 Elsevier Ltd. All rights reserved.

  10. Preparation and evaluation of a novel bioactive glass/lysozyme/PLGA composite microsphere.

    Science.gov (United States)

    Liu, Hongfei; Shi, Shuangshuang; Cao, Jin; Ji, Lijun; He, Yan; Xi, Jumei

    2015-03-01

    The objective of this study was to fabricate a novel nano-bioceramics incorporated lysozyme poly (d, l-lactide-co-glycolide) (PLGA) microsphere. The nano-bioceramics was used as a biodegradable and sustained-release antacid to stabilize the lysozyme in the drug release process. First, the nano-bioceramics were prepared by sol-gel method, and then were characterized by energy dispersive X-ray analysis, dynamic light scattering and in vitro degradation test. Second, the lysozyme PLGA microsphere incorporated with nano-bioceramic was fabricated by the S/W/O/W emulsion solvent evaporation method. The microsphere was characterized by scanning electron microscopy (SEM), Fourier transform infrared (FTIR) spectroscopy and UV circular dichroism (UV CD). Finally the in vitro drug release and bioactivity test was carried out. The composition of the nano-bioceramics was 58% SiO2, 36% CaO, 6% P2O5, and the average particle size was 295 nm. The nano-bioceramics incorporated lysozyme PLGA microspheres were prepared by the multi-emulsion method. The SEM results showed that the bioceramics was uniformly distributed in the PLGA microsphere. Results from in vitro lysozyme release test exhibited a prolonged release time for 1month. The FTIR and UVCD results suggested that the lysozyme in the drug release process had a similar secondary structure conformation to the native one. The Micrococcus lysodeikticus test showed that the microspheres incorporated with bioceramics provided long-term protein stability against the acidic environment resulted from PLGA's degradates and more than 90% of the lysozyme released over the 1 month period was preserved in a bioactive form. A novel bioceramics incorporated lysozyme PLGA microsphere was prepared with potentials for sustained protein release formulation.

  11. Characterization of Bioactive Recombinant Human Lysozyme Expressed in Milk of Cloned Transgenic Cattle

    OpenAIRE

    Yang, Bin; Wang, Jianwu; Tang, Bo; Liu, Yufang; Guo, Chengdong; Yang, Penghua; Yu, Tian; Li, Rong; Zhao, Jianmin; Zhang, Lei; Dai, Yunping; Li, Ning

    2011-01-01

    BACKGROUND: There is great potential for using transgenic technology to improve the quality of cow milk and to produce biopharmaceuticals within the mammary gland. Lysozyme, a bactericidal protein that protects human infants from microbial infections, is highly expressed in human milk but is found in only trace amounts in cow milk. METHODOLOGY/PRINCIPAL FINDINGS: We have produced 17 healthy cloned cattle expressing recombinant human lysozyme using somatic cell nuclear transfer. In this study,...

  12. Laser ablation of lysozyme with UV, visible and infrared femto- and nanosecond pulses

    OpenAIRE

    Schou, Jørgen; Canulescu, Stela; Matei, Andreea; Cazzaniga, Andrea Carlo; Constantinescu, Catalin; Amoruso, S.; Wang, X.; Bruzzese, R.; Dinescu, M.

    2013-01-01

    Lysozyme is an interesting molecule for laser ablation of organic materials, because the ablation has been comprehensively studied, it is a medium heavy molecule with a mass of 14305 Da, which can be detected by standard techniques, and because it is used as a bactericidal protein in the food industry. Lysozyme molecules do not absorb energy for wavelengths above 310 nm, but nevertheless there is a strong mass loss by ablation for laser irradiation in the visible regime. The total ablation yi...

  13. Pluronic-lysozyme conjugates as anti-adhesive and antibacterial bifunctional polymers for surface coating

    OpenAIRE

    Muszanska, Agnieszka K.; Busscher, Henk J.; Herrmann, Andreas; van der Mei, Henny C.; Norde, Willem

    2011-01-01

    This paper describes the preparation and characterization of polymer protein conjugates composed of a synthetic triblock copolymer with a central polypropylene oxide (PPO) block and two terminal polyethylene oxide (PEO) segments, Pluronic F-127, and the antibacterial enzyme lysozyme attached to the telechelic groups of the PEO chains. Covalent conjugation of lysozyme proceeded via reductive amination of aldehyde functionalized PEO blocks (CHO-Pluronic) and the amine groups of the lysine resid...

  14. Combining structure and dynamics: non-denaturing high-pressure effect on lysozyme in solution

    OpenAIRE

    Ortore, Maria Grazia; Spinozzi, Francesco; Mariani, Paolo; Paciaroni, Alessandro; Barbosa, Leandro R. S.; Amenitsch, Heinz; Steinhart, Milos; Ollivier, Jacques; Russo, Daniela

    2009-01-01

    Small-angle X-ray scattering (SAXS) and elastic and quasi-elastic neutron scattering techniques were used to investigate the high-pressure-induced changes on interactions, the low-resolution structure and the dynamics of lysozyme in solution. SAXS data, analysed using a global-fit procedure based on a new approach for hydrated protein form factor description, indicate that lysozyme completely maintains its globular structure up to 1500 bar, but significant modifications in the protein–protein...

  15. Protein-Protein Interaction on Lysozyme Crystallization Revealed by Rotational Diffusion Analysis

    OpenAIRE

    Takahashi, Daisuke; Nishimoto, Etsuko; Murase, Tadashi; Yamashita, Shoji

    2008-01-01

    Intermolecular interactions between protein molecules diffusing in various environments underlie many biological processes as well as control protein crystallization, which is a crucial step in x-ray protein structure determinations. Protein interactions were investigated through protein rotational diffusion analysis. First, it was confirmed that tetragonal lysozyme crystals containing fluorescein-tagged lysozyme were successfully formed with the same morphology as that of native protein. Usi...

  16. Lysozyme gene activity in chicken macrophages is controlled by positive and negative regulatory elements.

    OpenAIRE

    Steiner, C; Muller, M; Baniahmad, A; Renkawitz, R

    1987-01-01

    The chicken lysozyme gene is constitutively active in macrophages and under the control of steroid hormones in the oviduct. To investigate which DNA elements are involved in the control of its expression in macrophages we performed transient DNA transfer experiments with two different types of plasmids: 5'-deletion mutants of the upstream region of the chicken lysozyme gene and different fragments from this area in front of the thymidine kinase promoter (herpes simplex virus), each placed in ...

  17. Analysis of an approach to oviduct-specific expression of modified chicken lysozyme genes.

    Science.gov (United States)

    Shimizu, Mamiko; Losos, Jan K; Gibbins, Ann M Verrinder

    2005-02-01

    The -2.7 kb enhancer (E) element of the chicken lysozyme gene domain appears to govern expression of the gene in macrophages but not in oviduct tubular gland cells, the only other site of lysozyme expression. The ultimate goal of our research was to determine whether lysozyme domain variants could be developed that would mainly be expressed in the oviduct so that transgenic birds could be produced that would deposit exogenous protein in the egg white. Accordingly, precise mutations were made by poxvirus-mediated gene targeting in FEF/PU.1 and CCAAT/enhancer-binding protein (C/EBP) transcription factor binding sites in the -2.7 kb E of cloned copies of a specific lysozyme gene variant that includes a hydrophobic pentapeptide tail encoding sequence inserted immediately prior to the stop codon. This variant contains the entire lysozyme domain and is cloned in a lambda bacteriophage vector (lambdaDIILys-HT); the novel tail sequence enables distinction in cell-based expression systems between transcripts of the variant and those of the endogenous gene. These various lysozyme domain mutants, in bacteriophage vector form, were tested for expression in cultured chicken blastodermal cells cotransfected with plasmids encoding the transcription factors C/EBP and v-Myb. In the absence of these plasmids, barely detectable levels of endogenous lysozyme gene transcription resulted in the blastodermal cells. In the presence of the plasmids, however, transcripts of the endogenous gene could be detected as well as varying levels (as evaluated by quantitative real-time PCR) of transcripts of all of the lysozyme domain mutants. These results are discussed in the context of the known role and occurrence of various transcription factors involved in gene expression in differentiating macrophage cells. The ultimate test of expression of the variants in macrophages vs. oviduct cells will be to use them to produce transgenic birds.

  18. An entomological and seroepidemiological study of the vectorial-transmission risk of Chagas disease in the coast of northern Chile.

    Science.gov (United States)

    González, C R; Reyes, C; Canals, A; Parra, A; Muñoz, X; Rodríguez, K

    2015-12-01

    Four species of triatomines are known from Chile: Triatoma infestans Klug, Mepraia spinolai Porter, M. gajardoi Frías, Henry & González, and M. parapatrica Frías (Hemiptera: Reduviidae), the last three are endemic. The geographical distribution of M. gajardoi includes the coastal areas in the north of Chile between 18° and 21°S, an area with both a resident workforce and summer-season visitors. A study was developed to assess the risk of vectorial transmission of Chagas disease by M. gajardoi in hut settlements on the coast of the Tarapacá Region, in particular in Caleta San Marcos and Caleta Río Seco. The study comprised fingerstick sampling of 95 persons, venous samples from 29 domestic dogs and capture of 52 triatomines, from both fishing coves. The samples were analysed by enzyme-linked immunosorbent assay (ELISA) and polymerase chain reaction (PCR) techniques. The results show that, of the total number of persons studied, 100% were negative for Trypanosoma cruzi Chagas (Trypanosomatida: Trypanosomatidae) antibodies, 10.34% of canids were positive for the antibody and 5.8% of M. gajardoi were infected to the PCR technique. The presence of this species in areas close to human settlements constitutes a risk to human populations established on the coast of northern Chile. © 2015 The Royal Entomological Society.

  19. Photopolarization of Fucus zygotes is determined by time sensitive vectorial addition of environmental cues during axis amplification

    Directory of Open Access Journals (Sweden)

    Kenny Arthur Bogaert

    2015-02-01

    Full Text Available Fucoid zygotes have been extensively used to study cell polarization and asymmetrical cell division. Fertilized eggs are responsive to different environmental cues (e.g. light, gravity for a long period before the polarity is fixed and the cells germinate accordingly. First, it is commonly believed that the direction and sense of the polarization vector are established simultaneously as indicated by the formation of an F-actin patch. Secondly, upon reorientation of the zygote, a new polar gradient is formed and it is assumed that the position of the future rhizoid pole is only influenced by the latter. Here we tested these two hypotheses investigating photopolarization in Fucus zygotes by reorienting zygotes 90° relative to a unilateral light source at different time points during the first cell cycle. We conclude that fixation of direction and sense of the polarization vector is indeed established simultaneously. However, the experiments yielded a distribution of polarization axes that cannot be explained if only the last environmental cue is supposed to determine the polarization axis. We conclude that our observations, together with published findings, can only be explained by assuming imprinting of the different polarization vectors and their integration as a vectorial sum at the moment of axis fixation. This way cells will average different serially perceived cues resulting in a polarization vector representative of the dynamic intertidal environment, instead of betting exclusively on the perceived vector at the moment of axis fixation.

  20. A full vectorial contrast source inversion scheme for three-dimensional acoustic imaging of both compressibility and density profiles.

    Science.gov (United States)

    van Dongen, Koen W A; Wright, William M D

    2007-03-01

    Imaging the two acoustic medium parameters density and compressibility requires the use of both the acoustic pressure and velocity wave fields, described via integral equations. Imaging is based on solving for the unknown medium parameters using known measured scattered wave fields, and it is difficult to solve this ill-posed inverse problem directly using a conjugate gradient inversion scheme. Here, a contrast source inversion method is used in which the contrast sources, defined via the product of changes in compressibility and density with the pressure and velocity wave fields, respectively, are computed iteratively. After each update of the contrast sources, an update of the medium parameters is obtained. Total variation as multiplicative regularization is used to minimize blurring in the reconstructed contrasts. The method successfully reconstructed three-dimensional contrast profiles based on changes in both density and compressibility, using synthetic data both with and without 50% white noise. The results were compared with imaging based only on the pressure wave field, where speed of sound profiles were solely based on changes in compressibility. It was found that the results improved significantly by using the full vectorial method when changes in speed of sound depended on changes in both compressibility and density.