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Sample records for bacterial proteins

  1. Bacterial Protein-Tyrosine Kinases

    DEFF Research Database (Denmark)

    Shi, Lei; Kobir, Ahasanul; Jers, Carsten

    2010-01-01

    in exopolysaccharide production, virulence, DNA metabolism, stress response and other key functions of the bacterial cell. BY-kinases act through autophosphorylation (mainly in exopolysaccharide production) and phosphorylation of other proteins, which have in most cases been shown to be activated by tyrosine......Bacteria and Eukarya share essentially the same family of protein-serine/threonine kinases, also known as the Hanks-type kinases. However, when it comes to protein-tyrosine phosphorylation, bacteria seem to have gone their own way. Bacterial protein-tyrosine kinases (BY-kinases) are bacterial...... and highlighted their importance in bacterial physiology. Having no orthologues in Eukarya, BY-kinases are receiving a growing attention from the biomedical field, since they represent a particularly promising target for anti-bacterial drug design....

  2. Bacterial Ice Crystal Controlling Proteins

    Science.gov (United States)

    Lorv, Janet S. H.; Rose, David R.; Glick, Bernard R.

    2014-01-01

    Across the world, many ice active bacteria utilize ice crystal controlling proteins for aid in freezing tolerance at subzero temperatures. Ice crystal controlling proteins include both antifreeze and ice nucleation proteins. Antifreeze proteins minimize freezing damage by inhibiting growth of large ice crystals, while ice nucleation proteins induce formation of embryonic ice crystals. Although both protein classes have differing functions, these proteins use the same ice binding mechanisms. Rather than direct binding, it is probable that these protein classes create an ice surface prior to ice crystal surface adsorption. Function is differentiated by molecular size of the protein. This paper reviews the similar and different aspects of bacterial antifreeze and ice nucleation proteins, the role of these proteins in freezing tolerance, prevalence of these proteins in psychrophiles, and current mechanisms of protein-ice interactions. PMID:24579057

  3. Bacterial binding to extracellular proteins - in vitro adhesion

    DEFF Research Database (Denmark)

    Schou, C.; Fiehn, N.-E.

    1999-01-01

    Viridans streptococci, bacterial adherence, extracellular matrix proteins, surface receptors, endocarditis......Viridans streptococci, bacterial adherence, extracellular matrix proteins, surface receptors, endocarditis...

  4. C-reactive protein and bacterial meningitis

    DEFF Research Database (Denmark)

    Gerdes, Lars Ulrik; Jørgensen, P E; Nexø, E

    1998-01-01

    The aim of the study was to review published articles on the diagnostic accuracy of C-reactive protein (CRP) tests with cerebrospinal fluid and serum in diagnosing bacterial meningitis. The literature from 1980 and onwards was searched using the electronic databases of MEDLINE, and we used summary...... measured in serum, and 4 in which it had been measured in both cerebrospinal fluid and serum. The odds ratio for bacterial meningitis versus aseptic meningitis for a positive CRP test with cerebrospinal fluid was estimated at 241 (95% confidence interval [CI]: 59-980), and the central tendencies.......06-0.08, respectively, the post-test probability of not having bacterial meningitis given a negative test is very high (> or = 97%), in the range of a pre-test probability (prevalence of bacterial meningitis) from 10 to 30%, whereas the post-test probability of bacterial meningitis given a positive test is considerably...

  5. Protein phosphorylation and bacterial chemotaxis

    International Nuclear Information System (INIS)

    Hess, J.F.; Bourret, R.B.; Oosawa, K.; Simon, M.I.; Matsumura, P.

    1988-01-01

    Bacteria are able to respond to changes in concentration of a large variety of chemicals and to changes in physical parameters, including viscosity, osmolarity, and temperature, by swimming toward a more favorable location (for review, see Stewart and Dahlquist 1987). Most chemotactic responses are mediated by a series of transmembrane receptor proteins that interact with or bind specific chemicals and thus monitor environmental conditions. Over the past 10 years, work in a number of laboratories has resulted in the identification and characterization of many of the genes and proteins required for the signal transduction process. The authors postulated that rapid and transient covalent modification of the chemotaxis gene products could function to transmit information from the receptor by regulating protein-protein interaction between the chemotaxis gene products. To test this idea, the authors purified the proteins corresponding to the cheA, cheY, cheZ, cheW, and cheB genes and tested the purified polypeptides to determine whether they could be covalently modified and whether they would interact with each other in vitro

  6. Subcellular sites for bacterial protein export

    NARCIS (Netherlands)

    Campo, Nathalie; Tjalsma, Harold; Buist, Girbe; Stepniak, Dariusz; Meijer, Michel; Veenhuis, Marten; Westermann, Martin; Müller, Jörg P.; Bron, Sierd; Kok, Jan; Kuipers, Oscar P.; Jongbloed, Jan D.H.

    2004-01-01

    Most bacterial proteins destined to leave the cytoplasm are exported to extracellular compartments or imported into the cytoplasmic membrane via the highly conserved SecA-YEG pathway. In the present studies, the subcellular distributions of core components of this pathway, SecA and SecY, and of the

  7. Subcellular sites for bacterial protein export.

    NARCIS (Netherlands)

    Campo, N.; Tjalsma, H.; Buist, G.; Stepniak, D.; Meijer, M.; Veenhuis, M.; Westermann, M.; Muller, J.P.; Bron, S.; Kok, J.; Kuipers, O.P.; Jongbloed, J.D.

    2004-01-01

    Most bacterial proteins destined to leave the cytoplasm are exported to extracellular compartments or imported into the cytoplasmic membrane via the highly conserved SecA-YEG pathway. In the present studies, the subcellular distributions of core components of this pathway, SecA and SecY, and of the

  8. Protein malnutrition and metronidazole induced intestinal bacterial ...

    African Journals Online (AJOL)

    This study was designed to assess the effects of protein malnutrition (PM) associated with antibiotic on growth weight, cecal bacterial overgrowth and enterobacteria translocation. Eighteen Gnotobiotic young Wistar rats (135 ± 2.35 g) were treated orally with antibiotic and submitted to dietary restriction based on maize diet ...

  9. Fluorescent sensors based on bacterial fusion proteins

    International Nuclear Information System (INIS)

    Mateu, Batirtze Prats; Pum, Dietmar; Sleytr, Uwe B; Toca-Herrera, José L; Kainz, Birgit

    2014-01-01

    Fluorescence proteins are widely used as markers for biomedical and technological purposes. Therefore, the aim of this project was to create a fluorescent sensor, based in the green and cyan fluorescent protein, using bacterial S-layers proteins as scaffold for the fluorescent tag. We report the cloning, expression and purification of three S-layer fluorescent proteins: SgsE-EGFP, SgsE-ECFP and SgsE-13aa-ECFP, this last containing a 13-amino acid rigid linker. The pH dependence of the fluorescence intensity of the S-layer fusion proteins, monitored by fluorescence spectroscopy, showed that the ECFP tag was more stable than EGFP. Furthermore, the fluorescent fusion proteins were reassembled on silica particles modified with cationic and anionic polyelectrolytes. Zeta potential measurements confirmed the particle coatings and indicated their colloidal stability. Flow cytometry and fluorescence microscopy showed that the fluorescence of the fusion proteins was pH dependent and sensitive to the underlying polyelectrolyte coating. This might suggest that the fluorescent tag is not completely exposed to the bulk media as an independent moiety. Finally, it was found out that viscosity enhanced the fluorescence intensity of the three fluorescent S-layer proteins. (paper)

  10. Identifying Bacterial Immune Evasion Proteins Using Phage Display.

    Science.gov (United States)

    Fevre, Cindy; Scheepmaker, Lisette; Haas, Pieter-Jan

    2017-01-01

    Methods aimed at identification of immune evasion proteins are mainly rely on in silico prediction of sequence, structural homology to known evasion proteins or use a proteomics driven approach. Although proven successful these methods are limited by a low efficiency and or lack of functional identification. Here we describe a high-throughput genomic strategy to functionally identify bacterial immune evasion proteins using phage display technology. Genomic bacterial DNA is randomly fragmented and ligated into a phage display vector that is used to create a phage display library expressing bacterial secreted and membrane bound proteins. This library is used to select displayed bacterial secretome proteins that interact with host immune components.

  11. Anticancer Activity of Bacterial Proteins and Peptides.

    Science.gov (United States)

    Karpiński, Tomasz M; Adamczak, Artur

    2018-04-30

    Despite much progress in the diagnosis and treatment of cancer, tumour diseases constitute one of the main reasons of deaths worldwide. The side effects of chemotherapy and drug resistance of some cancer types belong to the significant current therapeutic problems. Hence, searching for new anticancer substances and medicines are very important. Among them, bacterial proteins and peptides are a promising group of bioactive compounds and potential anticancer drugs. Some of them, including anticancer antibiotics (actinomycin D, bleomycin, doxorubicin, mitomycin C) and diphtheria toxin, are already used in the cancer treatment, while other substances are in clinical trials (e.g., p28, arginine deiminase ADI) or tested in in vitro research. This review shows the current literature data regarding the anticancer activity of proteins and peptides originated from bacteria: antibiotics, bacteriocins, enzymes, nonribosomal peptides (NRPs), toxins and others such as azurin, p28, Entap and Pep27anal2. The special attention was paid to the still poorly understood active substances obtained from the marine sediment bacteria. In total, 37 chemical compounds or groups of compounds with antitumor properties have been described in the present article.

  12. Production of bacterial protein from sugar cane bagasse pith

    Energy Technology Data Exchange (ETDEWEB)

    Molina, O E; Callieri, D A.S.; Perotti de Galvez, N

    1980-01-01

    Bacterial protein was produced during the fermentation of sugar cane bagasse pith (BP) by a mixture of cellulolytic bacteria, one of them being a species of Cellulomonas. If the BP were treated with 1% NaOH prior to fermentation, the liquor could be used twice more without affecting the yield of bacterial protein. After that, the liquor became too dark and impaired the subsequent washing of BP. If the concentration of N (as NaN0/sub 3/) in the fermentation medium were raised, the conversion factor to protein was lowered, but the amount of protein formed per L per h and the ratio of protein to BP became higher. The evolution of pH, the dry matter content, cellulolytic activity, and protein yield were all affected by the type of N source used. The yield of bacterial protein can probably be increased by automatically controlling the pH and dissolved O levels of the culture.

  13. Computational Analysis of Uncharacterized Proteins of Environmental Bacterial Genome

    Science.gov (United States)

    Coxe, K. J.; Kumar, M.

    2017-12-01

    Betaproteobacteria strain CB is a gram-negative bacterium in the phylum Proteobacteria and are found naturally in soil and water. In this complex environment, bacteria play a key role in efficiently eliminating the organic material and other pollutants from wastewater. To investigate the process of pollutant removal from wastewater using bacteria, it is important to characterize the proteins encoded by the bacterial genome. Our study combines a number of bioinformatics tools to predict the function of unassigned proteins in the bacterial genome. The genome of Betaproteobacteria strain CB contains 2,112 proteins in which function of 508 proteins are unknown, termed as uncharacterized proteins (UPs). The localization of the UPs with in the cell was determined and the structure of 38 UPs was accurately predicted. These UPs were predicted to belong to various classes of proteins such as enzymes, transporters, binding proteins, signal peptides, transmembrane proteins and other proteins. The outcome of this work will help better understand wastewater treatment mechanism.

  14. Hijacking Complement Regulatory Proteins for Bacterial Immune Evasion.

    Science.gov (United States)

    Hovingh, Elise S; van den Broek, Bryan; Jongerius, Ilse

    2016-01-01

    The human complement system plays an important role in the defense against invading pathogens, inflammation and homeostasis. Invading microbes, such as bacteria, directly activate the complement system resulting in the formation of chemoattractants and in effective labeling of the bacteria for phagocytosis. In addition, formation of the membrane attack complex is responsible for direct killing of Gram-negative bacteria. In turn, bacteria have evolved several ways to evade complement activation on their surface in order to be able to colonize and invade the human host. One important mechanism of bacterial escape is attraction of complement regulatory proteins to the microbial surface. These molecules are present in the human body for tight regulation of the complement system to prevent damage to host self-surfaces. Therefore, recruitment of complement regulatory proteins to the bacterial surface results in decreased complement activation on the microbial surface which favors bacterial survival. This review will discuss recent advances in understanding the binding of complement regulatory proteins to the bacterial surface at the molecular level. This includes, new insights that have become available concerning specific conserved motives on complement regulatory proteins that are favorable for microbial binding. Finally, complement evasion molecules are of high importance for vaccine development due to their dominant role in bacterial survival, high immunogenicity and homology as well as their presence on the bacterial surface. Here, the use of complement evasion molecules for vaccine development will be discussed.

  15. Demodex-associated bacterial proteins induce neutrophil activation.

    LENUS (Irish Health Repository)

    2012-02-01

    Background: Patients with rosacea demonstrate a higher density of Demodex mites in their skin than controls. A bacterium isolated from a Demodex mite from a patient with papulopustular rosacea (PPR) was previously shown to provoke an immune response in patients with PPR or ocular rosacea thus suggesting a possible role for bacterial proteins in the etiology of this condition. Objectives: To examine the response of neutrophils to proteins derived from a bacterium isolated from a Demodex mite. Methods: Bacterial cells were lysed and proteins were partially purified by AKTA-FPLC. Isolated neutrophils were exposed to bacterial proteins and monitored for alterations in migration, degranulation and cytokine production. Results: Neutrophils exposed to proteins from Bacillus cells demonstrated increased levels of migration and elevated release of MMP-9, an enzyme known to degrade collagen and cathelicidin, an antimicrobial peptide. In addition neutrophils exposed to the bacterial proteins demonstrated elevated rates of Il-8 and TNF-alpha production. Conclusions: Proteins produced by a bacterium isolated from a Demodex mite have the ability to increase the migration, degranulation and cytokine production abilities of neutrophils. These results suggest that bacteria may play a role in the inflammatory erythema associated with rosacea.

  16. Rehosting of Bacterial Chaperones for High-Quality Protein Production▿

    Science.gov (United States)

    Martínez-Alonso, Mónica; Toledo-Rubio, Verónica; Noad, Rob; Unzueta, Ugutz; Ferrer-Miralles, Neus; Roy, Polly; Villaverde, Antonio

    2009-01-01

    Coproduction of DnaK/DnaJ in Escherichia coli enhances solubility but promotes proteolytic degradation of their substrates, minimizing the yield of unstable polypeptides. Higher eukaryotes have orthologs of DnaK/DnaJ but lack the linked bacterial proteolytic system. By coexpression of DnaK and DnaJ in insect cells with inherently misfolding-prone recombinant proteins, we demonstrate simultaneous improvement of soluble protein yield and quality and proteolytic stability. Thus, undesired side effects of bacterial folding modulators can be avoided by appropriate rehosting in heterologous cell expression systems. PMID:19820142

  17. Structural Aspects of Bacterial Outer Membrane Protein Assembly.

    Science.gov (United States)

    Calmettes, Charles; Judd, Andrew; Moraes, Trevor F

    2015-01-01

    The outer membrane of Gram-negative bacteria is predominantly populated by β-Barrel proteins and lipid anchored proteins that serve a variety of biological functions. The proper folding and assembly of these proteins is essential for bacterial viability and often plays a critical role in virulence and pathogenesis. The β-barrel assembly machinery (Bam) complex is responsible for the proper assembly of β-barrels into the outer membrane of Gram-negative bacteria, whereas the localization of lipoproteins (Lol) system is required for proper targeting of lipoproteins to the outer membrane.

  18. Blocking of bacterial biofilm formation by a fish protein coating

    DEFF Research Database (Denmark)

    Vejborg, Rebecca Munk; Klemm, Per

    2008-01-01

    Bacterial biofilm formation on inert surfaces is a significant health and economic problem in a wide range of environmental, industrial, and medical areas. Bacterial adhesion is generally a prerequisite for this colonization process and, thus, represents an attractive target for the development......, this proteinaceous coating is characterized with regards to its biofilm-reducing properties by using a range of urinary tract infectious isolates with various pathogenic and adhesive properties. The antiadhesive coating significantly reduced or delayed biofilm formation by all these isolates under every condition...... examined. The biofilm-reducing activity did, however, vary depending on the substratum physicochemical characteristics and the environmental conditions studied. These data illustrate the importance of protein conditioning layers with respect to bacterial biofilm formation and suggest that antiadhesive...

  19. Recombinant Expression Screening of P. aeruginosa Bacterial Inner Membrane Proteins

    Directory of Open Access Journals (Sweden)

    Jeffery Constance J

    2010-11-01

    Full Text Available Abstract Background Transmembrane proteins (TM proteins make up 25% of all proteins and play key roles in many diseases and normal physiological processes. However, much less is known about their structures and molecular mechanisms than for soluble proteins. Problems in expression, solubilization, purification, and crystallization cause bottlenecks in the characterization of TM proteins. This project addressed the need for improved methods for obtaining sufficient amounts of TM proteins for determining their structures and molecular mechanisms. Results Plasmid clones were obtained that encode eighty-seven transmembrane proteins with varying physical characteristics, for example, the number of predicted transmembrane helices, molecular weight, and grand average hydrophobicity (GRAVY. All the target proteins were from P. aeruginosa, a gram negative bacterial opportunistic pathogen that causes serious lung infections in people with cystic fibrosis. The relative expression levels of the transmembrane proteins were measured under several culture growth conditions. The use of E. coli strains, a T7 promoter, and a 6-histidine C-terminal affinity tag resulted in the expression of 61 out of 87 test proteins (70%. In this study, proteins with a higher grand average hydrophobicity and more transmembrane helices were expressed less well than less hydrophobic proteins with fewer transmembrane helices. Conclusions In this study, factors related to overall hydrophobicity and the number of predicted transmembrane helices correlated with the relative expression levels of the target proteins. Identifying physical characteristics that correlate with protein expression might aid in selecting the "low hanging fruit", or proteins that can be expressed to sufficient levels using an E. coli expression system. The use of other expression strategies or host species might be needed for sufficient levels of expression of transmembrane proteins with other physical

  20. Bacterial protein meal in diets for pigs and minks

    DEFF Research Database (Denmark)

    Hellwing, Anne Louise Frydendahl; Tauson, Anne-Helene; Skrede, Anders

    2007-01-01

    The effect of increasing the dietary content of bacterial protein meal (BPM) on protein turnover rate, and on nucleic acid and creatinine metabolism in growing minks and pigs was investigated in two experiments. In each experiment, 16 animals were allocated to four experimental diets. The diets...... containing no BPM served as controls, i.e. for minks diet M1, for pigs P1; the experimental diets contained increasing levels of BPM to replace fish meal (minks) or soybean meal (pigs), so that up to 17% (P2), 20% (M2), 35% (P3), 40% (M3), 52% (P4), and 60% (M4) of digestible N was BPM derived. Protein...... turnover rate was measured by means of the end-product method using [15N]glycine as tracer and urinary nitrogen as end-product. In minks, protein flux, synthesis, and breakdown increased significantly with increasing dietary BPM. In pigs, diet had no observed effect on protein turnover rate. The intake...

  1. Bacterial S-layer protein coupling to lipids

    DEFF Research Database (Denmark)

    Weygand, M.; Wetzer, B.; Pum, D.

    1999-01-01

    structure before and after protein recrystallization shows minimal reorganization of the lipid chains. By contrast, the lipid headgroups show major rearrangements. For the B. sphaericus CCM2177 protein underneath DPPE monolayers, x-ray reflectivity data suggest that amino acid side chains intercalate......The coupling of bacterial surface (S)-layer proteins to lipid membranes is studied in molecular detail for proteins from Bacillus sphaericus CCM2177 and B. coagulans E38-66 recrystallized at dipalmitoylphosphatidylethanolamine (DPPE) monolayers on aqueous buffer. A comparison of the monolayer...... the lipid headgroups at least to the phosphate moieties, and probably further beyond. The number of electrons in the headgroup region increases by more than four per lipid. Analysis of the changes of the deduced electron density profiles in terms of a molecular interpretation shows...

  2. Packaging protein drugs as bacterial inclusion bodies for therapeutic applications

    Directory of Open Access Journals (Sweden)

    Villaverde Antonio

    2012-06-01

    Full Text Available Abstract A growing number of insights on the biology of bacterial inclusion bodies (IBs have revealed intriguing utilities of these protein particles. Since they combine mechanical stability and protein functionality, IBs have been already exploited in biocatalysis and explored for bottom-up topographical modification in tissue engineering. Being fully biocompatible and with tuneable bio-physical properties, IBs are currently emerging as agents for protein delivery into mammalian cells in protein-replacement cell therapies. So far, IBs formed by chaperones (heat shock protein 70, Hsp70, enzymes (catalase and dihydrofolate reductase, grow factors (leukemia inhibitory factor, LIF and structural proteins (the cytoskeleton keratin 14 have been shown to rescue exposed cells from a spectrum of stresses and restore cell functions in absence of cytotoxicity. The natural penetrability of IBs into mammalian cells (reaching both cytoplasm and nucleus empowers them as an unexpected platform for the controlled delivery of essentially any therapeutic polypeptide. Production of protein drugs by biopharma has been traditionally challenged by IB formation. However, a time might have arrived in which recombinant bacteria are to be engineered for the controlled packaging of therapeutic proteins as nanoparticulate materials (nanopills, for their extra- or intra-cellular release in medicine and cosmetics.

  3. C-REACTIVE PROTEIN IN BACTERIAL MENINGITIS: DOSE IT HELP TO DIFFERENTIATE BACTERIAL FROM VIRAL MENINGITIS?

    Directory of Open Access Journals (Sweden)

    AR EMAMI NAEINI

    2001-03-01

    Full Text Available Introduction. Central nervous system infections are among the most serious conditions in of medical practice. C-reactive Protein has recently been evaluated in terms of its ability to diffeccentiate bacterial from nonbacterial central nervous system inflammations.
    Methods. We studied the frequency of positive CRP in 61 patients who had signs of meningitis. All the specimens referred to one laboratory and were examined by Slide method.
    Results. Positive CRP was found in 97.6 percent of those who were finally diagnosed as bacterial meningitis. The frequency of CRP for other types of meningitis was 16.6 percent (P < 0.05.
    Discussion. In the absence of infection, CSF is free of CRP. Positive CRP may help to the differentiate the different types of meningitis.

  4. Impact of fluorescent protein fusions on the bacterial flagellar motor.

    Science.gov (United States)

    Heo, M; Nord, A L; Chamousset, D; van Rijn, E; Beaumont, H J E; Pedaci, F

    2017-10-03

    Fluorescent fusion proteins open a direct and unique window onto protein function. However, they also introduce the risk of perturbation of the function of the native protein. Successful applications of fluorescent fusions therefore rely on a careful assessment and minimization of the side effects, but such insight is still lacking for many applications. This is particularly relevant in the study of the internal dynamics of motor proteins, where both the chemical and mechanical reaction coordinates can be affected. Fluorescent proteins fused to the stator of the Bacterial Flagellar Motor (BFM) have previously been used to unveil the motor subunit dynamics. Here we report the effects on single motors of three fluorescent proteins fused to the stators, all of which altered BFM behavior. The torque generated by individual stators was reduced while their stoichiometry remained unaffected. MotB fusions decreased the switching frequency and induced a novel bias-dependent asymmetry in the speed in the two directions. These effects could be mitigated by inserting a linker at the fusion point. These findings provide a quantitative account of the effects of fluorescent fusions to the stator on BFM dynamics and their alleviation- new insights that advance the use of fluorescent fusions to probe the dynamics of protein complexes.

  5. Protein export through the bacterial flagellar type III export pathway.

    Science.gov (United States)

    Minamino, Tohru

    2014-08-01

    For construction of the bacterial flagellum, which is responsible for bacterial motility, the flagellar type III export apparatus utilizes both ATP and proton motive force across the cytoplasmic membrane and exports flagellar proteins from the cytoplasm to the distal end of the nascent structure. The export apparatus consists of a membrane-embedded export gate made of FlhA, FlhB, FliO, FliP, FliQ, and FliR and a water-soluble ATPase ring complex consisting of FliH, FliI, and FliJ. FlgN, FliS, and FliT act as substrate-specific chaperones that do not only protect their cognate substrates from degradation and aggregation in the cytoplasm but also efficiently transfer the substrates to the export apparatus. The ATPase ring complex facilitates the initial entry of the substrates into the narrow pore of the export gate. The export gate by itself is a proton-protein antiporter that uses the two components of proton motive force, the electric potential difference and the proton concentration difference, for different steps of the export process. A specific interaction of FlhA with FliJ located in the center of the ATPase ring complex allows the export gate to efficiently use proton motive force to drive protein export. The ATPase ring complex couples ATP binding and hydrolysis to its assembly-disassembly cycle for rapid and efficient protein export cycle. This article is part of a Special Issue entitled: Protein trafficking and secretion in bacteria. Guest Editors: Anastassios Economou and Ross Dalbey. © 2013 Elsevier B.V. All rights reserved.

  6. Extractable Bacterial Surface Proteins in Probiotic–Host Interaction

    Directory of Open Access Journals (Sweden)

    Fillipe L. R. do Carmo

    2018-04-01

    Full Text Available Some Gram-positive bacteria, including probiotic ones, are covered with an external proteinaceous layer called a surface-layer. Described as a paracrystalline layer and formed by the self-assembly of a surface-layer-protein (Slp, this optional structure is peculiar. The surface layer per se is conserved and encountered in many prokaryotes. However, the sequence of the corresponding Slp protein is highly variable among bacterial species, or even among strains of the same species. Other proteins, including surface layer associated proteins (SLAPs, and other non-covalently surface-bound proteins may also be extracted with this surface structure. They can be involved a various functions. In probiotic Gram-positives, they were shown by different authors and experimental approaches to play a role in key interactions with the host. Depending on the species, and sometime on the strain, they can be involved in stress tolerance, in survival within the host digestive tract, in adhesion to host cells or mucus, or in the modulation of intestinal inflammation. Future trends include the valorization of their properties in the formation of nanoparticles, coating and encapsulation, and in the development of new vaccines.

  7. Bacterial protein meal in diets for growing pigs

    DEFF Research Database (Denmark)

    Hellwing, Anne Louise Frydendahl; Tauson, Anne-Helene; Kjos, N.P.

    2007-01-01

    This experiment investigated the effects of increasing the dietary content of bacterial protein meal (BPM) on the protein and energy metabolism of pigs from weaning to a live weight of 80 kg. FOur litters with four castrated male pigs in each litter were used. The litters were divided into two...... blocks according to age. One pig from each litter was fed one of the four experimental diets. Soya-bean meal was replaced with BPM on the basis of digestible protein, and the BPM contents in the four diets were 0% (BP0), 5% (BP5), 10% (BP10) and 15% (BP15), corresponding to 0%, 17%, 35% and 52...... by inclusion level of BPM. Retention of energy was 620 (BP0), 696 (BP5), 613 (BP10) and 664 kJ/kg0.75 per day (BP15), the differences among diets being non-significant. The N-free respiratory quotient was similar on all diets. It was concluded that the overall protein and energy metabolism in growing pigs were...

  8. Bacterial flagellar capping proteins adopt diverse oligomeric states

    Energy Technology Data Exchange (ETDEWEB)

    Postel, Sandra; Deredge, Daniel; Bonsor, Daniel A.; Yu, Xiong; Diederichs, Kay; Helmsing, Saskia; Vromen, Aviv; Friedler, Assaf; Hust, Michael; Egelman, Edward H.; Beckett, Dorothy; Wintrode, Patrick L.; Sundberg, Eric J. (UV); (Braunschweig); (Maryland-MED); (Konstanz); (Maryland); (Hebrew)

    2016-09-24

    Flagella are crucial for bacterial motility and pathogenesis. The flagellar capping protein (FliD) regulates filament assembly by chaperoning and sorting flagellin (FliC) proteins after they traverse the hollow filament and exit the growing flagellum tip. In the absence of FliD, flagella are not formed, resulting in impaired motility and infectivity. Here, we report the 2.2 Å resolution X-ray crystal structure of FliD fromPseudomonas aeruginosa, the first high-resolution structure of any FliD protein from any bacterium. Using this evidence in combination with a multitude of biophysical and functional analyses, we find thatPseudomonasFliD exhibits unexpected structural similarity to other flagellar proteins at the domain level, adopts a unique hexameric oligomeric state, and depends on flexible determinants for oligomerization. Considering that the flagellin filaments on which FliD oligomers are affixed vary in protofilament number between bacteria, our results suggest that FliD oligomer stoichiometries vary across bacteria to complement their filament assemblies.

  9. C-reactive protein velocity to distinguish febrile bacterial infections from non-bacterial febrile illnesses in the emergency department

    OpenAIRE

    Paran, Yael; Yablecovitch, Doron; Choshen, Guy; Zeitlin, Ina; Rogowski, Ori; Ben-Ami, Ronen; Katzir, Michal; Saranga, Hila; Rosenzweig, Tovit; Justo, Dan; Orbach, Yaffa; Halpern, Pinhas; Berliner, Shlomo

    2009-01-01

    Introduction C-reactive protein (CRP) is a real-time and low-cost biomarker to distinguish febrile bacterial infections from non-bacterial febrile illnesses. We hypothesised that measuring the velocity of the biomarker instead of its absolute serum concentration could enhance its ability to differentiate between these two conditions. Methods We prospectively recruited adult patients (age ? 18 years) who presented to the emergency department with fever. We recorded their data regarding the ons...

  10. Ribosome reinitiation at leader peptides increases translation of bacterial proteins.

    Science.gov (United States)

    Korolev, Semen A; Zverkov, Oleg A; Seliverstov, Alexandr V; Lyubetsky, Vassily A

    2016-04-16

    Short leader genes usually do not encode stable proteins, although their importance in expression control of bacterial genomes is widely accepted. Such genes are often involved in the control of attenuation regulation. However, the abundance of leader genes suggests that their role in bacteria is not limited to regulation. Specifically, we hypothesize that leader genes increase the expression of protein-coding (structural) genes via ribosome reinitiation at the leader peptide in the case of a short distance between the stop codon of the leader gene and the start codon of the structural gene. For instance, in Actinobacteria, the frequency of leader genes at a distance of 10-11 bp is about 70 % higher than the mean frequency within the 1 to 65 bp range; and it gradually decreases as the range grows longer. A pronounced peak of this frequency-distance relationship is also observed in Proteobacteria, Bacteroidetes, Spirochaetales, Acidobacteria, the Deinococcus-Thermus group, and Planctomycetes. In contrast, this peak falls to the distance of 15-16 bp and is not very pronounced in Firmicutes; and no such peak is observed in cyanobacteria and tenericutes. Generally, this peak is typical for many bacteria. Some leader genes located close to a structural gene probably play a regulatory role as well.

  11. Hierarchical protein export mechanism of the bacterial flagellar type III protein export apparatus.

    Science.gov (United States)

    Minamino, Tohru

    2018-06-01

    The bacterial flagellum is supramolecular motility machinery consisting of the basal body, the hook and the filament. Flagellar proteins are translocated across the cytoplasmic membrane via a type III protein export apparatus, diffuse down the central channel of the growing structure and assemble at the distal end. Flagellar assembly begins with the basal body, followed by the hook and finally the filament. The completion of hook assembly is the most important morphological checkpoint of the sequential flagellar assembly process. When the hook reaches its mature length of about 55 nm in Salmonella enterica, the type III protein export apparatus switches export specificity from proteins required for the structure and assembly of the hook to those responsible for filament assembly, thereby terminating hook assembly and initiating filament assembly. Three flagellar proteins, namely FliK, FlhB and FlhA, are responsible for this substrate specificity switching. Upon completion of the switching event, interactions among FlhA, the cytoplasmic ATPase complex and flagellar type III export chaperones establish the assembly order of the filament at the hook tip. Here, we describe our current understanding of a hierarchical protein export mechanism used in flagellar type III protein export.

  12. DMPD: The role of Toll-like receptors and Nod proteins in bacterial infection. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 15476921 The role of Toll-like receptors and Nod proteins in bacterial infection. P...of Toll-like receptors and Nod proteins in bacterial infection. PubmedID 15476921 Title The role of Toll-like receptors and Nod prote...ins in bacterial infection. Authors Philpott DJ, Girardi

  13. Exploring the diversity of protein modifications: special bacterial phosphorylation systems

    DEFF Research Database (Denmark)

    Mijakovic, Ivan; Grangeasse, Christophe; Turgay, Kürşad

    2016-01-01

    Protein modifications not only affect protein homeostasis but can also establish new cellular protein functions and are important components of complex cellular signal sensing and transduction networks. Among these post-translational modifications, protein phosphorylation represents the one that ...

  14. Subversion of the Endocytic and Secretory Pathways by Bacterial Effector Proteins

    Directory of Open Access Journals (Sweden)

    Mary M. Weber

    2018-01-01

    Full Text Available Intracellular bacteria have developed numerous strategies to hijack host vesicular trafficking pathways to form their unique replicative niches. To promote intracellular replication, the bacteria must interact with host organelles and modulate host signaling pathways to acquire nutrients and membrane for the growing parasitophorous vacuole all while suppressing activation of the immune response. To facilitate host cell subversion, bacterial pathogens use specialized secretion systems to deliver bacterial virulence factors, termed effectors, into the host cell that mimic, agonize, and/or antagonize the function of host proteins. In this review we will discuss how bacterial effector proteins from Coxiella burnetii, Brucella abortus, Salmonella enterica serovar Typhimurium, Legionella pneumophila, Chlamydia trachomatis, and Orientia tsutsugamushi manipulate the endocytic and secretory pathways. Understanding how bacterial effector proteins manipulate host processes not only gives us keen insight into bacterial pathogenesis, but also enhances our understanding of how eukaryotic membrane trafficking is regulated.

  15. Imparting albumin-binding affinity to a human protein by mimicking the contact surface of a bacterial binding protein.

    Science.gov (United States)

    Oshiro, Satoshi; Honda, Shinya

    2014-04-18

    Attachment of a bacterial albumin-binding protein module is an attractive strategy for extending the plasma residence time of protein therapeutics. However, a protein fused with such a bacterial module could induce unfavorable immune reactions. To address this, we designed an alternative binding protein by imparting albumin-binding affinity to a human protein using molecular surface grafting. The result was a series of human-derived 6 helix-bundle proteins, one of which specifically binds to human serum albumin (HSA) with adequate affinity (KD = 100 nM). The proteins were designed by transferring key binding residues of a bacterial albumin-binding module, Finegoldia magna protein G-related albumin-binding domain (GA) module, onto the human protein scaffold. Despite 13-15 mutations, the designed proteins maintain the original secondary structure by virtue of careful grafting based on structural informatics. Competitive binding assays and thermodynamic analyses of the best binders show that the binding mode resembles that of the GA module, suggesting that the contacting surface of the GA module is mimicked well on the designed protein. These results indicate that the designed protein may act as an alternative low-risk binding module to HSA. Furthermore, molecular surface grafting in combination with structural informatics is an effective approach for avoiding deleterious mutations on a target protein and for imparting the binding function of one protein onto another.

  16. EEVD motif of heat shock cognate protein 70 contributes to bacterial uptake by trophoblast giant cells

    Directory of Open Access Journals (Sweden)

    Kim Suk

    2009-12-01

    Full Text Available Abstract Background The uptake of abortion-inducing pathogens by trophoblast giant (TG cells is a key event in infectious abortion. However, little is known about phagocytic functions of TG cells against the pathogens. Here we show that heat shock cognate protein 70 (Hsc70 contributes to bacterial uptake by TG cells and the EEVD motif of Hsc70 plays an important role in this. Methods Brucella abortus and Listeria monocytogenes were used as the bacterial antigen in this study. Recombinant proteins containing tetratricopeptide repeat (TPR domains were constructed and confirmation of the binding capacity to Hsc70 was assessed by ELISA. The recombinant TPR proteins were used for investigation of the effect of TPR proteins on bacterial uptake by TG cells and on pregnancy in mice. Results The monoclonal antibody that inhibits bacterial uptake by TG cells reacted with the EEVD motif of Hsc70. Bacterial TPR proteins bound to the C-terminal of Hsc70 through its EEVD motif and this binding inhibited bacterial uptake by TG cells. Infectious abortion was also prevented by blocking the EEVD motif of Hsc70. Conclusions Our results demonstrate that surface located Hsc70 on TG cells mediates the uptake of pathogenic bacteria and proteins containing the TPR domain inhibit the function of Hsc70 by binding to its EEVD motif. These molecules may be useful in the development of methods for preventing infectious abortion.

  17. An ER-directed fusion protein comprising a bacterial subtilisin ...

    African Journals Online (AJOL)

    nausch

    subtilase tag was fused to human interleukin 6 (IL6) and transiently expressed in Nicotiana ..... MP, tobacco mosaic virus (TMV) movement protein; TVCV-3'-NTR, TVCV-3' untranslated .... on the degradation pattern of heterologous proteins.

  18. Impact of fluorescent protein fusions on the bacterial flagellar motor

    NARCIS (Netherlands)

    Heo, M.; Nord, A. L.; Chamousset, D.; van Rijn, E.; Beaumont, H.J.E.; Pedaci, F.

    2017-01-01

    Fluorescent fusion proteins open a direct and unique window onto protein function. However, they also introduce the risk of perturbation of the function of the native protein. Successful applications of fluorescent fusions therefore rely on a careful assessment and minimization of the side

  19. The diagnostic value of c-reactive protein estimation in differentiating bacterial from viral meningitis

    International Nuclear Information System (INIS)

    Sheikh, A.

    2001-01-01

    Objective: To evaluate the efficacy of serum and CSF C-reactive protein (C-rp) in differentiating bacterial from viral meningitis. Design: An observational, respective hospital-based study. Place and duration of study: It was conducted at the Department of Medicine and Department of Pediatrics, Shaikh Zayed Postgraduate Medical Institute Lahore, Over a Period of one year between march, 1999 and March, 2000. Subject and Methods: A randomized group of thirty patients, who presented with clinical features, suggestive of meningitis, were included in the study. C-reactive protein determinations were performed by latex agglutination method on the serum and cerebrospinal fluid (CSF) of these patients. Results: In the present study, c-reactive protein was found to be a more sensitive test for differentiating bacterial from non-bacterial meningitis on initial examination than the usual conventional methods used to diagnose bacterial meningitis. CSF C-reactive protein had a greater sensitivity (92% as compared to serum C-reactive protein (71%). Conclusion: C-reactive protein determination in CSF was found to be a useful indicator of bacterial meningitis that can be used to distinguish it from viral meningitis. (author)

  20. Protein Adsorption and Its Role in Bacterial Film Development

    Science.gov (United States)

    1989-06-27

    only the secondary antibody conjugated to alkaline phosphatase was used. Combined Amino Acids as Measured by HPLC We are interested in a simple, direct...specific assay for chitin that relies on the lectin, wheat germ agglutinin (WGA). Lectins are a general class of proteins that bind to carbohydrates. The...protein; 2) a new method for measuring combined amino acids (includes proteins) in seawater was shown to measure higher concentration than the old

  1. Measurement of the incorporation rates of four amino acids into proteins for estimating bacterial production.

    Science.gov (United States)

    Servais, P

    1995-03-01

    In aquatic ecosystems, [(3)H]thymidine incorporation into bacterial DNA and [(3)H]leucine incorporation into proteins are usually used to estimate bacterial production. The incorporation rates of four amino acids (leucine, tyrosine, lysine, alanine) into proteins of bacteria were measured in parallel on natural freshwater samples from the basin of the river Meuse (Belgium). Comparison of the incorporation into proteins and into the total macromolecular fraction showed that these different amino acids were incorporated at more than 90% into proteins. From incorporation measurements at four subsaturated concentrations (range, 2-77 nm), the maximum incorporation rates were determined. Strong correlations (r > 0.91 for all the calculated correlations) were found between the maximum incorporation rates of the different tested amino acids over a range of two orders of magnitude of bacterial activity. Bacterial production estimates were calculated using theoretical and experimental conversion factors. The productions calculated from the incorporation rates of the four amino acids were in good concordance, especially when the experimental conversion factors were used (slope range, 0.91-1.11, and r > 0.91). This study suggests that the incorporation of various amino acids into proteins can be used to estimate bacterial production.

  2. Effect of bacterial protein meal on protein and energy metabolism in growing chickens

    DEFF Research Database (Denmark)

    Hellwing, Anne Louise Frydendahl; Tauson, Anne-Helene; Skrede, Anders

    2006-01-01

    This experiment investigates the effect of increasing the dietary content of bacterial protein meal (BPM) on the protein and energy metabolism, and carcass chemical composition of growing chickens. Seventy-two Ross male chickens were allocated to four diets, each in three replicates with 0% (D0), 2...... (period 1), 5 chickens (period 2), and one chicken (periods 3-5). After each balance period, one chicken in each cage was killed and the carcass weight was recorded. Chemical Analyses were performed on the carcasses from periods, 1, 3, and 5. Weight gain, feed intake, and feed conversion rate were found...... to be similar for all diets. Chickens on D0 retained 1.59 g N·kg-°75·d-¹, respectively. This was probably caused by the higher nitrogen content of D0. Neither the HE (p=0.92) nor the retention of energy (P=0.88) were affected by diet. Carcass composition was similar between diets, in line with the values...

  3. Discovering the bacterial circular proteins : bacteriocins, cyanobactins, and pilins

    NARCIS (Netherlands)

    Montalban-Lopez, Manuel; Sanchez-Hidalgo, Marina; Cebrian, Ruben; Maqueda, Mercedes

    2012-01-01

    Over recent years, several examples of natural ribosomally synthesized circular proteins and peptides from diverse organisms have been described. They are a group of proteins for which the precursors must be post-translationally modified to join the N and C termini with a peptide bond. This feature

  4. Methyl-accepting protein associated with bacterial sensory phodopsin I

    International Nuclear Information System (INIS)

    Spudich, E.N.; Hasselbacher, C.A.; Spudich, J.L.

    1988-01-01

    In vivo radiolabeling of Halaobacterium halobium phototaxis mutants and revertants with L-[methyl- 3 H] methionine implicated seven methyl-accepting protein bands with apparent molecular masses from 65 to 150 kilodaltons (kDa) in adaptation of the organism to chemo and photo stimuli, and one of these (94 kDa) was specifically implicated in photoaxis. The lability of the radiolabeled bands to mild base treatment indicated the the methyl linkages are carboxylmethylesters, as is the case in the eubacterial chemotaxis receptor-transducers. The 94-kDa protein was present in increased amounts in an overproducer of the apoprotein of sensory rhodopsin I, one of two retinal-containing photoaxis receptors in H. halobium. It was absent in a strain the contained sensory rhodopsin II and that lacked sensory rhodopsin I and was also absent in a mutant that lacked both photoreceptors. Based in the role of methyl-accepting proteins in chemotaxis in other bacteria, we suggest that the 94-kDa protein is the signal transducer for sensory rhodopsin I. By [ 3 H]retinal labeling studies, we previously identified a 25-kDa retinal-binding polypeptide that was derived from photochemically reactive sensory rhodopsin I. When H. halobium membranes containing sensory rhodopsin I were treated by a procedure that stably reduced [ 3 H] retinal onto the 25-kDa apoprotein, a 94-kDa protein was also found to be radiolabeled. Protease digestion confirmed that the 94-kDa retinal-labeled protein was the same as the methyl-accepting protein that was suggested above to be the siginal transducer for sensory rhodopsin I. Possible models are that the 25- and 94-kDa proteins are tightly interacting components of the photosensory signaling machinery or that both are forms of sensory rhodopsin I

  5. Excretion of purine base derivatives after intake of bacterial protein meal in pigs

    DEFF Research Database (Denmark)

    Hellwing, Anne Louise Frydendahl; Tauson, Anne-Helene; Skrede, A.

    2007-01-01

    Bacterial protein meal has a high content ofprotein but also of RNA and DNA. Sixteen barrows were allocated to four diets containing increasing levels of bacterial protein meal (BPM), from weaning to 80 kg live weight, to evaluate whether the RNA and DNA contents of BPM influenced the retention...... of nitrogen. It was hypothesised that an increased intake of RNA and DNA would lead to an increased urinary excretion of purine base derivatives and increased plasma concentrations. Retention of nitrogen was unaffected by dietary content of BPM (P=0.08) and the urinary excretion of purine base derivatives...

  6. Bacterial Serine/Threonine Protein Kinases in Host-Pathogen Interactions*

    Science.gov (United States)

    Canova, Marc J.; Molle, Virginie

    2014-01-01

    In bacterial pathogenesis, monitoring and adapting to the dynamically changing environment in the host and an ability to disrupt host immune responses are critical. The virulence determinants of pathogenic bacteria include the sensor/signaling proteins of the serine/threonine protein kinase (STPK) family that have a dual role of sensing the environment and subverting specific host defense processes. STPKs can sense a wide range of signals and coordinate multiple cellular processes to mount an appropriate response. Here, we review some of the well studied bacterial STPKs that are essential virulence factors and that modify global host responses during infection. PMID:24554701

  7. Bacterial serine/threonine protein kinases in host-pathogen interactions.

    Science.gov (United States)

    Canova, Marc J; Molle, Virginie

    2014-04-04

    In bacterial pathogenesis, monitoring and adapting to the dynamically changing environment in the host and an ability to disrupt host immune responses are critical. The virulence determinants of pathogenic bacteria include the sensor/signaling proteins of the serine/threonine protein kinase (STPK) family that have a dual role of sensing the environment and subverting specific host defense processes. STPKs can sense a wide range of signals and coordinate multiple cellular processes to mount an appropriate response. Here, we review some of the well studied bacterial STPKs that are essential virulence factors and that modify global host responses during infection.

  8. Reversals and collisions optimize protein exchange in bacterial swarms

    Energy Technology Data Exchange (ETDEWEB)

    Amiri, Aboutaleb; Harvey, Cameron; Buchmann, Amy; Christley, Scott; Shrout, Joshua D.; Aranson, Igor S.; Alber, Mark

    2017-03-01

    Swarming groups of bacteria coordinate their behavior by self-organizing as a population to move over surfaces in search of nutrients and optimal niches for colonization. Many open questions remain about the cues used by swarming bacteria to achieve this self-organization. While chemical cue signaling known as quorum sensing is well-described, swarming bacteria often act and coordinate on time scales that could not be achieved via these extracellular quorum sensing cues. Here, cell-cell contact-dependent protein exchange is explored as amechanism of intercellular signaling for the bacterium Myxococcus xanthus. A detailed biologically calibrated computational model is used to study how M. xanthus optimizes the connection rate between cells and maximizes the spread of an extracellular protein within the population. The maximum rate of protein spreading is observed for cells that reverse direction optimally for swarming. Cells that reverse too slowly or too fast fail to spread extracellular protein efficiently. In particular, a specific range of cell reversal frequencies was observed to maximize the cell-cell connection rate and minimize the time of protein spreading. Furthermore, our findings suggest that predesigned motion reversal can be employed to enhance the collective behavior of biological synthetic active systems.

  9. Bacteriocin Isolated From Halomon sp.: A Bacterial Ding Protein?

    International Nuclear Information System (INIS)

    Atirah Azemin; Klappa, P.; Mohd Shahir Shamsir Omar

    2015-01-01

    A marine halophile, Halomonas sp. strain M3 was isolated from Straits of Johor, Malaysia and produce bacteriocin CC that acts as bacteriostatic agent. Characterisation of the bacterium showed that optimal growth and bacteriocin production is at ambient temperature, pH of 8-8.5 in nutrient broth medium supplemented with 2.9 % w/v NaCI to mimic saltwater conditions. The stability studies indicated that bacteriocin CC is heat-labile (35-50 degree Celsius) and was stable over 2 years when stored in 0.02 M Tris- HCI with 30-60 % glycerol at 4 degree Celsius. A loss of activity was detected after proteolytic enzymes treatment, indicating the proteinaceous nature of the antimicrobial compound. The amino acid sequence of bacteriocin CC was obtained by Edman degradation and MALDI-TOF analysis, showed the characteristic sequence of a DING protein (D-I-N-G-G-G-A-T-L-P-Q-A-LY- Q) in size 38.9-kDa at pI of 6.8. These proteins constitute a conserved and widely distributed set of proteins found in all kingdoms with ligand-binding activities and hydrolytic enzyme, suggesting a possible role in cell signalling and bio mineralization in DING isolates. Intriguingly, DING proteins also have been involved as an anti-tumour agent in humans. Thus, bacteriocin CC as DING protein family members should be further studied to investigate its potential as a novel antimicrobial agent. (author)

  10. Single-Molecule Studies of Bacterial Protein Translocation

    NARCIS (Netherlands)

    Kedrov, Alexej; Kusters, Ilja; Driessen, Arnold J. M.

    2013-01-01

    In prokaryotes, a large share of the proteins are secreted from the cell through a process that requires their translocation across the cytoplasmic membrane. This process is mediated by the universally conserved Sec system with homologues in the endoplasmic reticulum and thylakoid membranes of

  11. Insight into bacterial virulence mechanisms against host immune response via the Yersinia pestis-human protein-protein interaction network.

    Science.gov (United States)

    Yang, Huiying; Ke, Yuehua; Wang, Jian; Tan, Yafang; Myeni, Sebenzile K; Li, Dong; Shi, Qinghai; Yan, Yanfeng; Chen, Hui; Guo, Zhaobiao; Yuan, Yanzhi; Yang, Xiaoming; Yang, Ruifu; Du, Zongmin

    2011-11-01

    A Yersinia pestis-human protein interaction network is reported here to improve our understanding of its pathogenesis. Up to 204 interactions between 66 Y. pestis bait proteins and 109 human proteins were identified by yeast two-hybrid assay and then combined with 23 previously published interactions to construct a protein-protein interaction network. Topological analysis of the interaction network revealed that human proteins targeted by Y. pestis were significantly enriched in the proteins that are central in the human protein-protein interaction network. Analysis of this network showed that signaling pathways important for host immune responses were preferentially targeted by Y. pestis, including the pathways involved in focal adhesion, regulation of cytoskeleton, leukocyte transendoepithelial migration, and Toll-like receptor (TLR) and mitogen-activated protein kinase (MAPK) signaling. Cellular pathways targeted by Y. pestis are highly relevant to its pathogenesis. Interactions with host proteins involved in focal adhesion and cytoskeketon regulation pathways could account for resistance of Y. pestis to phagocytosis. Interference with TLR and MAPK signaling pathways by Y. pestis reflects common characteristics of pathogen-host interaction that bacterial pathogens have evolved to evade host innate immune response by interacting with proteins in those signaling pathways. Interestingly, a large portion of human proteins interacting with Y. pestis (16/109) also interacted with viral proteins (Epstein-Barr virus [EBV] and hepatitis C virus [HCV]), suggesting that viral and bacterial pathogens attack common cellular functions to facilitate infections. In addition, we identified vasodilator-stimulated phosphoprotein (VASP) as a novel interaction partner of YpkA and showed that YpkA could inhibit in vitro actin assembly mediated by VASP.

  12. Influence of protein deposition on bacterial adhesion to contact lenses.

    Science.gov (United States)

    Subbaraman, Lakshman N; Borazjani, Roya; Zhu, Hua; Zhao, Zhenjun; Jones, Lyndon; Willcox, Mark D P

    2011-08-01

    The aim of the study is to determine the adhesion of Gram positive and Gram negative bacteria onto conventional hydrogel (CH) and silicone hydrogel (SH) contact lens materials with and without lysozyme, lactoferrin, and albumin coating. Four lens types (three SH-balafilcon A, lotrafilcon B, and senofilcon A; one CH-etafilcon A) were coated with lysozyme, lactoferrin, or albumin (uncoated lenses acted as controls) and then incubated in Staphylococcus aureus (Saur 31) or either of two strains of Pseudomonas aeruginosa (Paer 6294 and 6206) for 24 h at 37 °C. The total counts of the adhered bacteria were determined using the H-thymidine method and viable counts by counting the number of colony-forming units on agar media. All three strains adhered significantly lower to uncoated etafilcon A lenses compared with uncoated SH lenses (p 0.05). Lactoferrin coating on lenses increased binding (total and viable counts) of Saur 31 (p lenses showed significantly higher total counts (p lenses. Albumin coating of lenses increased binding (total and viable counts) of all three strains (p lenses does not possess antibacterial activity against certain bacterial strains, whereas lactoferrin possess an antibacterial effect against strains of P. aeruginosa.

  13. The human-bacterial pathogen protein interaction networks of Bacillus anthracis, Francisella tularensis, and Yersinia pestis.

    Directory of Open Access Journals (Sweden)

    Matthew D Dyer

    2010-08-01

    Full Text Available Bacillus anthracis, Francisella tularensis, and Yersinia pestis are bacterial pathogens that can cause anthrax, lethal acute pneumonic disease, and bubonic plague, respectively, and are listed as NIAID Category A priority pathogens for possible use as biological weapons. However, the interactions between human proteins and proteins in these bacteria remain poorly characterized leading to an incomplete understanding of their pathogenesis and mechanisms of immune evasion.In this study, we used a high-throughput yeast two-hybrid assay to identify physical interactions between human proteins and proteins from each of these three pathogens. From more than 250,000 screens performed, we identified 3,073 human-B. anthracis, 1,383 human-F. tularensis, and 4,059 human-Y. pestis protein-protein interactions including interactions involving 304 B. anthracis, 52 F. tularensis, and 330 Y. pestis proteins that are uncharacterized. Computational analysis revealed that pathogen proteins preferentially interact with human proteins that are hubs and bottlenecks in the human PPI network. In addition, we computed modules of human-pathogen PPIs that are conserved amongst the three networks. Functionally, such conserved modules reveal commonalities between how the different pathogens interact with crucial host pathways involved in inflammation and immunity.These data constitute the first extensive protein interaction networks constructed for bacterial pathogens and their human hosts. This study provides novel insights into host-pathogen interactions.

  14. Entry and exit of bacterial outer membrane proteins.

    Science.gov (United States)

    Misra, Rajeev

    2015-08-01

    The sites of new outer membrane protein (OMP) deposition and the fate of pre-existing OMPs are still enigmatic despite numerous concerted efforts. Rassam et al. identified mid-cell regions as the primary entry points for new OMP insertion in clusters, driving the pre-existing OMP clusters towards cell poles for long-term storage. Copyright © 2015 Elsevier Ltd. All rights reserved.

  15. Molecular mechanism of pore creation in bacterial membranes by amyloid proteins

    International Nuclear Information System (INIS)

    Tsigelny, I F; Sharikov, Y; Miller, M A; Masliah, E

    2009-01-01

    This study explores the mechanism of pore creation in cellular membranes by MccE92 bacterial proteins. The results of this study are then compared with the mechanism of alpha-synuclein (aS)-based pore formation in mammalian cells, and its role in Parkinson's disease.

  16. Artificial acceleration of mammalian cell reprogramming by bacterial proteins.

    Science.gov (United States)

    Ikeda, Takashi; Uchiyama, Ikuo; Iwasaki, Mio; Sasaki, Tetsuhiko; Nakagawa, Masato; Okita, Keisuke; Masui, Shinji

    2017-10-01

    The molecular mechanisms of cell reprogramming and differentiation involve various signaling factors. Small molecule compounds have been identified to artificially influence these factors through interacting cellular proteins. Although such small molecule compounds are useful to enhance reprogramming and differentiation and to show the mechanisms that underlie these events, the screening usually requires a large number of compounds to identify only a very small number of hits (e.g., one hit among several tens of thousands of compounds). Here, we show a proof of concept that xenospecific gene products can affect the efficiency of cell reprogramming to pluripotency. Thirty genes specific for the bacterium Wolbachia pipientis were forcibly expressed individually along with reprogramming factors (Oct4, Sox2, Klf4 and c-Myc) that can generate induced pluripotent stem cells in mammalian cells, and eight were found to affect the reprogramming efficiency either positively or negatively (hit rate 26.7%). Mechanistic analysis suggested one of these proteins interacted with cytoskeleton to promote reprogramming. Our results raise the possibility that xenospecific gene products provide an alternative way to study the regulatory mechanism of cell identity. © 2017 Molecular Biology Society of Japan and John Wiley & Sons Australia, Ltd.

  17. Effect of Dietary Protein Levels on Composition of Odorous Compounds and Bacterial Ecology in Pig Manure

    Directory of Open Access Journals (Sweden)

    Sungback Cho

    2015-09-01

    Full Text Available This study was performed to investigate the effect of different levels of dietary crude protein (CP on composition of odorous compounds and bacterial communities in pig manure. A total of 48 male pigs (average initial body weight 45 kg fed diets containing three levels of dietary CP (20%, 17.5%, and 15% and their slurry samples were collected from the pits under the floor every week for one month. Changes in composition of odorous compounds and bacterial communities were analyzed by gas chromatography and 454 FLX titanium pyrosequencing systems, respectively. Levels of phenols, indoles, short chain fatty acid and branched chain fatty acid were lowest (p<0.05 in CP 15% group among three CP levels. Relative abundance of Bacteroidetes phylum and bacterial genera including Leuconostoc, Bacillus, Atopostipes, Peptonphilus, Ruminococcaceae_uc, Bacteroides, and Pseudomonas was lower (p<0.05 in CP 15% than in CP 20% group. There was a positive correlation (p<0.05 between odorous compounds and bacterial genera: phenol, indole, iso-butyric acid, and iso-valeric acid with Atopostipes, p-cresol and skatole with Bacteroides, acetic acid and butyric acid with AM982595_g of Porphyromonadaceae family, and propionic acid with Tissierella. Taken together, administration of 15% CP showed less production of odorous compounds than 20% CP group and this result might be associated with the changes in bacterial communities especially whose roles in protein metabolism.

  18. Effect of sulfur analogue of lysine on bacterial protein biosynthesis

    International Nuclear Information System (INIS)

    Tanaka, Hidehiko; Soda, Kenji.

    1976-01-01

    S-(beta-Aminoethyl)-L-cysteine, a sulfur analogue of lysine inhibited strongly growth of Escherichia coli A-19, and weakly that of Corynebacterium sp. isolated from soil, but did not inhibit growth of Aerobacter aerogenes. In Corynebacterium sp. the inhibitory effect was markedly enhanced in the presence of L-threonine. The inhibition of growth by S-(beta-aminoethyl)-L-cysteine was rapidly reversed by the addition of L-lysine. S-(beta-Aminoethyl)-L-cysteine inhibited protein synthesis and the activity of lysyl-tRNA synthetase from E. coli and A. aerogenes. All the other lysine analogues tested inhibited the activity of enzyme, but S-(beta-aminoethyl)-L-cysteine derivatives, S-(beta-N-acetyl-aminoethyl)-L-cysteine and S-(beta-aminoethyl)-alpha-N-acetyl-L-cysteine were not effective. (auth.)

  19. Determining and comparing protein function in Bacterial genome sequences

    DEFF Research Database (Denmark)

    Vesth, Tammi Camilla

    of this class have very little homology to other known genomes making functional annotation based on sequence similarity very difficult. Inspired in part by this analysis, an approach for comparative functional annotation was created based public sequenced genomes, CMGfunc. Functionally related groups......In November 2013, there was around 21.000 different prokaryotic genomes sequenced and publicly available, and the number is growing daily with another 20.000 or more genomes expected to be sequenced and deposited by the end of 2014. An important part of the analysis of this data is the functional...... annotation of genes – the descriptions assigned to genes that describe the likely function of the encoded proteins. This process is limited by several factors, including the definition of a function which can be more or less specific as well as how many genes can actually be assigned a function based...

  20. Differential protein expression in alligator leukocytes in response to bacterial lipopolysaccharide injection.

    Science.gov (United States)

    Merchant, Mark; Kinney, Clint; Sanders, Paige

    2009-12-01

    Blood was collected from three juvenile alligators (Alligator mississippiensis) before, and again 24h after, injection with bacterial lipopolysaccharide (LPS). The leukocytes were collected from both samples, and the proteins were extracted. Each group of proteins was labeled with a different fluorescent dye and the differences in protein expression were analyzed by two dimensional differential in-gel expressions (2D-DIGE). The proteins which appeared to be increased or decreased by treatment with LPS were selected and analyzed by MALDI-TOF to determine mass and LC-MS/MS to acquire the partial protein sequences. The peptide sequences were compared to the NCBI protein sequence database to determine homology with other sequences from other species. Several proteins of interest appeared to be increased upon LPS stimulation. Proteins with homology to human transgelin-2, fish glucose-6-phosphate dehydrogenase, amphibian α-enolase, alligator lactate dehydrogenase, fish ubiquitin-activating enzyme, and fungal β-tubulin were also increased after LPS injection. Proteins with homology to fish vimentin 4, murine heterogeneous nuclear ribonucleoprotein A3, and avian calreticulin were found to be decreased in response to LPS. In addition, five proteins, four of which were up-regulated (827, 560, 512, and 650%) and one that exhibited repressed expression (307%), did not show homology to any protein in the database, and thus may represent newly discovered proteins. We are using this biochemical approach to isolate and characterize alligator proteins with potential relevant immune function.

  1. Channel crossing: how are proteins shipped across the bacterial plasma membrane?

    Science.gov (United States)

    Collinson, Ian; Corey, Robin A; Allen, William J

    2015-10-05

    The structure of the first protein-conducting channel was determined more than a decade ago. Today, we are still puzzled by the outstanding problem of protein translocation--the dynamic mechanism underlying the consignment of proteins across and into membranes. This review is an attempt to summarize and understand the energy transducing capabilities of protein-translocating machines, with emphasis on bacterial systems: how polypeptides make headway against the lipid bilayer and how the process is coupled to the free energy associated with ATP hydrolysis and the transmembrane protein motive force. In order to explore how cargo is driven across the membrane, the known structures of the protein-translocation machines are set out against the background of the historic literature, and in the light of experiments conducted in their wake. The paper will focus on the bacterial general secretory (Sec) pathway (SecY-complex), and its eukaryotic counterpart (Sec61-complex), which ferry proteins across the membrane in an unfolded state, as well as the unrelated Tat system that assembles bespoke channels for the export of folded proteins. © 2015 The Authors.

  2. Bacterial single-stranded DNA-binding proteins are phosphorylated on tyrosine

    DEFF Research Database (Denmark)

    Mijakovic, Ivan; Petranovic, Dina; Macek, B

    2006-01-01

    for phosphotyrosine-containing proteins in Streptomyces griseus by immunoaffinity chromatography identified bacterial SSBs as a novel target of bacterial tyrosine kinases. Since genes encoding protein-tyrosine kinases (PTKs) have not been recognized in streptomycetes, and SSBs from Streptomyces coelicolor (Sc......SSB) and Bacillus subtilis (BsSSB) share 38.7% identity, we used a B.subtilis protein-tyrosine kinase YwqD to phosphorylate two cognate SSBs (BsSSB and YwpH) in vitro. We demonstrate that in vivo phosphorylation of B.subtilis SSB occurs on tyrosine residue 82, and this reaction is affected antagonistically...... by kinase YwqD and phosphatase YwqE. Phosphorylation of B.subtilis SSB increased binding almost 200-fold to single-stranded DNA in vitro. Tyrosine phosphorylation of B.subtilis, S.coelicolor and Escherichia coli SSBs occured while they were expressed in E.coli, indicating that tyrosine phosphorylation...

  3. Pathogenic Leptospira species express surface-exposed proteins belonging to the bacterial immunoglobulin superfamily

    Science.gov (United States)

    Matsunaga, James; Barocchi, Michele A.; Croda, Julio; Young, Tracy A.; Sanchez, Yolanda; Siqueira, Isadora; Bolin, Carole A.; Reis, Mitermayer G.; Riley, Lee W.; Haake, David A.; Ko, Albert I.

    2005-01-01

    Summary Proteins with bacterial immunoglobulin-like (Big) domains, such as the Yersinia pseudotuberculosis invasin and Escherichia coli intimin, are surface-expressed proteins that mediate host mammalian cell invasion or attachment. Here, we report the identification and characterization of a new family of Big domain proteins, referred to as Lig (leptospiral Ig-like) proteins, in pathogenic Leptospira. Screening of L. interrogans and L. kirschneri expression libraries with sera from leptospirosis patients identified 13 lambda phage clones that encode tandem repeats of the 90 amino acid Big domain. Two lig genes, designated ligA and ligB, and one pseudo-gene, ligC, were identified. The ligA and ligB genes encode amino-terminal lipoprotein signal peptides followed by 10 or 11 Big domain repeats and, in the case of ligB, a unique carboxy-terminal non-repeat domain. The organization of ligC is similar to that of ligB but contains mutations that disrupt the reading frame. The lig sequences are present in pathogenic but not saprophytic Leptospira species. LigA and LigB are expressed by a variety of virulent leptospiral strains. Loss of Lig protein and RNA transcript expression is correlated with the observed loss of virulence during culture attenuation of pathogenic strains. High-pressure freeze substitution followed by immunocytochemical electron microscopy confirmed that the Lig proteins were localized to the bacterial surface. Immunoblot studies with patient sera found that the Lig proteins are a major antigen recognized during the acute host infection. These observations demonstrate that the Lig proteins are a newly identified surface protein of pathogenic Leptospira, which by analogy to other bacterial immunoglobulin superfamily virulence factors, may play a role in host cell attachment and invasion during leptospiral pathogenesis. PMID:12890019

  4. Subcellular localization for Gram positive and Gram negative bacterial proteins using linear interpolation smoothing model.

    Science.gov (United States)

    Saini, Harsh; Raicar, Gaurav; Dehzangi, Abdollah; Lal, Sunil; Sharma, Alok

    2015-12-07

    Protein subcellular localization is an important topic in proteomics since it is related to a protein׳s overall function, helps in the understanding of metabolic pathways, and in drug design and discovery. In this paper, a basic approximation technique from natural language processing called the linear interpolation smoothing model is applied for predicting protein subcellular localizations. The proposed approach extracts features from syntactical information in protein sequences to build probabilistic profiles using dependency models, which are used in linear interpolation to determine how likely is a sequence to belong to a particular subcellular location. This technique builds a statistical model based on maximum likelihood. It is able to deal effectively with high dimensionality that hinders other traditional classifiers such as Support Vector Machines or k-Nearest Neighbours without sacrificing performance. This approach has been evaluated by predicting subcellular localizations of Gram positive and Gram negative bacterial proteins. Copyright © 2015 Elsevier Ltd. All rights reserved.

  5. Oxidative stress and S-100B protein in children with bacterial meningitis

    Directory of Open Access Journals (Sweden)

    Hamed Enas A

    2009-10-01

    Full Text Available Abstract Background Bacterial meningitis is often associated with cerebral compromise which may be responsible for neurological sequelae in nearly half of the survivors. Little is known about the mechanisms of CNS involvement in bacterial meningitis. Several studies have provided substantial evidence for the key role of nitric oxide (NO and reactive oxygen species in the complex pathophysiology of bacterial meningitis. Methods In the present study, serum and CSF levels of NO, lipid peroxide (LPO (mediators for oxidative stress and lipid peroxidation; total thiol, superoxide dismutase (SOD (antioxidant mediators and S-100B protein (mediator of astrocytes activation and injury, were investigated in children with bacterial meningitis (n = 40. Albumin ratio (CSF/serum is a marker of blood-CSF barriers integrity, while mediator index (mediator ratio/albumin ratio is indicative of intrathecal synthesis. Results Compared to normal children (n = 20, patients had lower serum albumin but higher NO, LPO, total thiol, SOD and S-100B. The ratios and indices of NO and LPO indicate blood-CSF barriers dysfunction, while the ratio of S-100B indicates intrathecal synthesis. Changes were marked among patients with positive culture and those with neurological complications. Positive correlation was found between NO index with CSF WBCs (r = 0.319, p Conclusion This study suggests that loss of integrity of brain-CSF barriers, oxidative stress and S-100B may contribute to the severity and neurological complications of bacterial meningitis.

  6. Rapid and widely disseminated acute phase protein response after experimental bacterial infection of pigs

    DEFF Research Database (Denmark)

    Skovgaard, Kerstin; Mortensen, Shila; Boye, Mette

    2009-01-01

    The acute phase protein response is a well-described generalized early host response to tissue injury, inflammation and infection, observed as pronounced changes in the concentrations of a number of circulating serum proteins. The biological function of this response and its interplay with other...... parts of innate host defence reactions remain somewhat elusive. In order to gain new insight into this early host defence response in the context of bacterial infection we studied gene expression changes in peripheral lymphoid tissues as compared to hepatic expression changes, 14-18 h after lung...... with measurements of interleukin-6 and selected acute phase proteins in serum. C-reactive protein and serum amyloid A were clearly induced 14-18 h after infection. Extrahepatic expression of acute phase proteins was found to be dramatically altered as a result of the lung infection with an extrahepatic acute phase...

  7. Effective non-denaturing purification method for improving the solubility of recombinant actin-binding proteins produced by bacterial expression.

    Science.gov (United States)

    Chung, Jeong Min; Lee, Sangmin; Jung, Hyun Suk

    2017-05-01

    Bacterial expression is commonly used to produce recombinant and truncated mutant eukaryotic proteins. However, heterologous protein expression may render synthesized proteins insoluble. The conventional method used to express a poorly soluble protein, which involves denaturation and refolding, is time-consuming and inefficient. There are several non-denaturing approaches that can increase the solubility of recombinant proteins that include using different bacterial cell strains, altering the time of induction, lowering the incubation temperature, and employing different detergents for purification. In this study, we compared several non-denaturing protocols to express and purify two insoluble 34 kDa actin-bundling protein mutants. The solubility of the mutant proteins was not affected by any of the approaches except for treatment with the detergent sarkosyl. These results indicate that sarkosyl can effectively improve the solubility of insoluble proteins during bacterial expression. Copyright © 2016. Published by Elsevier Inc.

  8. The use of fed batch approaches to maximise yields in bacterial fermentation and protein expression

    International Nuclear Information System (INIS)

    McLean, A.

    2001-01-01

    A fermentation facility for the scale up of bacterial and yeast fermentations has been set up at the University of Queensland under the auspices of the ARC Special Research Centre for Functional and Applied Genomics. A major application is the production of recombinant proteins for determination of tertiary structures by X-ray crystallography or nuclear magnetic resonance. For this purpose, large amounts of protein arc needed and the yield from a single fermentation run is crucial to success within constrained laboratory budgets. To achieve maximal yields we are optimising fed batch approaches in bacterial fermentation. Fed batch offers many advantages over batch cultures. Coupled with the ability to monitor online the internal conditions of the fermentation including pH and dissolved oxygen and stirrer cascading functions it is possible to ensure that the nutritional environment of the microorganism is optimised for its growth and or for optimal protein expression. The poster will describe some of our experience in setting up fed batch fermentations and successful applications of fed batches to increasing protein yield. It will also outline services that are available to academic groups outside the University of Queensland For structure determination and functional studies, the production of radiolabelled proteins can also be an advantage. We will describe initial experiments aimed at coupling the principles of fed batch fermentation to the introduction of carbon or nitrogen isotopes into the recombinant protein

  9. Protein deposition and its effect on bacterial adhesion to contact lenses.

    Science.gov (United States)

    Omali, Negar Babaei; Zhu, Hua; Zhao, Zhenjun; Willcox, Mark D P

    2013-06-01

    Bacterial adhesion to contact lenses is believed to be the initial step for the development of several adverse reactions that occur during lens wear such as microbial keratitis. This study examined the effect of combinations of proteins on the adhesion of bacteria to contact lenses. Unworn balafilcon A and senofilcon A lenses were soaked in commercially available pure protein mixtures to achieve the same amount of various proteins as found ex vivo. These lenses were then exposed to Pseudomonas aeruginosa and Staphylococcus aureus. Following incubation, the numbers of P. aeruginosa or S. aureus that adhered to the lenses were measured. The possible effect of proteins on bacterial growth was investigated by incubating bacteria in medium containing protein. Although there was a significant (p lenses soaked in the lysozyme/lactoferrin combination, the protein adhered to lenses did not alter the adhesion of any other strains of P. aeruginosa or S. aureus (p > 0.05). Growth of S. aureus 031 (p 0.05). Adsorption of amounts of lysozyme and lactoferrin or lipocalin equivalent to those extracted from worn contact lenses did not affect the adhesion of most strains of S. aureus or P. aeruginosa to lens surfaces.

  10. Brillouin spectroscopy as a new method of screening for increased CSF total protein during bacterial meningitis.

    Science.gov (United States)

    Steelman, Zachary; Meng, Zhaokai; Traverso, Andrew J; Yakovlev, Vladislav V

    2015-05-01

    Bacterial meningitis is a disease of pronounced clinical significance, especially in the developing world. Immediate treatment with antibiotics is essential, and no single test can provide a conclusive diagnosis. It is well established that elevated total protein in cerebrospinal fluid (CSF) is associated with bacterial meningitis. Brillouin spectroscopy is a widely used optical technique for noninvasive determination of the elastic moduli of materials. We found that elevated protein levels in CSF alter the fluid elasticity sufficiently to be measurable by Brillouin spectroscopy, with model healthy and diseased fluids distinguishable to marked significance (P = 0.014), which increases with sample concentration by dialysis. Typical raw output of a 2-stage VIPA Brillouin spectrometer: inelastically scattered Brillouin peaks (arrows) and elastically scattered incident radiation (center cross). © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  11. In vitro estimation of rumen protein degradability using 35S to label the bacterial mass

    International Nuclear Information System (INIS)

    Khristov, A.; Aleksandrov, S.; Aleksiev, I.

    1994-01-01

    An experiment was carried out in order to simplify a previously developed 15 N-method for in vitro estimation of rumen protein degradability. Casein (Cas), whole soybeans (Sb) heated at 120 o C for 20 min (SbTherm) and sunflower (Sfl) were incubated at 39 o C for 4 hours in a water bathshaker with the following media: McDougall's buffer, strained and enriched with particle associated bacteria rumen fluid (2:1), rapidly (maltose, sucrose, glucose) and more slowly (pectin, soluble starch) degradable carbohydrates with final concentration of 815 mg/100 ml and 21.7 μCi/100 ml of 35 S (from Na 2 35 SO 4 ). After the incubation had been ceased, a bacterial fraction was isolated through differential centrifugation and specific activity of bacterial (Bac) and high speed total solids (TS) nitrogen was measured. The ratio was used to calculate bacterial mass in TS and through the Kjeldahl nitrogen concentration in TS - the net bacterial growth (against control vessels without protein). The level of ammonia-N in the supernate after blank correction was used to find the ammonia-N released from protein degradation. The data showed that the rate (and extend) of degradation for the Cas (as a standard protein) was lower compared to those obtained through the 15 N-method but it was higher than the rate derived through another in vitro method. The Cas equivalent of the Sb was higher than the figure we found in a previous experiment with solvent extracted soybean meal suggesting that the 35 S-method underestimated the degradability of the Cas. After being tested on a wider range of foodstuffs, the proposed 35 S-method might be considered as an alternative procedure which is less laborous than the 15 N-method. (author)

  12. Looping and clustering model for the organization of protein-DNA complexes on the bacterial genome

    Science.gov (United States)

    Walter, Jean-Charles; Walliser, Nils-Ole; David, Gabriel; Dorignac, Jérôme; Geniet, Frédéric; Palmeri, John; Parmeggiani, Andrea; Wingreen, Ned S.; Broedersz, Chase P.

    2018-03-01

    The bacterial genome is organized by a variety of associated proteins inside a structure called the nucleoid. These proteins can form complexes on DNA that play a central role in various biological processes, including chromosome segregation. A prominent example is the large ParB-DNA complex, which forms an essential component of the segregation machinery in many bacteria. ChIP-Seq experiments show that ParB proteins localize around centromere-like parS sites on the DNA to which ParB binds specifically, and spreads from there over large sections of the chromosome. Recent theoretical and experimental studies suggest that DNA-bound ParB proteins can interact with each other to condense into a coherent 3D complex on the DNA. However, the structural organization of this protein-DNA complex remains unclear, and a predictive quantitative theory for the distribution of ParB proteins on DNA is lacking. Here, we propose the looping and clustering model, which employs a statistical physics approach to describe protein-DNA complexes. The looping and clustering model accounts for the extrusion of DNA loops from a cluster of interacting DNA-bound proteins that is organized around a single high-affinity binding site. Conceptually, the structure of the protein-DNA complex is determined by a competition between attractive protein interactions and loop closure entropy of this protein-DNA cluster on the one hand, and the positional entropy for placing loops within the cluster on the other. Indeed, we show that the protein interaction strength determines the ‘tightness’ of the loopy protein-DNA complex. Thus, our model provides a theoretical framework for quantitatively computing the binding profiles of ParB-like proteins around a cognate (parS) binding site.

  13. Structural analysis of a set of proteins resulting from a bacterial genomics project.

    Science.gov (United States)

    Badger, J; Sauder, J M; Adams, J M; Antonysamy, S; Bain, K; Bergseid, M G; Buchanan, S G; Buchanan, M D; Batiyenko, Y; Christopher, J A; Emtage, S; Eroshkina, A; Feil, I; Furlong, E B; Gajiwala, K S; Gao, X; He, D; Hendle, J; Huber, A; Hoda, K; Kearins, P; Kissinger, C; Laubert, B; Lewis, H A; Lin, J; Loomis, K; Lorimer, D; Louie, G; Maletic, M; Marsh, C D; Miller, I; Molinari, J; Muller-Dieckmann, H J; Newman, J M; Noland, B W; Pagarigan, B; Park, F; Peat, T S; Post, K W; Radojicic, S; Ramos, A; Romero, R; Rutter, M E; Sanderson, W E; Schwinn, K D; Tresser, J; Winhoven, J; Wright, T A; Wu, L; Xu, J; Harris, T J R

    2005-09-01

    The targets of the Structural GenomiX (SGX) bacterial genomics project were proteins conserved in multiple prokaryotic organisms with no obvious sequence homolog in the Protein Data Bank of known structures. The outcome of this work was 80 structures, covering 60 unique sequences and 49 different genes. Experimental phase determination from proteins incorporating Se-Met was carried out for 45 structures with most of the remainder solved by molecular replacement using members of the experimentally phased set as search models. An automated tool was developed to deposit these structures in the Protein Data Bank, along with the associated X-ray diffraction data (including refined experimental phases) and experimentally confirmed sequences. BLAST comparisons of the SGX structures with structures that had appeared in the Protein Data Bank over the intervening 3.5 years since the SGX target list had been compiled identified homologs for 49 of the 60 unique sequences represented by the SGX structures. This result indicates that, for bacterial structures that are relatively easy to express, purify, and crystallize, the structural coverage of gene space is proceeding rapidly. More distant sequence-structure relationships between the SGX and PDB structures were investigated using PDB-BLAST and Combinatorial Extension (CE). Only one structure, SufD, has a truly unique topology compared to all folds in the PDB. Copyright 2005 Wiley-Liss, Inc.

  14. Phosphoproteome analysis of streptomyces development reveals extensive protein phosphorylation accompanying bacterial differentiation

    DEFF Research Database (Denmark)

    Manteca, Angel; Ye, Juanying; Sánchez, Jesús

    2011-01-01

    Streptomycetes are bacterial species that undergo a complex developmental cycle that includes programmed cell death (PCD) events and sporulation. They are widely used in biotechnology because they produce most clinically relevant secondary metabolites. Although Streptomyces coelicolor is one...... events were detected during the presporulation and sporulation stages (80%). Most of these phosphorylations were not reported before in Streptomyces, and included sporulation factors, transcriptional regulators, protein kinases and other regulatory proteins. Several of the identified phosphorylated...... proteins, FtsZ, DivIVA, and FtsH2, were previously demonstrated to be involved in the sporulation process. We thus established for the first time the widespread occurrence and dynamic features of Ser/Thr/Tyr protein phosphorylation in a bacteria species and also revealed a previously unrecognized...

  15. The role of bacterial antizyme: From an inhibitory protein to AtoC transcriptional regulator

    Directory of Open Access Journals (Sweden)

    Kyriakidis Dimitrios A

    2004-06-01

    Full Text Available Abstract This review considers the role of bacterial antizyme in the regulation of polyamine biosynthesis and gives new perspectives on the involvement of antizyme in other significant cellular mechanisms. Antizyme is a protein molecule induced by the end product of the enzymic reaction that it inhibits, in a non-competitive manner. The bacterial ornithine decarboxylase is regulated by nucleotides, phosphorylation and antizyme. The inhibition of ornithine decarboxylase by antizyme can be relieved to different degrees by DNA or by a variety of synthetic nucleic acid polymers, attributed to a specific interaction between nucleic acid and antizyme. Recently, this interplay between bacterial antizyme and nucleic acid was determined by discerning an additional function to antizyme that proved to be the atoC gene product, encoding the response regulator of the bacterial two-component system AtoS-AtoC. The gene located just upstream of atoC encodes the sensor kinase, named AtoS, that modulates AtoC activity. AtoC regulates expression of atoDAEB operon which is involved in short-chain fatty acid metabolism. Antizyme is thus referred to as AtoC, functioning both as a post-translational and transcriptional regulator. Also, the AtoS-AtoC signal transduction system in E. coli has a positive regulatory role on poly-(R-3-hydroxybutyrate biosynthesis. The properties and gene structural similarities of antizymes from different organisms were compared. It was revealed that conserved domains are present mostly in the C-domain of all antizymes. BLAST analysis of the E. coli antizyme protein (AtoC showed similarities around 69–58% among proteobacteria, g-proteobacteria, enterobacteria and the thermophilic bacterium Thermus thermophilus. A working hypothesis is proposed for the metabolic role of antizyme (AtoC describing the significant biological implications of this protein molecule. Whether antizymes exist to other enzymes in different tissues, meeting the

  16. Trafficking and processing of bacterial proteins by mammalian cells: Insights from chondroitinase ABC.

    Science.gov (United States)

    Muir, Elizabeth; Raza, Mansoor; Ellis, Clare; Burnside, Emily; Love, Fiona; Heller, Simon; Elliot, Matthew; Daniell, Esther; Dasgupta, Debayan; Alves, Nuno; Day, Priscilla; Fawcett, James; Keynes, Roger

    2017-01-01

    There is very little reported in the literature about the relationship between modifications of bacterial proteins and their secretion by mammalian cells that synthesize them. We previously reported that the secretion of the bacterial enzyme Chondroitinase ABC by mammalian cells requires the strategic removal of at least three N-glycosylation sites. The aim of this study was to determine if it is possible to enhance the efficacy of the enzyme as a treatment for spinal cord injury by increasing the quantity of enzyme secreted or by altering its cellular location. To determine if the efficiency of enzyme secretion could be further increased, cells were transfected with constructs encoding the gene for chondroitinase ABC modified for expression by mammalian cells; these contained additional modifications of strategic N-glycosylation sites or alternative signal sequences to direct secretion of the enzyme from the cells. We show that while removal of certain specific N-glycosylation sites enhances enzyme secretion, N-glycosylation of at least two other sites, N-856 and N-773, is essential for both production and secretion of active enzyme. Furthermore, we find that the signal sequence directing secretion also influences the quantity of enzyme secreted, and that this varies widely amongst the cell types tested. Last, we find that replacing the 3'UTR on the cDNA encoding Chondroitinase ABC with that of β-actin is sufficient to target the enzyme to the neuronal growth cone when transfected into neurons. This also enhances neurite outgrowth on an inhibitory substrate. Some intracellular trafficking pathways are adversely affected by cryptic signals present in the bacterial gene sequence, whilst unexpectedly others are required for efficient secretion of the enzyme. Furthermore, targeting chondroitinase to the neuronal growth cone promotes its ability to increase neurite outgrowth on an inhibitory substrate. These findings are timely in view of the renewed prospects for

  17. Meat, dairy and plant proteins alter bacterial composition of rat gut bacteria.

    Science.gov (United States)

    Zhu, Yingying; Lin, Xisha; Zhao, Fan; Shi, Xuebin; Li, He; Li, Yingqiu; Zhu, Weiyun; Xu, Xinglian; Li, Chunbao; Lu, Chunbao; Zhou, Guanghong

    2015-10-14

    Long-term consumption of red meat has been considered a potential risk to gut health, but this is based on clinic investigations, excessive intake of fat, heme and some injurious compounds formed during cooking or additions to processed meat products. Whether intake of red meat protein affects gut bacteria and the health of the host remains unclear. In this work, we compared the composition of gut bacteria in the caecum, by sequencing the V4-V5 region of 16S ribosomal RNA gene, obtained from rats fed with proteins from red meat (beef and pork), white meat (chicken and fish) and other sources (casein and soy). The results showed significant differences in profiles of gut bacteria between the six diet groups. Rats fed with meat proteins had a similar overall structure of caecal bacterial communities separated from those fed non-meat proteins. The beneficial genus Lactobacillus was higher in the white meat than in the red meat or non-meat protein groups. Also, rats fed with meat proteins and casein had significantly lower levels of lipopolysaccharide-binding proteins, suggesting that the intake of meat proteins may maintain a more balanced composition of gut bacteria, thereby reducing the antigen load and inflammatory response in the host.

  18. Meat, dairy and plant proteins alter bacterial composition of rat gut bacteria

    Science.gov (United States)

    Zhu, Yingying; Lin, Xisha; Zhao, Fan; Shi, Xuebin; Li, He; Li, Yingqiu; Zhu, Weiyun; Xu, Xinglian; Lu, Chunbao; Zhou, Guanghong

    2015-01-01

    Long-term consumption of red meat has been considered a potential risk to gut health, but this is based on clinic investigations, excessive intake of fat, heme and some injurious compounds formed during cooking or additions to processed meat products. Whether intake of red meat protein affects gut bacteria and the health of the host remains unclear. In this work, we compared the composition of gut bacteria in the caecum, by sequencing the V4-V5 region of 16S ribosomal RNA gene, obtained from rats fed with proteins from red meat (beef and pork), white meat (chicken and fish) and other sources (casein and soy). The results showed significant differences in profiles of gut bacteria between the six diet groups. Rats fed with meat proteins had a similar overall structure of caecal bacterial communities separated from those fed non-meat proteins. The beneficial genus Lactobacillus was higher in the white meat than in the red meat or non-meat protein groups. Also, rats fed with meat proteins and casein had significantly lower levels of lipopolysaccharide-binding proteins, suggesting that the intake of meat proteins may maintain a more balanced composition of gut bacteria, thereby reducing the antigen load and inflammatory response in the host. PMID:26463271

  19. STUDIES ON THE BACTERIOPHAGE OF D'HERELLE : IX. EVIDENCE OF HYDROLYSIS OF BACTERIAL PROTEIN DURING LYSIS.

    Science.gov (United States)

    Hetler, D M; Bronfenbrenner, J

    1928-07-31

    1. During the process of lysis by bacteriophage, there is an appreciable increase in the amount of free amino acid present in the culture. 2. The increase of free amino acid is due to hydrolysis of bacterial protein.

  20. Mechanisms of Host-Pathogen Protein Complex Formation and Bacterial Immune Evasion of Streptococcus suis Protein Fhb.

    Science.gov (United States)

    Li, Xueqin; Liu, Peng; Gan, Shuzhen; Zhang, Chunmao; Zheng, Yuling; Jiang, Yongqiang; Yuan, Yuan

    2016-08-12

    Streptococcus suis serotype 2 (S. suis 2)-induced sepsis and meningitis are often accompanied by bacteremia. The evasion of polymorphonuclear leukocyte-mediated phagocytic clearance is central to the establishment of bacteremia caused by S. suis 2 and is facilitated by the ability of factor H (FH)-binding protein (Fhb) to bind FH on the bacterial surface, thereby impeding alternative pathway complement activation and phagocytic clearance. Here, C3b/C3d was found to bind to Fhb, along with FH, forming a large immune complex. The formation of this immune complex was mediated by domain II of Fhb via electrostatic and hydrophobic interactions, which, to our knowledge, is a new type of interaction. Interestingly, Fhb was found to be associated with the cell envelope and also present in the culture supernatant, where secreted Fhb inhibited complement activation via interactions with domain II, thereby enhancing antiphagocytic clearance by polymorphonuclear leukocytes. Thus, Fhb is a multifunctional bacterial protein, which binds host complement component C3 as well as FH and interferes with innate immune recognition in a secret protein manner. S. suis 2 therefore appears to have developed a new strategy to combat host innate immunity and enhance survival in host blood. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  1. Mechanisms of Host-Pathogen Protein Complex Formation and Bacterial Immune Evasion of Streptococcus suis Protein Fhb*

    Science.gov (United States)

    Li, Xueqin; Liu, Peng; Gan, Shuzhen; Zhang, Chunmao; Zheng, Yuling; Jiang, Yongqiang; Yuan, Yuan

    2016-01-01

    Streptococcus suis serotype 2 (S. suis 2)-induced sepsis and meningitis are often accompanied by bacteremia. The evasion of polymorphonuclear leukocyte-mediated phagocytic clearance is central to the establishment of bacteremia caused by S. suis 2 and is facilitated by the ability of factor H (FH)-binding protein (Fhb) to bind FH on the bacterial surface, thereby impeding alternative pathway complement activation and phagocytic clearance. Here, C3b/C3d was found to bind to Fhb, along with FH, forming a large immune complex. The formation of this immune complex was mediated by domain II of Fhb via electrostatic and hydrophobic interactions, which, to our knowledge, is a new type of interaction. Interestingly, Fhb was found to be associated with the cell envelope and also present in the culture supernatant, where secreted Fhb inhibited complement activation via interactions with domain II, thereby enhancing antiphagocytic clearance by polymorphonuclear leukocytes. Thus, Fhb is a multifunctional bacterial protein, which binds host complement component C3 as well as FH and interferes with innate immune recognition in a secret protein manner. S. suis 2 therefore appears to have developed a new strategy to combat host innate immunity and enhance survival in host blood. PMID:27342778

  2. Targeting Bacterial Dsb Proteins for the Development of Anti-Virulence Agents

    Directory of Open Access Journals (Sweden)

    Roxanne P. Smith

    2016-07-01

    Full Text Available Recent years have witnessed a dramatic increase in bacterial antimicrobial resistance and a decline in the development of novel antibiotics. New therapeutic strategies are urgently needed to combat the growing threat posed by multidrug resistant bacterial infections. The Dsb disulfide bond forming pathways are potential targets for the development of antimicrobial agents because they play a central role in bacterial pathogenesis. In particular, the DsbA/DsbB system catalyses disulfide bond formation in a wide array of virulence factors, which are essential for many pathogens to establish infections and cause disease. These redox enzymes are well placed as antimicrobial targets because they are taxonomically widespread, share low sequence identity with human proteins, and many years of basic research have provided a deep molecular understanding of these systems in bacteria. In this review, we discuss disulfide bond catalytic pathways in bacteria and their significance in pathogenesis. We also review the use of different approaches to develop inhibitors against Dsb proteins as potential anti-virulence agents, including fragment-based drug discovery, high-throughput screening and other structure-based drug discovery methods.

  3. Protein corona between nanoparticles and bacterial proteins in activated sludge: Characterization and effect on nanoparticle aggregation.

    Science.gov (United States)

    Zhang, Peng; Xu, Xiao-Yan; Chen, You-Peng; Xiao, Meng-Qian; Feng, Bo; Tian, Kai-Xun; Chen, Yue-Hui; Dai, You-Zhi

    2018-02-01

    In this work, the protein coronas of activated sludge proteins on TiO 2 nanoparticles (TNPs) and ZnO nanoparticles (ZNPs) were characterized. The proteins with high affinity to TNPs and ZNPs were identified by shotgun proteomics, and their effects of on the distributions of TNPs and ZNPs in activated sludge were concluded. In addition, the effects of protein coronas on the aggregations of TNPs and ZNPs were evaluated. Thirty and nine proteins with high affinities to TNPs and ZNPs were identified, respectively. The proteomics and adsorption isotherms demonstrated that activated sludge had a higher affinity to TNPs than to ZNPs. The aggregation percentages of ZNPs at 35, 53, and 106 mg/L of proteins were 13%, 14%, and 18%, respectively, whereas those of TNPs were 21%, 30%, 41%, respectively. The proteins contributed to ZNPs aggregation by dissolved Zn ion-bridging, whereas the increasing protein concentrations enhanced the TNPs aggregation through macromolecule bridging flocculation. Copyright © 2017 Elsevier Ltd. All rights reserved.

  4. Modulating bacterial and gut mucosal interactions with engineered biofilm matrix proteins.

    Science.gov (United States)

    Duraj-Thatte, Anna M; Praveschotinunt, Pichet; Nash, Trevor R; Ward, Frederick R; Joshi, Neel S

    2018-02-22

    Extracellular appendages play a significant role in mediating communication between bacteria and their host. Curli fibers are a class of bacterial fimbria that is highly amenable to engineering. We demonstrate the use of engineered curli fibers to rationally program interactions between bacteria and components of the mucosal epithelium. Commensal E. coli strains were engineered to produce recombinant curli fibers fused to the trefoil family of human cytokines. Biofilms formed from these strains bound more mucins than those producing wild-type curli fibers, and modulated mucin rheology as well. When treated with bacteria producing the curli-trefoil fusions mammalian cells behaved identically in terms of their migration behavior as when they were treated with the corresponding soluble trefoil factors. Overall, this demonstrates the potential utility of curli fibers as a scaffold for the display of bioactive domains and an untapped approach to rationally modulating host-microbe interactions using bacterial matrix proteins.

  5. A reproducible and scalable procedure for preparing bacterial extracts for cell-free protein synthesis.

    Science.gov (United States)

    Katsura, Kazushige; Matsuda, Takayoshi; Tomabechi, Yuri; Yonemochi, Mayumi; Hanada, Kazuharu; Ohsawa, Noboru; Sakamoto, Kensaku; Takemoto, Chie; Shirouzu, Mikako

    2017-11-01

    Cell-free protein synthesis is a useful method for preparing proteins for functional or structural analyses. However, batch-to-batch variability with regard to protein synthesis activity remains a problem for large-scale production of cell extract in the laboratory. To address this issue, we have developed a novel procedure for large-scale preparation of bacterial cell extract with high protein synthesis activity. The developed procedure comprises cell cultivation using a fermentor, harvesting and washing of cells by tangential flow filtration, cell disruption with high-pressure homogenizer and continuous diafiltration. By optimizing and combining these methods, ∼100 ml of the cell extract was prepared from 150 g of Escherichia coli cells. The protein synthesis activities, defined as the yield of protein per unit of absorbance at 260 nm of the cell extract, were shown to be reproducible, and the average activity of several batches was twice that obtained using a previously reported method. In addition, combinatorial use of the high-pressure homogenizer and diafiltration increased the scalability, indicating that the cell concentration at disruption varies from 0.04 to 1 g/ml. Furthermore, addition of Gam protein and examinations of the N-terminal sequence rendered the extract prepared here useful for rapid screening with linear DNA templates. © The Authors 2017. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.

  6. Chirality Switching by Martensitic Transformation in Protein Cylindrical Crystals: Application to Bacterial Flagella

    Science.gov (United States)

    Komai, Ricardo Kiyohiro

    Martensitic transformations provide unique engineering properties that, when designed properly, become important parts of new technology. Martensitic transformations have been studied for many years in traditional alloys (iron, steel, titanium, etc.), however there is still much to be learned in regards to these transformations in biological materials. Olson and Hartman showed in 1982 that these transformations are also observed in bacterial flagella and T4 bacteriophage viral sheaths, allowing for propulsion of bacteria in a fluid environment and, for the virus, is responsible for the infection mechanism. This work demonstrates, using the bacterial flagella as an example, that these transformations can be modelled using thermodynamic methods that are also used to model the transformations in alloys. This thesis work attempts to explain the transformations that occur in bacterial flagella, which are capable of small strain, highly reversible martensitic transformations. The first stress/temperature phase diagrams of these flagella were created by adding the mechanical energy of the transformation of the flagella to limited chemical thermodynamics information of the transformation. Mechanical energy is critical to the transformation process because the bacterial body applies a torque to the radius of the flagella. Finally, work has begun and will be completed in regards to understanding the kinetics of the transformation of the flagella. The motion of the transformation interface can be predicted by using a Landau-Ginzburg model. The crystallography of the transformation in bacterial flagella is also being computed to determine the invariant lines of transformation that occur within this cylindrical crystal. This work has shown that it is possible to treat proteins in a similar manner that alloys are treated when using thermodynamic modelling. Much can be learned from translating what is known regarding phase transformations in hard material systems to soft, organic

  7. Structural studies of bacterial transcriptional regulatory proteins by multidimensional heteronuclear NMR

    Energy Technology Data Exchange (ETDEWEB)

    Volkman, Brian Finley [Univ. of California, Berkeley, CA (United States)

    1995-02-01

    Nuclear magnetic resonance spectroscopy was used to elucidate detailed structural information for peptide and protein molecules. A small peptide was designed and synthesized, and its three-dimensional structure was calculated using distance information derived from two-dimensional NMR measurements. The peptide was used to induce antibodies in mice, and the cross-reactivity of the antibodies with a related protein was analyzed with enzyme-linked immunosorbent assays. Two proteins which are involved in regulation of transcription in bacteria were also studied. The ferric uptake regulation (Fur) protein is a metal-dependent repressor which controls iron uptake in bacteria. Two- and three-dimensional NMR techniques, coupled with uniform and selective isotope labeling allowed the nearly complete assignment of the resonances of the metal-binding domain of the Fur protein. NTRC is a transcriptional enhancer binding protein whose N-terminal domain is a "receiver domain" in the family of "two-component" regulatory systems. Phosphorylation of the N-terminal domain of NTRC activates the initiation of transcription of aeries encoding proteins involved in nitrogen regulation. Three- and four-dimensional NMR spectroscopy methods have been used to complete the resonance assignments and determine the solution structure of the N-terminal receiver domain of the NTRC protein. Comparison of the solution structure of the NTRC receiver domain with the crystal structures of the homologous protein CheY reveals a very similar fold, with the only significant difference being the position of helix 4 relative to the rest of the protein. The determination of the structure of the NTRC receiver domain is the first step toward understanding a mechanism of signal transduction which is common to many bacterial regulatory systems.

  8. Comparison of structure, function and regulation of plant cold shock domain proteins to bacterial and animal cold shock domain proteins.

    Science.gov (United States)

    Chaikam, Vijay; Karlson, Dale T

    2010-01-01

    The cold shock domain (CSD) is among the most ancient and well conserved nucleic acid binding domains from bacteria to higher animals and plants. The CSD facilitates binding to RNA, ssDNA and dsDNA and most functions attributed to cold shock domain proteins are mediated by this nucleic acid binding activity. In prokaryotes, cold shock domain proteins only contain a single CSD and are termed cold shock proteins (Csps). In animal model systems, various auxiliary domains are present in addition to the CSD and are commonly named Y-box proteins. Similar to animal CSPs, plant CSPs contain auxiliary C-terminal domains in addition to their N-terminal CSD. Cold shock domain proteins have been shown to play important roles in development and stress adaptation in wide variety of organisms. In this review, the structure, function and regulation of plant CSPs are compared and contrasted to the characteristics of bacterial and animal CSPs. [BMB reports 2010; 43(1): 1-8].

  9. Growth performance and carcase quality in broiler chickens fed on bacterial protein grown on natural gas.

    Science.gov (United States)

    Øverland, M; Schøyen, H F; Skrede, A

    2010-10-01

    1. The effects of increasing concentrations (0, 40, 80 or 120 g/kg) of bacterial protein meal (BPM) and bacterial protein autolysate (BPA) grown on natural gas on growth performance and carcase quality in broiler chickens were examined. 2. Adding BPM to diets reduced feed intake and improved gain: feed from 0 to 21 d and overall to 35 d, but did not significantly affect weight gain compared to the soybean meal based control diet. 3. Increasing concentrations of BPA significantly reduced growth rate, feed intake, gain: feed, carcase weight and dressing percentage, but significantly increased carcase dry matter, fat and energy content. 4. Adding BPM to diets had no effect on viscosity of diets and jejunal digesta, and minor effects on litter quality, whereas BPA increased the viscosity of diets and jejunal digesta, improved litter quality at 21 d, but decreased litter quality at 32 d. 5. To conclude, broiler chickens performed better on a BPM product with intact proteins than on an autolysate with ruptured cell walls and a high content of free amino acids and low molecular-weight peptides.

  10. A Simple and Rapid Method for Preparing a Cell-Free Bacterial Lysate for Protein Synthesis.

    Directory of Open Access Journals (Sweden)

    Nitzan Krinsky

    Full Text Available Cell-free protein synthesis (CFPS systems are important laboratory tools that are used for various synthetic biology applications. Here, we present a simple and inexpensive laboratory-scale method for preparing a CFPS system from E. coli. The procedure uses basic lab equipment, a minimal set of reagents, and requires less than one hour to process the bacterial cell mass into a functional S30-T7 extract. BL21(DE3 and MRE600 E. coli strains were used to prepare the S30-T7 extract. The CFPS system was used to produce a set of fluorescent and therapeutic proteins of different molecular weights (up to 66 kDa. This system was able to produce 40-150 μg-protein/ml, with variations depending on the plasmid type, expressed protein and E. coli strain. Interestingly, the BL21-based CFPS exhibited stability and increased activity at 40 and 45°C. To the best of our knowledge, this is the most rapid and affordable lab-scale protocol for preparing a cell-free protein synthesis system, with high thermal stability and efficacy in producing therapeutic proteins.

  11. A Simple and Rapid Method for Preparing a Cell-Free Bacterial Lysate for Protein Synthesis

    Science.gov (United States)

    Kaduri, Maya; Shainsky-Roitman, Janna; Goldfeder, Mor; Ivanir, Eran; Benhar, Itai; Shoham, Yuval; Schroeder, Avi

    2016-01-01

    Cell-free protein synthesis (CFPS) systems are important laboratory tools that are used for various synthetic biology applications. Here, we present a simple and inexpensive laboratory-scale method for preparing a CFPS system from E. coli. The procedure uses basic lab equipment, a minimal set of reagents, and requires less than one hour to process the bacterial cell mass into a functional S30-T7 extract. BL21(DE3) and MRE600 E. coli strains were used to prepare the S30-T7 extract. The CFPS system was used to produce a set of fluorescent and therapeutic proteins of different molecular weights (up to 66 kDa). This system was able to produce 40–150 μg-protein/ml, with variations depending on the plasmid type, expressed protein and E. coli strain. Interestingly, the BL21-based CFPS exhibited stability and increased activity at 40 and 45°C. To the best of our knowledge, this is the most rapid and affordable lab-scale protocol for preparing a cell-free protein synthesis system, with high thermal stability and efficacy in producing therapeutic proteins. PMID:27768741

  12. Structural and sequence analysis of imelysin-like proteins implicated in bacterial iron uptake.

    Directory of Open Access Journals (Sweden)

    Qingping Xu

    Full Text Available Imelysin-like proteins define a superfamily of bacterial proteins that are likely involved in iron uptake. Members of this superfamily were previously thought to be peptidases and were included in the MEROPS family M75. We determined the first crystal structures of two remotely related, imelysin-like proteins. The Psychrobacter arcticus structure was determined at 2.15 Å resolution and contains the canonical imelysin fold, while higher resolution structures from the gut bacteria Bacteroides ovatus, in two crystal forms (at 1.25 Å and 1.44 Å resolution, have a circularly permuted topology. Both structures are highly similar to each other despite low sequence similarity and circular permutation. The all-helical structure can be divided into two similar four-helix bundle domains. The overall structure and the GxHxxE motif region differ from known HxxE metallopeptidases, suggesting that imelysin-like proteins are not peptidases. A putative functional site is located at the domain interface. We have now organized the known homologous proteins into a superfamily, which can be separated into four families. These families share a similar functional site, but each has family-specific structural and sequence features. These results indicate that imelysin-like proteins have evolved from a common ancestor, and likely have a conserved function.

  13. Bacterial actin MreB assembles in complex with cell shape protein RodZ.

    Science.gov (United States)

    van den Ent, Fusinita; Johnson, Christopher M; Persons, Logan; de Boer, Piet; Löwe, Jan

    2010-03-17

    Bacterial actin homologue MreB is required for cell shape maintenance in most non-spherical bacteria, where it assembles into helical structures just underneath the cytoplasmic membrane. Proper assembly of the actin cytoskeleton requires RodZ, a conserved, bitopic membrane protein that colocalises to MreB and is essential for cell shape determination. Here, we present the first crystal structure of bacterial actin engaged with a natural partner and provide a clear functional significance of the interaction. We show that the cytoplasmic helix-turn-helix motif of Thermotoga maritima RodZ directly interacts with monomeric as well as filamentous MreB and present the crystal structure of the complex. In vitro and in vivo analyses of mutant T. maritima and Escherichia coli RodZ validate the structure and reveal the importance of the MreB-RodZ interaction in the ability of cells to propagate as rods. Furthermore, the results elucidate how the bacterial actin cytoskeleton might be anchored to the membrane to help constrain peptidoglycan synthesis in the periplasm.

  14. Cooperative working of bacterial chromosome replication proteins generated by a reconstituted protein expression system

    Science.gov (United States)

    Fujiwara, Kei; Katayama, Tsutomu; Nomura, Shin-ichiro M.

    2013-01-01

    Replication of all living cells relies on the multirounds flow of the central dogma. Especially, expression of DNA replication proteins is a key step to circulate the processes of the central dogma. Here we achieved the entire sequential transcription–translation–replication process by autonomous expression of chromosomal DNA replication machineries from a reconstituted transcription–translation system (PURE system). We found that low temperature is essential to express a complex protein, DNA polymerase III, in a single tube using the PURE system. Addition of the 13 genes, encoding initiator, DNA helicase, helicase loader, RNA primase and DNA polymerase III to the PURE system gave rise to a DNA replication system by a coupling manner. An artificial genetic circuit demonstrated that the DNA produced as a result of the replication is able to provide genetic information for proteins, indicating the in vitro central dogma can sequentially undergo two rounds. PMID:23737447

  15. Differential Modulation of Transcription Factors and Cytoskeletal Proteins in Prostate Carcinoma Cells by a Bacterial Lactone

    Directory of Open Access Journals (Sweden)

    Senthil R. Kumar

    2018-01-01

    Full Text Available The present study tested the effect of a bacterial lactone N-(3-oxododecanoyl-homoserine lactone (C12-HSL on the cytoskeletal and transcriptional genes and proteins in prostate adenocarcinoma (PA cells (DU145 and LNCaP and prostate small cell neuroendocrine carcinoma (SCNC PC3 cells including their cellular viability and apoptosis. Our data indicate that cell migration and colony formation were affected in the presence of C12-HSL. C12-HSL induced apoptosis and altered viability of both PA and SCNC cells in a concentration dependent manner as measured by fluorescence and chemiluminescence assays. Compared to PCa cells, noncancerous prostate epithelial cells (RWPE1 were resistant to modification by C12-HSL. Further, the viability of PC3 cells in 3D matrix was suppressed by C12-HSL treatment as detected using calcein AM fluorescence in situ. C12-HSL treatment induced cytoskeletal associated protein expression of vinculin and RhoC, which may have implications in cancer cell motility, adhesion, and metastasis. IQGAP protein expression was reduced in DU145 and RWPE1 cells in the presence of C12-HSL. C12-HSL decreased STAT3 phosphorylation in DU145 cells but increased STAT1 protein phosphorylation in PC3 and LNCaP cells. Overall, these studies indicate that C12-HSL can trigger changes in transcription factors and cytoskeletal proteins and thereby modulate growth and migration properties of PCa cells.

  16. Nitrogen and energy balance in growing mink (Mustela vison) fed different levels of bacterial protein meal produced with natural gas

    DEFF Research Database (Denmark)

    Hellwing, Anne Louise Frydendahl; Tauson, Anne-Helene; Ahlstrøm, Øystein

    2005-01-01

    The objective of this study was to estimate the effect of increasing the dietary content of bacterial protein meal (BPM) on energy and protein metabolism in growing mink kits. Sixteen male mink kits of the standard brown genotype were randomly fed one of four diets: A control (Diet III) and 60% (...

  17. Blood parameters in growing pigs fed increasing levels of bacterial protein meal

    DEFF Research Database (Denmark)

    Hellwing, Anne Louise Frydendahl; Tauson, Anne-Helene; Skrede, Anders

    2007-01-01

    The experiment investigated the effects of increasing dietary levels of bacterial protein meal (BPM) on various blood parameters reflecting protein and fat metabolism, liver function, and purine base metabolism in growing pigs. Sixteen barrows were allocated to four different experimental diets....... The control diet was based on soybean meal. In the other three diets soybean meal was replaced with increasing levels of BPM, approximately 17%, 35%, and 50% of the nitrogen being derived from BPM. Blood samples from the jugular vein were taken when the body weights of the pigs were approximately 10 kg, 21 kg......, 45 kg, and 77 kg. The blood parameters reflecting fat metabolism and liver funtion were not affected by diet. Both the plasma albumin and uric acid concentrations tended to decrease (P = 0.07 and 0.01, respectively) with increasing dietary BPM content, whereas the plasma glucose concentration tended...

  18. Trafficking and processing of bacterial proteins by mammalian cells: Insights from chondroitinase ABC.

    Directory of Open Access Journals (Sweden)

    Elizabeth Muir

    Full Text Available There is very little reported in the literature about the relationship between modifications of bacterial proteins and their secretion by mammalian cells that synthesize them. We previously reported that the secretion of the bacterial enzyme Chondroitinase ABC by mammalian cells requires the strategic removal of at least three N-glycosylation sites. The aim of this study was to determine if it is possible to enhance the efficacy of the enzyme as a treatment for spinal cord injury by increasing the quantity of enzyme secreted or by altering its cellular location.To determine if the efficiency of enzyme secretion could be further increased, cells were transfected with constructs encoding the gene for chondroitinase ABC modified for expression by mammalian cells; these contained additional modifications of strategic N-glycosylation sites or alternative signal sequences to direct secretion of the enzyme from the cells. We show that while removal of certain specific N-glycosylation sites enhances enzyme secretion, N-glycosylation of at least two other sites, N-856 and N-773, is essential for both production and secretion of active enzyme. Furthermore, we find that the signal sequence directing secretion also influences the quantity of enzyme secreted, and that this varies widely amongst the cell types tested. Last, we find that replacing the 3'UTR on the cDNA encoding Chondroitinase ABC with that of β-actin is sufficient to target the enzyme to the neuronal growth cone when transfected into neurons. This also enhances neurite outgrowth on an inhibitory substrate.Some intracellular trafficking pathways are adversely affected by cryptic signals present in the bacterial gene sequence, whilst unexpectedly others are required for efficient secretion of the enzyme. Furthermore, targeting chondroitinase to the neuronal growth cone promotes its ability to increase neurite outgrowth on an inhibitory substrate. These findings are timely in view of the renewed

  19. Serum C-reactive protein as a diagnostic biomarker in dogs with bacterial respiratory diseases.

    Science.gov (United States)

    Viitanen, S J; Laurila, H P; Lilja-Maula, L I; Melamies, M A; Rantala, M; Rajamäki, M M

    2014-01-01

    C-reactive protein (CRP) is a major acute-phase protein in dogs. Serum concentrations are low in healthy animals, but increase rapidly after inflammatory stimuli. The aim of the study was to investigate CRP concentrations in various respiratory diseases of dogs and to determine if CRP can be used as a biomarker in the diagnosis of bacterial respiratory diseases. A total of 106 privately owned dogs with respiratory diseases (17 with bacterial tracheobronchitis [BTB], 20 with chronic bronchitis [CB], 20 with eosinophilic bronchopneumopathy [EBP], 12 with canine idiopathic pulmonary fibrosis [CIPF], 15 with cardiogenic pulmonary edema [CPE], and 22 with bacterial pneumonia [BP]) and 72 healthy controls. The study was conducted as a prospective cross-sectional observational study. CRP was measured in serum samples. Diagnosis was confirmed by clinical and laboratory findings, diagnostic imaging, and selected diagnostic methods such as cytological and microbiological analysis of respiratory samples, echocardiography, and histopathology. Dogs with BP had significantly higher CRP concentrations (median, 121 mg/L; interquartile range, 68-178 mg/L) than dogs with BTB (23, 15-38, P = .0003), CB (13, 8-14, P < .0001), EBP (5, 5-15, P < .0001), CIPF (17, 10-20, P < .0001), or CPE (19, 13-32, P < .0001) and healthy controls (14, 8-20, P < .0001). Dogs with BTB had significantly higher CRP concentrations than dogs with CB (P = .001) or EBP (P < .0001) and healthy controls (P = .029). These results indicate that CRP has potential for use as an additional biomarker, especially in the diagnostics of BP. Copyright © 2013 by the American College of Veterinary Internal Medicine.

  20. De novo generation of infectious prions with bacterially expressed recombinant prion protein.

    Science.gov (United States)

    Zhang, Zhihong; Zhang, Yi; Wang, Fei; Wang, Xinhe; Xu, Yuanyuan; Yang, Huaiyi; Yu, Guohua; Yuan, Chonggang; Ma, Jiyan

    2013-12-01

    The prion hypothesis is strongly supported by the fact that prion infectivity and the pathogenic conformer of prion protein (PrP) are simultaneously propagated in vitro by the serial protein misfolding cyclic amplification (sPMCA). However, due to sPMCA's enormous amplification power, whether an infectious prion can be formed de novo with bacterially expressed recombinant PrP (rPrP) remains to be satisfactorily resolved. To address this question, we performed unseeded sPMCA with rPrP in a laboratory that has never been exposed to any native prions. Two types of proteinase K (PK)-resistant and self-perpetuating recombinant PrP conformers (rPrP-res) with PK-resistant cores of 17 or 14 kDa were generated. A bioassay revealed that rPrP-res(17kDa) was highly infectious, causing prion disease in wild-type mice with an average survival time of about 172 d. In contrast, rPrP-res(14kDa) completely failed to induce any disease. Our findings reveal that sPMCA is sufficient to initiate various self-perpetuating PK-resistant rPrP conformers, but not all of them possess in vivo infectivity. Moreover, generating an infectious prion in a prion-free environment establishes that an infectious prion can be formed de novo with bacterially expressed rPrP.

  1. Motion of single MreB bacterial actin proteins in Caulobacter show treadmilling in vivo

    Science.gov (United States)

    Moerner, W. E.; Kim, Soyeon; Gitai, Zemer; Kinkhabwala, Anika; McAdams, Harley; Shapiro, Lucy

    2006-03-01

    Ensemble imaging of a bacterial actin homologue, the MreB protein, suggests that the MreB proteins form a dynamic filamentous spiral along the long axis of the cell in Caulobacter crescentus. MreB contracts and expands along the cell axis and plays an important role in cell shape and polarity maintenance, as well as chromosome segregation and translocation of the origin of replication during cell division. In this study we investigated the real-time polymerization of MreB in Caulobacter crescentus using single-molecule fluorescence imaging. With time-lapse imaging, polymerized MreB could be distinguished from cytoplasmic MreB monomers, because single monomeric MreB showed fast motion characteristic of Brownian diffusion, while single polymerized MreB displayed slow, directed motion. This directional movement of labeled MreB in the growing polymer implies that treadmilling is the predominant mechanism in MreB filament formation. These single-molecule imaging experiments provide the first available information on the velocity of bacterial actin polymerization in a living cell.

  2. The participation of outer membrane proteins in the bacterial sensitivity to nanosilver

    Directory of Open Access Journals (Sweden)

    Anna Kędziora

    2016-06-01

    Full Text Available The presented study is to analyze the participation of outer membrane proteins of Gram- negative bacteria in sensitivity to silver nanomaterials. The mechanism of interaction of silver with the bacterial cell is best described in this group of microorganisms. There are several theories regarding the effectiveness of antimicrobial ions and nanosilver, and at the indicated differences in the way they work. Outer membrane proteins of Gram-negative bacteria are involved in the procurement of silver from the environment and contribute to the development mechanisms of resistance to nanometals. They are measurable parameter in the field of cell phenotypic response to the presence of Gram-negative bacteria in the environment silver nanoforms: its properties, chemical composition, content or times of action. Proteomic methods (including two dimensional electrophoresis and MALDI‑TOF MS are therefore relevant techniques for determining the susceptibility of bacteria to silver and the changes taking place in the outer membrane under the influence: uptime/exposure and physical and chemical parameters of silver nanomaterials. Many products containing nanosilver is still in the research phase in terms of physico‑chemical characteristics and biological activity, others have been already implemented in many industries. During the very fast nanotechnology developing and introduction to the market products based on the nanosilver the bacterial answer to nanosilver is needed.

  3. The bacterial cell cycle checkpoint protein Obg and its role in programmed cell death

    Directory of Open Access Journals (Sweden)

    Liselot Dewachter

    2016-03-01

    Full Text Available The phenomenon of programmed cell death (PCD, in which cells initiate their own demise, is not restricted to multicellular organisms. Unicellular organisms, both eukaryotes and prokaryotes, also possess pathways that mediate PCD. We recently identified a PCD mechanism in Escherichia coli that is triggered by a mutant isoform of the essential GTPase ObgE (Obg of E. coli. Importantly, the PCD pathway mediated by mutant Obg (Obg* differs fundamentally from other previously described bacterial PCD pathways and thus constitutes a new mode of PCD. ObgE was previously proposed to act as a cell cycle checkpoint protein able to halt cell division. The implication of ObgE in the regulation of PCD further increases the similarity between this protein and eukaryotic cell cycle regulators that are capable of doing both. Moreover, since Obg is conserved in eukaryotes, the elucidation of this cell death mechanism might contribute to the understanding of PCD in higher organisms. Additionally, if Obg*-mediated PCD is conserved among different bacterial species, it will be a prime target for the development of innovative antibacterials that artificially induce this pathway.

  4. Differential repair of etheno-DNA adducts by bacterial and human AlkB proteins.

    Science.gov (United States)

    Zdżalik, Daria; Domańska, Anna; Prorok, Paulina; Kosicki, Konrad; van den Born, Erwin; Falnes, Pål Ø; Rizzo, Carmelo J; Guengerich, F Peter; Tudek, Barbara

    2015-06-01

    AlkB proteins are evolutionary conserved Fe(II)/2-oxoglutarate-dependent dioxygenases, which remove alkyl and highly promutagenic etheno(ɛ)-DNA adducts, but their substrate specificity has not been fully determined. We developed a novel assay for the repair of ɛ-adducts by AlkB enzymes using oligodeoxynucleotides with a single lesion and specific DNA glycosylases and AP-endonuclease for identification of the repair products. We compared the repair of three ɛ-adducts, 1,N(6)-ethenoadenine (ɛA), 3,N(4)-ethenocytosine (ɛC) and 1,N(2)-ethenoguanine (1,N(2)-ɛG) by nine bacterial and two human AlkBs, representing four different structural groups defined on the basis of conserved amino acids in the nucleotide recognition lid, engaged in the enzyme binding to the substrate. Two bacterial AlkB proteins, MT-2B (from Mycobacterium tuberculosis) and SC-2B (Streptomyces coelicolor) did not repair these lesions in either double-stranded (ds) or single-stranded (ss) DNA. Three proteins, RE-2A (Rhizobium etli), SA-2B (Streptomyces avermitilis), and XC-2B (Xanthomonas campestris) efficiently removed all three lesions from the DNA substrates. Interestingly, XC-2B and RE-2A are the first AlkB proteins shown to be specialized for ɛ-adducts, since they do not repair methylated bases. Three other proteins, EcAlkB (Escherichia coli), SA-1A, and XC-1B removed ɛA and ɛC from ds and ssDNA but were inactive toward 1,N(2)-ɛG. SC-1A repaired only ɛA with the preference for dsDNA. The human enzyme ALKBH2 repaired all three ɛ-adducts in dsDNA, while only ɛA and ɛC in ssDNA and repair was less efficient in ssDNA. ALKBH3 repaired only ɛC in ssDNA. Altogether, we have shown for the first time that some AlkB proteins, namely ALKBH2, RE-2A, SA-2B and XC-2B can repair 1,N(2)-ɛG and that ALKBH3 removes only ɛC from ssDNA. Our results also suggest that the nucleotide recognition lid is not the sole determinant of the substrate specificity of AlkB proteins. Copyright © 2015 Elsevier B

  5. Bacterial-based systems for expression and purification of recombinant Lassa virus proteins of immunological relevance

    Directory of Open Access Journals (Sweden)

    Cashman Kathleen A

    2008-06-01

    Full Text Available Abstract Background There is a significant requirement for the development and acquisition of reagents that will facilitate effective diagnosis, treatment, and prevention of Lassa fever. In this regard, recombinant Lassa virus (LASV proteins may serve as valuable tools in diverse antiviral applications. Bacterial-based systems were engineered for expression and purification of recombinant LASV nucleoprotein (NP, glycoprotein 1 (GP1, and glycoprotein 2 (GP2. Results Full-length NP and the ectodomains of GP1 and GP2 were generated as maltose-binding protein (MBP fusions in the Rosetta strains of Escherichia coli (E. coli using pMAL-c2x vectors. Average fusion protein yields per liter of culture for MBP-NP, MBP-GP1, and MBP-GP2 were 10 mg, 9 mg, and 9 mg, respectively. Each protein was captured from cell lysates using amylose resin, cleaved with Factor Xa, and purified using size-exclusion chromatography (SEC. Fermentation cultures resulted in average yields per liter of 1.6 mg, 1.5 mg, and 0.7 mg of purified NP, GP1 and GP2, respectively. LASV-specific antibodies in human convalescent sera specifically detected each of the purified recombinant LASV proteins, highlighting their utility in diagnostic applications. In addition, mouse hyperimmune ascitic fluids (MHAF against a panel of Old and New World arenaviruses demonstrated selective cross reactivity with LASV proteins in Western blot and enzyme-linked immunosorbent assay (ELISA. Conclusion These results demonstrate the potential for developing broadly reactive immunological assays that employ all three arenaviral proteins individually and in combination.

  6. Surface Proteins of Lactococcus lactis: Bacterial Resources for Muco-adhesion in the Gastrointestinal Tract

    Directory of Open Access Journals (Sweden)

    Muriel Mercier-Bonin

    2017-11-01

    Full Text Available Food and probiotic bacteria, in particular lactic acid bacteria, are ingested in large amounts by humans and are part of the transient microbiota which is increasingly considered to be able to impact the resident microbiota and thus possibly the host health. The lactic acid bacterium Lactococcus lactis is extensively used in starter cultures to produce dairy fermented food. Also because of a generally recognized as safe status, L. lactis has been considered as a possible vehicle to deliver in vivo therapeutic molecules with anti-inflammatory properties in the gastrointestinal tract. One of the key factors that may favor health effects of beneficial bacteria to the host is their capacity to colonize transiently the gut, notably through close interactions with mucus, which covers and protects the intestinal epithelium. Several L. lactis strains have been shown to exhibit mucus-binding properties and bacterial surface proteins have been identified as key determinants of such capacity. In this review, we describe the different types of surface proteins found in L. lactis, with a special focus on mucus-binding proteins and pili. We also review the different approaches used to investigate the adhesion of L. lactis to mucus, and particularly to mucins, one of its major components, and we present how these approaches allowed revealing the role of surface proteins in muco-adhesion.

  7. The host antimicrobial peptide Bac71-35 binds to bacterial ribosomal proteins and inhibits protein synthesis.

    Science.gov (United States)

    Mardirossian, Mario; Grzela, Renata; Giglione, Carmela; Meinnel, Thierry; Gennaro, Renato; Mergaert, Peter; Scocchi, Marco

    2014-12-18

    Antimicrobial peptides (AMPs) are molecules from innate immunity with high potential as novel anti-infective agents. Most of them inactivate bacteria through pore formation or membrane barrier disruption, but others cross the membrane without damages and act inside the cells, affecting vital processes. However, little is known about their intracellular bacterial targets. Here we report that Bac71-35, a proline-rich AMP belonging to the cathelicidin family, can reach high concentrations (up to 340 μM) inside the E. coli cytoplasm. The peptide specifically and completely inhibits in vitro translation in the micromolar concentration range. Experiments of incorporation of radioactive precursors in macromolecules with E. coli cells confirmed that Bac71-35 affects specifically protein synthesis. Ribosome coprecipitation and crosslinking assays showed that the peptide interacts with ribosomes, binding to a limited subset of ribosomal proteins. Overall, these results indicate that the killing mechanism of Bac71-35 is based on a specific block of protein synthesis. Copyright © 2014 Elsevier Ltd. All rights reserved.

  8. Co-ordinate synthesis and protein localization in a bacterial organelle by the action of a penicillin-binding-protein.

    Science.gov (United States)

    Hughes, H Velocity; Lisher, John P; Hardy, Gail G; Kysela, David T; Arnold, Randy J; Giedroc, David P; Brun, Yves V

    2013-12-01

    Organelles with specialized form and function occur in diverse bacteria. Within the Alphaproteobacteria, several species extrude thin cellular appendages known as stalks, which function in nutrient uptake, buoyancy and reproduction. Consistent with their specialization, stalks maintain a unique molecular composition compared with the cell body, but how this is achieved remains to be fully elucidated. Here we dissect the mechanism of localization of StpX, a stalk-specific protein in Caulobacter crescentus. Using a forward genetics approach, we identify a penicillin-binding-protein, PbpC, which is required for the localization of StpX in the stalk. We show that PbpC acts at the stalked cell pole to anchor StpX to rigid components of the outer membrane of the elongating stalk, concurrent with stalk synthesis. Stalk-localized StpX in turn functions in cellular responses to copper and zinc, suggesting that the stalk may contribute to metal homeostasis in Caulobacter. Together, these results identify a novel role for a penicillin-binding-protein in compartmentalizing a bacterial organelle it itself helps create, raising the possibility that cell wall-synthetic enzymes may broadly serve not only to synthesize the diverse shapes of bacteria, but also to functionalize them at the molecular level. © 2013 John Wiley & Sons Ltd.

  9. Stealth proteins: in silico identification of a novel protein family rendering bacterial pathogens invisible to host immune defense.

    Directory of Open Access Journals (Sweden)

    Peter Sperisen

    2005-11-01

    Full Text Available There are a variety of bacterial defense strategies to survive in a hostile environment. Generation of extracellular polysaccharides has proved to be a simple but effective strategy against the host's innate immune system. A comparative genomics approach led us to identify a new protein family termed Stealth, most likely involved in the synthesis of extracellular polysaccharides. This protein family is characterized by a series of domains conserved across phylogeny from bacteria to eukaryotes. In bacteria, Stealth (previously characterized as SacB, XcbA, or WefC is encoded by subsets of strains mainly colonizing multicellular organisms, with evidence for a protective effect against the host innate immune defense. More specifically, integrating all the available information about Stealth proteins in bacteria, we propose that Stealth is a D-hexose-1-phosphoryl transferase involved in the synthesis of polysaccharides. In the animal kingdom, Stealth is strongly conserved across evolution from social amoebas to simple and complex multicellular organisms, such as Dictyostelium discoideum, hydra, and human. Based on the occurrence of Stealth in most Eukaryotes and a subset of Prokaryotes together with its potential role in extracellular polysaccharide synthesis, we propose that metazoan Stealth functions to regulate the innate immune system. Moreover, there is good reason to speculate that the acquisition and spread of Stealth could be responsible for future epidemic outbreaks of infectious diseases caused by a large variety of eubacterial pathogens. Our in silico identification of a homologous protein in the human host will help to elucidate the causes of Stealth-dependent virulence. At a more basic level, the characterization of the molecular and cellular function of Stealth proteins may shed light on fundamental mechanisms of innate immune defense against microbial invasion.

  10. Stealth Proteins: In Silico Identification of a Novel Protein Family Rendering Bacterial Pathogens Invisible to Host Immune Defense.

    Directory of Open Access Journals (Sweden)

    2005-11-01

    Full Text Available There are a variety of bacterial defense strategies to survive in a hostile environment. Generation of extracellular polysaccharides has proved to be a simple but effective strategy against the host's innate immune system. A comparative genomics approach led us to identify a new protein family termed Stealth, most likely involved in the synthesis of extracellular polysaccharides. This protein family is characterized by a series of domains conserved across phylogeny from bacteria to eukaryotes. In bacteria, Stealth (previously characterized as SacB, XcbA, or WefC is encoded by subsets of strains mainly colonizing multicellular organisms, with evidence for a protective effect against the host innate immune defense. More specifically, integrating all the available information about Stealth proteins in bacteria, we propose that Stealth is a D-hexose-1-phosphoryl transferase involved in the synthesis of polysaccharides. In the animal kingdom, Stealth is strongly conserved across evolution from social amoebas to simple and complex multicellular organisms, such as Dictyostelium discoideum, hydra, and human. Based on the occurrence of Stealth in most Eukaryotes and a subset of Prokaryotes together with its potential role in extracellular polysaccharide synthesis, we propose that metazoan Stealth functions to regulate the innate immune system. Moreover, there is good reason to speculate that the acquisition and spread of Stealth could be responsible for future epidemic outbreaks of infectious diseases caused by a large variety of eubacterial pathogens. Our in silico identification of a homologous protein in the human host will help to elucidate the causes of Stealth-dependent virulence. At a more basic level, the characterization of the molecular and cellular function of Stealth proteins may shed light on fundamental mechanisms of innate immune defense against microbial invasion.

  11. Novel cyclic di-GMP effectors of the YajQ protein family control bacterial virulence.

    Directory of Open Access Journals (Sweden)

    Shi-qi An

    2014-10-01

    Full Text Available Bis-(3',5' cyclic di-guanylate (cyclic di-GMP is a key bacterial second messenger that is implicated in the regulation of many critical processes that include motility, biofilm formation and virulence. Cyclic di-GMP influences diverse functions through interaction with a range of effectors. Our knowledge of these effectors and their different regulatory actions is far from complete, however. Here we have used an affinity pull-down assay using cyclic di-GMP-coupled magnetic beads to identify cyclic di-GMP binding proteins in the plant pathogen Xanthomonas campestris pv. campestris (Xcc. This analysis identified XC_3703, a protein of the YajQ family, as a potential cyclic di-GMP receptor. Isothermal titration calorimetry showed that the purified XC_3703 protein bound cyclic di-GMP with a high affinity (K(d∼2 µM. Mutation of XC_3703 led to reduced virulence of Xcc to plants and alteration in biofilm formation. Yeast two-hybrid and far-western analyses showed that XC_3703 was able to interact with XC_2801, a transcription factor of the LysR family. Mutation of XC_2801 and XC_3703 had partially overlapping effects on the transcriptome of Xcc, and both affected virulence. Electromobility shift assays showed that XC_3703 positively affected the binding of XC_2801 to the promoters of target virulence genes, an effect that was reversed by cyclic di-GMP. Genetic and functional analysis of YajQ family members from the human pathogens Pseudomonas aeruginosa and Stenotrophomonas maltophilia showed that they also specifically bound cyclic di-GMP and contributed to virulence in model systems. The findings thus identify a new class of cyclic di-GMP effector that regulates bacterial virulence.

  12. Photorhabdus adhesion modification protein (Pam) binds extracellular polysaccharide and alters bacterial attachment.

    Science.gov (United States)

    Jones, Robert T; Sanchez-Contreras, Maria; Vlisidou, Isabella; Amos, Matthew R; Yang, Guowei; Muñoz-Berbel, Xavier; Upadhyay, Abhishek; Potter, Ursula J; Joyce, Susan A; Ciche, Todd A; Jenkins, A Toby A; Bagby, Stefan; Ffrench-Constant, Richard H; Waterfield, Nicholas R

    2010-05-12

    Photorhabdus are Gram-negative nematode-symbiotic and insect-pathogenic bacteria. The species Photorhabdus asymbiotica is able to infect humans as well as insects. We investigated the secreted proteome of a clinical isolate of P. asymbiotica at different temperatures in order to identify proteins relevant to the infection of the two different hosts. A comparison of the proteins secreted by a clinical isolate of P. asymbiotica at simulated insect (28 degrees C) and human (37 degrees C) temperatures led to the identification of a small and highly abundant protein, designated Pam, that is only secreted at the lower temperature. The pam gene is present in all Photorhabdus strains tested and shows a high level of conservation across the whole genus, suggesting it is both ancestral to the genus and probably important to the biology of the bacterium. The Pam protein shows limited sequence similarity to the 13.6 kDa component of a binary toxin of Bacillus thuringiensis. Nevertheless, injection or feeding of heterologously produced Pam showed no insecticidal activity to either Galleria mellonella or Manduca sexta larvae. In bacterial colonies, Pam is associated with an extracellular polysaccharide (EPS)-like matrix, and modifies the ability of wild-type cells to attach to an artificial surface. Interestingly, Surface Plasmon Resonance (SPR) binding studies revealed that the Pam protein itself has adhesive properties. Although Pam is produced throughout insect infection, genetic knockout does not affect either insect virulence or the ability of P. luminescens to form a symbiotic association with its host nematode, Heterorhabditis bacteriophora. We studied a highly abundant protein, Pam, which is secreted in a temperature-dependent manner in P. asymbiotica. Our findings indicate that Pam plays an important role in enhancing surface attachment in insect blood. Its association with exopolysaccharide suggests it may exert its effect through mediation of EPS properties. Despite

  13. LocateP: Genome-scale subcellular-location predictor for bacterial proteins

    Directory of Open Access Journals (Sweden)

    Zhou Miaomiao

    2008-03-01

    current tools especially where the N-terminally anchored and the SPIase-cleaved secreted proteins are concerned. Overall, the accuracy of LocateP was always higher than 90%. LocateP was then used to predict the SCLs of all proteins encoded by completed Gram-positive bacterial genomes. The results are stored in the database LocateP-DB http://www.cmbi.ru.nl/locatep-db1. Conclusion LocateP is by far the most accurate and detailed protein SCL predictor for Gram-positive bacteria currently available.

  14. Lactoferrin binding protein B - a bi-functional bacterial receptor protein.

    Directory of Open Access Journals (Sweden)

    Nicholas K H Ostan

    2017-03-01

    Full Text Available Lactoferrin binding protein B (LbpB is a bi-lobed outer membrane-bound lipoprotein that comprises part of the lactoferrin (Lf receptor complex in Neisseria meningitidis and other Gram-negative pathogens. Recent studies have demonstrated that LbpB plays a role in protecting the bacteria from cationic antimicrobial peptides due to large regions rich in anionic residues in the C-terminal lobe. Relative to its homolog, transferrin-binding protein B (TbpB, there currently is little evidence for its role in iron acquisition and relatively little structural and biophysical information on its interaction with Lf. In this study, a combination of crosslinking and deuterium exchange coupled to mass spectrometry, information-driven computational docking, bio-layer interferometry, and site-directed mutagenesis was used to probe LbpB:hLf complexes. The formation of a 1:1 complex of iron-loaded Lf and LbpB involves an interaction between the Lf C-lobe and LbpB N-lobe, comparable to TbpB, consistent with a potential role in iron acquisition. The Lf N-lobe is also capable of binding to negatively charged regions of the LbpB C-lobe and possibly other sites such that a variety of higher order complexes are formed. Our results are consistent with LbpB serving dual roles focused primarily on iron acquisition when exposed to limited levels of iron-loaded Lf on the mucosal surface and effectively binding apo Lf when exposed to high levels at sites of inflammation.

  15. Structure of an essential bacterial protein YeaZ (TM0874) from Thermotoga maritima at 2.5 Å resolution

    International Nuclear Information System (INIS)

    Xu, Qingping; McMullan, Daniel; Jaroszewski, Lukasz; Krishna, S. Sri; Elsliger, Marc-André; Yeh, Andrew P.; Abdubek, Polat; Astakhova, Tamara; Axelrod, Herbert L.; Carlton, Dennis; Chiu, Hsiu-Ju; Clayton, Thomas; Duan, Lian; Feuerhelm, Julie; Grant, Joanna; Han, Gye Won; Jin, Kevin K.; Klock, Heath E.; Knuth, Mark W.; Miller, Mitchell D.; Morse, Andrew T.; Nigoghossian, Edward; Okach, Linda; Oommachen, Silvya; Paulsen, Jessica; Reyes, Ron; Rife, Christopher L.; Bedem, Henry van den; Hodgson, Keith O.; Wooley, John; Deacon, Ashley M.; Godzik, Adam; Lesley, Scott A.; Wilson, Ian A.

    2009-01-01

    The crystal structure of an essential bacterial protein, YeaZ, from T. maritima identifies an interface that potentially mediates protein–protein interaction. YeaZ is involved in a protein network that is essential for bacteria. The crystal structure of YeaZ from Thermotoga maritima was determined to 2.5 Å resolution. Although this protein belongs to a family of ancient actin-like ATPases, it appears that it has lost the ability to bind ATP since it lacks some key structural features that are important for interaction with ATP. A conserved surface was identified, supporting its role in the formation of protein complexes

  16. Super Resolution Fluorescence Microscopy and Tracking of Bacterial Flotillin (Reggie Paralogs Provide Evidence for Defined-Sized Protein Microdomains within the Bacterial Membrane but Absence of Clusters Containing Detergent-Resistant Proteins.

    Directory of Open Access Journals (Sweden)

    Felix Dempwolff

    2016-06-01

    Full Text Available Biological membranes have been proposed to contain microdomains of a specific lipid composition, in which distinct groups of proteins are clustered. Flotillin-like proteins are conserved between pro-and eukaryotes, play an important function in several eukaryotic and bacterial cells, and define in vertebrates a type of so-called detergent-resistant microdomains. Using STED microscopy, we show that two bacterial flotillins, FloA and FloT, form defined assemblies with an average diameter of 85 to 110 nm in the model bacterium Bacillus subtilis. Interestingly, flotillin microdomains are of similar size in eukaryotic cells. The soluble domains of FloA form higher order oligomers of up to several hundred kDa in vitro, showing that like eukaryotic flotillins, bacterial assemblies are based in part on their ability to self-oligomerize. However, B. subtilis paralogs show significantly different diffusion rates, and consequently do not colocalize into a common microdomain. Dual colour time lapse experiments of flotillins together with other detergent-resistant proteins in bacteria show that proteins colocalize for no longer than a few hundred milliseconds, and do not move together. Our data reveal that the bacterial membrane contains defined-sized protein domains rather than functional microdomains dependent on flotillins. Based on their distinct dynamics, FloA and FloT confer spatially distinguishable activities, but do not serve as molecular scaffolds.

  17. Communication: Microsecond dynamics of the protein and water affect electron transfer in a bacterial bc1 complex

    Science.gov (United States)

    Martin, Daniel R.; Matyushov, Dmitry V.

    2015-04-01

    Cross-membrane electron transport between cofactors localized in proteins of mitochondrial respiration and bacterial photosynthesis is the source of all biological energy. The statistics and dynamics of nuclear fluctuations in these protein/membrane/water heterogeneous systems are critical for their energetic efficiency. The results of 13 μs of atomistic molecular dynamics simulations of the membrane-bound bc1 bacterial complex are analyzed here. The reaction is affected by a broad spectrum of nuclear modes, with the slowest dynamics in the range of time-scales ˜0.1-1.6 μs contributing half of the reaction reorganization energy. Two reorganization energies are required to describe protein electron transfer due to dynamical arrest of protein conformations on the observation window. This mechanistic distinction allows significant lowering of activation barriers for reactions in proteins.

  18. Communication: Microsecond dynamics of the protein and water affect electron transfer in a bacterial bc1 complex

    International Nuclear Information System (INIS)

    Martin, Daniel R.; Matyushov, Dmitry V.

    2015-01-01

    Cross-membrane electron transport between cofactors localized in proteins of mitochondrial respiration and bacterial photosynthesis is the source of all biological energy. The statistics and dynamics of nuclear fluctuations in these protein/membrane/water heterogeneous systems are critical for their energetic efficiency. The results of 13 μs of atomistic molecular dynamics simulations of the membrane-bound bc 1 bacterial complex are analyzed here. The reaction is affected by a broad spectrum of nuclear modes, with the slowest dynamics in the range of time-scales ∼0.1-1.6 μs contributing half of the reaction reorganization energy. Two reorganization energies are required to describe protein electron transfer due to dynamical arrest of protein conformations on the observation window. This mechanistic distinction allows significant lowering of activation barriers for reactions in proteins

  19. Communication: Microsecond dynamics of the protein and water affect electron transfer in a bacterial bc{sub 1} complex

    Energy Technology Data Exchange (ETDEWEB)

    Martin, Daniel R.; Matyushov, Dmitry V., E-mail: dmitrym@asu.edu [Department of Physics and Department of Chemistry and Biochemistry, Arizona State University, P.O. Box 871504, Tempe, Arizona 85287 (United States)

    2015-04-28

    Cross-membrane electron transport between cofactors localized in proteins of mitochondrial respiration and bacterial photosynthesis is the source of all biological energy. The statistics and dynamics of nuclear fluctuations in these protein/membrane/water heterogeneous systems are critical for their energetic efficiency. The results of 13 μs of atomistic molecular dynamics simulations of the membrane-bound bc{sub 1} bacterial complex are analyzed here. The reaction is affected by a broad spectrum of nuclear modes, with the slowest dynamics in the range of time-scales ∼0.1-1.6 μs contributing half of the reaction reorganization energy. Two reorganization energies are required to describe protein electron transfer due to dynamical arrest of protein conformations on the observation window. This mechanistic distinction allows significant lowering of activation barriers for reactions in proteins.

  20. Cross-phosphorylation of bacterial serine/threonine and tyrosine protein kinases on key regulatory residues

    Directory of Open Access Journals (Sweden)

    Lei eShi

    2014-09-01

    Full Text Available Bacteria possess protein serine/threonine and tyrosine kinases which resemble eukaryal kinases in their capacity to phosphorylate multiple substrates. We hypothesized that the analogy might extend further, and bacterial kinases may also undergo mutual phosphorylation and activation, which is currently considered as a hallmark of eukaryal kinase networks. In order to test this hypothesis, we explored the capacity of all members of four different classes of serine/threonine and tyrosine kinases present in the firmicute model organism Bacillus subtilis to phosphorylate each other in vitro and interact with each other in vivo. The interactomics data suggested a high degree of connectivity among all types of kinases, while phosphorylation assays revealed equally wide-spread cross-phosphorylation events. Our findings suggest that the Hanks-type kinases PrkC, PrkD and YabT exhibit the highest capacity to phosphorylate other B. subtilis kinases, while the BY-kinase PtkA and the two-component-like kinases RsbW and SpoIIAB show the highest propensity to be phosphorylated by other kinases. Analysis of phosphorylated residues on several selected recipient kinases suggests that most cross-phosphorylation events concern key regulatory residues. Therefore, cross-phosphorylation events are very likely to influence the capacity of recipient kinases to phosphorylate substrates downstream in the signal transduction cascade. We therefore conclude that bacterial serine/threonine and tyrosine kinases probably engage in a network-type behavior previously described only in eukaryal cells.

  1. Dynamic Filament Formation by a Divergent Bacterial Actin-Like ParM Protein.

    Directory of Open Access Journals (Sweden)

    Anthony J Brzoska

    Full Text Available Actin-like proteins (Alps are a diverse family of proteins whose genes are abundant in the chromosomes and mobile genetic elements of many bacteria. The low-copy-number staphylococcal multiresistance plasmid pSK41 encodes ParM, an Alp involved in efficient plasmid partitioning. pSK41 ParM has previously been shown to form filaments in vitro that are structurally dissimilar to those formed by other bacterial Alps. The mechanistic implications of these differences are not known. In order to gain insights into the properties and behavior of the pSK41 ParM Alp in vivo, we reconstituted the parMRC system in the ectopic rod-shaped host, E. coli, which is larger and more genetically amenable than the native host, Staphylococcus aureus. Fluorescence microscopy showed a functional fusion protein, ParM-YFP, formed straight filaments in vivo when expressed in isolation. Strikingly, however, in the presence of ParR and parC, ParM-YFP adopted a dramatically different structure, instead forming axial curved filaments. Time-lapse imaging and selective photobleaching experiments revealed that, in the presence of all components of the parMRC system, ParM-YFP filaments were dynamic in nature. Finally, molecular dissection of the parMRC operon revealed that all components of the system are essential for the generation of dynamic filaments.

  2. Optimized localization of bacterial infections with technetium-99m labelled human immunoglobulin after protein charge selection

    International Nuclear Information System (INIS)

    Welling, M.; Feitsma, H.I.J.; Calame, W.; Ensing, G.J.; Goedemans, W.; Pauwels, E.K.J.

    1994-01-01

    To improve the scintigraphic detection of bacterial infections a protein charge-purified fraction of polyclonal human immunoglobulin was applied as a radiopharmaceutical. This purification was achieved by attaching the immunoglobulin to an anion-exchanger column and by obtaining the column-bound fraction with buffer. The binding to bacteria in vitro and the target to non-target ratios of an experimental thigh infection with Staphylococcus aureus or Klebsiella pneumoniae in mice were evaluated to compare the purified and the unpurified immunoglobulin. The percentage of binding to all gram-positive and gram-negative bacteria used in this study was significantly (P 99m Tc-labelled protein charge-purified polyclonal human immunoglobulin was administered intravenously. At all time intervals the target (infected thighs) to non-target (non-infected thighs) ratios for both infections were significantly higher (P 99m Tc-labelled protein charge-purified immunoglobulin localizes both a gram-positive and a gram-negative thigh infection more intensely and faster than 99m Tc-labelled unpurified immunoglobulin. (orig.)

  3. Quantitative phosphoproteomics reveals the role of protein arginine phosphorylation in the bacterial stress response.

    Science.gov (United States)

    Schmidt, Andreas; Trentini, Débora Broch; Spiess, Silvia; Fuhrmann, Jakob; Ammerer, Gustav; Mechtler, Karl; Clausen, Tim

    2014-02-01

    Arginine phosphorylation is an emerging protein modification implicated in the general stress response of Gram-positive bacteria. The modification is mediated by the arginine kinase McsB, which phosphorylates and inactivates the heat shock repressor CtsR. In this study, we developed a mass spectrometric approach accounting for the peculiar chemical properties of phosphoarginine. The improved methodology was used to analyze the dynamic changes in the Bacillus subtilis arginine phosphoproteome in response to different stress situations. Quantitative analysis showed that a B. subtilis mutant lacking the YwlE arginine phosphatase accumulated a strikingly large number of arginine phosphorylations (217 sites in 134 proteins), however only a minor fraction of these sites was increasingly modified during heat shock or oxidative stress. The main targets of McsB-mediated arginine phosphorylation comprise central factors of the stress response system including the CtsR and HrcA heat shock repressors, as well as major components of the protein quality control system such as the ClpCP protease and the GroEL chaperonine. These findings highlight the impact of arginine phosphorylation in orchestrating the bacterial stress response.

  4. Functional display of ice nucleation protein InaZ on the surface of bacterial ghosts.

    Science.gov (United States)

    Kassmannhuber, Johannes; Rauscher, Mascha; Schöner, Lea; Witte, Angela; Lubitz, Werner

    2017-09-03

    In a concept study the ability to induce heterogeneous ice formation by Bacterial Ghosts (BGs) from Escherichia coli carrying ice nucleation protein InaZ from Pseudomonas syringae in their outer membrane was investigated by a droplet-freezing assay of ultra-pure water. As determined by the median freezing temperature and cumulative ice nucleation spectra it could be demonstrated that both the living recombinant E. coli and their corresponding BGs functionally display InaZ on their surface. Under the production conditions chosen both samples belong to type II ice-nucleation particles inducing ice formation at a temperature range of between -5.6 °C and -6.7 °C, respectively. One advantage for the application of such BGs over their living recombinant mother bacteria is that they are non-living native cell envelopes retaining the biophysical properties of ice nucleation and do no longer represent genetically modified organisms (GMOs).

  5. High-resolution diffraction from crystals of a membrane-protein complex: bacterial outer membrane protein OmpC complexed with the antibacterial eukaryotic protein lactoferrin

    International Nuclear Information System (INIS)

    Sundara Baalaji, N.; Acharya, K. Ravi; Singh, T. P.; Krishnaswamy, S.

    2005-01-01

    Crystals of the complex formed between the bacterial membrane protein OmpC and the antibacterial protein lactoferrin suitable for high-resolution structure determination have been obtained. The crystals belong to the hexagonal space group P6, with unit-cell parameters a = b = 116.3, c = 152.4 Å. Crystals of the complex formed between the outer membrane protein OmpC from Escherichia coli and the eukaryotic antibacterial protein lactoferrin from Camelus dromedarius (camel) have been obtained using a detergent environment. Initial data processing suggests that the crystals belong to the hexagonal space group P6, with unit-cell parameters a = b = 116.3, c = 152.4 Å, α = β = 90, γ = 120°. This indicated a Matthews coefficient (V M ) of 3.3 Å 3 Da −1 , corresponding to a possible molecular complex involving four molecules of lactoferrin and two porin trimers in the unit cell (4832 amino acids; 533.8 kDa) with 63% solvent content. A complete set of diffraction data was collected to 3 Å resolution at 100 K. Structure determination by molecular replacement is in progress. Structural study of this first surface-exposed membrane-protein complex with an antibacterial protein will provide insights into the mechanism of action of OmpC as well as lactoferrin

  6. A bacterial view of the periodic table: genes and proteins for toxic inorganic ions.

    Science.gov (United States)

    Silver, Simon; Phung, Le T

    2005-12-01

    Essentially all bacteria have genes for toxic metal ion resistances and these include those for Ag+, AsO2-, AsO4(3-), Cd2+ Co2+, CrO4(2-), Cu2+, Hg2+, Ni2+, Pb2+, TeO3(2-), Tl+ and Zn2+. The largest group of resistance systems functions by energy-dependent efflux of toxic ions. Fewer involve enzymatic transformations (oxidation, reduction, methylation, and demethylation) or metal-binding proteins (for example, metallothionein SmtA, chaperone CopZ and periplasmic silver binding protein SilE). Some of the efflux resistance systems are ATPases and others are chemiosmotic ion/proton exchangers. For example, Cd2+-efflux pumps of bacteria are either inner membrane P-type ATPases or three polypeptide RND chemiosmotic complexes consisting of an inner membrane pump, a periplasmic-bridging protein and an outer membrane channel. In addition to the best studied three-polypeptide chemiosmotic system, Czc (Cd2+, Zn2+, and Co2), others are known that efflux Ag+, Cu+, Ni2+, and Zn2+. Resistance to inorganic mercury, Hg2+ (and to organomercurials, such as CH3Hg+ and phenylmercury) involve a series of metal-binding and membrane transport proteins as well as the enzymes mercuric reductase and organomercurial lyase, which overall convert more toxic to less toxic forms. Arsenic resistance and metabolizing systems occur in three patterns, the widely-found ars operon that is present in most bacterial genomes and many plasmids, the more recently recognized arr genes for the periplasmic arsenate reductase that functions in anaerobic respiration as a terminal electron acceptor, and the aso genes for the periplasmic arsenite oxidase that functions as an initial electron donor in aerobic resistance to arsenite.

  7. A bacterial cysteine protease effector protein interferes with photosynthesis to suppress plant innate immune responses.

    Science.gov (United States)

    Rodríguez-Herva, José J; González-Melendi, Pablo; Cuartas-Lanza, Raquel; Antúnez-Lamas, María; Río-Alvarez, Isabel; Li, Ziduo; López-Torrejón, Gema; Díaz, Isabel; Del Pozo, Juan C; Chakravarthy, Suma; Collmer, Alan; Rodríguez-Palenzuela, Pablo; López-Solanilla, Emilia

    2012-05-01

    The bacterial pathogen Pseudomonas syringae pv tomato DC3000 suppresses plant innate immunity with effector proteins injected by a type III secretion system (T3SS). The cysteine protease effector HopN1, which reduces the ability of DC3000 to elicit programmed cell death in non-host tobacco, was found to also suppress the production of defence-associated reactive oxygen species (ROS) and callose when delivered by Pseudomonas fluorescens heterologously expressing a P. syringae T3SS. Purified His(6) -tagged HopN1 was used to identify tomato PsbQ, a member of the oxygen evolving complex of photosystem II (PSII), as an interacting protein. HopN1 localized to chloroplasts and both degraded PsbQ and inhibited PSII activity in chloroplast preparations, whereas a HopN1(D299A) non-catalytic mutant lost these abilities. Gene silencing of NtPsbQ in tobacco compromised ROS production and programmed cell death by DC3000. Our data reveal PsbQ as a contributor to plant immunity responses and a target for pathogen suppression. © 2012 Blackwell Publishing Ltd.

  8. Blood parameters in growing pigs fed increasing levels of bacterial protein meal

    Directory of Open Access Journals (Sweden)

    Tauson Anne-Helene

    2007-11-01

    Full Text Available Abstract The experiment investigated the effects of increasing dietary levels of bacterial protein meal (BPM on various blood parameters reflecting protein and fat metabolism, liver function, and purine base metabolism in growing pigs. Sixteen barrows were allocated to four different experimental diets. The control diet was based on soybean meal. In the other three diets soybean meal was replaced with increasing levels of BPM, approximately 17%, 35%, and 50% of the nitrogen being derived from BPM. Blood samples from the jugular vein were taken when the body weights of the pigs were approximately 10 kg, 21 kg, 45 kg, and 77 kg. The blood parameters reflecting fat metabolism and liver function were not affected by diet. Both the plasma albumin and uric acid concentrations tended to decrease (P = 0.07 and 0.01, respectively with increasing dietary BPM content, whereas the plasma glucose concentration tended to increase (P = 0.07 with increasing dietary BPM content. It was concluded that up to 50% of the nitrogen could be derived from BPM without affecting metabolic function, as reflected in the measured blood parameters.

  9. Coordination of genomic structure and transcription by the main bacterial nucleoid-associated protein HU

    Science.gov (United States)

    Berger, Michael; Farcas, Anca; Geertz, Marcel; Zhelyazkova, Petya; Brix, Klaudia; Travers, Andrew; Muskhelishvili, Georgi

    2010-01-01

    The histone-like protein HU is a highly abundant DNA architectural protein that is involved in compacting the DNA of the bacterial nucleoid and in regulating the main DNA transactions, including gene transcription. However, the coordination of the genomic structure and function by HU is poorly understood. Here, we address this question by comparing transcript patterns and spatial distributions of RNA polymerase in Escherichia coli wild-type and hupA/B mutant cells. We demonstrate that, in mutant cells, upregulated genes are preferentially clustered in a large chromosomal domain comprising the ribosomal RNA operons organized on both sides of OriC. Furthermore, we show that, in parallel to this transcription asymmetry, mutant cells are also impaired in forming the transcription foci—spatially confined aggregations of RNA polymerase molecules transcribing strong ribosomal RNA operons. Our data thus implicate HU in coordinating the global genomic structure and function by regulating the spatial distribution of RNA polymerase in the nucleoid. PMID:20010798

  10. Photorhabdus adhesion modification protein (Pam) binds extracellular polysaccharide and alters bacterial attachment

    LENUS (Irish Health Repository)

    Jones, Robert T

    2010-05-12

    Abstract Background Photorhabdus are Gram-negative nematode-symbiotic and insect-pathogenic bacteria. The species Photorhabdus asymbiotica is able to infect humans as well as insects. We investigated the secreted proteome of a clinical isolate of P. asymbiotica at different temperatures in order to identify proteins relevant to the infection of the two different hosts. Results A comparison of the proteins secreted by a clinical isolate of P. asymbiotica at simulated insect (28°C) and human (37°C) temperatures led to the identification of a small and highly abundant protein, designated Pam, that is only secreted at the lower temperature. The pam gene is present in all Photorhabdus strains tested and shows a high level of conservation across the whole genus, suggesting it is both ancestral to the genus and probably important to the biology of the bacterium. The Pam protein shows limited sequence similarity to the 13.6 kDa component of a binary toxin of Bacillus thuringiensis. Nevertheless, injection or feeding of heterologously produced Pam showed no insecticidal activity to either Galleria mellonella or Manduca sexta larvae. In bacterial colonies, Pam is associated with an extracellular polysaccharide (EPS)-like matrix, and modifies the ability of wild-type cells to attach to an artificial surface. Interestingly, Surface Plasmon Resonance (SPR) binding studies revealed that the Pam protein itself has adhesive properties. Although Pam is produced throughout insect infection, genetic knockout does not affect either insect virulence or the ability of P. luminescens to form a symbiotic association with its host nematode, Heterorhabditis bacteriophora. Conclusions We studied a highly abundant protein, Pam, which is secreted in a temperature-dependent manner in P. asymbiotica. Our findings indicate that Pam plays an important role in enhancing surface attachment in insect blood. Its association with exopolysaccharide suggests it may exert its effect through mediation of

  11. Photorhabdus adhesion modification protein (Pam binds extracellular polysaccharide and alters bacterial attachment

    Directory of Open Access Journals (Sweden)

    Joyce Susan A

    2010-05-01

    Full Text Available Abstract Background Photorhabdus are Gram-negative nematode-symbiotic and insect-pathogenic bacteria. The species Photorhabdus asymbiotica is able to infect humans as well as insects. We investigated the secreted proteome of a clinical isolate of P. asymbiotica at different temperatures in order to identify proteins relevant to the infection of the two different hosts. Results A comparison of the proteins secreted by a clinical isolate of P. asymbiotica at simulated insect (28°C and human (37°C temperatures led to the identification of a small and highly abundant protein, designated Pam, that is only secreted at the lower temperature. The pam gene is present in all Photorhabdus strains tested and shows a high level of conservation across the whole genus, suggesting it is both ancestral to the genus and probably important to the biology of the bacterium. The Pam protein shows limited sequence similarity to the 13.6 kDa component of a binary toxin of Bacillus thuringiensis. Nevertheless, injection or feeding of heterologously produced Pam showed no insecticidal activity to either Galleria mellonella or Manduca sexta larvae. In bacterial colonies, Pam is associated with an extracellular polysaccharide (EPS-like matrix, and modifies the ability of wild-type cells to attach to an artificial surface. Interestingly, Surface Plasmon Resonance (SPR binding studies revealed that the Pam protein itself has adhesive properties. Although Pam is produced throughout insect infection, genetic knockout does not affect either insect virulence or the ability of P. luminescens to form a symbiotic association with its host nematode, Heterorhabditis bacteriophora. Conclusions We studied a highly abundant protein, Pam, which is secreted in a temperature-dependent manner in P. asymbiotica. Our findings indicate that Pam plays an important role in enhancing surface attachment in insect blood. Its association with exopolysaccharide suggests it may exert its effect

  12. Mutations in the Bacterial Ribosomal Protein L3 and Their Association with Antibiotic Resistance

    Science.gov (United States)

    Klitgaard, Rasmus N.; Ntokou, Eleni; Nørgaard, Katrine; Biltoft, Daniel; Hansen, Lykke H.; Trædholm, Nicolai M.; Kongsted, Jacob

    2015-01-01

    Different groups of antibiotics bind to the peptidyl transferase center (PTC) in the large subunit of the bacterial ribosome. Resistance to these groups of antibiotics has often been linked with mutations or methylations of the 23S rRNA. In recent years, there has been a rise in the number of studies where mutations have been found in the ribosomal protein L3 in bacterial strains resistant to PTC-targeting antibiotics but there is often no evidence that these mutations actually confer antibiotic resistance. In this study, a plasmid exchange system was used to replace plasmid-carried wild-type genes with mutated L3 genes in a chromosomal L3 deletion strain. In this way, the essential L3 gene is available for the bacteria while allowing replacement of the wild type with mutated L3 genes. This enables investigation of the effect of single mutations in Escherichia coli without a wild-type L3 background. Ten plasmid-carried mutated L3 genes were constructed, and their effect on growth and antibiotic susceptibility was investigated. Additionally, computational modeling of the impact of L3 mutations in E. coli was used to assess changes in 50S structure and antibiotic binding. All mutations are placed in the loops of L3 near the PTC. Growth data show that 9 of the 10 mutations were well accepted in E. coli, although some of them came with a fitness cost. Only one of the mutants exhibited reduced susceptibility to linezolid, while five exhibited reduced susceptibility to tiamulin. PMID:25845869

  13. Structural, kinetic and proteomic characterization of acetyl phosphate-dependent bacterial protein acetylation.

    Directory of Open Access Journals (Sweden)

    Misty L Kuhn

    Full Text Available The emerging view of Nε-lysine acetylation in eukaryotes is of a relatively abundant post-translational modification (PTM that has a major impact on the function, structure, stability and/or location of thousands of proteins involved in diverse cellular processes. This PTM is typically considered to arise by the donation of the acetyl group from acetyl-coenzyme A (acCoA to the ε-amino group of a lysine residue that is reversibly catalyzed by lysine acetyltransferases and deacetylases. Here, we provide genetic, mass spectrometric, biochemical and structural evidence that Nε-lysine acetylation is an equally abundant and important PTM in bacteria. Applying a recently developed, label-free and global mass spectrometric approach to an isogenic set of mutants, we detected acetylation of thousands of lysine residues on hundreds of Escherichia coli proteins that participate in diverse and often essential cellular processes, including translation, transcription and central metabolism. Many of these acetylations were regulated in an acetyl phosphate (acP-dependent manner, providing compelling evidence for a recently reported mechanism of bacterial Nε-lysine acetylation. These mass spectrometric data, coupled with observations made by crystallography, biochemistry, and additional mass spectrometry showed that this acP-dependent acetylation is both non-enzymatic and specific, with specificity determined by the accessibility, reactivity and three-dimensional microenvironment of the target lysine. Crystallographic evidence shows acP can bind to proteins in active sites and cofactor binding sites, but also potentially anywhere molecules with a phosphate moiety could bind. Finally, we provide evidence that acP-dependent acetylation can impact the function of critical enzymes, including glyceraldehyde-3-phosphate dehydrogenase, triosephosphate isomerase, and RNA polymerase.

  14. Host and bacterial proteins that repress recruitment of LC3 to Shigella early during infection.

    Directory of Open Access Journals (Sweden)

    Leigh A Baxt

    Full Text Available Shigella spp. are intracytosolic gram-negative pathogens that cause disease by invasion and spread through the colonic mucosa, utilizing host cytoskeletal components to form propulsive actin tails. We have previously identified the host factor Toca-1 as being recruited to intracellular S. flexneri and being required for efficient bacterial actin tail formation. We show that at early times during infection (40 min., the type three-secreted effector protein IcsB recruits Toca-1 to intracellular bacteria and that recruitment of Toca-1 is associated with repression of recruitment of LC3, as well as with repression of recruitment of the autophagy marker NDP52, around these intracellular bacteria. LC3 is best characterized as a marker of autophagosomes, but also marks phagosomal membranes in the process LC3-associated phagocytosis. IcsB has previously been demonstrated to be required for S. flexneri evasion of autophagy at late times during infection (4-6 hr by inhibiting binding of the autophagy protein Atg5 to the Shigella surface protein IcsA (VirG. Our results suggest that IcsB and Toca-1 modulation of LC3 recruitment restricts LC3-associated phagocytosis and/or LC3 recruitment to vacuolar membrane remnants. Together with published results, our findings suggest that IcsB inhibits innate immune responses in two distinct ways, first, by inhibiting LC3-associated phagocytosis and/or LC3 recruitment to vacuolar membrane remnants early during infection, and second, by inhibiting autophagy late during infection.

  15. Generation of Nanobodies against SlyD and development of tools to eliminate this bacterial contaminant from recombinant proteins.

    Science.gov (United States)

    Hu, Yaozhong; Romão, Ema; Vertommen, Didier; Vincke, Cécile; Morales-Yánez, Francisco; Gutiérrez, Carlos; Liu, Changxiao; Muyldermans, Serge

    2017-09-01

    The gene for a protein domain, derived from a tumor marker, fused to His tag codons and under control of a T7 promotor was expressed in E. coli strain BL21 (DE3). The recombinant protein was purified from cell lysates through immobilized metal affinity chromatography and size-exclusion chromatography. A contaminating bacterial protein was consistently co-purified, even using stringent washing solutions containing 50 or 100 mM imidazole. Immunization of a dromedary with this contaminated protein preparation, and the subsequent generation and panning of the immune Nanobody library yielded several Nanobodies of which 2/3 were directed against the bacterial contaminant, reflecting the immunodominance of this protein to steer the dromedary immune response. Affinity adsorption of this contaminant using one of our specific Nanobodies followed by mass spectrometry identified the bacterial contaminant as FKBP-type peptidyl-prolyl cis-trans isomerase (SlyD) from E. coli. This SlyD protein contains in its C-terminal region 14 histidines in a stretch of 31 amino acids, which explains its co-purification on Ni-NTA resin. This protein is most likely present to varying extents in all recombinant protein preparations after immobilized metal affinity chromatography. Using our SlyD-specific Nb 5 we generated an immune-complex that could be removed either by immunocapturing or by size exclusion chromatography. Both methods allow us to prepare a recombinant protein sample where the SlyD contaminant was quantitatively eliminated. Copyright © 2017 Elsevier Inc. All rights reserved.

  16. Surfactant protein D augments bacterial association but attenuates major histocompatibility complex class II presentation of bacterial antigens

    DEFF Research Database (Denmark)

    Hansen, Søren; Lo, Bernice; Evans, Kathy

    2006-01-01

    Development of dementia, including Alzheimer's disease (AD), is associated with lipid dysregulation and inflammation. As the host defense lectin surfactant protein D (SP-D) has multiple effects in lipid homeostasis and inflammation, the correlation between SP-D concentrations and development of d...

  17. Bacterial effector binding to ribosomal protein s3 subverts NF-kappaB function.

    Directory of Open Access Journals (Sweden)

    Xiaofei Gao

    2009-12-01

    Full Text Available Enteric bacterial pathogens cause food borne disease, which constitutes an enormous economic and health burden. Enterohemorrhagic Escherichia coli (EHEC causes a severe bloody diarrhea following transmission to humans through various means, including contaminated beef and vegetable products, water, or through contact with animals. EHEC also causes a potentially fatal kidney disease (hemolytic uremic syndrome for which there is no effective treatment or prophylaxis. EHEC and other enteric pathogens (e.g., enteropathogenic E. coli (EPEC, Salmonella, Shigella, Yersinia utilize a type III secretion system (T3SS to inject virulence proteins (effectors into host cells. While it is known that T3SS effectors subvert host cell function to promote diarrheal disease and bacterial transmission, in many cases, the mechanisms by which these effectors bind to host proteins and disrupt the normal function of intestinal epithelial cells have not been completely characterized. In this study, we present evidence that the E. coli O157:H7 nleH1 and nleH2 genes encode T3SS effectors that bind to the human ribosomal protein S3 (RPS3, a subunit of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kappaB transcriptional complexes. NleH1 and NleH2 co-localized with RPS3 in the cytoplasm, but not in cell nuclei. The N-terminal region of both NleH1 and NleH2 was required for binding to the N-terminus of RPS3. NleH1 and NleH2 are autophosphorylated Ser/Thr protein kinases, but their binding to RPS3 is independent of kinase activity. NleH1, but not NleH2, reduced the nuclear abundance of RPS3 without altering the p50 or p65 NF-kappaB subunits or affecting the phosphorylation state or abundance of the inhibitory NF-kappaB chaperone IkappaBalpha NleH1 repressed the transcription of a RPS3/NF-kappaB-dependent reporter plasmid, but did not inhibit the transcription of RPS3-independent reporters. In contrast, NleH2 stimulated RPS3-dependent transcription, as well

  18. A bacterial E3 ubiquitin ligase targets a host protein kinase to disrupt plant immunity.

    Science.gov (United States)

    Rosebrock, Tracy R; Zeng, Lirong; Brady, Jennifer J; Abramovitch, Robert B; Xiao, Fangming; Martin, Gregory B

    2007-07-19

    Many bacterial pathogens of plants and animals use a type III secretion system to deliver diverse virulence-associated 'effector' proteins into the host cell. The mechanisms by which these effectors act are mostly unknown; however, they often promote disease by suppressing host immunity. One type III effector, AvrPtoB, expressed by the plant pathogen Pseudomonas syringae pv. tomato, has a carboxy-terminal domain that is an E3 ubiquitin ligase. Deletion of this domain allows an amino-terminal region of AvrPtoB (AvrPtoB(1-387)) to be detected by certain tomato varieties leading to immunity-associated programmed cell death. Here we show that a host kinase, Fen, physically interacts with AvrPtoB(1-387 )and is responsible for activating the plant immune response. The AvrPtoB E3 ligase specifically ubiquitinates Fen and promotes its degradation in a proteasome-dependent manner. This degradation leads to disease susceptibility in Fen-expressing tomato lines. Various wild species of tomato were found to exhibit immunity in response to AvrPtoB(1-387 )and not to full-length AvrPtoB. Thus, by acquiring an E3 ligase domain, AvrPtoB has thwarted a highly conserved host resistance mechanism.

  19. Growh performance, nitrogen balance and urinary purine derivatives in growing-furring mink (Mustela vison) fed bacterial protein produced from natural gas

    DEFF Research Database (Denmark)

    Ahlstrøm, Ø.; Tauson, Anne-Helene; Hellwing, Anne Louise Frydendahl

    2006-01-01

    A bacterial protein meal (BPM), containing 70% crude protein and produced on natural gas, was evaluated versus fish meal as protein source for mink in the growing-furring period (June 29-November 26). BPM, rich in nucleic acids, accounted for 0 (control), 20 and 40% of dietary crude protein...

  20. Novel protein interactions with an actin homolog (MreB) of Helicobacter pylori determined by bacterial two-hybrid system.

    Science.gov (United States)

    Zepeda Gurrola, Reyna Cristina; Fu, Yajuan; Rodríguez Luna, Isabel Cristina; Benítez Cardoza, Claudia Guadalupe; López López, María de Jesús; López Vidal, Yolanda; Gutíerrez, Germán Rubén Aguilar; Rodríguez Pérez, Mario A; Guo, Xianwu

    2017-08-01

    The bacterium Helicobacter pylori infects more than 50% of the world population and causes several gastroduodenal diseases, including gastric cancer. Nevertheless, we still need to explore some protein interactions that may be involved in pathogenesis. MreB, an actin homolog, showed some special characteristics in previous studies, indicating that it could have different functions. Protein functions could be realized via protein-protein interactions. In the present study, the MreB protein from H. pylori 26695 fused with two tags 10×His and GST in tandem was overexpressed and purified from Escherchia coli. The purified recombinant protein was used to perform a pull-down assay with H. pylori 26695 cell lysate. The pulled-down proteins were identified by mass spectrometry (MALDI-TOF), in which the known important proteins related to morphogenesis were absent but several proteins related to pathogenesis process were observed. The bacterial two-hybrid system was further used to evaluate the protein interactions and showed that new interactions of MreB respectively with VacA, UreB, HydB, HylB and AddA were confirmed but the interaction MreB-MreC was not validated. These results indicated that the protein MreB in H. pylori has a distinct interactome, does not participate in cell morphogenesis via MreB-MreC but could be related to pathogenesis. Copyright © 2017 Elsevier GmbH. All rights reserved.

  1. An efficient tag derived from the common epitope of tospoviral NSs proteins for monitoring recombinant proteins expressed in both bacterial and plant systems.

    Science.gov (United States)

    Cheng, Hao-Wen; Chen, Kuan-Chun; Raja, Joseph A J; Li, Jian-Xian; Yeh, Shyi-Dong

    2013-04-15

    NSscon (23 aa), a common epitope in the gene silencing suppressor NSs proteins of the members of the Watermelon silver mottle virus (WSMoV) serogroup, was previously identified. In this investigation, we expressed different green fluorescent protein (GFP)-fused deletions of NSscon in bacteria and reacted with NSscon monoclonal antibody (MAb). Our results indicated that the core 9 amino acids, "(109)KFTMHNQIF(117)", denoted as "nss", retain the reactivity of NSscon. In bacterial pET system, four different recombinant proteins labeled with nss, either at N- or C-extremes, were readily detectable without position effects, with sensitivity superior to that for the polyhistidine-tag. When the nss-tagged Zucchini yellow mosaic virus (ZYMV) helper component-protease (HC-Pro) and WSMoV nucleocapsid protein were transiently expressed by agroinfiltration in tobacco, they were readily detectable and the tag's possible efficacy for gene silencing suppression was not noticed. Co-immunoprecipitation of nss-tagged and non-tagged proteins expressed from bacteria confirmed the interaction of potyviral HC-Pro and coat protein. Thus, we conclude that this novel nss sequence is highly valuable for tagging recombinant proteins in both bacterial and plant expression systems. Copyright © 2013 Elsevier B.V. All rights reserved.

  2. Structure of the complex between teicoplanin and a bacterial cell-wall peptide: use of a carrier-protein approach

    International Nuclear Information System (INIS)

    Economou, Nicoleta J.; Zentner, Isaac J.; Lazo, Edwin; Jakoncic, Jean; Stojanoff, Vivian; Weeks, Stephen D.; Grasty, Kimberly C.; Cocklin, Simon; Loll, Patrick J.

    2013-01-01

    Using a carrier-protein strategy, the structure of teicoplanin bound to its bacterial cell-wall target has been determined. The structure reveals the molecular determinants of target recognition, flexibility in the antibiotic backbone and intrinsic radiation sensitivity of teicoplanin. Multidrug-resistant bacterial infections are commonly treated with glycopeptide antibiotics such as teicoplanin. This drug inhibits bacterial cell-wall biosynthesis by binding and sequestering a cell-wall precursor: a d-alanine-containing peptide. A carrier-protein strategy was used to crystallize the complex of teicoplanin and its target peptide by fusing the cell-wall peptide to either MBP or ubiquitin via native chemical ligation and subsequently crystallizing the protein–peptide–antibiotic complex. The 2.05 Å resolution MBP–peptide–teicoplanin structure shows that teicoplanin recognizes its ligand through a combination of five hydrogen bonds and multiple van der Waals interactions. Comparison of this teicoplanin structure with that of unliganded teicoplanin reveals a flexibility in the antibiotic peptide backbone that has significant implications for ligand recognition. Diffraction experiments revealed an X-ray-induced dechlorination of the sixth amino acid of the antibiotic; it is shown that teicoplanin is significantly more radiation-sensitive than other similar antibiotics and that ligand binding increases radiosensitivity. Insights derived from this new teicoplanin structure may contribute to the development of next-generation antibacterials designed to overcome bacterial resistance

  3. Utility of the serum C-reactive protein for detection of occult bacterial infection in children.

    Science.gov (United States)

    Isaacman, Daniel J; Burke, Bonnie L

    2002-09-01

    To assess the utility of serum C-reactive protein (CRP) as a screen for occult bacterial infection in children. Febrile children ages 3 to 36 months who visited an urban children's hospital emergency department and received a complete blood cell count and blood culture as part of their evaluation were prospectively enrolled from February 2, 2000, through May 30, 2001. Informed consent was obtained for the withdrawal of an additional 1-mL aliquot of blood for use in CRP evaluation. Logistic regression and receiver operator characteristic (ROC) curves were modeled for each predictor to identify optimal test values, and were compared using likelihood ratio tests. Two hundred fifty-six patients were included in the analysis, with a median age of 15.3 months (range, 3.1-35.2 months) and median temperature at triage 40.0 degrees C (range, 39.0 degrees C-41.3 degrees C). Twenty-nine (11.3%) cases of occult bacterial infection (OBI) were identified, including 17 cases of pneumonia, 9 cases of urinary tract infection, and 3 cases of bacteremia. The median white blood cell count in this data set was 12.9 x 10(3)/ micro L [corrected] (range, 3.6-39.1 x10(3)/ micro L) [corrected], the median absolute neutrophil count (ANC) was 7.12 x 10(3)/L [corrected] (range, 0.56-28.16 x10(3)/L) [corrected], and the median CRP level was 1.7 mg/dL (range, 0.2-43.3 mg/dL). The optimal cut-off point for CRP in this data set (4.4 mg/dL) achieved a sensitivity of 63% and a specificity of 81% for detection of OBI in this population. Comparing models using cut-off values from individual laboratory predictors (ANC, white blood cell count, and CRP) that maximized sensitivity and specificity revealed that a model using an ANC of 10.6 x10(3)/L [corrected] (sensitivity, 69%; specificity, 79%) was the best predictive model. Adding CRP to the model insignificantly increased sensitivity to 79%, while significantly decreasing specificity to 50%. Active monitoring of emergency department blood cultures

  4. Biological characterization of a new radioactive labeling reagent for bacterial penicillin-binding proteins

    International Nuclear Information System (INIS)

    Preston, D.A.; Wu, C.Y.; Blaszczak, L.C.; Seitz, D.E.; Halligan, N.G.

    1990-01-01

    Radiolabeled penicillin G is widely used as the imaging agent in penicillin-binding protein (PBP) assays. The disadvantages of most forms of labeled penicillin G are instability on storage and the long exposure times usually required for autoradiography or fluorography of electrophoretic gels. We investigated the utility of radioiodinated penicillin V as an alternative reagent. Radioiodination of p-(trimethylstannyl)penicillin V with [ 125 I]Na, using a modification of the chloramine-T method, is simple, high yielding, and site specific. We demonstrated the general equivalence of commercially obtained [ 3 H]penicillin G and locally synthesized [ 125 I]penicillin V (IPV) in their recognition of bacterial PBPs. Profiles of PBPs in membranes from Bacteroides fragilis, Escherichia coli, Providencia rettgeri, Staphylococcus aureus, Streptococcus pyogenes, Enterococcus faecalis, and Enterococcus faecium labeled with IPV or [3H]penicillin G were virtually identical. Use of IPV as the imaging agent in competition experiments for determination of the affinities of various beta-lactam antibiotics for the PBPs of E. coli yielded results similar to those obtained in experiments with [ 3 H]penicillin G. Dried electrophoretic gels from typical PBP experiments, using IPV at 37.3 Ci/mmol and 30 micrograms/ml, exposed X-ray film in 8 to 24 h. The stability of IPV on storage at 4 degrees C was inversely proportional to specific activity. At 37.3 Ci/mmol and 60 micrograms/ml, IPV retained useful activity for at least 60 days at 4 degrees C. IPV represents a practical and stable reagent for rapid PBP assays

  5. Coevolved Mutations Reveal Distinct Architectures for Two Core Proteins in the Bacterial Flagellar Motor.

    Directory of Open Access Journals (Sweden)

    Alessandro Pandini

    Full Text Available Switching of bacterial flagellar rotation is caused by large domain movements of the FliG protein triggered by binding of the signal protein CheY to FliM. FliG and FliM form adjacent multi-subunit arrays within the basal body C-ring. The movements alter the interaction of the FliG C-terminal (FliGC "torque" helix with the stator complexes. Atomic models based on the Salmonella entrovar C-ring electron microscopy reconstruction have implications for switching, but lack consensus on the relative locations of the FliG armadillo (ARM domains (amino-terminal (FliGN, middle (FliGM and FliGC as well as changes during chemotaxis. The generality of the Salmonella model is challenged by the variation in motor morphology and response between species. We studied coevolved residue mutations to determine the unifying elements of switch architecture. Residue interactions, measured by their coevolution, were formalized as a network, guided by structural data. Our measurements reveal a common design with dedicated switch and motor modules. The FliM middle domain (FliMM has extensive connectivity most simply explained by conserved intra and inter-subunit contacts. In contrast, FliG has patchy, complex architecture. Conserved structural motifs form interacting nodes in the coevolution network that wire FliMM to the FliGC C-terminal, four-helix motor module (C3-6. FliG C3-6 coevolution is organized around the torque helix, differently from other ARM domains. The nodes form separated, surface-proximal patches that are targeted by deleterious mutations as in other allosteric systems. The dominant node is formed by the EHPQ motif at the FliMMFliGM contact interface and adjacent helix residues at a central location within FliGM. The node interacts with nodes in the N-terminal FliGc α-helix triad (ARM-C and FliGN. ARM-C, separated from C3-6 by the MFVF motif, has poor intra-network connectivity consistent with its variable orientation revealed by structural data. ARM

  6. Manipulating the glycosylation pathway in bacterial and lower eukaryotes for production of therapeutic proteins

    DEFF Research Database (Denmark)

    Anyaogu, Diana Chinyere; Mortensen, Uffe Hasbro

    2015-01-01

    The medical use of pharmaceutical proteins is rapidly increasing and cheap, fast and efficient production is therefore attractive. Microbial production hosts are promising candidates for development and production of pharmaceutical proteins. However, as most therapeutic proteins are secreted...... to produce proteins with humanlike glycan structures setting the stage for production of pharmaceutical proteins in bacteria, yeasts and algae....

  7. Bacterial Protein Characterization of Streptococcus agalactiae by SDS-page Method for Subclinical Mastitis Irradiated Vaccine Materials in Dairy Cattle

    International Nuclear Information System (INIS)

    Tuasikal, B.J.; Wibawan, I.W.T.; Pasaribu, F.H; Estuningsih, S.

    2012-01-01

    A study have been conducted to isolate and characterize bacterial protein S. agalactiae, which is antigenic and can be used to test immunogenicity of vaccine in order to manufacture irradiated mastitis (inflammation of the udder) vaccine in ruminant. The study aims to determine the Molecular Weight (MW) bacterial protein S. agalactiae irradiation, which can be used to test the nature of its antigenic caharacteristic. The character of S. agalactiae antigenic stimulates antibody induction of the immune system, in which case is the body's defense system against mastitis disease in cattle. In this study, irradiation of gamma ray is used to attenuate the pathogenicity of bacteria by reducing S. agalactiae antigenic characteristic. Previous research, in irradiation dose orientation before antigenic protein isolation of S. agalactiae, indicated that irradiation lethal dose to 50% (LD 50 ) is 17 Gy. The characterization of S. agalactiae bacteria isolate using SDS-page method results in no significance different between irradiated and non-irradiated group, which indicated by MW range 75 - 100 kDa base on marker standard which used, or 99 kDa by the linier equation of Y = 11,60 - 0.05X (where Y = bands distance; X = MW standard protein); r 2 = 0.99. In conclusion, 17 Gy irradiation dose does not impair antigenic property of S. agalactiae and therefore, can be applied to produce base material of irradiated vaccine for mastitis. (author)

  8. S-layer proteins from Lactobacillus sp. inhibit bacterial infection by blockage of DC-SIGN cell receptor.

    Science.gov (United States)

    Prado Acosta, Mariano; Ruzal, Sandra M; Cordo, Sandra M

    2016-11-01

    Many species of Lactobacillus sp. possess Surface(s) layer proteins in their envelope. Among other important characteristics S-layer from Lactobacillus acidophilus binds to the cellular receptor DC-SIGN (Dendritic Cell-Specific Intercellular adhesion molecule-3-Grabbing Non-integrin; CD209), which is involved in adhesion and infection of several families of bacteria. In this report we investigate the activity of new S-layer proteins from the Lactobacillus family (Lactobacillus acidophilus, Lactobacillus brevis, Lactobacillus helveticus and Lactobacillus kefiri) over the infection of representative microorganisms important to human health. After the treatment of DC-SIGN expressing cells with these proteins, we were able to diminish bacterial infection by up to 79% in both gram negative and mycobacterial models. We discovered that pre-treatment of the bacteria with S-layers from Lactobacillus acidophilus and Lactobacillus brevis reduced bacteria viability but also prevent infection by the pathogenic bacteria. We also proved the importance of the glycosylation of the S-layer from Lactobacillus kefiri in the binding to the receptor and thus inhibition of infection. This novel characteristic of the S-layers proteins may contribute to the already reported pathogen exclusion activity for these Lactobacillus probiotic strains; and might be also considered as a novel enzymatic antimicrobial agents to inhibit bacterial infection and entry to host cells. Copyright © 2016 Elsevier B.V. All rights reserved.

  9. A census of membrane-bound and intracellular signal transduction proteins in bacteria: bacterial IQ, extroverts and introverts.

    Science.gov (United States)

    Galperin, Michael Y

    2005-06-14

    Analysis of complete microbial genomes showed that intracellular parasites and other microorganisms that inhabit stable ecological niches encode relatively primitive signaling systems, whereas environmental microorganisms typically have sophisticated systems of environmental sensing and signal transduction. This paper presents results of a comprehensive census of signal transduction proteins--histidine kinases, methyl-accepting chemotaxis receptors, Ser/Thr/Tyr protein kinases, adenylate and diguanylate cyclases and c-di-GMP phosphodiesterases--encoded in 167 bacterial and archaeal genomes, sequenced by the end of 2004. The data have been manually checked to avoid false-negative and false-positive hits that commonly arise during large-scale automated analyses and compared against other available resources. The census data show uneven distribution of most signaling proteins among bacterial and archaeal phyla. The total number of signal transduction proteins grows approximately as a square of genome size. While histidine kinases are found in representatives of all phyla and are distributed according to the power law, other signal transducers are abundant in certain phylogenetic groups but virtually absent in others. The complexity of signaling systems differs even among closely related organisms. Still, it usually can be correlated with the phylogenetic position of the organism, its lifestyle, and typical environmental challenges it encounters. The number of encoded signal transducers (or their fraction in the total protein set) can be used as a measure of the organism's ability to adapt to diverse conditions, the 'bacterial IQ', while the ratio of transmembrane receptors to intracellular sensors can be used to define whether the organism is an 'extrovert', actively sensing the environmental parameters, or an 'introvert', more concerned about its internal homeostasis. Some of the microorganisms with the highest IQ, including the current leader Wolinella succinogenes

  10. A census of membrane-bound and intracellular signal transduction proteins in bacteria: Bacterial IQ, extroverts and introverts

    Directory of Open Access Journals (Sweden)

    Galperin Michael Y

    2005-06-01

    Full Text Available Abstract Background Analysis of complete microbial genomes showed that intracellular parasites and other microorganisms that inhabit stable ecological niches encode relatively primitive signaling systems, whereas environmental microorganisms typically have sophisticated systems of environmental sensing and signal transduction. Results This paper presents results of a comprehensive census of signal transduction proteins – histidine kinases, methyl-accepting chemotaxis receptors, Ser/Thr/Tyr protein kinases, adenylate and diguanylate cyclases and c-di-GMP phosphodiesterases – encoded in 167 bacterial and archaeal genomes, sequenced by the end of 2004. The data have been manually checked to avoid false-negative and false-positive hits that commonly arise during large-scale automated analyses and compared against other available resources. The census data show uneven distribution of most signaling proteins among bacterial and archaeal phyla. The total number of signal transduction proteins grows approximately as a square of genome size. While histidine kinases are found in representatives of all phyla and are distributed according to the power law, other signal transducers are abundant in certain phylogenetic groups but virtually absent in others. Conclusion The complexity of signaling systems differs even among closely related organisms. Still, it usually can be correlated with the phylogenetic position of the organism, its lifestyle, and typical environmental challenges it encounters. The number of encoded signal transducers (or their fraction in the total protein set can be used as a measure of the organism's ability to adapt to diverse conditions, the 'bacterial IQ', while the ratio of transmembrane receptors to intracellular sensors can be used to define whether the organism is an 'extrovert', actively sensing the environmental parameters, or an 'introvert', more concerned about its internal homeostasis. Some of the microorganisms with the

  11. Nanocomposited coatings produced by laser-assisted process to prevent silicone hydogels from protein fouling and bacterial contamination

    International Nuclear Information System (INIS)

    Huang, Guobang; Chen, Yi; Zhang, Jin

    2016-01-01

    Graphical abstract: Nanocomposited-coating was deposited on silicone hydrogel by using the matrix-assisted pulsed laser evaporation (MAPLE) process. The ZnO–PEG nanocomposited coating reduces over 50% protein absorption on silicone hydrogel, and can inhibit the bacterial growth efficiently. - Highlights: • We developed a nanocomposited coating to prevent silicone hydrogel from biofouling. • Matrix-assisted pulsed laser evaporation can deposit inorganic–organic nanomaterials. • The designed nanocomposited coating reduces protein absorption by over 50%. • The designed nanocomposited coating shows significant antimicrobial efficiency. - Abstract: Zinc oxide (ZnO) nanoparticles incorporating with polyethylene glycol (PEG) were deposited together on the surface of silicone hydrogel through matrix-assisted pulsed laser evaporation (MAPLE). In this process, frozen nanocomposites (ZnO–PEG) in isopropanol were irradiated under a pulsed Nd:YAG laser at 532 nm for 1 h. Our results indicate that the MAPLE process is able to maintain the chemical backbone of polymer and prevent the nanocomposite coating from contamination. The ZnO–PEG nanocomposited coating reduces over 50% protein absorption on silicone hydrogel. The cytotoxicity study shows that the ZnO–PEG nanocomposites deposited on silicone hydrogels do not impose the toxic effect on mouse NIH/3T3 cells. In addition, MAPLE-deposited ZnO–PEG nanocomposites can inhibit the bacterial growth significantly.

  12. Expression of the recombinant bacterial outer surface protein A in tobacco chloroplasts leads to thylakoid localization and loss of photosynthesis.

    Science.gov (United States)

    Hennig, Anna; Bonfig, Katharina; Roitsch, Thomas; Warzecha, Heribert

    2007-11-01

    Bacterial lipoproteins play crucial roles in host-pathogen interactions and pathogenesis and are important targets for the immune system. A prominent example is the outer surface protein A (OspA) of Borrelia burgdorferi, which has been efficiently used as a vaccine for the prevention of Lyme disease. In a previous study, OspA could be produced in tobacco chloroplasts in a lipidated and immunogenic form. To further explore the potential of chloroplasts for the production of bacterial lipoproteins, the role of the N-terminal leader sequence was investigated. The amount of recombinant OspA could be increased up to ten-fold by the variation of the insertion site in the chloroplast genome. Analysis of OspA mutants revealed that replacement of the invariant cysteine residue as well as deletion of the leader sequence abolishes palmitolyation of OspA. Also, decoration of OspA with an N-terminal eukaryotic lipidation motif does not lead to palmitoylation in chloroplasts. Strikingly, the bacterial signal peptide of OspA efficiently targets the protein to thylakoids, and causes a mutant phenotype. Plants accumulating OspA at 10% total soluble protein could not grow without exogenously supplied sugars and rapidly died after transfer to soil under greenhouse conditions. The plants were found to be strongly affected in photosystem II, as revealed by the analyses of temporal and spatial dynamics of photosynthetic activity by chlorophyll fluorescence imaging. Thus, overexpression of OspA in chloroplasts is limited by its concentration-dependent interference with essential functions of chloroplastic membranes required for primary metabolism.

  13. Development and characterization of bacterial cellulose reinforced biocomposite films based on protein from buckwheat distiller's dried grains.

    Science.gov (United States)

    Wang, Xuejiao; Ullah, Niamat; Sun, Xuchun; Guo, Yan; Chen, Lin; Li, Zhixi; Feng, Xianchao

    2017-03-01

    Biocomposite films were manufactured by combining protein extracted from buckwheat distiller's dried grains with bacterial cellulose (BC). The film microstructures showed that BC is compatible with protein matrix and endows the film with high rigidity. Differential scanning calorimetry (DSC) showed that BC can promote thermal stability of the composite films. BC promoted the transition from a Newtonian to a non-Newtonian fluid and the shear thinning behavior of protein-BC solution. Fourier Transform Infrared (FTIR) spectroscopy showed the main functional groups' absorption peaks shifted to lower wavenumbers. Results of both FTIR and viscosity analysis proved the formation of intermolecular interactions through hydrogen bonds. These bonds affected film characteristics such as moisture content (MC), water solubility (WS), and water vapor permeability (WVP), which decreased with addition of BC. The WVP (6.68±0.78-5.95±0.54×10 -10 gm/Pasm 2 ) of the films were lower than other protein films. Tensile strength (TS) values of films containing 1.8% and 2.0% BC (14.98±0.97 and 15.03±2.04MPa) were significantly higher than that of pure protein films (4.26±0.66MPa). Combination of proteins extracted from a waste product and BC led to composite films with low water vapor permeability and excellent mechanical properties. Copyright © 2016 Elsevier B.V. All rights reserved.

  14. Characterization of a synthetic bacterial self-destruction device for programmed cell death and for recombinant proteins release

    Directory of Open Access Journals (Sweden)

    Lupotto Manuel

    2011-06-01

    Full Text Available Abstract Background Bacterial cell lysis is a widely studied mechanism that can be achieved through the intracellular expression of phage native lytic proteins. This mechanism can be exploited for programmed cell death and for gentle cell disruption to release recombinant proteins when in vivo secretion is not feasible. Several genetic parts for cell lysis have been developed and their quantitative characterization is an essential step to enable the engineering of synthetic lytic systems with predictable behavior. Results Here, a BioBrick™ lysis device present in the Registry of Standard Biological Parts has been quantitatively characterized. Its activity has been measured in E. coli by assembling the device under the control of a well characterized N-3-oxohexanoyl-L-homoserine lactone (HSL -inducible promoter and the transfer function, lysis dynamics, protein release capability and genotypic and phenotypic stability of the device have been evaluated. Finally, its modularity was tested by assembling the device to a different inducible promoter, which can be triggered by heat induction. Conclusions The studied device is suitable for recombinant protein release as 96% of the total amount of the intracellular proteins was successfully released into the medium. Furthermore, it has been shown that the device can be assembled to different input devices to trigger cell lysis in response to a user-defined signal. For this reason, this lysis device can be a useful tool for the rational design and construction of complex synthetic biological systems composed by biological parts with known and well characterized function. Conversely, the onset of mutants makes this device unsuitable for the programmed cell death of a bacterial population.

  15. The bacterial defensin resistance protein MprF consists of separable domains for lipid lysinylation and antimicrobial peptide repulsion.

    Directory of Open Access Journals (Sweden)

    Christoph M Ernst

    2009-11-01

    Full Text Available Many bacterial pathogens achieve resistance to defensin-like cationic antimicrobial peptides (CAMPs by the multiple peptide resistance factor (MprF protein. MprF plays a crucial role in Staphylococcus aureus virulence and it is involved in resistance to the CAMP-like antibiotic daptomycin. MprF is a large membrane protein that modifies the anionic phospholipid phosphatidylglycerol with l-lysine, thereby diminishing the bacterial affinity for CAMPs. Its widespread occurrence recommends MprF as a target for novel antimicrobials, although the mode of action of MprF has remained incompletely understood. We demonstrate that the hydrophilic C-terminal domain and six of the fourteen proposed trans-membrane segments of MprF are sufficient for full-level lysyl-phosphatidylglycerol (Lys-PG production and that several conserved amino acid positions in MprF are indispensable for Lys-PG production. Notably, Lys-PG production did not lead to efficient CAMP resistance and most of the Lys-PG remained in the inner leaflet of the cytoplasmic membrane when the large N-terminal hydrophobic domain of MprF was absent, indicating a crucial role of this protein part. The N-terminal domain alone did not confer CAMP resistance or repulsion of the cationic test protein cytochrome c. However, when the N-terminal domain was coexpressed with the Lys-PG synthase domain either in one protein or as two separate proteins, full-level CAMP resistance was achieved. Moreover, only coexpression of the two domains led to efficient Lys-PG translocation to the outer leaflet of the membrane and to full-level cytochrome c repulsion, indicating that the N-terminal domain facilitates the flipping of Lys-PG. Thus, MprF represents a new class of lipid-biosynthetic enzymes with two separable functional domains that synthesize Lys-PG and facilitate Lys-PG translocation. Our study unravels crucial details on the molecular basis of an important bacterial immune evasion mechanism and it may help

  16. Strategies for production of active eukaryotic proteins in bacterial expression system

    Institute of Scientific and Technical Information of China (English)

    Orawan Khow; Sunutcha Suntrarachun

    2012-01-01

    Bacteria have long been the favorite expression system for recombinant protein production. However, the flaw of the system is that insoluble and inactive proteins are co-produced due to codon bias, protein folding, phosphorylation, glycosylation, mRNA stability and promoter strength. Factors are cited and the methods to convert to soluble and active proteins are described, for example a tight control of Escherichia coli milieu, refolding from inclusion body and through fusion technology.

  17. Assessment of heavy metal bioavailability in contaminated sediments and soils using green fluorescent protein-based bacterial biosensors

    International Nuclear Information System (INIS)

    Liao, V.H.-C.; Chien, M.-T.; Tseng, Y.-Y.; Ou, K.-L.

    2006-01-01

    A green fluorescent protein (GFP)-based bacterial biosensor Escherichia coli DH5α (pVLCD1) was developed based on the expression of gfp under the control of the cad promoter and the cadC gene of Staphylococcus aureus plasmid pI258. DH5α (pVLCD1) mainly responded to Cd(II), Pb(II), and Sb(III), the lowest detectable concentrations being 0.1 nmol L -1 , 10 nmol L -1 , and 0.1 nmol L -1 , respectively, with 2 h exposure. The biosensor was field-tested to measure the relative bioavailability of the heavy metals in contaminated sediments and soil samples. The results showed that the majority of heavy metals remained adsorbed to soil particles: Cd(II)/Pb(II) was only partially available to the biosensor in soil-water extracts. Our results demonstrate that the GFP-based bacterial biosensor is useful and applicable in determining the bioavailability of heavy metals with high sensitivity in contaminated sediment and soil samples and suggests a potential for its inexpensive application in environmentally relevant sample tests. - Nonpathogenic GFP-based bacterial biosensor is applicable in determining the bioavailability of heavy metals in environmental samples

  18. The level of neuron-specific enolase and S-100 protein in the cerebrospinal fluid of patients with acute bacterial meningitis

    Directory of Open Access Journals (Sweden)

    A. V. Sokhan

    2016-08-01

    Full Text Available Aim. To evaluate the diagnostic and prognostic role of neuron-specific enolase (NSE and S-100 protein levels in cerebrospinal fluid (CSF of patients with acute bacterial meningitis in the course of the disease. Materials and Methods. 54 cases of acute bacterial meningitis were analyzed, among them – 26 with pneumococcal and 28 with meningococcal etiology. Patients were divided into groups depending on the severity and etiology of disease. In addition to routine laboratory methods, we analyzed the CSF levels of S-100 protein and NSE at admission and after 10 – 12 days of treatment. 12 patients with acute respiratory infections and meningism were examined as a comparison group. Results. In all patients with acute bacterial meningitis CSF NSE and protein S-100 levels were significantly higher than in the control group (P <0,05. CSF neuro specific proteins level was in direct dependence on severity of the disease, and in patients with severe disease was significantly higher than in patients with moderate severity and in the control group (P <0,01. After 10 – 12 days of treatment, the level of the NSE and S-100 protein decreased, but in severe cases was still higher than in the control group (P <0,05. Conclusions. Increased cerebrospinal fluid NSE and S – 100 protein levels shows the presence and value of neurons and glial cells damage in patients with acute bacterial meningitis. CSF S-100 protein and neuron-specific enolase levels help to determine the severity of neurons destruction and glial cells in patients with acute bacterial meningitis. Level of neurospecific protein is in direct proportion to the severity of the disease and is the highest in patients with severe cases (P<0,05. It confirms the diagnostic and prognostic value of CSF neurospecific protein determination in patients with bacterial meningitis.

  19. Bacterial production of site specific {sup 13}C labeled phenylalanine and methodology for high level incorporation into bacterially expressed recombinant proteins

    Energy Technology Data Exchange (ETDEWEB)

    Ramaraju, Bhargavi; McFeeters, Hana; Vogler, Bernhard; McFeeters, Robert L., E-mail: robert.mcfeeters@uah.edu [University of Alabama in Huntsville, Department of Chemistry (United States)

    2017-01-15

    Nuclear magnetic resonance spectroscopy studies of ever larger systems have benefited from many different forms of isotope labeling, in particular, site specific isotopic labeling. Site specific {sup 13}C labeling of methyl groups has become an established means of probing systems not amenable to traditional methodology. However useful, methyl reporter sites can be limited in number and/or location. Therefore, new complementary site specific isotope labeling strategies are valuable. Aromatic amino acids make excellent probes since they are often found at important interaction interfaces and play significant structural roles. Aromatic side chains have many of the same advantages as methyl containing amino acids including distinct {sup 13}C chemical shifts and multiple magnetically equivalent {sup 1}H positions. Herein we report economical bacterial production and one-step purification of phenylalanine with {sup 13}C incorporation at the Cα, Cγ and Cε positions, resulting in two isolated {sup 1}H-{sup 13}C spin systems. We also present methodology to maximize incorporation of phenylalanine into recombinantly overexpressed proteins in bacteria and demonstrate compatibility with ILV-methyl labeling. Inexpensive, site specific isotope labeled phenylalanine adds another dimension to biomolecular NMR, opening new avenues of study.

  20. A secretory system for bacterial production of high-profile protein targets

    DEFF Research Database (Denmark)

    Kotzsch, Alexander; Vernet, Erik; Hammarström, Martin

    2011-01-01

    Escherichia coli represents a robust, inexpensive expression host for the production of recombinant proteins. However, one major limitation is that certain protein classes do not express well in a biologically relevant form using standard expression approaches in the cytoplasm of E. coli. To impr......Escherichia coli represents a robust, inexpensive expression host for the production of recombinant proteins. However, one major limitation is that certain protein classes do not express well in a biologically relevant form using standard expression approaches in the cytoplasm of E. coli...... membrane protein F (OmpF) and osmotically inducible protein Y (OsmY). Based on the results of this initial study, we carried out an extended expression screen employing the OsmY fusion and multiple constructs of a more diverse set of human proteins. Using this high-throughput compatible system, we clearly...

  1. Xylo-Oligosaccharides and Inulin Affect Genotoxicity and Bacterial Populations Differently in a Human Colonic Simulator Challenged with Soy Protein

    Science.gov (United States)

    Christophersen, Claus T.; Petersen, Anne; Licht, Tine R.; Conlon, Michael A.

    2013-01-01

    High dietary intakes of some protein sources, including soy protein, can increase colonic DNA damage in animals, whereas some carbohydrates attenuate this. We investigated whether inulin and xylo-oligosaccharides (XOS) could be protective against DNA strand breaks by adding them to a human colonic simulator consisting of a proximal vessel (PV) (pH 5.5) and a distal vessel (DV) (pH 6.8) inoculated with human faeces and media containing soy protein. Genotoxicity of the liquid phase and microbial population changes in the vessels were measured. Soy protein (3%) was fermented with 1% low amylose cornstarch for 10 day followed by soy protein with 1% XOS or 1% inulin for 10 day. Inulin did not alter genotoxicity but XOS significantly reduced PV genotoxicity and increased DV genotoxicity. Inulin and XOS significantly increased butyrate concentration in the DV but not PV. Numbers of the key butyrate-producing bacterium Faecalibacterium prausnitzii were significantly increased in the PV and DV by inulin but significantly decreased by XOS in both vessels. Other bacteria examined were also significantly impacted by the carbohydrate treatments or by the vessel (i.e., pH). There was a significant overall inverse correlation between levels of damage induced by the ferments and levels of sulphate-reducing bacteria, Bacteroides fragilis, and acetate. In conclusion, dietary XOS can potentially modulate the genotoxicity of the colonic environment and specific bacterial groups and short chain fatty acids may mediate this. PMID:24064573

  2. Positioning cell wall synthetic complexes by the bacterial morphogenetic proteins MreB and MreD.

    Science.gov (United States)

    White, Courtney L; Kitich, Aleksandar; Gober, James W

    2010-05-01

    In Caulobacter crescentus, intact cables of the actin homologue, MreB, are required for the proper spatial positioning of MurG which catalyses the final step in peptidoglycan precursor synthesis. Similarly, in the periplasm, MreC controls the spatial orientation of the penicillin binding proteins and a lytic transglycosylase. We have now found that MreB cables are required for the organization of several other cytosolic murein biosynthetic enzymes such as MraY, MurB, MurC, MurE and MurF. We also show these proteins adopt a subcellular pattern of localization comparable to MurG, suggesting the existence of cytoskeletal-dependent interactions. Through extensive two-hybrid analyses, we have now generated a comprehensive interaction map of components of the bacterial morphogenetic complex. In the cytosol, this complex contains both murein biosynthetic enzymes and morphogenetic proteins, including RodA, RodZ and MreD. We show that the integral membrane protein, MreD, is essential for lateral peptidoglycan synthesis, interacts with the precursor synthesizing enzymes MurG and MraY, and additionally, determines MreB localization. Our results suggest that the interdependent localization of MreB and MreD functions to spatially organize a complex of peptidoglycan precursor synthesis proteins, which is required for propagation of a uniform cell shape and catalytically efficient peptidoglycan synthesis.

  3. The Utility of Acute-Phase Proteins in the Assessment of Treatment Response in Dogs With Bacterial Pneumonia

    DEFF Research Database (Denmark)

    Viitanen, S. J.; Lappalainen, A. K.; Christensen, M. B.

    2017-01-01

    Background: Acute-phase proteins (APPs) are sensitive markers of inflammation, and serum C-reactive protein (CRP) recently has been shown to be a useful diagnostic marker in dogs with bacterial pneumonia (BP). In humans with community-acquired pneumonia, APPs also have great utility as follow......-up markers aiding in the assessment of treatment response. Objectives: The aim of our study was to investigate the applicability of APPs as markers of treatment response in dogs with BP. Animals: Nineteen dogs diagnosed with BP and 64 healthy dogs. Methods: The study was conducted as a prospective...... longitudinal observational study. Serum CRP, serum amyloid A (SAA), and haptoglobin concentrations were followed during a natural course of BP. Normalization of serum CRP was used to guide the duration of antibiotic treatment (treatment was stopped 5–7 days after CRP normalized) in 8 of 17 dogs surviving...

  4. Procalcitonin and C-reactive protein as markers of bacterial infection in patients with solid tumours

    DEFF Research Database (Denmark)

    Diness, Laura V; Maraldo, Maja V; Mortensen, Christiane E

    2014-01-01

    INTRODUCTION: The diagnosis of bacterial infections in patients with solid tumours can be difficult as both the tumour and its treatment can cause symptoms and signs similar to those of infections. Many patients with solid tumours therefore receive antibiotic treatment without having a bacterial......, but with no signs of infection. RESULTS: Of the 41 admitted patients, 25 were classified as having an infection (either microbiologically or radioo-gically verified). Among the 25 cases with infection, PCT was within the normal range in 11 cases and only elevated in 14. As nearly half of the patients with infection...... had PCT within the normal range, PCT is not suited to exclude an infection. CRP was elevated in 20 patients out of the 25. CONCLUSION: PCT within the normal range cannot exclude an infection and does not appear to be superior to CRP to exclude an infection in patients with solid tumours. FUNDING...

  5. Novel bacterial gas sensor proteins with transition metal-containing prosthetic groups as active sites.

    Science.gov (United States)

    Aono, Shigetoshi

    2012-04-01

    Gas molecules function as signaling molecules in many biological regulatory systems responsible for transcription, chemotaxis, and other complex physiological processes. Gas sensor proteins play a crucial role in regulating such biological systems in response to gas molecules. New sensor proteins that sense oxygen or nitric oxide have recently been found, and they have been characterized by X-ray crystallographic and/or spectroscopic analysis. It has become clear that the interaction between a prosthetic group and gas molecules triggers dynamic structural changes in the protein backbone when a gas sensor protein senses gas molecules. Gas sensor proteins employ novel mechanisms to trigger conformational changes in the presence of a gas. In gas sensor proteins that have iron-sulfur clusters as active sites, the iron-sulfur clusters undergo structural changes, which trigger a conformational change. Heme-based gas sensor proteins reconstruct hydrogen-bonding networks around the heme and heme-bound ligand. Gas sensor proteins have two functional states, on and off, which are active and inactive, respectively, for subsequent signal transduction in response to their physiological effector molecules. To fully understand the structure-function relationships of gas sensor proteins, it is vital to perform X-ray crystal structure analyses of full-length proteins in both the on and off states.

  6. Engineering bacterial surface displayed human norovirus capsid proteins: A novel system to explore interaction between norovirus and ligands

    Directory of Open Access Journals (Sweden)

    Mengya eNiu

    2015-12-01

    Full Text Available Human noroviruses (HuNoVs are major contributors to acute nonbacterial gastroenteritis outbreaks. Many aspects of HuNoVs are poorly understood due to both the current inability to culture HuNoVs, and the lack of efficient small animal models. Surrogates for HuNoVs, such as recombinant viral like particles (VLPs expressed in eukaryotic system or P particles expressed in prokaryotic system, have been used for studies in immunology and interaction between the virus and its receptors. However, it is difficult to use VLPs or P particles to collect or isolate potential ligands binding to these recombinant capsid proteins. In this study, a new strategy was used to collect HuNoVs binding ligands through the use of ice nucleation protein (INP to display recombinant capsid proteins of HuNoVs on bacterial surfaces. The viral protein-ligand complex could be easily separated by a low speed centrifugation step. This system was also used to explore interaction between recombinant capsid proteins of HuNoVs and their receptors. In this system, the VP1 capsid encoding gene (ORF2 and the protruding domain (P domain encoding gene (3’ terminal fragment of ORF2 of HuNoVs GI.1 and GII.4 were fused with 5’ terminal fragment of ice nucleation protein encoding gene (inaQn. The results demonstrated that the recombinant VP1 and P domains of HuNoVs were expressed and anchored on the surface of Escherichia coli BL21 cells after the bacteria were transformed with the corresponding plasmids. Both cell surface displayed VP1 and P domains could be recognized by HuNoVs specific antibodies and interact with the viral histo-blood group antigens receptors. In both cases, displayed P domains had better binding abilities than VP1. This new strategy of using displayed HuNoVs capsid proteins on the bacterial surface could be utilized to separate HuNoVs binding components from complex samples, to investigate interaction between the virus and its receptors, as well as to develop an

  7. Thionin-D4E1 chimeric protein protects plants against bacterial infections

    Science.gov (United States)

    Stover, Eddie W; Gupta, Goutam; Hao, Guixia

    2017-08-08

    The generation of a chimeric protein containing a first domain encoding either a pro-thionon or thionin, a second domain encoding D4E1 or pro-D4E1, and a third domain encoding a peptide linker located between the first domain and second domain is described. Either the first domain or the second domain is located at the amino terminal of the chimeric protein and the other domain (second domain or first domain, respectively) is located at the carboxyl terminal. The chimeric protein has antibacterial activity. Genetically altered plants and their progeny expressing a polynucleotide encoding the chimeric protein resist diseases caused by bacteria.

  8. Studies on the interaction of riboflavin 5'-phosphate with proteins with special attention to bacterial bioluminescence

    NARCIS (Netherlands)

    Gast, R.

    1978-01-01

    The central theme of this thesis is the interaction of FMN with proteins. For one of the proteins studied, the enzyme luciferase from bacteria, further investigations were done on the process of light emission.
    In chapter 2 and 3 studies are reported on the binding of FMN with relatively

  9. Structural and Functional Characterization of an Ancient Bacterial Transglutaminase Sheds Light on the Minimal Requirements for Protein Cross-Linking.

    Science.gov (United States)

    Fernandes, Catarina G; Plácido, Diana; Lousa, Diana; Brito, José A; Isidro, Anabela; Soares, Cláudio M; Pohl, Jan; Carrondo, Maria A; Archer, Margarida; Henriques, Adriano O

    2015-09-22

    Transglutaminases are best known for their ability to catalyze protein cross-linking reactions that impart chemical and physical resilience to cellular structures. Here, we report the crystal structure and characterization of Tgl, a transglutaminase from the bacterium Bacillus subtilis. Tgl is produced during sporulation and cross-links the surface of the highly resilient spore. Tgl-like proteins are found only in spore-forming bacteria of the Bacillus and Clostridia classes, indicating an ancient origin. Tgl is a single-domain protein, produced in active form, and the smallest transglutaminase characterized to date. We show that Tgl is structurally similar to bacterial cell wall endopeptidases and has an NlpC/P60 catalytic core, thought to represent the ancestral unit of the cysteine protease fold. We show that Tgl functions through a unique partially redundant catalytic dyad formed by Cys116 and Glu187 or Glu115. Strikingly, the catalytic Cys is insulated within a hydrophobic tunnel that traverses the molecule from side to side. The lack of similarity of Tgl to other transglutaminases together with its small size suggests that an NlpC/P60 catalytic core and insulation of the active site during catalysis may be essential requirements for protein cross-linking.

  10. Avoiding acidic region streaking in two-dimensional gel electrophoresis: case study with two bacterial whole cell protein extracts.

    Science.gov (United States)

    Roy, Arnab; Varshney, Umesh; Pal, Debnath

    2014-09-01

    Acidic region streaking (ARS) is one of the lacunae in two-dimensional gel electrophoresis (2DE) of bacterial proteome. This streaking is primarily caused by nucleic acid (NuA) contamination and poses major problem in the downstream processes like image analysis and protein identification. Although cleanup and nuclease digestion are practiced as remedial options, these strategies may incur loss in protein recovery and perform incomplete removal of NuA. As a result, ARS has remained a common observation across publications, including the recent ones. In this work, we demonstrate how ultrasound wave can be used to shear NuA in plain ice-cooled water, facilitating the elimination of ARS in the 2DE gels without the need for any additional sample cleanup tasks. In combination with a suitable buffer recipe, IEF program and frequent paper-wick changing approach, we are able to reproducibly demonstrate the production of clean 2DE gels with improved protein recovery and negligible or no ARS. We illustrate our procedure using whole cell protein extracts from two diverse organisms, Escherichia coli and Mycobacterium smegmatis. Our designed protocols are straightforward and expected to provide good 2DE gels without ARS, with comparable times and significantly lower cost.

  11. Procalcitonin and C-reactive protein cannot differentiate bacterial or viral infection in COPD exacerbation requiring emergency department visits

    Directory of Open Access Journals (Sweden)

    Chang CH

    2015-04-01

    Full Text Available Chih-Hao Chang,1 Kuo-Chien Tsao,2,3 Han-Chung Hu,1,4 Chung-Chi Huang,1,4 Kuo-Chin Kao,1,4 Ning-Hung Chen,1,4 Cheng-Ta Yang,1,4 Ying-Huang Tsai,4,5 Meng-Jer Hsieh4,51Department of Pulmonary and Critical Care Medicine, Linkou Chang-Gung Memorial Hospital, Chang-Gung Medical Foundation, Chang-Gung University College of Medicine, Taoyuan, Taiwan; 2Department of Laboratory Medicine, Linkou Chang-Gung Memorial Hospital, Chang-Gung Medical Foundation; 3Department of Medical Biotechnology and Laboratory Science, Chang Gung University, Taoyuan, Taiwan; 4Department of Respiratory Therapy, Chang-Gung University, Taoyuan, Taiwan; 5Department of Pulmonary and Critical Care Medicine, Chiayi Chang-Gung Memorial Hospital, Chang-Gung Medical Foundation, Puzi City, TaiwanBackground: Viral and bacterial infections are the most common causes of chronic obstructive pulmonary disease (COPD exacerbations. Whether serum inflammatory markers can differentiate bacterial from virus infection in patients with COPD exacerbation requiring emergency department (ED visits remains controversial.Methods: Viral culture and polymerase chain reaction (PCR were used to identify the viruses in the oropharynx of patients with COPD exacerbations. The bacteria were identified by the semiquantitative culture of the expectorated sputum. The peripheral blood white blood cell (WBC counts, serum C-reactive protein (CRP, procalcitonin (PCT, and clinical symptoms were compared among patients with different types of infections.Results: Viruses were isolated from 16 (22.2% of the 72 patients enrolled. The most commonly identified viruses were parainfluenza type 3, influenza A, and rhinovirus. A total of 30 (41.7% patients had positive bacterial cultures, with the most commonly found bacteria being Haemophilus influenzae and Haemophilus parainfluenzae. Five patients (6.9% had both positive sputum cultures and virus identification. The WBC, CRP, and PCT levels of the bacteria-positive and bacteria

  12. Changes in the protein fraction of Merluccius bilinearis muscle under lactic acid bacterial fermentation using a Lactobacillus Acidophilus starter culture (ESP

    Directory of Open Access Journals (Sweden)

    Luis J. Elizondo

    2016-03-01

    Full Text Available The effect of lactic acid bacterial fermentation on the protein fraction of Merluccius bilinearis muscle was evaluated. The non-protein fraction increased progressively with corresponding decreases in the percentage protein (dry weight indicating proteolytic activity during fermentation. Significant increases in the percentages of the amino acids cystine, isoleucine, phenylalanine and tyrosine were observed after two months of fermentation. Percentages of arginine decreased significantly after one week and again after two months of fermentation.

  13. Changes in the protein fraction of Merluccius bilinearis muscle under lactic acid bacterial fermentation using a Lactobacillus Acidophilus starter culture (ESP)

    OpenAIRE

    Elizondo, Luis J.

    2016-01-01

    The effect of lactic acid bacterial fermentation on the protein fraction of Merluccius bilinearis muscle was evaluated. The non-protein fraction increased progressively with corresponding decreases in the percentage protein (dry weight) indicating proteolytic activity during fermentation. Significant increases in the percentages of the amino acids cystine, isoleucine, phenylalanine and tyrosine were observed after two months of fermentation. Percentages of arginine decreased significantly aft...

  14. Novel Bacterial Proteins and Lipids Reveal the Diversity of Triterpenoid Biomarker Synthesis

    Science.gov (United States)

    Wei, J. H.; Banta, A. B.; Gill, C. C. C.; Giner, J. L.; Welander, P. V.

    2017-12-01

    Lipids preserved in sediments and rocks function as organic biomarkers providing evidence for the types of organisms that lived in ancient environments. We use a combined approach utilizing comparative genomics, molecular biology, and lipid analysis to discover novel cyclic triteprenoid lipids and their biosynthetic pathways in bacteria. Here, we present two cases of bacterial synthesis of pentacylic triterpenols previously thought to be indicative of eukaryotes, which address current incongruities in the fossil record. Cyclic triterpenoid lipids, such as hopanoids and sterols, are generally associated with bacteria and eukaryotes, respectively. The pentacyclic triterpenoid tetrahymanol, first discovered in the ciliate Tetrahymena pyriformis, and its diagenetic product gammacerane, have been previously interpreted as markers for eukaryotes and linked to water column stratification. Yet the occurrence of tetrahymanol in bacteria implies our knowledge of extant tetrahymanol producers is not complete. Through comparative genomics we identified a new gene required for tetrahymanol synthesis in the bacterium Methylomicrobium alcaliphilum. This gene encodes a novel enzyme, Tetrahymanol synthase (THS), that synthesizes tetrahymanol from the hopanoid diploptene demonstrating a pathway for tetrahymanol production in bacteria distinct from that in eukaryotes. We bionformatically identified THS homologs in 104 bacterial genomes and 472 metagenomes, implying a great diversity of tetrahymanol producers. Lipids of the arborane class, such as iso-arborinol, are commonly found in modern angiosperms. Arobranes are synthesized by the enzyme oxidosqualene cyclase (OSC), which in plants can form both tetra and pentacyclic molecules. While bacteria are known to produce tetracyclic sterol compounds, bacterial synthesis of pentacyclic arborane class triterpenols of this class were previously undiscovered. We have identified a bacterium, Eudoraea adriatica, whose OSC synthesizes

  15. Multi-location gram-positive and gram-negative bacterial protein subcellular localization using gene ontology and multi-label classifier ensemble.

    Science.gov (United States)

    Wang, Xiao; Zhang, Jun; Li, Guo-Zheng

    2015-01-01

    It has become a very important and full of challenge task to predict bacterial protein subcellular locations using computational methods. Although there exist a lot of prediction methods for bacterial proteins, the majority of these methods can only deal with single-location proteins. But unfortunately many multi-location proteins are located in the bacterial cells. Moreover, multi-location proteins have special biological functions capable of helping the development of new drugs. So it is necessary to develop new computational methods for accurately predicting subcellular locations of multi-location bacterial proteins. In this article, two efficient multi-label predictors, Gpos-ECC-mPLoc and Gneg-ECC-mPLoc, are developed to predict the subcellular locations of multi-label gram-positive and gram-negative bacterial proteins respectively. The two multi-label predictors construct the GO vectors by using the GO terms of homologous proteins of query proteins and then adopt a powerful multi-label ensemble classifier to make the final multi-label prediction. The two multi-label predictors have the following advantages: (1) they improve the prediction performance of multi-label proteins by taking the correlations among different labels into account; (2) they ensemble multiple CC classifiers and further generate better prediction results by ensemble learning; and (3) they construct the GO vectors by using the frequency of occurrences of GO terms in the typical homologous set instead of using 0/1 values. Experimental results show that Gpos-ECC-mPLoc and Gneg-ECC-mPLoc can efficiently predict the subcellular locations of multi-label gram-positive and gram-negative bacterial proteins respectively. Gpos-ECC-mPLoc and Gneg-ECC-mPLoc can efficiently improve prediction accuracy of subcellular localization of multi-location gram-positive and gram-negative bacterial proteins respectively. The online web servers for Gpos-ECC-mPLoc and Gneg-ECC-mPLoc predictors are freely accessible

  16. Ribosomal proteins as biomarkers for bacterial identification by mass spectrometry in the clinical microbiology laboratory.

    Science.gov (United States)

    Suarez, Stéphanie; Ferroni, Agnès; Lotz, Aurélie; Jolley, Keith A; Guérin, Philippe; Leto, Julie; Dauphin, Brunhilde; Jamet, Anne; Maiden, Martin C J; Nassif, Xavier; Armengaud, Jean

    2013-09-01

    Whole-cell matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) is a rapid method for identification of microorganisms that is increasingly used in microbiology laboratories. This identification is based on the comparison of the tested isolate mass spectrum with reference databases. Using Neisseria meningitidis as a model organism, we showed that in one of the available databases, the Andromas database, 10 of the 13 species-specific biomarkers correspond to ribosomal proteins. Remarkably, one biomarker, ribosomal protein L32, was subject to inter-strain variability. The analysis of the ribosomal protein patterns of 100 isolates for which whole genome sequences were available, confirmed the presence of inter-strain variability in the molecular weight of 29 ribosomal proteins, thus establishing a correlation between the sequence type (ST) and/or clonal complex (CC) of each strain and its ribosomal protein pattern. Since the molecular weight of three of the variable ribosomal proteins (L30, L31 and L32) was included in the spectral window observed by MALDI-TOF MS in clinical microbiology, i.e., 3640-12000 m/z, we were able by analyzing the molecular weight of these three ribosomal proteins to classify each strain in one of six subgroups, each of these subgroups corresponding to specific STs and/or CCs. Their detection by MALDI-TOF allows therefore a quick typing of N. meningitidis isolates. © 2013 Elsevier B.V. All rights reserved.

  17. Intracellular Protein Delivery System Using a Target-Specific Repebody and Translocation Domain of Bacterial Exotoxin.

    Science.gov (United States)

    Kim, Hee-Yeon; Kang, Jung Ae; Ryou, Jeong-Hyun; Lee, Gyeong Hee; Choi, Dae Seong; Lee, Dong Eun; Kim, Hak-Sung

    2017-11-17

    With the high efficacy of protein-based therapeutics and plenty of intracellular drug targets, cytosolic protein delivery in a cell-specific manner has attracted considerable attention in the field of precision medicine. Herein, we present an intracellular protein delivery system based on a target-specific repebody and the translocation domain of Pseudomonas aeruginosa exotoxin A. The delivery platform was constructed by genetically fusing an EGFR-specific repebody as a targeting moiety to the translocation domain, while a protein cargo was fused to the C-terminal end of the delivery platform. The delivery platform was revealed to efficiently translocate a protein cargo to the cytosol in a target-specific manner. We demonstrate the utility and potential of the delivery platform by showing a remarkable tumor regression with negligible toxicity in a xenograft mice model when gelonin was used as the cytotoxic protein cargo. The present platform can find wide applications to the cell-selective cytosolic delivery of diverse proteins in many areas.

  18. GTPase activity, structure, and mechanical properties of filaments assembled from bacterial cytoskeleton protein MreB.

    Science.gov (United States)

    Esue, Osigwe; Wirtz, Denis; Tseng, Yiider

    2006-02-01

    MreB, a major component of the recently discovered bacterial cytoskeleton, displays a structure homologous to its eukaryotic counterpart actin. Here, we study the assembly and mechanical properties of Thermotoga maritima MreB in the presence of different nucleotides in vitro. We found that GTP, not ADP or GDP, can mediate MreB assembly into filamentous structures as effectively as ATP. Upon MreB assembly, both GTP and ATP release the gamma phosphate at similar rates. Therefore, MreB is an equally effective ATPase and GTPase. Electron microscopy and quantitative rheology suggest that the morphologies and micromechanical properties of filamentous ATP-MreB and GTP-MreB are similar. In contrast, mammalian actin assembly is favored in the presence of ATP over GTP. These results indicate that, despite high structural homology of their monomers, T. maritima MreB and actin filaments display different assembly, morphology, micromechanics, and nucleotide-binding specificity. Furthermore, the biophysical properties of T. maritima MreB filaments, including high rigidity and propensity to form bundles, suggest a mechanism by which MreB helical structure may be involved in imposing a cylindrical architecture on rod-shaped bacterial cells.

  19. Phagocytosis escape by a Staphylococcus aureus protein that connects complement and coagulation proteins at the bacterial surface.

    Directory of Open Access Journals (Sweden)

    Ya-Ping Ko

    Full Text Available Upon contact with human plasma, bacteria are rapidly recognized by the complement system that labels their surface for uptake and clearance by phagocytic cells. Staphylococcus aureus secretes the 16 kD Extracellular fibrinogen binding protein (Efb that binds two different plasma proteins using separate domains: the Efb N-terminus binds to fibrinogen, while the C-terminus binds complement C3. In this study, we show that Efb blocks phagocytosis of S. aureus by human neutrophils. In vitro, we demonstrate that Efb blocks phagocytosis in plasma and in human whole blood. Using a mouse peritonitis model we show that Efb effectively blocks phagocytosis in vivo, either as a purified protein or when produced endogenously by S. aureus. Mutational analysis revealed that Efb requires both its fibrinogen and complement binding residues for phagocytic escape. Using confocal and transmission electron microscopy we show that Efb attracts fibrinogen to the surface of complement-labeled S. aureus generating a 'capsule'-like shield. This thick layer of fibrinogen shields both surface-bound C3b and antibodies from recognition by phagocytic receptors. This information is critical for future vaccination attempts, since opsonizing antibodies may not function in the presence of Efb. Altogether we discover that Efb from S. aureus uniquely escapes phagocytosis by forming a bridge between a complement and coagulation protein.

  20. Biosensing for the Environment and Defence: Aqueous Uranyl Detection Using Bacterial Surface Layer Proteins

    Directory of Open Access Journals (Sweden)

    David J.R. Conroy

    2010-05-01

    Full Text Available The fabrication of novel uranyl (UO22+ binding protein based sensors is reported. The new biosensor responds to picomolar levels of aqueous uranyl ions within minutes using Lysinibacillus sphaericus JG-A12 S-layer protein tethered to gold electrodes. In comparison to traditional self assembled monolayer based biosensors the porous bioconjugated layer gave greater stability, longer electrode life span and a denser protein layer. Biosensors responded specifically to UO22+ ions and showed minor interference from Ni2+, Cs+, Cd2+ and Co2+. Chemical modification of JG-A12 protein phosphate and carboxyl groups prevented UO22+ binding, showing that both moieties are involved in the recognition to UO22+.

  1. Bacterial Genome Editing Strategy for Control of Transcription and Protein Stability

    DEFF Research Database (Denmark)

    Lauritsen, Ida; Martinez, Virginia; Ronda, Carlotta

    2018-01-01

    In molecular biology and cell factory engineering, tools that enable control of protein production and stability are highly important. Here, we describe protocols for tagging genes in Escherichia coli allowing for inducible degradation and transcriptional control of any soluble protein of interest....... The underlying molecular biology is based on the two cross-kingdom tools CRISPRi and the N-end rule for protein degradation. Genome editing is performed with the CRMAGE technology and randomization of the translational initiation region minimizes the polar effects of tag insertion. The approach has previously...... been applied for targeting proteins originating from essential operon-located genes and has potential to serve as a universal synthetic biology tool....

  2. The primary structure of rat liver ribosomal protein L37. Homology with yeast and bacterial ribosomal proteins.

    Science.gov (United States)

    Lin, A; McNally, J; Wool, I G

    1983-09-10

    The covalent structure of the rat liver 60 S ribosomal subunit protein L37 was determined. Twenty-four tryptic peptides were purified and the sequence of each was established; they accounted for all 111 residues of L37. The sequence of the first 30 residues of L37, obtained previously by automated Edman degradation of the intact protein, provided the alignment of the first 9 tryptic peptides. Three peptides (CN1, CN2, and CN3) were produced by cleavage of protein L37 with cyanogen bromide. The sequence of CN1 (65 residues) was established from the sequence of secondary peptides resulting from cleavage with trypsin and chymotrypsin. The sequence of CN1 in turn served to order tryptic peptides 1 through 14. The sequence of CN2 (15 residues) was determined entirely by a micromanual procedure and allowed the alignment of tryptic peptides 14 through 18. The sequence of the NH2-terminal 28 amino acids of CN3 (31 residues) was determined; in addition the complete sequences of the secondary tryptic and chymotryptic peptides were done. The sequence of CN3 provided the order of tryptic peptides 18 through 24. Thus the sequence of the three cyanogen bromide peptides also accounted for the 111 residues of protein L37. The carboxyl-terminal amino acids were identified after carboxypeptidase A treatment. There is a disulfide bridge between half-cystinyl residues at positions 40 and 69. Rat liver ribosomal protein L37 is homologous with yeast YP55 and with Escherichia coli L34. Moreover, there is a segment of 17 residues in rat L37 that occurs, albeit with modifications, in yeast YP55 and in E. coli S4, L20, and L34.

  3. Recognition and cleavage of 5-methylcytosine DNA by bacterial SRA-HNH proteins

    OpenAIRE

    Han, Tiesheng; Yamada-Mabuchi, Megumu; Zhao, Gong; Li, Li; liu, Guang; Ou, Hong-Yu; Deng, Zixin; Zheng, Yu; He, Xinyi

    2015-01-01

    SET and RING-finger-associated (SRA) domain is involved in establishment and maintenance of DNA methylation in eukaryotes. Proteins containing SRA domains exist in mammals, plants, even microorganisms. It has been established that mammalian SRA domain recognizes 5-methylcytosine (5mC) through a base-flipping mechanism. Here, we identified and characterized two SRA domain-containing proteins with the common domain architecture of N-terminal SRA domain and C-terminal HNH nuclease domain, Sco533...

  4. Bacterial infections in patients with liver cirrhosis: clinical characteristics and the role of C-reactive protein

    Science.gov (United States)

    Deutsch, Melanie; Manolakopoulos, Spilios; Andreadis, Ioannis; Giannaris, Markos; Kontos, George; Kranidioti, Hariklia; Pirounaki, Maria; Koskinas, John

    2018-01-01

    Background: The diagnosis of bacterial infection in cirrhotic patients may be difficult, because of the absence of classical signs such as fever and raised white blood cell count. The role of C-reactive protein (CRP) in this context has not been clearly defined. Methods: Clinical and laboratory characteristics of 210 consecutive cirrhotic patients with (n=100) or without (n=110) bacterial infection were compared with a control group of non-cirrhotic patients with infection (n=106). Results: Significantly fewer patients with cirrhosis had a body temperature ≥37°C when presenting with bacterial infection (56% cirrhotic vs. 85.5% non-cirrhotic patients, P=0.01). Mean leukocyte count was 6.92 × 103/mm3 in patients with cirrhosis and infection, 5.75 × 103/mm3 (P=0.02) in cirrhotic patients without infection, and 11.28 × 103/mm3 in non-cirrhotic patients with infection (P10 mg/L indicated the presence of infection with a sensitivity of 68%, a specificity of 84.5% and an area under the receiver operating characteristic curve of 0.8197. CRP cutoff level differed according to the severity of the liver disease: Child-Pugh score (CPS) A: 21.3 mg/L, B: 17 mg/L, and C: 5.78 mg/L. Conclusions: CRP at admission could help diagnose infection in cirrhotic patients. Since the severity of liver disease seems to affect the CRP values, lower CRP levels might indicate infection. Clinical suspicion is necessary to avoid delay in diagnosis and initiate antibiotic treatment. PMID:29333070

  5. Bacterial infections in patients with liver cirrhosis: clinical characteristics and the role of C-reactive protein.

    Science.gov (United States)

    Deutsch, Melanie; Manolakopoulos, Spilios; Andreadis, Ioannis; Giannaris, Markos; Kontos, George; Kranidioti, Hariklia; Pirounaki, Maria; Koskinas, John

    2018-01-01

    The diagnosis of bacterial infection in cirrhotic patients may be difficult, because of the absence of classical signs such as fever and raised white blood cell count. The role of C-reactive protein (CRP) in this context has not been clearly defined. Clinical and laboratory characteristics of 210 consecutive cirrhotic patients with (n=100) or without (n=110) bacterial infection were compared with a control group of non-cirrhotic patients with infection (n=106). Significantly fewer patients with cirrhosis had a body temperature ≥37°C when presenting with bacterial infection (56% cirrhotic vs. 85.5% non-cirrhotic patients, P=0.01). Mean leukocyte count was 6.92 × 10 3 /mm 3 in patients with cirrhosis and infection, 5.75 × 10 3 /mm 3 (P=0.02) in cirrhotic patients without infection, and 11.28 × 10 3 /mm 3 in non-cirrhotic patients with infection (P10 mg/L indicated the presence of infection with a sensitivity of 68%, a specificity of 84.5% and an area under the receiver operating characteristic curve of 0.8197. CRP cutoff level differed according to the severity of the liver disease: Child-Pugh score (CPS) A: 21.3 mg/L, B: 17 mg/L, and C: 5.78 mg/L. CRP at admission could help diagnose infection in cirrhotic patients. Since the severity of liver disease seems to affect the CRP values, lower CRP levels might indicate infection. Clinical suspicion is necessary to avoid delay in diagnosis and initiate antibiotic treatment.

  6. Bacterial periplasmic sialic acid-binding proteins exhibit a conserved binding site

    Energy Technology Data Exchange (ETDEWEB)

    Gangi Setty, Thanuja [Institute for Stem Cell Biology and Regenerative Medicine, NCBS Campus, GKVK Post, Bangalore, Karnataka 560 065 (India); Cho, Christine [Carver College of Medicine, University of Iowa, Iowa City, IA 52242-1109 (United States); Govindappa, Sowmya [Institute for Stem Cell Biology and Regenerative Medicine, NCBS Campus, GKVK Post, Bangalore, Karnataka 560 065 (India); Apicella, Michael A. [Carver College of Medicine, University of Iowa, Iowa City, IA 52242-1109 (United States); Ramaswamy, S., E-mail: ramas@instem.res.in [Institute for Stem Cell Biology and Regenerative Medicine, NCBS Campus, GKVK Post, Bangalore, Karnataka 560 065 (India)

    2014-07-01

    Structure–function studies of sialic acid-binding proteins from F. nucleatum, P. multocida, V. cholerae and H. influenzae reveal a conserved network of hydrogen bonds involved in conformational change on ligand binding. Sialic acids are a family of related nine-carbon sugar acids that play important roles in both eukaryotes and prokaryotes. These sialic acids are incorporated/decorated onto lipooligosaccharides as terminal sugars in multiple bacteria to evade the host immune system. Many pathogenic bacteria scavenge sialic acids from their host and use them for molecular mimicry. The first step of this process is the transport of sialic acid to the cytoplasm, which often takes place using a tripartite ATP-independent transport system consisting of a periplasmic binding protein and a membrane transporter. In this paper, the structural characterization of periplasmic binding proteins from the pathogenic bacteria Fusobacterium nucleatum, Pasteurella multocida and Vibrio cholerae and their thermodynamic characterization are reported. The binding affinities of several mutations in the Neu5Ac binding site of the Haemophilus influenzae protein are also reported. The structure and the thermodynamics of the binding of sugars suggest that all of these proteins have a very well conserved binding pocket and similar binding affinities. A significant conformational change occurs when these proteins bind the sugar. While the C1 carboxylate has been identified as the primary binding site, a second conserved hydrogen-bonding network is involved in the initiation and stabilization of the conformational states.

  7. Bacterial periplasmic sialic acid-binding proteins exhibit a conserved binding site

    International Nuclear Information System (INIS)

    Gangi Setty, Thanuja; Cho, Christine; Govindappa, Sowmya; Apicella, Michael A.; Ramaswamy, S.

    2014-01-01

    Structure–function studies of sialic acid-binding proteins from F. nucleatum, P. multocida, V. cholerae and H. influenzae reveal a conserved network of hydrogen bonds involved in conformational change on ligand binding. Sialic acids are a family of related nine-carbon sugar acids that play important roles in both eukaryotes and prokaryotes. These sialic acids are incorporated/decorated onto lipooligosaccharides as terminal sugars in multiple bacteria to evade the host immune system. Many pathogenic bacteria scavenge sialic acids from their host and use them for molecular mimicry. The first step of this process is the transport of sialic acid to the cytoplasm, which often takes place using a tripartite ATP-independent transport system consisting of a periplasmic binding protein and a membrane transporter. In this paper, the structural characterization of periplasmic binding proteins from the pathogenic bacteria Fusobacterium nucleatum, Pasteurella multocida and Vibrio cholerae and their thermodynamic characterization are reported. The binding affinities of several mutations in the Neu5Ac binding site of the Haemophilus influenzae protein are also reported. The structure and the thermodynamics of the binding of sugars suggest that all of these proteins have a very well conserved binding pocket and similar binding affinities. A significant conformational change occurs when these proteins bind the sugar. While the C1 carboxylate has been identified as the primary binding site, a second conserved hydrogen-bonding network is involved in the initiation and stabilization of the conformational states

  8. Two-step membrane binding by the bacterial SRP receptor enable efficient and accurate Co-translational protein targeting.

    Science.gov (United States)

    Hwang Fu, Yu-Hsien; Huang, William Y C; Shen, Kuang; Groves, Jay T; Miller, Thomas; Shan, Shu-Ou

    2017-07-28

    The signal recognition particle (SRP) delivers ~30% of the proteome to the eukaryotic endoplasmic reticulum, or the bacterial plasma membrane. The precise mechanism by which the bacterial SRP receptor, FtsY, interacts with and is regulated at the target membrane remain unclear. Here, quantitative analysis of FtsY-lipid interactions at single-molecule resolution revealed a two-step mechanism in which FtsY initially contacts membrane via a Dynamic mode, followed by an SRP-induced conformational transition to a Stable mode that activates FtsY for downstream steps. Importantly, mutational analyses revealed extensive auto-inhibitory mechanisms that prevent free FtsY from engaging membrane in the Stable mode; an engineered FtsY pre-organized into the Stable mode led to indiscriminate targeting in vitro and disrupted FtsY function in vivo. Our results show that the two-step lipid-binding mechanism uncouples the membrane association of FtsY from its conformational activation, thus optimizing the balance between the efficiency and fidelity of co-translational protein targeting.

  9. Procalcitonin, C-reactive protein and serum lactate dehydrogenase in the diagnosis of bacterial sepsis, SIRS and systemic candidiasis.

    Science.gov (United States)

    Miglietta, Fabio; Faneschi, Maria Letizia; Lobreglio, Giambattista; Palumbo, Claudio; Rizzo, Adriana; Cucurachi, Marco; Portaccio, Gerolamo; Guerra, Francesco; Pizzolante, Maria

    2015-09-01

    The aim of this study was to evaluate procalcitonin (PCT), C-reactive protein (CRP), platelet count (PLT) and serum lactate dehydrogenase (LDH) as early markers for diagnosis of SIRS, bacterial sepsis and systemic candidiasis in intensive care unit (ICU) patients. Based on blood culture results, the patients were divided into a sepsis group (70 patients), a SIRS group (42 patients) and a systemic candidiasis group (33 patients). PCT, CRP, LDH and PLT levels were measured on day 0 and on day 2 from the sepsis symptom onset. PCT levels were higher in Gram negative sepsis than those in Gram positive sepsis, although the P value between the two subgroups is not significant (P=0.095). Bacterial sepsis group had higher PCT and CRP levels compared with the systemic candidiasis group, whereas PLT and LDH levels showed similar levels in these two subgroups. The AUC for PCT (AUC: 0.892, P candidiasis groups (P=0.093 N.S.). In conclusion, PCT can be used as a preliminary marker in the event of clinical suspicion of systemic candidiasis; however, low PCT levels (candidiasis and SIRS groups.

  10. Purification of CD47-streptavidin fusion protein from bacterial lysate using biotin-agarose affinity chromatography.

    Science.gov (United States)

    Salehi, Nasrin; Peng, Ching-An

    2016-07-08

    CD47 is a widely expressed transmembrane glycoprotein that modulates the activity of a plethora of immune cells via its extracellular domain. Therefore, CD47 plays important roles in the regulation of immune responses and may serve as targets for the development of immunotherapeutic agents. To make sure CD47 functionality is intact under the process of protein conjugation, CD47-streptavidin fusion protein was expressed and purified because it can easily bind to biotin-tagged materials via the unique biotin-streptavidin affinity. In this study, gene sequences of CD47 extracellular domain (CD47ECD) and core streptavidin (coreSA) with a total 834 bp were inserted into pET20b plasmid to construct recombinant plasmid encoding CD47-SA fusion gene. After bacteria transformation, the CD47-SA fusion protein was expressed by isopropyl-β-d-thiogalactopyranoside (IPTG) induction. The collected bacteria lysate was loaded on biotinylated agarose to proceed the purification of CD47-SA fusion protein. Due to the unexpected high affinity between biotin and coreSA, standard washing and elution approaches (e.g., varying pH, using biotin, and applying guanidine hydrochloride) reported for biotin-streptavidin affinity chromatography were not able to separate the target fusion protein. Instead, using low concentration of the non-ionic detergent Triton X-100 followed with alkaline buffer could efficiently weaken the binding between biotin and coreSA, thereby eluting out CD47-SA fusion protein from the biotin agarose column. The purified CD47-SA fusion protein was further characterized by molecular biology methods and its antiphagocytic functionality was confirmed by the phagocytosis assay. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:949-958, 2016. © 2016 American Institute of Chemical Engineers.

  11. Functional characterization of Kaposi's sarcoma-associated herpesvirus small capsid protein by bacterial artificial chromosome-based mutagenesis

    International Nuclear Information System (INIS)

    Sathish, Narayanan; Yuan Yan

    2010-01-01

    A systematic investigation of interactions amongst KSHV capsid proteins was undertaken in this study to comprehend lesser known KSHV capsid assembly mechanisms. Interestingly the interaction patterns of the KSHV small capsid protein, ORF65 suggested its plausible role in viral capsid assembly pathways. Towards further understanding this, ORF65-null recombinant mutants (BAC-Δ65 and BAC-stop65) employing a bacterial artificial chromosome (BAC) system were generated. No significant difference was found in both overall viral gene expression and lytic DNA replication between stable monolayers of 293T-BAC36 (wild-type) and 293T-BAC-ORF65-null upon induction with 12-O-tetradecanoylphorbol-13-acetate, though the latter released 30-fold fewer virions to the medium than 293T-BAC36 cells. Sedimentation profiles of capsid proteins of ORF65-null recombinant mutants were non-reflective of their organization into the KSHV capsids and were also undetectable in cytoplasmic extracts compared to noticeable levels in nuclear extracts. These observations collectively suggested the pivotal role of ORF65 in the KSHV capsid assembly processes.

  12. HupB Is a Bacterial Nucleoid-Associated Protein with an Indispensable Eukaryotic-Like Tail

    Directory of Open Access Journals (Sweden)

    Joanna Hołówka

    2017-11-01

    Full Text Available In bacteria, chromosomal DNA must be efficiently compacted to fit inside the small cell compartment while remaining available for the proteins involved in replication, segregation, and transcription. Among the nucleoid-associated proteins (NAPs responsible for maintaining this highly organized and yet dynamic chromosome structure, the HU protein is one of the most conserved and highly abundant. HupB, a homologue of HU, was recently identified in mycobacteria. This intriguing mycobacterial NAP is composed of two domains: an N-terminal domain that resembles bacterial HU, and a long and distinctive C-terminal domain that contains several PAKK/KAAK motifs, which are characteristic of the H1/H5 family of eukaryotic histones. In this study, we analyzed the in vivo binding of HupB on the chromosome scale. By using PALM (photoactivated localization microscopy and ChIP-Seq (chromatin immunoprecipitation followed by deep sequencing, we observed that the C-terminal domain is indispensable for the association of HupB with the nucleoid. Strikingly, the in vivo binding of HupB displayed a bias from the origin (oriC to the terminus (ter of the mycobacterial chromosome (numbers of binding sites decreased toward ter. We hypothesized that this binding mode reflects a role for HupB in organizing newly replicated oriC regions. Thus, HupB may be involved in coordinating replication with chromosome segregation.

  13. Proteomic analysis of growth phase-dependent expression of Legionella pneumophila proteins which involves regulation of bacterial virulence traits.

    Directory of Open Access Journals (Sweden)

    Tsuyoshi Hayashi

    Full Text Available Legionella pneumophila, which is a causative pathogen of Legionnaires' disease, expresses its virulent traits in response to growth conditions. In particular, it is known to become virulent at a post-exponential phase in vitro culture. In this study, we performed a proteomic analysis of differences in expression between the exponential phase and post-exponential phase to identify candidates associated with L. pneumophila virulence using 2-Dimentional Fluorescence Difference Gel Electrophoresis (2D-DIGE combined with Matrix-Assisted Laser Desorption/Ionization-Mass Spectrometry (MALDI-TOF-MS. Of 68 identified proteins that significantly differed in expression between the two growth phases, 64 were up-regulated at a post-exponential phase. The up-regulated proteins included enzymes related to glycolysis, ketone body biogenesis and poly-3-hydroxybutyrate (PHB biogenesis, suggesting that L. pneumophila may utilize sugars and lipids as energy sources, when amino acids become scarce. Proteins related to motility (flagella components and twitching motility-associated proteins were also up-regulated, predicting that they enhance infectivity of the bacteria in host cells under certain conditions. Furthermore, 9 up-regulated proteins of unknown function were found. Two of them were identified as novel bacterial factors associated with hemolysis of sheep red blood cells (SRBCs. Another 2 were found to be translocated into macrophages via the Icm/Dot type IV secretion apparatus as effector candidates in a reporter assay with Bordetella pertussis adenylate cyclase. The study will be helpful for virulent analysis of L. pneumophila from the viewpoint of physiological or metabolic modulation dependent on growth phase.

  14. BacHbpred: Support Vector Machine Methods for the Prediction of Bacterial Hemoglobin-Like Proteins

    Directory of Open Access Journals (Sweden)

    MuthuKrishnan Selvaraj

    2016-01-01

    Full Text Available The recent upsurge in microbial genome data has revealed that hemoglobin-like (HbL proteins may be widely distributed among bacteria and that some organisms may carry more than one HbL encoding gene. However, the discovery of HbL proteins has been limited to a small number of bacteria only. This study describes the prediction of HbL proteins and their domain classification using a machine learning approach. Support vector machine (SVM models were developed for predicting HbL proteins based upon amino acid composition (AC, dipeptide composition (DC, hybrid method (AC + DC, and position specific scoring matrix (PSSM. In addition, we introduce for the first time a new prediction method based on max to min amino acid residue (MM profiles. The average accuracy, standard deviation (SD, false positive rate (FPR, confusion matrix, and receiver operating characteristic (ROC were analyzed. We also compared the performance of our proposed models in homology detection databases. The performance of the different approaches was estimated using fivefold cross-validation techniques. Prediction accuracy was further investigated through confusion matrix and ROC curve analysis. All experimental results indicate that the proposed BacHbpred can be a perspective predictor for determination of HbL related proteins. BacHbpred, a web tool, has been developed for HbL prediction.

  15. A peptide export-import control circuit modulating bacterial development regulates protein phosphatases of the phosphorelay.

    Science.gov (United States)

    Perego, M

    1997-08-05

    The phosphorelay signal transduction system activates developmental transcription in sporulation of Bacillus subtilis by phosphorylation of aspartyl residues of the Spo0F and Spo0A response regulators. The phosphorylation level of these response regulators is determined by the opposing activities of protein kinases and protein aspartate phosphatases that interpret positive and negative signals for development in a signal integration circuit. The RapA protein aspartate phosphatase of the phosphorelay is regulated by a peptide that directly inhibits its activity. This peptide is proteolytically processed from an inactive pre-inhibitor protein encoded in the phrA gene. The pre-inhibitor is cleaved by the protein export apparatus to a putative pro-inhibitor that is further processed to the active inhibitor peptide and internalized by the oligopeptide permease. This export-import circuit is postulated to be a mechanism for timing phosphatase activity where the processing enzymes regulate the rate of formation of the active inhibitor. The processing events may, in turn, be controlled by a regulatory hierarchy. Chromosome sequencing has revealed several other phosphatase-prepeptide gene pairs in B. subtilis, suggesting that the use of this mechanism may be widespread in signal transduction.

  16. Characterization of Enzymatic Activity of MlrB and MlrC Proteins Involved in Bacterial Degradation of Cyanotoxins Microcystins.

    Science.gov (United States)

    Dziga, Dariusz; Zielinska, Gabriela; Wladyka, Benedykt; Bochenska, Oliwia; Maksylewicz, Anna; Strzalka, Wojciech; Meriluoto, Jussi

    2016-03-16

    Bacterial degradation of toxic microcystins produced by cyanobacteria is a common phenomenon. However, our understanding of the mechanisms of these processes is rudimentary. In this paper several novel discoveries regarding the action of the enzymes of the mlr cluster responsible for microcystin biodegradation are presented using recombinant proteins. In particular, the predicted active sites of the recombinant MlrB and MlrC were analyzed using functional enzymes and their inactive muteins. A new degradation intermediate, a hexapeptide derived from linearized microcystins by MlrC, was discovered. Furthermore, the involvement of MlrA and MlrB in further degradation of the hexapeptides was confirmed and a corrected biochemical pathway of microcystin biodegradation has been proposed.

  17. Characterization of Enzymatic Activity of MlrB and MlrC Proteins Involved in Bacterial Degradation of Cyanotoxins Microcystins

    Directory of Open Access Journals (Sweden)

    Dariusz Dziga

    2016-03-01

    Full Text Available Bacterial degradation of toxic microcystins produced by cyanobacteria is a common phenomenon. However, our understanding of the mechanisms of these processes is rudimentary. In this paper several novel discoveries regarding the action of the enzymes of the mlr cluster responsible for microcystin biodegradation are presented using recombinant proteins. In particular, the predicted active sites of the recombinant MlrB and MlrC were analyzed using functional enzymes and their inactive muteins. A new degradation intermediate, a hexapeptide derived from linearized microcystins by MlrC, was discovered. Furthermore, the involvement of MlrA and MlrB in further degradation of the hexapeptides was confirmed and a corrected biochemical pathway of microcystin biodegradation has been proposed.

  18. Bacterial mitosis: Partitioning protein ParA oscillates in spiral-shaped structures and positions plasmids at mid-cell

    DEFF Research Database (Denmark)

    Ebersbach, G.; Gerdes, Kenn

    2004-01-01

    The par2 locus of Escherichia coli plasmid pB171 encodes oscillating ATPase ParA, DNA binding protein ParB and two cis-acting DNA regions to which ParB binds (parC1 and parC2). Three independent techniques were used to investigate the subcellular localization of plasmids carrying par2. In cells......A-GFP oscillated in spiral-shaped structures. Amino acid substitutions in ParA simultaneously abolished ParA spiral formation, oscillation and either plasmid localization or plasmid separation at mid-cell. Therefore, our results suggest that ParA spirals position plasmids at the middle of the bacterial nucleoid...

  19. TARSyn: Tunable Antibiotic Resistance Devices Enabling Bacterial Synthetic Evolution and Protein Production

    DEFF Research Database (Denmark)

    Rennig, Maja; Martinez, Virginia; Mirzadeh, Kiavash

    2018-01-01

    Evolution can be harnessed to optimize synthetic biology designs. A prominent example is recombinant protein production-a dominating theme in biotechnology for more than three decades. Typically, a protein coding sequence (cds) is recombined with genetic elements, such as promoters, ribosome...... and allows expression levels in large clone libraries to be probed using a simple cell survival assay on the respective antibiotic. The power of the approach is demonstrated by substantially increasing production of two commercially interesting proteins, a Nanobody and an Affibody. The method is a simple......-level expression-an example of synthetic evolution. However, manual screening limits the ability to assay expression levels of all putative sequences in the libraries. Here we have solved this bottleneck by designing a collection of translational coupling devices based on a RNA secondary structure. Exchange...

  20. Activation of phagocytic cells by Staphylococcus epidermidis biofilms: effects of extracellular matrix proteins and the bacterial stress protein GroEL on netosis and MRP-14 release.

    Science.gov (United States)

    Dapunt, Ulrike; Gaida, Matthias M; Meyle, Eva; Prior, Birgit; Hänsch, Gertrud M

    2016-07-01

    The recognition and phagocytosis of free-swimming (planktonic) bacteria by polymorphonuclear neutrophils have been investigated in depth. However, less is known about the neutrophil response towards bacterial biofilms. Our previous work demonstrated that neutrophils recognize activating entities within the extracellular polymeric substance (EPS) of biofilms (the bacterial heat shock protein GroEL) and that this process does not require opsonization. Aim of this study was to evaluate the release of DNA by neutrophils in response to biofilms, as well as the release of the inflammatory cytokine MRP-14. Neutrophils were stimulated with Staphylococcus epidermidis biofilms, planktonic bacteria, extracted EPS and GroEL. Release of DNA and of MRP-14 was evaluated. Furthermore, tissue samples from patients suffering from biofilm infections were collected and evaluated by histology. MRP-14 concentration in blood samples was measured. We were able to show that biofilms, the EPS and GroEL induce DNA release. MRP-14 was only released after stimulation with EPS, not GroEL. Histology of tissue samples revealed MRP-14 positive cells in association with neutrophil infiltration and MRP-14 concentration was elevated in blood samples of patients suffering from biofilm infections. Our data demonstrate that neutrophil-activating entities are present in the EPS and that GroEL induces DNA release by neutrophils. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  1. In-Culture Cross-Linking of Bacterial Cells Reveals Large-Scale Dynamic Protein-Protein Interactions at the Peptide Level.

    Science.gov (United States)

    de Jong, Luitzen; de Koning, Edward A; Roseboom, Winfried; Buncherd, Hansuk; Wanner, Martin J; Dapic, Irena; Jansen, Petra J; van Maarseveen, Jan H; Corthals, Garry L; Lewis, Peter J; Hamoen, Leendert W; de Koster, Chris G

    2017-07-07

    Identification of dynamic protein-protein interactions at the peptide level on a proteomic scale is a challenging approach that is still in its infancy. We have developed a system to cross-link cells directly in culture with the special lysine cross-linker bis(succinimidyl)-3-azidomethyl-glutarate (BAMG). We used the Gram-positive model bacterium Bacillus subtilis as an exemplar system. Within 5 min extensive intracellular cross-linking was detected, while intracellular cross-linking in a Gram-negative species, Escherichia coli, was still undetectable after 30 min, in agreement with the low permeability in this organism for lipophilic compounds like BAMG. We were able to identify 82 unique interprotein cross-linked peptides with cross-links occur in assemblies involved in transcription and translation. Several of these interactions are new, and we identified a binding site between the δ and β' subunit of RNA polymerase close to the downstream DNA channel, providing a clue into how δ might regulate promoter selectivity and promote RNA polymerase recycling. Our methodology opens new avenues to investigate the functional dynamic organization of complex protein assemblies involved in bacterial growth. Data are available via ProteomeXchange with identifier PXD006287.

  2. The T4 Phage DNA Mimic Protein Arn Inhibits the DNA Binding Activity of the Bacterial Histone-like Protein H-NS*

    Science.gov (United States)

    Ho, Chun-Han; Wang, Hao-Ching; Ko, Tzu-Ping; Chang, Yuan-Chih; Wang, Andrew H.-J.

    2014-01-01

    The T4 phage protein Arn (Anti restriction nuclease) was identified as an inhibitor of the restriction enzyme McrBC. However, until now its molecular mechanism remained unclear. In the present study we used structural approaches to investigate biological properties of Arn. A structural analysis of Arn revealed that its shape and negative charge distribution are similar to dsDNA, suggesting that this protein could act as a DNA mimic. In a subsequent proteomic analysis, we found that the bacterial histone-like protein H-NS interacts with Arn, implying a new function. An electrophoretic mobility shift assay showed that Arn prevents H-NS from binding to the Escherichia coli hns and T4 p8.1 promoters. In vitro gene expression and electron microscopy analyses also indicated that Arn counteracts the gene-silencing effect of H-NS on a reporter gene. Because McrBC and H-NS both participate in the host defense system, our findings suggest that T4 Arn might knock down these mechanisms using its DNA mimicking properties. PMID:25118281

  3. Role of bacterial virulence proteins in Agrobacterium-mediated transformation of Aspergillus awamori

    NARCIS (Netherlands)

    Michielse, C.B.; Ram, A.F.J.; Hooykaas, P.J.J.; Hondel, C.A.M.J.J. van den

    2004-01-01

    The Agrobacterium-mediated transformation of Aspergillus awamori was optimized using defined co-cultivation conditions, which resulted in a reproducible and efficient transformation system. Optimal co-cultivation conditions were used to study the role of Agrobacterium tumefaciens virulence proteins

  4. Crosslinking of a Peritrophic Matrix Protein Protects Gut Epithelia from Bacterial Exotoxins

    Science.gov (United States)

    Shibata, Toshio; Maki, Kouki; Hadano, Jinki; Fujikawa, Takumi; Kitazaki, Kazuki; Koshiba, Takumi; Kawabata, Shun-ichiro

    2015-01-01

    Transglutaminase (TG) catalyzes protein-protein crosslinking, which has important and diverse roles in vertebrates and invertebrates. Here we demonstrate that Drosophila TG crosslinks drosocrystallin, a peritrophic matrix protein, to form a stable fiber structure on the gut peritrophic matrix. RNA interference (RNAi) of the TG gene was highly lethal in flies and induced apoptosis of gut epithelial cells after oral infection with Pseudomonas entomophila. Moreover, AprA, a metalloprotease secreted by P. entomophila, digested non-crosslinked drosocrystallin fibers, but not drosocrystallin fibers crosslinked by TG. In vitro experiments using recombinant drosocrystallin and monalysin proteins demonstrated that monalysin, a pore-forming exotoxin of P. entomophila, was adsorbed on the crosslinked drosocrystallin fibers in the presence of P. entomophila culture supernatant. In addition, gut-specific TG-RNAi flies had a shorter lifespan than control flies after ingesting P. entomophila, whereas the lifespan after ingesting AprA-knockout P. entomophila was at control levels. We conclude that drosocrystallin fibers crosslinked by TG, but not non-crosslinked drosocrystallin fibers, form an important physical barrier against exotoxins of invading pathogenic microbes. PMID:26506243

  5. Crosslinking of a Peritrophic Matrix Protein Protects Gut Epithelia from Bacterial Exotoxins.

    Directory of Open Access Journals (Sweden)

    Toshio Shibata

    2015-10-01

    Full Text Available Transglutaminase (TG catalyzes protein-protein crosslinking, which has important and diverse roles in vertebrates and invertebrates. Here we demonstrate that Drosophila TG crosslinks drosocrystallin, a peritrophic matrix protein, to form a stable fiber structure on the gut peritrophic matrix. RNA interference (RNAi of the TG gene was highly lethal in flies and induced apoptosis of gut epithelial cells after oral infection with Pseudomonas entomophila. Moreover, AprA, a metalloprotease secreted by P. entomophila, digested non-crosslinked drosocrystallin fibers, but not drosocrystallin fibers crosslinked by TG. In vitro experiments using recombinant drosocrystallin and monalysin proteins demonstrated that monalysin, a pore-forming exotoxin of P. entomophila, was adsorbed on the crosslinked drosocrystallin fibers in the presence of P. entomophila culture supernatant. In addition, gut-specific TG-RNAi flies had a shorter lifespan than control flies after ingesting P. entomophila, whereas the lifespan after ingesting AprA-knockout P. entomophila was at control levels. We conclude that drosocrystallin fibers crosslinked by TG, but not non-crosslinked drosocrystallin fibers, form an important physical barrier against exotoxins of invading pathogenic microbes.

  6. Heterologously expressed bacterial and human multidrug resistance proteins confer cadmium resistance to Escherichia coli

    NARCIS (Netherlands)

    Achard-Joris, M; van Saparoea, HBV; Driessen, AJM; Bourdineaud, JP; Bourdineaud, Jean-Paul

    2005-01-01

    The human MDR1 gene is induced by cadmium exposure although no resistance to this metal is observed in human cells overexpressing hMDR1. To access the role of MDR proteins in cadmium resistance, human MDR1, Lactococcus lactis lmrA, and Oenococcus oeni omrA were expressed in an Escherichia coli tolC

  7. Expression, secretion and antigenic variation of bacterial S-layer proteins

    NARCIS (Netherlands)

    Boot, H.J.; Pouwels, P.H.

    1996-01-01

    The function of the S-layer, a regularly arranged structure on the outside of numerous bacteria, appears to be different for bacteria living in different environments. Almost no similarity exists between the primary sequences of S-proteins, although their amino acid composition is comparable.

  8. Enterococcus faecalis zinc-responsive proteins mediate bacterial defence against zinc overload, lysozyme and oxidative stress

    NARCIS (Netherlands)

    C. Abrantes, Marta; Kok, Jan; de Fatima Silva Lopes, Maria

    2014-01-01

    Two Enterococcus faecalis genes encoding the P-type ATPase EF1400 and the putative SapB protein EF0759 were previously shown to be strongly upregulated in the presence of high concentrations of zinc. In the present work, we showed that a Zn(2+)-responsive DNA-binding motif (zim) is present in the

  9. Characterization of two bacterial hydroxynitrile lyases with high similarity to cupin superfamily proteins

    NARCIS (Netherlands)

    Hussain, Z.; Wiedner, R.; Steiner, K.; Hajek, T.; Avi, M.; Hecher, B.; Sessitsch, A.; Schwab, H.

    2012-01-01

    Hydroxynitrile lyases (HNLs) catalyze the cleavage of cyanohydrins. In the reverse reaction, they catalyze the formation of carbon-carbon bonds by enantioselective condensation of hydrocyanic acid with carbonyls. In this study, we describe two proteins from endophytic bacteria that display activity

  10. Bacterial surface layer proteins as a novel capillary coating material for capillary electrophoretic separations

    Energy Technology Data Exchange (ETDEWEB)

    Moreno-Gordaliza, Estefanía, E-mail: emorenog@ucm.es [Division of Analytical Biosciences, Leiden Academic Centre for Drug Research, Universiteit Leiden, Einsteinweg 55, 2300, RA, Leiden (Netherlands); Department of Analytical Chemistry, Faculty of Chemistry, Universidad Complutense de Madrid, Avda. Complutense s/n, 28040, Madrid (Spain); Stigter, Edwin C.A. [Division of Analytical Biosciences, Leiden Academic Centre for Drug Research, Universiteit Leiden, Einsteinweg 55, 2300, RA, Leiden (Netherlands); Department of Molecular Cancer Research, Universitair Medisch Centrum Utrecht, Wilhelmina Kinder Ziekenhuis, Lundlaan 6, 3584, EA Utrecht (Netherlands); Lindenburg, Petrus W.; Hankemeier, Thomas [Division of Analytical Biosciences, Leiden Academic Centre for Drug Research, Universiteit Leiden, Einsteinweg 55, 2300, RA, Leiden (Netherlands)

    2016-06-07

    A novel concept for stable coating in capillary electrophoresis, based on recrystallization of surface layer proteins on hydrophobized fused silica capillaries, was demonstrated. Surface layer protein A (SlpA) from Lactobacillus acidophilus bacteria was extracted, purified and used for coating pre-silanized glass substrates presenting different surface wettabilities (either hydrophobic or hydrophilic). Contact angle determination on SlpA-coated hydrophobic silica slides showed that the surfaces turned to hydrophilic after coating (53 ± 5°), due to a protein monolayer formation by protein-surface hydrophobic interactions. Visualization by atomic force microscopy demonstrated the presence of a SlpA layer on methylated silica slides displaying a surface roughness of 0.44 ± 0.02 nm. Additionally, a protein layer was visualized by fluorescence microscopy in methylated silica capillaries coated with SlpA and fluorescein isothiocyanate-labeled. The SlpA-coating showed an outstanding stability, even after treatment with 20 mM NaOH (pH 12.3). The electroosmotic flow in coated capillaries showed a partial suppression at pH 7.50 (3.8 ± 0.5 10{sup −9} m{sup 2} V{sup −1} s{sup −1}) when compared with unmodified fused silica (5.9 ± 0.1 10{sup −8} m{sup 2} V{sup −1} s{sup −1}). To demonstrate the potential of this novel coating, the SlpA-coated capillaries were applied for the first time for electrophoretic separation, and proved to be very suitable for the isotachophoretic separation of lipoproteins in human serum. The separations showed a high degree of repeatability (absolute migration times with 1.1–1.8% coefficient-of-variation (CV) within a day) and 2–3% CV inter-capillary reproducibility. The capillaries were stable for more than 100 runs at pH 9.40, and showed to be an exceptional alternative for challenging electrophoretic separations at long-term use. - Highlights: • New coating using recrystallized surface-layer proteins on

  11. Coupling of anaerobic waste treatment to produce protein- and lipid-rich bacterial biomass

    Science.gov (United States)

    Steinberg, Lisa M.; Kronyak, Rachel E.; House, Christopher H.

    2017-11-01

    Future long-term manned space missions will require effective recycling of water and nutrients as part of a life support system. Biological waste treatment is less energy intensive than physicochemical treatment methods, yet anaerobic methanogenic waste treatment has been largely avoided due to slow treatment rates and safety issues concerning methane production. However, methane is generated during atmosphere regeneration on the ISS. Here we propose waste treatment via anaerobic digestion followed by methanotrophic growth of Methylococcus capsulatus to produce a protein- and lipid-rich biomass that can be directly consumed, or used to produce other high-protein food sources such as fish. To achieve more rapid methanogenic waste treatment, we built and tested a fixed-film, flow-through, anaerobic reactor to treat an ersatz wastewater. During steady-state operation, the reactor achieved a 97% chemical oxygen demand (COD) removal rate with an organic loading rate of 1740 g d-1 m-3 and a hydraulic retention time of 12.25 d. The reactor was also tested on three occasions by feeding ca. 500 g COD in less than 12 h, representing 50x the daily feeding rate, with COD removal rates ranging from 56-70%, demonstrating the ability of the reactor to respond to overfeeding events. While investigating the storage of treated reactor effluent at a pH of 12, we isolated a strain of Halomonas desiderata capable of acetate degradation under high pH conditions. We then tested the nutritional content of the alkaliphilic Halomonas desiderata strain, as well as the thermophile Thermus aquaticus, as supplemental protein and lipid sources that grow in conditions that should preclude pathogens. The M. capsulatus biomass consisted of 52% protein and 36% lipids, the H. desiderata biomass consisted of 15% protein and 7% lipids, and the Thermus aquaticus biomass consisted of 61% protein and 16% lipids. This work demonstrates the feasibility of rapid waste treatment in a compact reactor design

  12. Bacterial Production, Characterization and Protein Modeling of a Novel Monofuctional Isoform of FAD Synthase in Humans: An Emergency Protein?

    Directory of Open Access Journals (Sweden)

    Piero Leone

    2018-01-01

    Full Text Available FAD synthase (FADS, EC 2.7.7.2 is the last essential enzyme involved in the pathway of biosynthesis of Flavin cofactors starting from Riboflavin (Rf. Alternative splicing of the human FLAD1 gene generates different isoforms of the enzyme FAD synthase. Besides the well characterized isoform 1 and 2, other FADS isoforms with different catalytic domains have been detected, which are splice variants. We report the characterization of one of these novel isoforms, a 320 amino acid protein, consisting of the sole C-terminal 3′-phosphoadenosine 5′-phosphosulfate (PAPS reductase domain (named FADS6. This isoform has been previously detected in Riboflavin-Responsive (RR-MADD and Non-responsive Multiple Acyl-CoA Dehydrogenase Deficiency (MADD patients with frameshift mutations of FLAD1 gene. To functionally characterize the hFADS6, it has been over-expressed in Escherichia coli and purified with a yield of 25 mg·L−1 of cell culture. The protein has a monomeric form, it binds FAD and is able to catalyze FAD synthesis (kcat about 2.8 min−1, as well as FAD pyrophosphorolysis in a strictly Mg2+-dependent manner. The synthesis of FAD is inhibited by HgCl2. The enzyme lacks the ability to hydrolyze FAD. It behaves similarly to PAPS. Combining threading and ab-initio strategy a 3D structural model for such isoform has been built. The relevance to human physio-pathology of this FADS isoform is discussed.

  13. Expressing a bacterial mercuric ion binding protein in plant for phytoremediation of heavy metals.

    Science.gov (United States)

    Hsieh, Ju-Liang; Chen, Ching-Yi; Chiu, Meng-Hsuen; Chein, Mei-Fang; Chang, Jo-Shu; Endo, Ginro; Huang, Chieh-Chen

    2009-01-30

    A specific mercuric ion binding protein (MerP) originating from transposon TnMERI1 of Bacillus megaterium strain MB1 isolated from Minamata Bay displayed good adsorption capability for a variety of heavy metals. In this study, the Gram-positive MerP protein was expressed in transgenic Arabidopsis to create a model system for phytoremediation of heavy metals. Under control of an actin promoter, the transgenic Arabidpsis showed higher tolerance and accumulation capacity for mercury, cadium and lead when compared with the control plant. Results from confocal microscopy analysis also indicate that MerP was localized at the cell membrane and vesicles of plant cells. The developed transgenic plants possessing excellent metal-accumulative ability could have potential applications in decontamination of heavy metals.

  14. Subcutaneous immunization with inactivated bacterial components and purified protein of Escherichia coli, Fusobacterium necrophorum and Trueperella pyogenes prevents puerperal metritis in Holstein dairy cows.

    Science.gov (United States)

    Machado, Vinícius Silva; Bicalho, Marcela Luccas de Souza; Meira Junior, Enoch Brandão de Souza; Rossi, Rodolfo; Ribeiro, Bruno Leonardo; Lima, Svetlana; Santos, Thiago; Kussler, Arieli; Foditsch, Carla; Ganda, Erika Korzune; Oikonomou, Georgios; Cheong, Soon Hon; Gilbert, Robert Owen; Bicalho, Rodrigo Carvalho

    2014-01-01

    In this study we evaluate the efficacy of five vaccine formulations containing different combinations of proteins (FimH; leukotoxin, LKT; and pyolysin, PLO) and/or inactivated whole cells (Escherichia coli, Fusobacterium necrophorum, and Trueperella pyogenes) in preventing postpartum uterine diseases. Inactivated whole cells were produced using two genetically distinct strains of each bacterial species (E. coli, F. necrophorum, and T. pyogenes). FimH and PLO subunits were produced using recombinant protein expression, and LKT was recovered from culturing a wild F. necrophorum strain. Three subcutaneous vaccines were formulated: Vaccine 1 was composed of inactivated bacterial whole cells and proteins; Vaccine 2 was composed of proteins only; and Vaccine 3 was composed of inactivated bacterial whole cells only. Two intravaginal vaccines were formulated: Vaccine 4 was composed of inactivated bacterial whole cells and proteins; and Vaccine 5 was composed of PLO and LKT. To evaluate vaccine efficacy, a randomized clinical trial was conducted at a commercial dairy farm; 371 spring heifers were allocated randomly into one of six different treatments groups: control, Vaccine 1, Vaccine 2, Vaccine 3, Vaccine 4 and Vaccine 5. Late pregnant heifers assigned to one of the vaccine groups were each vaccinated twice: at 230 and 260 days of pregnancy. When vaccines were evaluated grouped as subcutaneous and intravaginal, the subcutaneous ones were found to significantly reduce the incidence of puerperal metritis. Additionally, subcutaneous vaccination significantly reduced rectal temperature at 6±1 days in milk. Reproduction was improved for cows that received subcutaneous vaccines. In general, vaccination induced a significant increase in serum IgG titers against all antigens, with subcutaneous vaccination again being more effective. In conclusion, subcutaneous vaccination with inactivated bacterial components and/or protein subunits of E. coli, F. necrophorum and T. pyogenes

  15. Structural basis of antifreeze activity of a bacterial multi-domain antifreeze protein.

    Directory of Open Access Journals (Sweden)

    Chen Wang

    Full Text Available Antifreeze proteins (AFPs enhance the survival of organisms inhabiting cold environments by affecting the formation and/or structure of ice. We report the crystal structure of the first multi-domain AFP that has been characterized. The two ice binding domains are structurally similar. Each consists of an irregular β-helix with a triangular cross-section and a long α-helix that runs parallel on one side of the β-helix. Both domains are stabilized by hydrophobic interactions. A flat plane on the same face of each domain's β-helix was identified as the ice binding site. Mutating any of the smaller residues on the ice binding site to bulkier ones decreased the antifreeze activity. The bulky side chain of Leu174 in domain A sterically hinders the binding of water molecules to the protein backbone, partially explaining why antifreeze activity by domain A is inferior to that of domain B. Our data provide a molecular basis for understanding differences in antifreeze activity between the two domains of this protein and general insight on how structural differences in the ice-binding sites affect the activity of AFPs.

  16. Novel electrospun polyvinylidene fluoride-graphene oxide-silver nanocomposite membranes with protein and bacterial antifouling characteristics

    Directory of Open Access Journals (Sweden)

    C. Liu

    2018-04-01

    Full Text Available We developed and fabricated novel polyvinylidene fluoride (PVDF-(0.5–2%Ag and PVDF-(0.5–2%Ag-1% graphene oxide (GO nanocomposite membranes with antifouling properties through electrospinning. Silver nanoparticles (AgNPs were in situ synthesized from silver nitrate precursor directly. The tensile properties, wetting, antifouling characteristics of pristine PVDF and its nanocomposite membranes were studied. Tensile tests showed that the addition of 0.5–2% AgNPs to PVDF improves its elastic modulus and tensile strength markedly. A further increase in both tensile modulus and strength of PVDF were obtained by hybridizing AgNPs with 1% GO. Water contact angle measurements revealed that the incorporation of AgNPs or AgNPs/GO nanofillers into PVDF decreases its degree of hydrophobicity. This led to the nanocomposite membranes having higher water flux permeation. In addition, AgNPs and AgNPs/GO fillers played a crucial role against protein and bacterial fouling of the resulting composite membranes. The antibacterial activities of electrospun nanocomposite membranes were assessed against Gram-negative Escherichia coli and Gram-positive Staphylococcus aureus. On the basis of water contact angle, water permeation flux and antifouling results, electrospun PVDF-2% Ag-GO composite membrane was found to exhibit excellent filtration performance, protein antifouling and bactericidal activities. Thus such a fibrous nanocomposite is considered as a high-potential membrane for water purification and disinfection applications.

  17. Inhibition of bacterial conjugation by phage M13 and its protein g3p: quantitative analysis and model.

    Directory of Open Access Journals (Sweden)

    Abraham Lin

    Full Text Available Conjugation is the main mode of horizontal gene transfer that spreads antibiotic resistance among bacteria. Strategies for inhibiting conjugation may be useful for preserving the effectiveness of antibiotics and preventing the emergence of bacterial strains with multiple resistances. Filamentous bacteriophages were first observed to inhibit conjugation several decades ago. Here we investigate the mechanism of inhibition and find that the primary effect on conjugation is occlusion of the conjugative pilus by phage particles. This interaction is mediated primarily by phage coat protein g3p, and exogenous addition of the soluble fragment of g3p inhibited conjugation at low nanomolar concentrations. Our data are quantitatively consistent with a simple model in which association between the pili and phage particles or g3p prevents transmission of an F plasmid encoding tetracycline resistance. We also observe a decrease in the donor ability of infected cells, which is quantitatively consistent with a reduction in pili elaboration. Since many antibiotic-resistance factors confer susceptibility to phage infection through expression of conjugative pili (the receptor for filamentous phage, these results suggest that phage may be a source of soluble proteins that slow the spread of antibiotic resistance genes.

  18. Bright blue-shifted fluorescent proteins with Cys in the GAF domain engineered from bacterial phytochromes: fluorescence mechanisms and excited-state dynamics

    NARCIS (Netherlands)

    Hontani, Yusaku; Shcherbakova, Daria M.; Baloban, Mikhail; Zhu, Jingyi; Verkhusha, Vladislav V.; Kennis, John T. M.

    2016-01-01

    Near-infrared fluorescent proteins (NIR FPs) engineered from bacterial phytochromes (BphPs) are of great interest for in vivo imaging. They utilize biliverdin (BV) as a chromophore, which is a heme degradation product, and therefore they are straightforward to use in mammalian tissues. Here, we

  19. Low ascitic fluid total protein levels is not associated to the development of spontaneous bacterial peritonitis in a cohort of 274 patients with cirrhosis

    DEFF Research Database (Denmark)

    Mo, Silje; Bendtsen, Flemming; Wiese, Signe Skovgaard

    2018-01-01

    Background: Spontaneous bacterial peritonitis (SBP) is a complication to decompensated cirrhosis. Fluoroquinolones may prevent SBP. However, predictive markers for SBP are wanted. Guidelines suggest that patients with ascitic fluid protein below 15 g/l receive fluoroquinolones to prevent SBP...

  20. Identification of the Dimer Exchange Interface of the Bacterial DNA Damage Response Protein UmuD.

    Science.gov (United States)

    Murison, David A; Timson, Rebecca C; Koleva, Bilyana N; Ordazzo, Michael; Beuning, Penny J

    2017-09-12

    The Escherichia coli SOS response, an induced DNA damage response pathway, confers survival on bacterial cells by providing accurate repair mechanisms as well as the potentially mutagenic pathway translesion synthesis (TLS). The umuD gene products are upregulated after DNA damage and play roles in both nonmutagenic and mutagenic aspects of the SOS response. Full-length UmuD is expressed as a homodimer of 139-amino-acid subunits, which eventually cleaves its N-terminal 24 amino acids to form UmuD'. The cleavage product UmuD' and UmuC form the Y-family polymerase DNA Pol V (UmuD' 2 C) capable of performing TLS. UmuD and UmuD' exist as homodimers, but their subunits can readily exchange to form UmuDD' heterodimers preferentially. Heterodimer formation is an essential step in the degradation pathway of UmuD'. The recognition sequence for ClpXP protease is located within the first 24 amino acids of full-length UmuD, and the partner of full-length UmuD, whether UmuD or UmuD', is degraded by ClpXP. To better understand the mechanism by which UmuD subunits exchange, we measured the kinetics of exchange of a number of fluorescently labeled single-cysteine UmuD variants as detected by Förster resonance energy transfer. Labeling sites near the dimer interface correlate with increased rates of exchange, indicating that weakening the dimer interface facilitates exchange, whereas labeling sites on the exterior decrease the rate of exchange. In most but not all cases, homodimer and heterodimer exchange exhibit similar rates, indicating that somewhat different molecular surfaces mediate homodimer exchange and heterodimer formation.

  1. A Bacterial Biosensor for Oxidative Stress Using the Constitutively Expressed Redox-Sensitive Protein roGFP2

    Directory of Open Access Journals (Sweden)

    Carlos R. Arias-Barreiro

    2010-06-01

    Full Text Available A highly specific, high throughput-amenable bacterial biosensor for chemically induced cellular oxidation was developed using constitutively expressed redox-sensitive green fluorescent protein roGFP2 in E. coli (E. coli-roGFP2. Disulfide formation between two key cysteine residues of roGFP2 was assessed using a double-wavelength ratiometric approach. This study demonstrates that only a few minutes were required to detect oxidation using E. coli-roGFP2, in contrast to conventional bacterial oxidative stress sensors. Cellular oxidation induced by hydrogen peroxide, menadione, sodium selenite, zinc pyrithione, triphenyltin and naphthalene became detectable after 10 seconds and reached the maxima between 80 to 210 seconds, contrary to Cd2+, Cu2+, Pb2+, Zn2+ and sodium arsenite, which induced the oxidation maximum immediately. The lowest observable effect concentrations (in ppm were determined as 1.0 x 10−7 (arsenite, 1.0 x 10−4 (naphthalene, 1.0 x 10−4 (Cu2+, 3.8 x 10−4 (H2O2, 1.0 x 10−3 (Cd2+, 1.0 x 10−3 (Zn2+, 1.0 x 10−2 (menadione, 1.0 (triphenyltin, 1.56 (zinc pyrithione, 3.1 (selenite and 6.3 (Pb2+, respectively. Heavy metal-induced oxidation showed unclear response patterns, whereas concentration-dependent sigmoid curves were observed for other compounds. In vivo GSH content and in vitro roGFP2 oxidation assays together with E. coli-roGFP2 results suggest that roGFP2 is sensitive to redox potential change and thiol modification induced by environmental stressors. Based on redox-sensitive technology, E. coli-roGFP2 provides a fast comprehensive detection system for toxicants that induce cellular oxidation.

  2. Protein and Bacterial Antifouling Behavior of Melt-Coextruded Nanofiber Mats.

    Science.gov (United States)

    Kim, Si-Eun; Zhang, Cong; Advincula, Abigail A; Baer, Eric; Pokorski, Jonathan K

    2016-04-13

    Antifouling surfaces are important for biomedical devices to prevent secondary infections and mitigate the effects of the foreign body response. Herein, we describe melt-coextruded poly(ε-caprolactone) (PCL) nanofiber mats grafted with antifouling polymers. Nonwoven PCL fiber mats are produced using a multilayered melt coextrusion process followed by high-pressure hydroentanglement to yield porous patches. The resulting fiber mats show submicrometer cross-sectional fiber dimensions and yield pore sizes that were nearly uniform, with a mean pore size of 1.6 ± 0.9 μm. Several antifouling polymers, including hydrophilic, zwitterionic, and amphipathic molecules, are grafted to the surface of the mats using a two-step procedure that includes photochemistry followed by the copper-catalyzed azide-alkyne cycloaddition reaction. Fiber mats are evaluated using separate adsorption tests for serum proteins and E. coli. The results indicate that poly(oligo(ethylene glycol) methyl ether methacrylate)-co-(trifluoroethyl methacrylate) (poly(OEGMEMA-co-TFEMA)) grafted mats exhibit approximately 85% less protein adhesion and 97% less E. coli adsorption when compared to unmodified PCL fibermats. In dynamic antifouling testing, the amphiphilic fluorous polymer surface shows the highest flux and highest rejection value of foulants. The work presented within has implications on the high-throughput production of antifouling microporous patches for medical applications.

  3. Secondary Structure Preferences of Mn2+ Binding Sites in Bacterial Proteins

    Directory of Open Access Journals (Sweden)

    Tatyana Aleksandrovna Khrustaleva

    2014-01-01

    Full Text Available 3D structures of proteins with coordinated Mn2+ ions from bacteria with low, average, and high genomic GC-content have been analyzed (149 PDB files were used. Major Mn2+ binders are aspartic acid (6.82% of Asp residues, histidine (14.76% of His residues, and glutamic acid (3.51% of Glu residues. We found out that the motif of secondary structure “beta strand-major binder-random coil” is overrepresented around all the three major Mn2+ binders. That motif may be followed by either alpha helix or beta strand. Beta strands near Mn2+ binding residues should be stable because they are enriched by such beta formers as valine and isoleucine, as well as by specific combinations of hydrophobic and hydrophilic amino acid residues characteristic to beta sheet. In the group of proteins from GC-rich bacteria glutamic acid residues situated in alpha helices frequently coordinate Mn2+ ions, probably, because of the decrease of Lys usage under the influence of mutational GC-pressure. On the other hand, the percentage of Mn2+ sites with at least one amino acid in the “beta strand-major binder-random coil” motif of secondary structure (77.88% does not depend on genomic GC-content.

  4. Bacterial intermediate filaments

    DEFF Research Database (Denmark)

    Charbon, Godefroid; Cabeen, M.; Jacobs-Wagner, C.

    2009-01-01

    Crescentin, which is the founding member of a rapidly growing family of bacterial cytoskeletal proteins, was previously proposed to resemble eukaryotic intermediate filament (IF) proteins based on structural prediction and in vitro polymerization properties. Here, we demonstrate that crescentin...

  5. Bacterial Adhesion & Blocking Bacterial Adhesion

    DEFF Research Database (Denmark)

    Vejborg, Rebecca Munk

    2008-01-01

    , which influence the transition from a planktonic lifestyle to a sessile lifestyle, have been studied. Protein conditioning film formation was found to influence bacterial adhesion and subsequent biofilm formation considerable, and an aqueous extract of fish muscle tissue was shown to significantly...... tract to the microbial flocs in waste water treatment facilities. Microbial biofilms may however also cause a wide range of industrial and medical problems, and have been implicated in a wide range of persistent infectious diseases, including implantassociated microbial infections. Bacterial adhesion...... is the first committing step in biofilm formation, and has therefore been intensely scrutinized. Much however, still remains elusive. Bacterial adhesion is a highly complex process, which is influenced by a variety of factors. In this thesis, a range of physico-chemical, molecular and environmental parameters...

  6. Is the C-terminal insertional signal in Gram-negative bacterial outer membrane proteins species-specific or not?

    Directory of Open Access Journals (Sweden)

    Paramasivam Nagarajan

    2012-09-01

    heterologous overexpression of almost all OMPs should be feasible in E. coli and other Gram-negative bacterial model organisms. This is relevant especially for biotechnology applications, where recombinant OMPs are used e.g. for the development of vaccines. For the species in which the motif is significantly different, we identify the residues mainly responsible for this difference that can now be changed in heterologous expression experiments to yield functional proteins.

  7. Decreased Bacterial Attachment and Protein Adsorption to Coatings Produced by Low Enegy Plasma Polymerization

    DEFF Research Database (Denmark)

    Andersen, T.E.; Kingshott, Peter; Benter, M.

    .figure) .and E. coli grown on uncoated silicone compared to PP-PVP coated silicone (right figure). Results from the flow chamber analysis shows PP-PVP to be very good at preventing E. coli colonization during prolonged growth in flow chamber. At this point other surfaces and bacteria remains to be tested...... adsorption and bacteria attachment/colonization. This is emphasized by the fact that long dwelling urinary catheters, which is a typical silicone medical device, causes 5% per day incidence of urinary tract infection [1,2]. A demand therefore exists for surface modifications providing the silicone material......-coated crystals were then treated with one of the plasma polymerized coatings. Adsorption of fibrinogen, human serum albumin or immunoglobulin G was measured using a QCM-D instrument [5] (model E4, Q-Sense AB, Vastra Frolunda, Sweden) using a solution of 50llg/1 protein in PBS buffer. Results and Discussion: Our...

  8. Etching of polymers, proteins and bacterial spores by atmospheric pressure DBD plasma in air

    Science.gov (United States)

    Kuzminova, A.; Kretková, T.; Kylián, O.; Hanuš, J.; Khalakhan, I.; Prukner, V.; Doležalová, E.; Šimek, M.; Biederman, H.

    2017-04-01

    Many studies proved that non-equilibrium discharges generated at atmospheric pressure are highly effective for the bio-decontamination of surfaces of various materials. One of the key processes that leads to a desired result is plasma etching and thus the evaluation of etching rates of organic materials is of high importance. However, the comparison of reported results is rather difficult if impossible as different authors use diverse sources of atmospheric plasma that are operated at significantly different operational parameters. Therefore, we report here on the systematic study of the etching of nine different common polymers that mimic the different structures of more complicated biological systems, bovine serum albumin (BSA) selected as the model protein and spores of Bacillus subtilis taken as a representative of highly resistant micro-organisms. The treatment of these materials was performed by means of atmospheric pressure dielectric barrier discharge (DBD) sustained in open air at constant conditions. All tested polymers, BSA and spores, were readily etched by DBD plasma. However, the measured etching rates were found to be dependent on the chemical structure of treated materials, namely on the presence of oxygen in the structure of polymers.

  9. Interaction between bacterial outer membrane proteins and periplasmic quality control factors: a kinetic partitioning mechanism.

    Science.gov (United States)

    Wu, Si; Ge, Xi; Lv, Zhixin; Zhi, Zeyong; Chang, Zengyi; Zhao, Xin Sheng

    2011-09-15

    The OMPs (outer membrane proteins) of Gram-negative bacteria have to be translocated through the periplasmic space before reaching their final destination. The aqueous environment of the periplasmic space and high permeability of the outer membrane engender such a translocation process inevitably challenging. In Escherichia coli, although SurA, Skp and DegP have been identified to function in translocating OMPs across the periplasm, their precise roles and their relationship remain to be elucidated. In the present paper, by using fluorescence resonance energy transfer and single-molecule detection, we have studied the interaction between the OMP OmpC and these periplasmic quality control factors. The results of the present study reveal that the binding rate of OmpC to SurA or Skp is much faster than that to DegP, which may lead to sequential interaction between OMPs and different quality control factors. Such a kinetic partitioning mechanism for the chaperone-substrate interaction may be essential for the quality control of the biogenesis of OMPs.

  10. Bacterial mitosis

    DEFF Research Database (Denmark)

    Møller-Jensen, Jakob; Borch, Jonas; Dam, Mette

    2003-01-01

    Bacterial DNA segregation takes place in an active and ordered fashion. In the case of Escherichia coli plasmid R1, the partitioning system (par) separates paired plasmid copies and moves them to opposite cell poles. Here we address the mechanism by which the three components of the R1 par system...... act together to generate the force required for plasmid movement during segregation. ParR protein binds cooperatively to the centromeric parC DNA region, thereby forming a complex that interacts with the filament-forming actin-like ParM protein in an ATP-dependent manner, suggesting that plasmid...

  11. High-temperature short-time pasteurisation of human breastmilk is efficient in retaining protein and reducing the bacterial count.

    Science.gov (United States)

    Klotz, Daniel; Joellenbeck, Mirjam; Winkler, Karl; Kunze, Mirjam; Huzly, Daniela; Hentschel, Roland

    2017-05-01

    Milk banks are advised to use Holder pasteurisation to inactivate the cytomegalovirus, but the process adversely affects the bioactive properties of human breastmilk. This study explored the antibacterial efficacy of an alternative high-temperature short-time (HTST) treatment of human breastmilk and its effect on marker proteins, compared with the Holder method. Breastmilk samples were obtained from 27 mothers with infants in a German neonatal intensive care unit. The samples were either heated to 62°C for five seconds using HTST or processed using Holder pasteurisation, at 63 ± 0.5°C for 30 minutes. Immunoglobulin A, lactoferrin, lysozyme, alkaline phosphatase and bile salt-stimulated lipase concentrations and bacterial colony-forming units/mL were measured before and after heating. HTST-treated samples retained higher rates of immunoglobulin A (95% versus 83%), alkaline phosphatase (6% versus 0%) and bile salt-stimulated lipase (0.8% versus 0.4%) than Holder pasteurisation samples (all p HTST treatment protocol retained some of the bioactive properties of human breastmilk and appeared to have similar antibacterial efficacy to Holder pasteurisation. ©2017 Foundation Acta Paediatrica. Published by John Wiley & Sons Ltd.

  12. Pseudomonas fluorescens filamentous hemagglutinin, an iron-regulated protein, is an important virulence factor that modulates bacterial pathogenicity

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    Yuan-yuan Sun

    2016-08-01

    Full Text Available Pseudomonas fluorescens is a common bacterial pathogen to a wide range of aquaculture animals including various species of fish. In this study, we employed proteomic analysis and identified filamentous hemagglutinin (FHA as an iron-responsive protein secreted by TSS, a pathogenic P. fluorescens isolate. In vitro study showed that compared to the wild type, the fha mutant TSSfha (i exhibited a largely similar vegetative growth profile but significantly retarded in the ability of biofilm growth and producing extracellular matrix, (ii displayed no apparent flagella and motility, (iii was defective in the attachment to host cells and unable to form self-aggregation, (iv displayed markedly reduced capacity of hemagglutination and surviving in host serum. In vivo infection analysis revealed that TSSfha was significantly attenuated in the ability of dissemination in fish tissues and inducing host mortality, and that antibody blocking of the natural FHA produced by the wild type TSS impaired the infectivity of the pathogen. Furthermore, when introduced into turbot as a subunit vaccine, recombinant FHA elicited a significant protection against lethal TSS challenge. Taken together, these results indicate for the first time that P. fluorescens FHA is a key virulence factor essential to multiple biological processes associated with pathogenicity.

  13. Intracellular crowding effects on the self-association of the bacterial cell division protein FtsZ.

    Science.gov (United States)

    Naddaf, Lamis; Sayyed-Ahmad, Abdallah

    2014-12-15

    The dimerization rate of the bacterial cell division protein FtsZ is strongly affected by the intracellular crowding. Yet the complexity of the intracellular environment makes it difficult to investigate via all-atom molecular dynamics or other detailed theoretical methods. We study the crowding effect on FtsZ dimerization which is the first step of an oligomerization process that results in more elaborate supramolecular structures. In particular, we consider the effect of intracellular crowding on the reaction rates, and their dependence on the different concentrations of crowding agents. We achieved this goal by using Brownian dynamics (BD) simulation techniques and a modified post-processing approach in which we decompose the rate constant in crowded media as a product of the rate constant in the dilute solution times a factor that incorporates the crowding effect. The latter factor accounts for the diffusion reduction and crowder induced energy. In addition we include the crowding effects on water viscosity in the BD simulations of crowded media. We finally show that biomolecular crowding has a considerable effect on the FtsZ dimerization by increasing the dimerization rate constant from 2.6×10(7)M(-1)s(-1) in the absence of crowders to 1.0×10(8)M(-1)s(-1) at crowding level of 0.30. Copyright © 2014 Elsevier Inc. All rights reserved.

  14. Incorporation of 15N and 14C into amino acids of bacterial and protozoal protein in the rumen of the cow on urea-rich feed

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    Eeva-Liisa Syväoja

    1979-01-01

    Full Text Available The utilization of the non-protein nitrogen and carbon of feed by rumen microorganisms for the synthesis of protein was studied by administering [U-14C] sucrose and 15NH4Cl to a cow on urea-rich, low-protein feed. By studying the labelling of the protozoa and bacteria and the amino acids isolated from them at intervals up to 48 hours afterwards, it was found that the bacteria synthesized amino acids from nonprotein nitrogen much more rapidly and effectively than the protozoa. Also the labelling of the carbon in the amino acids of the bacteria was more rapid than in the protozoa. In both protozoa and bacteria there was intracellular storage of [14C] sucrose. Of the bacterial amino acids the most vigorous 14C labelling was found in Glu, Arg, Lys, Val and Ala and the weakest labelling in Gly, His and Ser. Of the protozoal amino acids Ala, Asp, Glu, Leu and Lys had the highest labelling and Pro, Gly, His and Phe the lowest. In the bacterial protein the labelling of Pro and Arg was ten times that of the corresponding protozoal amino acids, and Asp, Ser and Ala four times. After the 15NH4Cl dose the half-life of 15N in the rumen fluid was estimated to be 3.3 h. Labelled ammonium nitrogen was about 11 —15 % of the bacterial nitrogen and 2—3 % of the protozoal nitrogen after 1 h. Of the protozoal amino acids Ala, Glu, Val, Asp and Met had the most vigorous labelling, and of the bacterial amino acids Glu, Asp, Ser, He and Tyr. The slowest incorporation of ammonium nitrogen was into His, Pro, Arg and Gly in both bacteria and protozoa. The labelling of the bacterial amino acids was approximately 7—8 times more vigorous than that of the protozoal amino acids. The labelling of Ala was only 4 times, and that of Val, Met and Glu 5 times more vigorous than with protozoal protein. The pathway of histidine synthesis seemed to be restricted in both bacteria and protozoa and therefore may be a limiting factor in protein synthesis, particularly in cows fed

  15. Endosulfan induced alteration in bacterial protein profile and RNA yield of Klebsiella sp. M3, Achromobacter sp. M6, and Rhodococcus sp. M2.

    Science.gov (United States)

    Singh, Madhu; Singh, Dileep Kumar

    2014-01-30

    Three bacterial strains identified as Klebsiella sp. M3, Achromobacter sp. M6 and Rhodococcus sp. M2 were isolated by soil enrichment with endosulfan followed by shake flask enrichment technique. They were efficiently degrading endosulfan in the NSM (non sulfur medium) broth. Degradation of endosulfan was faster with the cell free extract of bacterial cells grown in the sulfur deficient medium (NSM) supplemented with endosulfan than that of nutrient rich medium (Luria Bertani). In the cell free extract of NSM supplemented with endosulfan as sole sulfur source, a unique band was visualized on SDS-PAGE but not with magnesium sulfate as the sole sulfur source in NSM and LB with endosulfan. Expression of a unique polypeptide band was speculated to be induced by endosulfan under sulfur starved condition. These unique polypeptide bands were identified as OmpK35 protein, sulfate binding protein and outer membrane porin protein, respectively, in Klebsiella sp. M3, Achromobacter sp. M6 and Rhodococcus sp. M2. Endosulfan showed dose dependent negative effect on total RNA yield of bacterial strains in nutrient rich medium. Absence of plasmid DNA indicated the presence of endosulfan metabolizing gene on genomic DNA. Copyright © 2013 Elsevier B.V. All rights reserved.

  16. The pH dependence of polymerization and bundling by the essential bacterial cytoskeletal protein FtsZ.

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    Raúl Pacheco-Gómez

    Full Text Available There is a growing body of evidence that bacterial cell division is an intricate coordinated process of comparable complexity to that seen in eukaryotic cells. The dynamic assembly of Escherichia coli FtsZ in the presence of GTP is fundamental to its activity. FtsZ polymerization is a very attractive target for novel antibiotics given its fundamental and universal function. In this study our aim was to understand further the GTP-dependent FtsZ polymerization mechanism and our main focus is on the pH dependence of its behaviour. A key feature of this work is the use of linear dichroism (LD to follow the polymerization of FtsZ monomers into polymeric structures. LD is the differential absorption of light polarized parallel and perpendicular to an orientation direction (in this case that provided by shear flow. It thus readily distinguishes between FtsZ polymers and monomers. It also distinguishes FtsZ polymers and less well-defined aggregates, which light scattering methodologies do not. The polymerization of FtsZ over a range of pHs was studied by right-angled light scattering to probe mass of FtsZ structures, LD to probe real-time formation of linear polymeric fibres, a specially developed phosphate release assay to relate guanosine triphosphate (GTP hydrolysis to polymer formation, and electron microscopy (EM imaging of reaction products as a function of time and pH. We have found that lowering the pH from neutral to 6.5 does not change the nature of the FtsZ polymers in solution--it simply facilitates the polymerization so the fibres present are longer and more abundant. Conversely, lowering the pH to 6.0 has much the same effect as introducing divalent cations or the FtsZ-associated protein YgfE (a putative ZapA orthologue in E. coli--it stabilizes associations of protofilaments.

  17. Macromolecular interactions of the bacterial division FtsZ protein: from quantitative biochemistry and crowding to reconstructing minimal divisomes in the test tube.

    Science.gov (United States)

    Rivas, Germán; Alfonso, Carlos; Jiménez, Mercedes; Monterroso, Begoña; Zorrilla, Silvia

    2013-06-01

    The division of Escherichia coli is an essential process strictly regulated in time and space. It requires the association of FtsZ with other proteins to assemble a dynamic ring during septation, forming part of the functionally active division machinery, the divisome. FtsZ reversibly interacts with FtsA and ZipA at the cytoplasmic membrane to form a proto-ring, the first molecular assembly of the divisome, which is ultimately joined by the rest of the division-specific proteins. In this review we summarize the quantitative approaches used to study the activity, interactions, and assembly properties of FtsZ under well-defined solution conditions, with the aim of furthering our understanding of how the behavior of FtsZ is controlled by nucleotides and physiological ligands. The modulation of the association and assembly properties of FtsZ by excluded-volume effects, reproducing in part the natural crowded environment in which this protein has evolved to function, will be described. The subsequent studies on the reactivity of FtsZ in membrane-like systems using biochemical, biophysical, and imaging technologies are reported. Finally, we discuss the experimental challenges to be met to achieve construction of the minimum protein set needed to initiate bacterial division, without cells, in a cell-like compartment. This integrated approach, combining quantitative and synthetic strategies, will help to support (or dismiss) conclusions already derived from cellular and molecular analysis and to complete our understanding on how bacterial division works.

  18. Context-dependent protein folding of a virulence peptide in the bacterial and host environments: structure of an SycH–YopH chaperone–effector complex

    International Nuclear Information System (INIS)

    Vujanac, Milos; Stebbins, C. Erec

    2013-01-01

    The structure of a SycH–YopH chaperone–effector complex from Yersinia reveals the bacterial state of a protein that adopts different folds in the host and pathogen environments. Yersinia pestis injects numerous bacterial proteins into host cells through an organic nanomachine called the type 3 secretion system. One such substrate is the tyrosine phosphatase YopH, which requires an interaction with a cognate chaperone in order to be effectively injected. Here, the first crystal structure of a SycH–YopH complex is reported, determined to 1.9 Å resolution. The structure reveals the presence of (i) a nonglobular polypeptide in YopH, (ii) a so-called β-motif in YopH and (iii) a conserved hydrophobic patch in SycH that recognizes the β-motif. Biochemical studies establish that the β-motif is critical to the stability of this complex. Finally, since previous work has shown that the N-terminal portion of YopH adopts a globular fold that is functional in the host cell, aspects of how this polypeptide adopts radically different folds in the host and in the bacterial environments are analysed

  19. Effects of dietary protein levels and 2-methylbutyrate on ruminal fermentation, nutrient degradability, bacterial populations and urinary purine derivatives in Simmental steers.

    Science.gov (United States)

    Wang, C; Liu, Q; Guo, G; Huo, W J; Pei, C X; Zhang, S L; Yang, W Z

    2018-06-01

    The objective of this study was to evaluate the effects of dietary crude protein (CP) levels and 2-methylbutyrate (MB) supplementation on ruminal fermentation, bacterial populations, microbial enzyme activity and urinary excretion of purine derivatives (PD) in Simmental steers. Eight ruminally cannulated Simmental steers, averaging 18 months of age and 465 ± 8.6 kg of body weight (BW), were used in a replicated 4 × 4 Latin square design by a 2 × 2 factorial arrangement. Low protein (98.5 g CP/kg dry matter [LP] or high protein (128.7 g CP/kg dry matter [HP]) diets were fed with MB supplementation (0 g [MB-] or 16.8 g steer -1  day -1 [MB+]). Steers were fed a total mixed ration with dietary corn straw to concentrate ratio of 50:50 (dry matter [DM] basis). The CP × MB interaction was observed for ruminal total VFA, molar proportions of acetate and propionate, acetate to propionate ratio, ammonia-N, effective degradability of neutral detergent fibre (NDF) and CP, microbial enzyme activity, bacterial populations and total PD excretion (p Ruminal pH decreased (p ruminal total VFA concentration increased (p Ruminal ammonia-N content increased (p = .034) with increasing dietary CP level, but decreased (p = .012) with MB supplementation. The effective degradability of NDF and CP increased (p ruminal fermentation, nutrient degradability, microbial enzyme activity, ruminal bacterial populations and microbial protein synthesis improved with increasing dietary CP level or MB supplementation in steers. © 2017 Blackwell Verlag GmbH.

  20. The Bacterial Flagellar Type III Export Gate Complex Is a Dual Fuel Engine That Can Use Both H+ and Na+ for Flagellar Protein Export.

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    Tohru Minamino

    2016-03-01

    Full Text Available The bacterial flagellar type III export apparatus utilizes ATP and proton motive force (PMF to transport flagellar proteins to the distal end of the growing flagellar structure for self-assembly. The transmembrane export gate complex is a H+-protein antiporter, of which activity is greatly augmented by an associated cytoplasmic ATPase complex. Here, we report that the export gate complex can use sodium motive force (SMF in addition to PMF across the cytoplasmic membrane to drive protein export. Protein export was considerably reduced in the absence of the ATPase complex and a pH gradient across the membrane, but Na+ increased it dramatically. Phenamil, a blocker of Na+ translocation, inhibited protein export. Overexpression of FlhA increased the intracellular Na+ concentration in the presence of 100 mM NaCl but not in its absence, suggesting that FlhA acts as a Na+ channel. In wild-type cells, however, neither Na+ nor phenamil affected protein export, indicating that the Na+ channel activity of FlhA is suppressed by the ATPase complex. We propose that the export gate by itself is a dual fuel engine that uses both PMF and SMF for protein export and that the ATPase complex switches this dual fuel engine into a PMF-driven export machinery to become much more robust against environmental changes in external pH and Na+ concentration.

  1. The Bacterial Flagellar Type III Export Gate Complex Is a Dual Fuel Engine That Can Use Both H+ and Na+ for Flagellar Protein Export.

    Science.gov (United States)

    Minamino, Tohru; Morimoto, Yusuke V; Hara, Noritaka; Aldridge, Phillip D; Namba, Keiichi

    2016-03-01

    The bacterial flagellar type III export apparatus utilizes ATP and proton motive force (PMF) to transport flagellar proteins to the distal end of the growing flagellar structure for self-assembly. The transmembrane export gate complex is a H+-protein antiporter, of which activity is greatly augmented by an associated cytoplasmic ATPase complex. Here, we report that the export gate complex can use sodium motive force (SMF) in addition to PMF across the cytoplasmic membrane to drive protein export. Protein export was considerably reduced in the absence of the ATPase complex and a pH gradient across the membrane, but Na+ increased it dramatically. Phenamil, a blocker of Na+ translocation, inhibited protein export. Overexpression of FlhA increased the intracellular Na+ concentration in the presence of 100 mM NaCl but not in its absence, suggesting that FlhA acts as a Na+ channel. In wild-type cells, however, neither Na+ nor phenamil affected protein export, indicating that the Na+ channel activity of FlhA is suppressed by the ATPase complex. We propose that the export gate by itself is a dual fuel engine that uses both PMF and SMF for protein export and that the ATPase complex switches this dual fuel engine into a PMF-driven export machinery to become much more robust against environmental changes in external pH and Na+ concentration.

  2. Gaseous ligand selectivity of the H-NOX sensor protein from Shewanella oneidensis and comparison to those of other bacterial H-NOXs and soluble guanylyl cyclase.

    Science.gov (United States)

    Wu, Gang; Liu, Wen; Berka, Vladimir; Tsai, Ah-Lim

    2017-09-01

    To delineate the commonalities and differences in gaseous ligand discrimination among the heme-based sensors with Heme Nitric oxide/OXygen binding protein (H-NOX) scaffold, the binding kinetic parameters for gaseous ligands NO, CO, and O 2 , including K D , k on , and k off , of Shewanella oneidensis H-NOX (So H-NOX) were characterized in detail in this study and compared to those of previously characterized H-NOXs from Clostridium botulinum (Cb H-NOX), Nostoc sp. (Ns H-NOX), Thermoanaerobacter tengcongensis (Tt H-NOX), Vibrio cholera (Vc H-NOX), and human soluble guanylyl cyclase (sGC), an H-NOX analogue. The K D (NO) and K D (CO) of each bacterial H-NOX or sGC follow the "sliding scale rule"; the affinities of the bacterial H-NOXs for NO and CO vary in a small range but stronger than those of sGC by at least two orders of magnitude. On the other hand, each bacterial H-NOX exhibits different characters in the stability of its 6c NO complex, reactivity with secondary NO, stability of oxyferrous heme and autoxidation to ferric heme. A facile access channel for gaseous ligands is also identified, implying that ligand access has only minimal effect on gaseous ligand selectivity of H-NOXs or sGC. This comparative study of the binding parameters of the bacterial H-NOXs and sGC provides a basis to guide future new structural and functional studies of each specific heme sensor with the H-NOX protein fold. Copyright © 2017 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

  3. DnaK as Antibiotic Target: Hot Spot Residues Analysis for Differential Inhibition of the Bacterial Protein in Comparison with the Human HSP70.

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    Federica Chiappori

    Full Text Available DnaK, the bacterial homolog of human Hsp70, plays an important role in pathogens survival under stress conditions, like antibiotic therapies. This chaperone sequesters protein aggregates accumulated in bacteria during antibiotic treatment reducing the effect of the cure. Although different classes of DnaK inhibitors have been already designed, they present low specificity. DnaK is highly conserved in prokaryotes (identity 50-70%, which encourages the development of a unique inhibitor for many different bacterial strains. We used the DnaK of Acinetobacter baumannii as representative for our analysis, since it is one of the most important opportunistic human pathogens, exhibits a significant drug resistance and it has the ability to survive in hospital environments. The E.coli DnaK was also included in the analysis as reference structure due to its wide diffusion. Unfortunately, bacterial DnaK and human Hsp70 have an elevated sequence similarity. Therefore, we performed a differential analysis of DnaK and Hsp70 residues to identify hot spots in bacterial proteins that are not present in the human homolog, with the aim of characterizing the key pharmacological features necessary to design selective inhibitors for DnaK. Different conformations of DnaK and Hsp70 bound to known inhibitor-peptides for DnaK, and ineffective for Hsp70, have been analysed by molecular dynamics simulations to identify residues displaying stable and selective interactions with these peptides. Results achieved in this work show that there are some residues that can be used to build selective inhibitors for DnaK, which should be ineffective for the human Hsp70.

  4. Identification of a novel calcium binding motif based on the detection of sequence insertions in the animal peroxidase domain of bacterial proteins.

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    Saray Santamaría-Hernando

    Full Text Available Proteins of the animal heme peroxidase (ANP superfamily differ greatly in size since they have either one or two catalytic domains that match profile PS50292. The orf PP_2561 of Pseudomonas putida KT2440 that we have called PepA encodes a two-domain ANP. The alignment of these domains with those of PepA homologues revealed a variable number of insertions with the consensus G-x-D-G-x-x-[GN]-[TN]-x-D-D. This motif has also been detected in the structure of pseudopilin (pdb 3G20, where it was found to be involved in Ca(2+ coordination although a sequence analysis did not reveal the presence of any known calcium binding motifs in this protein. Isothermal titration calorimetry revealed that a peptide containing this consensus motif bound specifically calcium ions with affinities ranging between 33-79 µM depending on the pH. Microcalorimetric titrations of the purified N-terminal ANP-like domain of PepA revealed Ca(2+ binding with a K(D of 12 µM and stoichiometry of 1.25 calcium ions per protein monomer. This domain exhibited peroxidase activity after its reconstitution with heme. These data led to the definition of a novel calcium binding motif that we have termed PERCAL and which was abundantly present in animal peroxidase-like domains of bacterial proteins. Bacterial heme peroxidases thus possess two different types of calcium binding motifs, namely PERCAL and the related hemolysin type calcium binding motif, with the latter being located outside the catalytic domains and in their C-terminal end. A phylogenetic tree of ANP-like catalytic domains of bacterial proteins with PERCAL motifs, including single domain peroxidases, was divided into two major clusters, representing domains with and without PERCAL motif containing insertions. We have verified that the recently reported classification of bacterial heme peroxidases in two families (cd09819 and cd09821 is unrelated to these insertions. Sequences matching PERCAL were detected in all kingdoms of

  5. Identification of a novel calcium binding motif based on the detection of sequence insertions in the animal peroxidase domain of bacterial proteins.

    Science.gov (United States)

    Santamaría-Hernando, Saray; Krell, Tino; Ramos-González, María-Isabel

    2012-01-01

    Proteins of the animal heme peroxidase (ANP) superfamily differ greatly in size since they have either one or two catalytic domains that match profile PS50292. The orf PP_2561 of Pseudomonas putida KT2440 that we have called PepA encodes a two-domain ANP. The alignment of these domains with those of PepA homologues revealed a variable number of insertions with the consensus G-x-D-G-x-x-[GN]-[TN]-x-D-D. This motif has also been detected in the structure of pseudopilin (pdb 3G20), where it was found to be involved in Ca(2+) coordination although a sequence analysis did not reveal the presence of any known calcium binding motifs in this protein. Isothermal titration calorimetry revealed that a peptide containing this consensus motif bound specifically calcium ions with affinities ranging between 33-79 µM depending on the pH. Microcalorimetric titrations of the purified N-terminal ANP-like domain of PepA revealed Ca(2+) binding with a K(D) of 12 µM and stoichiometry of 1.25 calcium ions per protein monomer. This domain exhibited peroxidase activity after its reconstitution with heme. These data led to the definition of a novel calcium binding motif that we have termed PERCAL and which was abundantly present in animal peroxidase-like domains of bacterial proteins. Bacterial heme peroxidases thus possess two different types of calcium binding motifs, namely PERCAL and the related hemolysin type calcium binding motif, with the latter being located outside the catalytic domains and in their C-terminal end. A phylogenetic tree of ANP-like catalytic domains of bacterial proteins with PERCAL motifs, including single domain peroxidases, was divided into two major clusters, representing domains with and without PERCAL motif containing insertions. We have verified that the recently reported classification of bacterial heme peroxidases in two families (cd09819 and cd09821) is unrelated to these insertions. Sequences matching PERCAL were detected in all kingdoms of life.

  6. Identification of two proteins that interact with the Erp virulence factor from Mycobacterium tuberculosis by using the bacterial two-hybrid system

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    Cataldi Angel A

    2009-01-01

    Full Text Available Abstract Background The exported repetitive protein (erp gene encodes a secreted 36-kDa protein with a central domain containing several proline-glycine-leucine-threonine-serine (PGLTS repeats. It has been demonstrated that erp is a virulence-associated factor since the disruption of this gene impairs the growth of Mycobacterium bovis and Mycobacterium tuberculosis in mice. Results In order to elucidate the function of Erp we searched for Erp-binding proteins from M. tuberculosis by using a bacterial two-hybrid system. Our results indicate that Erp interacts specifically with two putative membrane proteins, Rv1417 and Rv2617c. Further analysis revealed that the latter two interact with each other, indicating that Rv1417, Rv2617c and Erp are connected through multiple interactions. While Rv1417 is disseminated in several Actinomycetales genera, orthologues of Rv2617c are exclusively present in members of the M. tuberculosis complex (MTC. The central and amino-terminal regions of Erp were determined to be involved in the interaction with Rv1417 and Rv2627c. Erp forms from Mycobacterium smegmatis and Mycobacterium leprae were not able to interact with Rv2617c in two-hybrid assays. Immunolocalization experiments showed that Rv1417 and Rv2617c are found on the cell membrane and Erp on the bacterial cell wall. Finally, comparative genomics and expression studies revealed a possible role of Rv1417 in riboflavin metabolism. Conclusion We identified interactive partners of Erp, an M. tuberculosis protein involved in virulence, which will be the focus of future investigation to decipher the function of the Erp family protein.

  7. The TLR2 Antagonist Staphylococcal Superantigen-Like Protein 3 Acts as a Virulence Factor to Promote Bacterial Pathogenicity in vivo.

    Science.gov (United States)

    Koymans, Kirsten J; Goldmann, Oliver; Karlsson, Christofer A Q; Sital, Wiedjai; Thänert, Robert; Bisschop, Adinda; Vrieling, Manouk; Malmström, Johan; van Kessel, Kok P M; de Haas, Carla J C; van Strijp, Jos A G; Medina, Eva

    2017-01-01

    Toll-like receptor (TLR) signaling is important in the initiation of immune responses and subsequent instigation of adaptive immunity. TLR2 recognizes bacterial lipoproteins and plays a central role in the host defense against bacterial infections, including those caused by Staphylococcus aureus. Many studies have demonstrated the importance of TLR2 in murine S. aureus infection. S. aureus evades TLR2 activation by secreting two proteins, staphylococcal superantigen-like protein 3 (SSL3) and 4 (SSL4). In this study, we demonstrate that antibodies against SSL3 and SSL4 are found in healthy individuals, indicating that humans are exposed to these proteins during S. aureus colonization or infection. To investigate the TLR2-antagonistic properties of SSL3 and SSL4, we compared the infection with wild-type and SSL3/4 knockout S. aureus strains in an intravenous murine infection model. Direct evaluation of the contribution of SSL3/4 to infection pathogenesis was hindered by the fact that the SSLs were not expressed in the murine system. To circumvent this limitation, an SSL3-overproducing strain (pLukM-SSL3) was generated, resulting in constitutive expression of SSL3. pLukM-SSL3 exhibited increased virulence compared to the parental strain in a murine model that was found to be TLR2 dependent. Altogether, these data indicate that SSL3 contributes to S. aureus virulence in vivo. © 2017 S. Karger AG, Basel.

  8. Rab11-family of interacting protein 2 associates with chlamydial inclusions through its Rab-binding domain and promotes bacterial multiplication.

    Science.gov (United States)

    Leiva, Natalia; Capmany, Anahí; Damiani, María Teresa

    2013-01-01

    Chlamydia trachomatis, an obligate intracellular pathogen, survives within host cells in a special compartment named 'inclusion' and takes advantage of host vesicular transport pathways for its growth and replication. Rab GTPases are key regulatory proteins of intracellular trafficking. Several Rabs, among them Rab11 and Rab14, are implicated in chlamydial development. FIP2, a member of the Rab11-Family of Interacting Proteins, presents at the C-terminus a Rab-binding domain that interacts with both Rab11 and Rab14. In this study, we determined and characterized the recruitment of endogenous and GFP-tagged FIP2 to the chlamydial inclusions. The recruitment of FIP2 is specific since other members of the Rab11-Family of Interacting Proteins do not associate with the chlamydial inclusions. The Rab-binding domain of FIP2 is essential for its association. Our results indicate that FIP2 binds to Rab11 at the chlamydial inclusion membrane through its Rab-binding domain. The presence of FIP2 at the chlamydial inclusion favours the recruitment of Rab14. Furthermore, our results show that FIP2 promotes inclusion development and bacterial replication. In agreement, the silencing of FIP2 decreases the bacterial progeny. C. trachomatis likely recruits FIP2 to hijack host intracellular trafficking to redirect vesicles full of nutrients towards the inclusion. © 2012 Blackwell Publishing Ltd.

  9. Steady-state levels of G-protein beta-subunit expression are regulated by treatment of cells with bacterial toxins

    International Nuclear Information System (INIS)

    Watkins, D.C.; Northup, J.K.; Malbon, C.C.

    1987-01-01

    Cultures of 3T3-L1 cells were incubated with either 10 ng/ml cholera toxin or 10 ng/ml pertussis toxin from 4 days prior to the initiation of differentiation and throughout the subsequent incubation. Toxin concentrations were sufficient to completely prevent the labelling of alpha-subunits with [ 32 P]NAD + and pertussis toxin and to prevent by more than 90% the labelling with [ 32 P]NAD + and cholera toxin in membranes prepared from these cells. Neither toxin prevented the differentiation to the adipocyte phenotype. Neither toxin prevented the increases in the relative amounts of G-proteins which occur upon differentiation. Both toxins dramatically decreased the amount of beta-subunits. As measured by quantitative immunoblotting with antisera specific for both the 35 kDa and 36 kDa beta-subunits, levels of beta-subunit were decreased by more than 50% of steady-state level of control cells. Thus, bacterial toxins which modifies G-protein alpha-subunits are capable of modulating the levels of beta-subunits in vivo. The basis for the regulation of G-protein subunit expression by bacterial toxins is under study

  10. Natural antigenic differences in the functionally equivalent extracellular DNABII proteins of bacterial biofilms provide a means for targeted biofilm therapeutics

    Science.gov (United States)

    Rocco, Christopher J.; Davey, Mary Ellen; Bakaletz, Lauren O.; Goodman, Steven D.

    2016-01-01

    SUMMARY Bacteria that persist in the oral cavity exist within complex biofilm communities. A hallmark of biofilms is the presence of an extracellular polymeric substance (EPS), which consists of polysaccharides, extracellular DNA (eDNA), and proteins, including the DNABII family of proteins. The removal of DNABII proteins from a biofilm results in the loss of structural integrity of the eDNA and the collapse of the biofilm structure. We examined the role of DNABII proteins in the biofilm structure of the periodontal pathogen Porphyromonas gingivalis and the oral commensal Streptococcus gordonii. Co-aggregation with oral streptococci is thought to facilitate the establishment of P. gingivalis within the biofilm community. We demonstrate that DNABII proteins are present in the EPS of both S. gordonii and P. gingivalis biofilms, and that these biofilms can be disrupted through the addition of antisera derived against their respective DNABII proteins. We provide evidence that both eDNA and DNABII proteins are limiting in S. gordonii but not in P. gingivalis biofilms. In addition, these proteins are capable of complementing one another functionally. We also found that while antisera derived against most DNABII proteins are capable of binding a wide variety of DNABII proteins, the P. gingivalis DNABII proteins are antigenically distinct. The presence of DNABII proteins in the EPS of these biofilms and the antigenic uniqueness of the P. gingivalis proteins provide an opportunity to develop therapies that are targeted to remove P. gingivalis and biofilms that contain P. gingivalis from the oral cavity. PMID:26988714

  11. Bacterial interactions with proteins and cells relevant to the development of life-threatening endocarditis studied by use of a quartz-crystal microbalance.

    Science.gov (United States)

    Krajewski, Stefanie; Rheinlaender, Johannes; Ries, Philip; Canjuga, Denis; Mack, Carmen; Scheideler, Lutz; Schäffer, Tilman E; Geis-Gerstorfer, Jürgen; Wendel, Hans-Peter; Rupp, Frank

    2014-05-01

    Implant-related infections are a major challenge in clinical routine because of severe complications, for example infective endocarditis (IE). The purpose of this study was to investigate the real-time interaction of S. gordonii with proteins and cells important in the development of IE, in a flow system, by use of a quartz-crystal microbalance (QCM). Acoustic sensors were biologically modified by preconditioning with sterile saliva, platelet-poor plasma (PPP), or platelet-rich plasma (PRP), followed then by perfusion of a bacterial suspension. After perfusion, additional fluorescence and scanning electron microscopic (SEM) studies were performed. The surface structure of S. gordonii was analyzed by atomic force microscopy (AFM). Compared with S. gordonii adhesion on the abiotic sensor surface following normal mass loading indicated by a frequency decrease, adhesion on saliva, PPP, or PRP-conditioned sensors resulted in an increase in frequency. Furthermore, adhesion induced slightly increased damping signals for saliva and PPP-coated sensors but a decrease upon bacterial adhesion to PRP, indicating the formation of a more rigid biofilm. Microscopic analysis confirmed the formation of dense and vital bacterial layers and the aggregation of platelets and bacteria. In conclusion, our study shows that the complex patterns of QCM output data observed are strongly dependent on the biological substrate and adhesion mechanisms of S. gordonii. Overall, QCM sheds new light on the pathways of such severe infections as IE.

  12. The structure of BVU2987 from Bacteroides vulgatus reveals a superfamily of bacterial periplasmic proteins with possible inhibitory function

    International Nuclear Information System (INIS)

    Das, Debanu; Finn, Robert D.; Carlton, Dennis; Miller, Mitchell D.; Abdubek, Polat; Astakhova, Tamara; Axelrod, Herbert L.; Bakolitsa, Constantina; Chen, Connie; Chiu, Hsiu-Ju; Chiu, Michelle; Clayton, Thomas; Deller, Marc C.; Duan, Lian; Ellrott, Kyle; Ernst, Dustin; Farr, Carol L.; Feuerhelm, Julie; Grant, Joanna C.; Grzechnik, Anna; Han, Gye Won; Jaroszewski, Lukasz; Jin, Kevin K.; Klock, Heath E.; Knuth, Mark W.; Kozbial, Piotr; Krishna, S. Sri; Kumar, Abhinav; Marciano, David; McMullan, Daniel; Morse, Andrew T.; Nigoghossian, Edward; Nopakun, Amanda; Okach, Linda; Puckett, Christina; Reyes, Ron; Rife, Christopher L.; Sefcovic, Natasha; Tien, Henry J.; Trame, Christine B.; Bedem, Henry van den; Weekes, Dana; Wooten, Tiffany; Xu, Qingping; Hodgson, Keith O.; Wooley, John; Elsliger, Marc-André; Deacon, Ashley M.; Godzik, Adam; Lesley, Scott A.; Wilson, Ian A.

    2010-01-01

    The crystal structure of the BVU2987 gene product from B. vulgatus (UniProt A6L4L1) reveals that members of the new Pfam family PF11396 (domain of unknown function; DUF2874) are similar to β-lactamase inhibitor protein and YpmB. Proteins that contain the DUF2874 domain constitute a new Pfam family PF11396. Members of this family have predominantly been identified in microbes found in the human gut and oral cavity. The crystal structure of one member of this family, BVU2987 from Bacteroides vulgatus, has been determined, revealing a β-lactamase inhibitor protein-like structure with a tandem repeat of domains. Sequence analysis and structural comparisons reveal that BVU2987 and other DUF2874 proteins are related to β-lactamase inhibitor protein, PepSY and SmpA-OmlA proteins and hence are likely to function as inhibitory proteins

  13. Intramammary Immunization of Pregnant Mice with Staphylococcal Protein A Reduces the Post-Challenge Mammary Gland Bacterial Load but Not Pathology.

    Directory of Open Access Journals (Sweden)

    Jully Gogoi-Tiwari

    Full Text Available Protein A, encoded by the spa gene, is one of the major immune evading MSCRAMM of S. aureus, demonstrated to be prevalent in a significant percentage of clinical bovine mastitis isolates in Australia. Given its' reported significance in biofilm formation and the superior performance of S. aureus biofilm versus planktonic vaccine in the mouse mastitis model, it was of interest to determine the immunogenicity and protective potential of Protein A as a potential vaccine candidate against bovine mastitis using the mouse mastitis model. Pregnant Balb/c mice were immunised with Protein A emulsified in an alum-based adjuvant by subcutaneous (s/c or intramammary (i/mam routes. While humoral immune response of mice post-immunization were determined using indirect ELISA, cell-mediated immune response was assessed by estimation of interferon-gamma (IFN-γ produced by protein A-stimulated splenocyte supernatants. Protective potential of Protein A against experimental mastitis was determined by challenge of immunized versus sham-vaccinated mice by i/mam route, based upon manifestation of clinical symptoms, total bacterial load and histopathological damage to mammary glands. Significantly (p<0.05 higher levels of IgG1 isotype were produced in mice immunized by the s/c route. In contrast, significantly higher levels of the antibody isotype IgG2a were produced in mice immunized by the i/mam route (p<0.05. There was significant reduction (p<0.05 in bacterial loads of the mammary glands of mice immunized by Protein A regardless of the route of immunization, with medium level of clinical symptoms observed up to day 3 post-challenge. However, Protein A vaccine failed to protect immunized mice post-challenge with biofilm producing encapsulated S. aureus via i/mam route, regardless of the route of immunization, as measured by the level of mammary tissue damage. It was concluded that, Protein A in its' native state was apparently not a suitable candidate for inclusion

  14. Addressing the Immunogenicity of the Cargo and of the Targeting Antibodies with a Focus on Deimmunized Bacterial Toxins and on Antibody-Targeted Human Effector Proteins

    Science.gov (United States)

    Grinberg, Yehudit; Benhar, Itai

    2017-01-01

    Third-generation immunotoxins are composed of a human, or humanized, targeting moiety, usually a monoclonal antibody or an antibody fragment, and a non-human effector molecule. Due to the non-human origin of the cytotoxic domain, these molecules stimulate potent anti-drug immune responses, which limit treatment options. Efforts are made to deimmunize such immunotoxins or to combine treatment with immunosuppression. An alternative approach is using the so-called “human cytotoxic fusion proteins”, in which antibodies are used to target human effector proteins. Here, we present three relevant approaches for reducing the immunogenicity of antibody-targeted protein therapeutics: (1) reducing the immunogenicity of the bacterial toxin, (2) fusing human cytokines to antibodies to generate immunocytokines and (3) addressing the immunogenicity of the targeting antibodies. PMID:28574434

  15. SCM, a novel M-like protein from Streptococcus canis, binds (mini)-plasminogen with high affinity and facilitates bacterial transmigration.

    Science.gov (United States)

    Fulde, Marcus; Rohde, Manfred; Hitzmann, Angela; Preissner, Klaus T; Nitsche-Schmitz, D Patric; Nerlich, Andreas; Chhatwal, Gursharan Singh; Bergmann, Simone

    2011-03-15

    Streptococcus canis is an important zoonotic pathogen capable of causing serious invasive diseases in domestic animals and humans. In the present paper we report the binding of human plasminogen to S. canis and the recruitment of proteolytically active plasmin on its surface. The binding receptor for plasminogen was identified as a novel M-like protein designated SCM (S. canis M-like protein). SPR (surface plasmon resonance) analyses, radioactive dot-blot analyses and heterologous expression on the surface of Streptococcus gordonii confirmed the plasminogen-binding capability of SCM. The binding domain was located within the N-terminus of SCM, which specifically bound to the C-terminal part of plasminogen (mini-plasminogen) comprising kringle domain 5 and the catalytic domain. In the presence of urokinase, SCM mediated plasminogen activation on the bacterial surface that was inhibited by serine protease inhibitors and lysine amino acid analogues. Surface-bound plasmin effectively degraded purified fibrinogen as well as fibrin clots, resulting in the dissolution of fibrin thrombi. Electron microscopic illustration and time-lapse imaging demonstrated bacterial transmigration through fibrinous thrombi. The present study has led, for the first time, to the identification of SCM as a novel receptor for (mini)-plasminogen mediating the fibrinolytic activity of S. canis.

  16. A lower isoelectric point increases signal sequence-mediated secretion of recombinant proteins through a bacterial ABC transporter.

    Science.gov (United States)

    Byun, Hyunjong; Park, Jiyeon; Kim, Sun Chang; Ahn, Jung Hoon

    2017-12-01

    Efficient protein production for industrial and academic purposes often involves engineering microorganisms to produce and secrete target proteins into the culture. Pseudomonas fluorescens has a TliDEF ATP-binding cassette transporter, a type I secretion system, which recognizes C-terminal LARD3 signal sequence of thermostable lipase TliA. Many proteins are secreted by TliDEF in vivo when recombined with LARD3, but there are still others that cannot be secreted by TliDEF even when LARD3 is attached. However, the factors that determine whether or not a recombinant protein can be secreted through TliDEF are still unknown. Here, we recombined LARD3 with several proteins and examined their secretion through TliDEF. We found that the proteins secreted via LARD3 are highly negatively charged with highly-acidic isoelectric points (pI) lower than 5.5. Attaching oligo-aspartate to lower the pI of negatively-charged recombinant proteins improved their secretion, and attaching oligo-arginine to negatively-charged proteins blocked their secretion by LARD3. In addition, negatively supercharged green fluorescent protein (GFP) showed improved secretion, whereas positively supercharged GFP did not secrete. These results disclosed that proteins' acidic pI and net negative charge are major factors that determine their secretion through TliDEF. Homology modeling for TliDEF revealed that TliD dimer forms evolutionarily-conserved positively-charged clusters in its pore and substrate entrance site, which also partially explains the pI dependence of the TliDEF-dependent secretions. In conclusion, lowering the isoelectric point improved LARD3-mediated protein secretion, both widening the range of protein targets for efficient production via secretion and signifying an important aspect of ABC transporter-mediated secretions. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  17. Bacterial Actins.

    Science.gov (United States)

    Izoré, Thierry; van den Ent, Fusinita

    2017-01-01

    A diverse set of protein polymers, structurally related to actin filaments contributes to the organization of bacterial cells as cytomotive or cytoskeletal filaments. This chapter describes actin homologs encoded by bacterial chromosomes. MamK filaments, unique to magnetotactic bacteria, help establishing magnetic biological compasses by interacting with magnetosomes. Magnetosomes are intracellular membrane invaginations containing biomineralized crystals of iron oxide that are positioned by MamK along the long-axis of the cell. FtsA is widespread across bacteria and it is one of the earliest components of the divisome to arrive at midcell, where it anchors the cell division machinery to the membrane. FtsA binds directly to FtsZ filaments and to the membrane through its C-terminus. FtsA shows altered domain architecture when compared to the canonical actin fold. FtsA's subdomain 1C replaces subdomain 1B of other members of the actin family and is located on the opposite side of the molecule. Nevertheless, when FtsA assembles into protofilaments, the protofilament structure is preserved, as subdomain 1C replaces subdomain IB of the following subunit in a canonical actin filament. MreB has an essential role in shape-maintenance of most rod-shaped bacteria. Unusually, MreB filaments assemble from two protofilaments in a flat and antiparallel arrangement. This non-polar architecture implies that both MreB filament ends are structurally identical. MreB filaments bind directly to membranes where they interact with both cytosolic and membrane proteins, thereby forming a key component of the elongasome. MreB filaments in cells are short and dynamic, moving around the long axis of rod-shaped cells, sensing curvature of the membrane and being implicated in peptidoglycan synthesis.

  18. Small proteins in cyanobacteria provide a paradigm for the functional analysis of the bacterial micro-proteome.

    Science.gov (United States)

    Baumgartner, Desiree; Kopf, Matthias; Klähn, Stephan; Steglich, Claudia; Hess, Wolfgang R

    2016-11-28

    Despite their versatile functions in multimeric protein complexes, in the modification of enzymatic activities, intercellular communication or regulatory processes, proteins shorter than 80 amino acids (μ-proteins) are a systematically underestimated class of gene products in bacteria. Photosynthetic cyanobacteria provide a paradigm for small protein functions due to extensive work on the photosynthetic apparatus that led to the functional characterization of 19 small proteins of less than 50 amino acids. In analogy, previously unstudied small ORFs with similar degrees of conservation might encode small proteins of high relevance also in other functional contexts. Here we used comparative transcriptomic information available for two model cyanobacteria, Synechocystis sp. PCC 6803 and Synechocystis sp. PCC 6714 for the prediction of small ORFs. We found 293 transcriptional units containing candidate small ORFs ≤80 codons in Synechocystis sp. PCC 6803, also including the known mRNAs encoding small proteins of the photosynthetic apparatus. From these transcriptional units, 146 are shared between the two strains, 42 are shared with the higher plant Arabidopsis thaliana and 25 with E. coli. To verify the existence of the respective μ-proteins in vivo, we selected five genes as examples to which a FLAG tag sequence was added and re-introduced them into Synechocystis sp. PCC 6803. These were the previously annotated gene ssr1169, two newly defined genes norf1 and norf4, as well as nsiR6 (nitrogen stress-induced RNA 6) and hliR1(high light-inducible RNA 1) , which originally were considered non-coding. Upon activation of expression via the Cu 2+. responsive petE promoter or from the native promoters, all five proteins were detected in Western blot experiments. The distribution and conservation of these five genes as well as their regulation of expression and the physico-chemical properties of the encoded proteins underline the likely great bandwidth of small protein

  19. The major outer membrane proteins of enterobacteriaceae. Their immunological relatedness and their possible role in bacterial opsonization

    NARCIS (Netherlands)

    Hofstra, Harmen

    1981-01-01

    This thesis deals with immunological investigations of the major outer membrane proteins of the Enterobacteriaceae as a new group of enterobacterial common envelope antigens, and with some aspects of the possible role of antibodies, prepared against these proteins, in host defense mechanisms. ...

  20. Monocyte CD64 or CD89 targeting by surfactant protein D/anti-Fc receptor mediates bacterial uptake.

    NARCIS (Netherlands)

    Tacken, P.J.; Batenburg, J.J.

    2006-01-01

    We recently showed that a chimeric protein, consisting of a recombinant fragment of human surfactant protein D (rfSP-D) coupled to a Fab' fragment directed against the human Fcalpha receptor (CD89), effectively targets pathogens recognized by SP-D to human neutrophils. The present study evaluates

  1. Bacterial CpG-DNA activates dendritic cells in vivo: T helper cell-independent cytotoxic T cell responses to soluble proteins.

    Science.gov (United States)

    Sparwasser, T; Vabulas, R M; Villmow, B; Lipford, G B; Wagner, H

    2000-12-01

    Receptors for conserved molecular patterns associated with microbial pathogens induce synthesis of co-stimulatory molecules and cytokines in immature dendritic cells (DC), as do antigen-reactive CD4 T helper cells via CD40 signaling. Once activated, antigen-presenting DC may activate CD8 T cell responses in a T helper cell-independent fashion. Using immunostimulatory CpG-oligonucleotides (ODN) mimicking bacterial CpG-DNA, we tested whether CpG-DNA bypasses the need for T helper cells in CTL responses towards proteins by directly activating antigen-presenting DC to transit into professional APC. We describe that immature DC in situ constitutively process soluble proteins and generate CD8 T cell determinants yet CD8 T cell responses remain abortive. Induction of primary antigen-specific CD8 cytotoxic T lymphocyte (CTL)-mediated responses becomes initiated in wild-type as well as T helper cell-deficient mice, provided soluble protein and CpG-ODN are draining into the same lymph node. Specifically we show that CpG-ODN trigger antigen-presenting immature DC within the draining lymph node to acutely up-regulate co-stimulatory molecules and produce IL-12. These results provide new insights for generating in vivo efficient CTL responses to soluble proteins which may influence vaccination strategies.

  2. Detailed analysis of RNA-protein interactions within the bacterial ribosomal protein L5/5S rRNA complex.

    Science.gov (United States)

    Perederina, Anna; Nevskaya, Natalia; Nikonov, Oleg; Nikulin, Alexei; Dumas, Philippe; Yao, Min; Tanaka, Isao; Garber, Maria; Gongadze, George; Nikonov, Stanislav

    2002-12-01

    The crystal structure of ribosomal protein L5 from Thermus thermophilus complexed with a 34-nt fragment comprising helix III and loop C of Escherichia coli 5S rRNA has been determined at 2.5 A resolution. The protein specifically interacts with the bulged nucleotides at the top of loop C of 5S rRNA. The rRNA and protein contact surfaces are strongly stabilized by intramolecular interactions. Charged and polar atoms forming the network of conserved intermolecular hydrogen bonds are located in two narrow planar parallel layers belonging to the protein and rRNA, respectively. The regions, including these atoms conserved in Bacteria and Archaea, can be considered an RNA-protein recognition module. Comparison of the T. thermophilus L5 structure in the RNA-bound form with the isolated Bacillus stearothermophilus L5 structure shows that the RNA-recognition module on the protein surface does not undergo significant changes upon RNA binding. In the crystal of the complex, the protein interacts with another RNA molecule in the asymmetric unit through the beta-sheet concave surface. This protein/RNA interface simulates the interaction of L5 with 23S rRNA observed in the Haloarcula marismortui 50S ribosomal subunit.

  3. Accuracy of C - Reactive protein as a bacterial infection marker in critically immunosuppressed patients: A systematic review and meta-analysis.

    Science.gov (United States)

    de Oliveira, Vanessa Martins; Moraes, Rafael Barberena; Stein, Airton Tetelbom; Wendland, Eliana Márcia

    2017-12-01

    There is a need for a better understanding of the role of C-reactive protein (CRP) as a valid marker for the detection of bacterial infections in critically immunosuppressed patients. A high negative predictive value of CRP is also needed to rule out sepsis and bacterial infections in immunocompetent patients. However, few studies have evaluated the performance of CRP in immunocompromised hosts. The aim of the present study was to evaluate the performance of CRP as a marker of infection in critically immunosuppressed patients. The inclusion criterion was immunosuppression for which CRP was used as a bacterial infection marker. Searches were performed in the Cochrane Register, MEDLINE, EMBASE, SCOPUS, Web OF Science, LILACS and CINAHL databases. We applied the Quality Assessment of Diagnostic Accuracy Studies tool 2 (QUADAS 2) to evaluate the quality of the articles and evaluated the test accuracy parameters using hierarchical summary receiver operating characteristic (HSROC) curves and bivariate random effect models. Only 13 of 21 studies produced quantitative results. We analyzed all studies using the random effects method (restricted maximum likelihood) and obtained a joint diagnostic odds ratio (DOR) of 3.04 (95% confidence interval [CI] 1.71-5.40) with heterogeneity (I 2 =91%, Q=181.48, p<0.001). Therefore, a bivariate model was applied. Analyzing the tuberculosis carrier, steroid user, or presence of opportunistic infection subgroups, as described in the proposal, was not possible due to the lack of information on these topics included in the articles. CRP appears to be a good screening tool for sepsis in critically immunosuppressed patients. Submitted PROSPERO 2015: CRD42015019329. Copyright © 2017 Elsevier Inc. All rights reserved.

  4. Bacterial motility complexes require the actin-like protein, MreB and the Ras homologue, MglA.

    Science.gov (United States)

    Mauriello, Emilia M F; Mouhamar, Fabrice; Nan, Beiyan; Ducret, Adrien; Dai, David; Zusman, David R; Mignot, Tâm

    2010-01-20

    Gliding motility in the bacterium Myxococcus xanthus uses two motility engines: S-motility powered by type-IV pili and A-motility powered by uncharacterized motor proteins and focal adhesion complexes. In this paper, we identified MreB, an actin-like protein, and MglA, a small GTPase of the Ras superfamily, as essential for both motility systems. A22, an inhibitor of MreB cytoskeleton assembly, reversibly inhibited S- and A-motility, causing rapid dispersal of S- and A-motility protein clusters, FrzS and AglZ. This suggests that the MreB cytoskeleton is involved in directing the positioning of these proteins. We also found that a DeltamglA motility mutant showed defective localization of AglZ and FrzS clusters. Interestingly, MglA-YFP localization mimicked both FrzS and AglZ patterns and was perturbed by A22 treatment, consistent with results indicating that both MglA and MreB bind to motility complexes. We propose that MglA and the MreB cytoskeleton act together in a pathway to localize motility proteins such as AglZ and FrzS to assemble the A-motility machineries. Interestingly, M. xanthus motility systems, like eukaryotic systems, use an actin-like protein and a small GTPase spatial regulator.

  5. On the Spatial Organization of mRNA, Plasmids, and Ribosomes in a Bacterial Host Overexpressing Membrane Proteins.

    Directory of Open Access Journals (Sweden)

    Lieke A van Gijtenbeek

    2016-12-01

    Full Text Available By using fluorescence imaging, we provide a time-resolved single-cell view on coupled defects in transcription, translation, and growth during expression of heterologous membrane proteins in Lactococcus lactis. Transcripts encoding poorly produced membrane proteins accumulate in mRNA-dense bodies at the cell poles, whereas transcripts of a well-expressed homologous membrane protein show membrane-proximal localization in a translation-dependent fashion. The presence of the aberrant polar mRNA foci correlates with cessation of cell division, which is restored once these bodies are cleared. In addition, activation of the heat-shock response and a loss of nucleoid-occluded ribosomes are observed. We show that the presence of a native-like N-terminal domain is key to SRP-dependent membrane localization and successful production of membrane proteins. The work presented gives new insights and detailed understanding of aberrant membrane protein biogenesis, which can be used for strategies to optimize membrane protein production.

  6. Bacterial meningitis

    NARCIS (Netherlands)

    Roos, Karen L.; van de Beek, Diederik

    2010-01-01

    Bacterial meningitis is a neurological emergency. Empiric antimicrobial and adjunctive therapy should be initiated as soon as a single set of blood cultures has been obtained. Clinical signs suggestive of bacterial meningitis include fever, headache, meningismus, vomiting, photophobia, and an

  7. Detailed analysis of RNA-protein interactions within the bacterial ribosomal protein L5/5S rRNA complex.

    OpenAIRE

    Perederina, Anna; Nevskaya, Natalia; Nikonov, Oleg; Nikulin, Alexei; Dumas, Philippe; Yao, Min; Tanaka, Isao; Garber, Maria; Gongadze, George; Nikonov, Stanislav

    2002-01-01

    The crystal structure of ribosomal protein L5 from Thermus thermophilus complexed with a 34-nt fragment comprising helix III and loop C of Escherichia coli 5S rRNA has been determined at 2.5 A resolution. The protein specifically interacts with the bulged nucleotides at the top of loop C of 5S rRNA. The rRNA and protein contact surfaces are strongly stabilized by intramolecular interactions. Charged and polar atoms forming the network of conserved intermolecular hydrogen bonds are located in ...

  8. Streptococcus mutans autolysin AtlA is a fibronectin-binding protein and contributes to bacterial survival in the bloodstream and virulence for infective endocarditis.

    Science.gov (United States)

    Jung, Chiau-Jing; Zheng, Quan-Hau; Shieh, Ya-Hsiung; Lin, Chi-Shuan; Chia, Jean-San

    2009-11-01

    Streptococcus mutans, a commensal of the human oral cavity, can survive in the bloodstream and cause infective endocarditis (IE). However, the virulence factors associated with this manifestation of disease are not known. Here, we demonstrate that AtlA, an autolysin of S. mutans is a newly identified fibronectin (Fn) binding protein and contributes to bacterial resistance to phagocytosis and survival in the bloodstream. Interestingly, prior exposure to plasma at low concentrations was sufficient to enhance bacterial survival in the circulation. Calcium ions at physiological plasma concentrations induced maturation of AtlA from the 104-90 kDa isoform resulting in increased Fn binding and resistance to phagocytosis. An isogenic mutant strain defective in AtlA expression exhibited reduced survival and virulence when tested in a rat model of IE compared with the wild-type and complemented strains. The data presented suggest that plasma components utilized by S. mutans enhanced survival in the circulation and AtlA is a virulence factor associated with infective endocarditis.

  9. Anti-protozoal and anti-bacterial antibiotics that inhibit protein synthesis kill cancer subtypes enriched for stem cell-like properties.

    Science.gov (United States)

    Cuyàs, Elisabet; Martin-Castillo, Begoña; Corominas-Faja, Bruna; Massaguer, Anna; Bosch-Barrera, Joaquim; Menendez, Javier A

    2015-01-01

    Key players in translational regulation such as ribosomes might represent powerful, but hitherto largely unexplored, targets to eliminate drug-refractory cancer stem cells (CSCs). A recent study by the Lisanti group has documented how puromycin, an old antibiotic derived from Streptomyces alboniger that inhibits ribosomal protein translation, can efficiently suppress CSC states in tumorspheres and monolayer cultures. We have used a closely related approach based on Biolog Phenotype Microarrays (PM), which contain tens of lyophilized antimicrobial drugs, to assess the chemosensitivity profiles of breast cancer cell lines enriched for stem cell-like properties. Antibiotics directly targeting active sites of the ribosome including emetine, puromycin and cycloheximide, inhibitors of ribosome biogenesis such as dactinomycin, ribotoxic stress agents such as daunorubicin, and indirect inhibitors of protein synthesis such as acriflavine, had the largest cytotoxic impact against claudin-low and basal-like breast cancer cells. Thus, biologically aggressive, treatment-resistant breast cancer subtypes enriched for stem cell-like properties exhibit exacerbated chemosensitivities to anti-protozoal and anti-bacterial antibiotics targeting protein synthesis. These results suggest that old/existing microbicides might be repurposed not only as new cancer therapeutics, but also might provide the tools and molecular understanding needed to develop second-generation inhibitors of ribosomal translation to eradicate CSC traits in tumor tissues.

  10. Structural Characterization and Oligomerization of the TssL Protein, a Component Shared by Bacterial Type VI and Type IVb Secretion Systems*

    Science.gov (United States)

    Durand, Eric; Zoued, Abdelrahim; Spinelli, Silvia; Watson, Paul J. H.; Aschtgen, Marie-Stéphanie; Journet, Laure; Cambillau, Christian; Cascales, Eric

    2012-01-01

    The Type VI secretion system (T6SS) is a macromolecular system distributed in Gram-negative bacteria, responsible for the secretion of effector proteins into target cells. The T6SS has a broad versatility as it can target both eukaryotic and prokaryotic cells. It is therefore involved in host pathogenesis or killing neighboring bacterial cells to colonize a new niche. At the architecture level, the T6SS core apparatus is composed of 13 proteins, which assemble in two subcomplexes. One of these subcomplexes, composed of subunits that share structural similarities with bacteriophage tail and baseplate components, is anchored to the cell envelope by the membrane subcomplex. This latter is constituted of at least three proteins, TssL, TssM, and TssJ. The crystal structure of the TssJ outer membrane lipoprotein and its interaction with the inner membrane TssM protein have been recently reported. TssL and TssM share sequence homology and characteristics with two components of the Type IVb secretion system (T4bSS), IcmH/DotU and IcmF, respectively. In this study, we report the crystal structure of the cytoplasmic domain of the TssL inner membrane protein from the enteroaggregative Escherichia coli Sci-1 T6SS. It folds as a hook-like structure composed of two three-helix bundles. Two TssL molecules associate to form a functional complex. Although the TssL trans-membrane segment is the main determinant of self-interaction, contacts between the cytoplasmic domains are required for TssL function. Based on sequence homology and secondary structure prediction, we propose that the TssL structure is the prototype for the members of the TssL and IcmH/DotU families. PMID:22371492

  11. Interaction of Prevotella intermedia strain 17 leucine-rich repeat domain protein AdpF with eukaryotic cells promotes bacterial internalization.

    Science.gov (United States)

    Sengupta, Dipanwita; Kang, Dae-Joong; Anaya-Bergman, Cecilia; Wyant, Tiana; Ghosh, Arnab K; Miyazaki, Hiroshi; Lewis, Janina P

    2014-06-01

    Prevotella intermedia is an oral bacterium implicated in a variety of oral diseases. Although internalization of this bacterium by nonphagocytic host cells is well established, the molecular players mediating the process are not well known. Here, the properties of a leucine-rich repeat (LRR) domain protein, designated AdpF, are described. This protein contains a leucine-rich region composed of 663 amino acid residues, and molecular modeling shows that it folds into a classical curved solenoid structure. The cell surface localization of recombinant AdpF (rAdpF) was confirmed by electron and confocal microscopy analyses. The recombinant form of this protein bound fibronectin in a dose-dependent manner. Furthermore, the protein was internalized by host cells, with the majority of the process accomplished within 30 min. The internalization of rAdpF was inhibited by nystatin, cytochalasin, latrunculin, nocodazole, and wortmannin, indicating that microtubules, microfilaments, and signal transduction are required for the invasion. It is noteworthy that preincubation of eukaryotic cells with AdpF increased P. intermedia 17 internalization by 5- and 10-fold for HeLa and NIH 3T3 fibroblast cell lines, respectively. The addition of the rAdpF protein was also very effective in inducing bacterial internalization into the oral epithelial cell line HN4, as well as into primary cells, including human oral keratinocytes (HOKs) and human umbilical vein endothelial cells (HUVECs). Finally, cells exposed to P. intermedia 17 internalized the bacteria more readily upon reinfection. Taken together, our data demonstrate that rAdpF plays a role in the internalization of P. intermedia 17 by a variety of host cells.

  12. Jasmonate induction of the monoterpene linalool confers resistance to rice bacterial blight and its biosynthesis is regulated by JAZ protein in rice.

    Science.gov (United States)

    Taniguchi, Shiduku; Hosokawa-Shinonaga, Yumi; Tamaoki, Daisuke; Yamada, Shoko; Akimitsu, Kazuya; Gomi, Kenji

    2014-02-01

    Jasmonic acid (JA) is involved in the regulation of host immunity in plants. Recently, we demonstrated that JA signalling has an important role in resistance to rice bacterial blight caused by Xanthomonas oryzae pv. oryzae (Xoo) in rice. Here, we report that many volatile compounds accumulate in response to exogenous application of JA, including the monoterpene linalool. Expression of linalool synthase was up-regulated by JA. Vapour treatment with linalool induced resistance to Xoo, and transgenic rice plants overexpressing linalool synthase were more resistance to Xoo, presumably due to the up-regulation of defence-related genes in the absence of any treatment. JA-induced accumulation of linalool was regulated by OsJAZ8, a rice jasmonate ZIM-domain protein involving the JA signalling pathway at the transcriptional level, suggesting that linalool plays an important role in JA-induced resistance to Xoo in rice. © 2013 John Wiley & Sons Ltd.

  13. Bacterial membrane proteomics.

    Science.gov (United States)

    Poetsch, Ansgar; Wolters, Dirk

    2008-10-01

    About one quarter to one third of all bacterial genes encode proteins of the inner or outer bacterial membrane. These proteins perform essential physiological functions, such as the import or export of metabolites, the homeostasis of metal ions, the extrusion of toxic substances or antibiotics, and the generation or conversion of energy. The last years have witnessed completion of a plethora of whole-genome sequences of bacteria important for biotechnology or medicine, which is the foundation for proteome and other functional genome analyses. In this review, we discuss the challenges in membrane proteome analysis, starting from sample preparation and leading to MS-data analysis and quantification. The current state of available proteomics technologies as well as their advantages and disadvantages will be described with a focus on shotgun proteomics. Then, we will briefly introduce the most abundant proteins and protein families present in bacterial membranes before bacterial membrane proteomics studies of the last years will be presented. It will be shown how these works enlarged our knowledge about the physiological adaptations that take place in bacteria during fine chemical production, bioremediation, protein overexpression, and during infections. Furthermore, several examples from literature demonstrate the suitability of membrane proteomics for the identification of antigens and different pathogenic strains, as well as the elucidation of membrane protein structure and function.

  14. Xylo-oligosaccharides and inulin affect genotoxicity and bacterial populations differently in a human colonic simulator challenged with soy protein

    DEFF Research Database (Denmark)

    Christophersen, C. T.; Petersen, Anne; Licht, Tine Rask

    2013-01-01

    High dietary intakes of some protein sources, including soy protein, can increase colonic DNA damage in animals, whereas some carbohydrates attenuate this. We investigated whether inulin and xylo-oligosaccharides (XOS) could be protective against DNA strand breaks by adding them to a human colonic...... cornstarch for 10 day followed by soy protein with 1% XOS or 1% inulin for 10 day. Inulin did not alter genotoxicity but XOS significantly reduced PV genotoxicity and increased DV genotoxicity. Inulin and XOS significantly increased butyrate concentration in the DV but not PV. Numbers of the key butyrate......-producing bacterium Faecalibacterium prausnitzii were significantly increased in the PV and DV by inulin but significantly decreased by XOS in both vessels. Other bacteria examined were also significantly impacted by the carbohydrate treatments or by the vessel (i.e., pH). There was a significant overall inverse...

  15. Bacterial incorporation of leucine into protein down to -20 degrees C with evidence for potential activity in sub-eutectic saline ice formations.

    Science.gov (United States)

    Junge, Karen; Eicken, Hajo; Swanson, Brian D; Deming, Jody W

    2006-06-01

    Direct evidence for metabolism in a variety of frozen environments has pushed temperature limits for bacterial activity to increasingly lower temperatures, so far to -20 degrees C. To date, the metabolic activities of marine psychrophilic bacteria, important components of sea-ice communities, have not been studied in laboratory culture, not in ice and not below -12 degrees C. We measured [3H]-leucine incorporation into macromolecules (further fractionated biochemically) by the marine psychrophilic bacterium Colwellia psychrerythraea strain 34H over a range of anticipated activity-permissive temperatures, from +13 to -20 degrees C, including expected negative controls at -80 and -196 degrees C. For incubation temperatures below -1 degrees C, the cell suspensions [all in artificial seawater (ASW)] were first quick-frozen in liquid nitrogen. We also examined the effect of added extracellular polymeric substances (EPS) on [3H]-leucine incorporation. Results showed that live cells of strain 34H incorporated substantial amounts of [3H]-leucine into TCA-precipitable material (primarily protein) down to -20 degrees C. At temperatures from -1 to -20 degrees C, rates were enhanced by EPS. No activity was detected in the killed controls for strain 34H (or in Escherichia coli controls), which included TCA-killed, heat-killed, and sodium azide- and chloramphenicol-treated samples. Surprisingly, evidence for low but significant rates of intracellular incorporation of [3H]-leucine into protein was observed for both ASW-only and EPS-amended (and live only) samples incubated at -80 and -196 degrees C. Mechanisms that could explain the latter results require further study, but the process of vitrification promoted by rapid freezing and the presence of salts and organic polymers may be relevant. Overall, distinguishing between intracellular and extracellular aspects of bacterial activity appears important to understanding behavior at sub-freezing temperatures.

  16. Influence of dietary protein and fructooligosaccharides on fecal fermentative end-products, fecal bacterial populations and apparent total tract digestibility in dogs.

    Science.gov (United States)

    Pinna, Carlo; Vecchiato, Carla Giuditta; Bolduan, Carmen; Grandi, Monica; Stefanelli, Claudio; Windisch, Wilhelm; Zaghini, Giuliano; Biagi, Giacomo

    2018-03-20

    Feeding dogs with diets rich in protein may favor putrefactive fermentations in the hindgut, negatively affecting the animal's intestinal environment. Conversely, prebiotics may improve the activity of health-promoting bacteria and prevent bacterial proteolysis in the colon. The aim of this study was to evaluate the effects of dietary supplementation with fructooligosaccharides (FOS) on fecal microbiota and apparent total tract digestibility (ATTD) in dogs fed kibbles differing in protein content. Twelve healthy adult dogs were used in a 4 × 4 replicated Latin Square design to determine the effects of four diets: 1) Low protein diet (LP, crude protein (CP) 229 g/kg dry matter (DM)); 2) High protein diet (HP, CP 304 g/kg DM); 3) Diet 1 + 1.5 g of FOS/kg; 4) Diet 2 + 1.5 g of FOS/kg. The diets contained silica at 5 g/kg as a digestion marker. Differences in protein content were obtained using different amounts of a highly digestible swine greaves meal. Each feeding period lasted 28 d, with a 12 d wash-out in between periods. Fecal samples were collected from dogs at 0, 21 and 28 d of each feeding period. Feces excreted during the last five days of each feeding period were collected and pooled in order to evaluate ATTD. Higher fecal ammonia concentrations were observed both when dogs received the HP diets (p < 0.001) and the supplementation with FOS (p < 0.05). The diets containing FOS resulted in greater ATTD of DM, Ca, Mg, Na, Zn, and Fe (p < 0.05) while HP diets were characterized by lower crude ash ATTD (p < 0.05). Significant interactions were observed between FOS and protein concentration in regards to fecal pH (p < 0.05), propionic acid (p < 0.05), acetic to propionic acid and acetic + n-butyric to propionic acid ratios (p < 0.01), bifidobacteria (p < 0.05) and ATTD of CP (p < 0.05) and Mn (p < 0.001). A relatively moderate increase of dietary protein resulted in higher concentrations of ammonia in

  17. Molecular organization in bacterial cell membranes. Specific labelling and topological distribution of glycoproteins and proteins in Streptomyces albus membranes

    Energy Technology Data Exchange (ETDEWEB)

    Larraga, V; Munoz, E [Consejo Superior de Investigaciones Cientificas, Madrid (Spain). Instituto de Biologia Celular

    1975-05-01

    The paper reports about an investigation into the question of the specific labelling and topological distribution of glycoproteins and proteins in Streptomyces albus membranes. The method of sample preparation is described: Tritium labelling of glycoproteins in protoplasts and membranes, iodination of proteins, trypsin treatment and polyacrylamide gel electrophoresis. The findings suggest an asymmetrical distribution of the glycoproteins in membranes and a weak accessibility to iodine label. A structural model of the plasma membranes of Streptomyces albus is proposed similar to the general 'fluid mosaic' model of Singer and Nicholson.

  18. Effects of corn-based diet starch content and corn particle size on lactation performance, digestibility, and bacterial protein flow in dairy cows.

    Science.gov (United States)

    Fredin, S M; Ferraretto, L F; Akins, M S; Bertics, S J; Shaver, R D

    2015-01-01

    An experiment was conducted to determine the effects of dietary starch content in corn-based diets and corn particle size on lactation performance, nutrient digestibility, and bacterial protein flow in dairy cows using the omasal and reticular sampling technique. Eight ruminally cannulated lactating multiparous Holstein cows were used in a replicated 4×4 Latin square design with a 2×2 factorial arrangement of treatments. Treatments were fine (FG; mean particle size=552µm) and coarse (CG; 1,270µm) ground dry shelled corn in normal- (NS) and reduced- (RS) starch diets fed as total mixed rations. The NS and RS rations contained 27 and 18% starch (dry matter basis), respectively, and were formulated by partially replacing corn with soy hull pellets. Mean dry matter intake was unaffected by treatment (23.2kg/d). Cows fed NS diets produced 1.9kg/d more milk and 0.06kg/d more milk protein compared with cows fed RS diets. Cows fed NSFG and RSCG diets produced more fat-corrected milk than did cows fed NSCG and RSFG diets. Milk urea concentration was decreased for cows fed NS diets (12.4mg/dL) compared with RS diets (13.5mg/dL). Ruminal digestibility of neutral detergent fiber (NDF; % of NDF intake) determined by the omasal sampling technique was increased in cows fed RS diets compared with NS diets (43.4 vs. 34.9%), and total-tract digestibility of NDF (% of NDF intake) was increased in cows fed RS diets compared with those fed NS diets (50.1 vs. 43.1%). Ruminal digestibility of starch (% of starch intake) determined by the omasal sampling technique was greater in cows fed NS diets compared with those fed RS diets (85.6 vs. 81.6%). Total-tract starch digestion was increased in cows fed RS diets compared with those fed NS diets (96.9 vs. 94.6%) and in cows fed FG diets compared with those fed CG diets (98.0 vs. 93.5%). Bacterial protein flow was unaffected by treatment. The omasal and reticular sampling techniques resulted in similar treatment effects for nutrient flow

  19. Evaluation of spectral libraries and sample preparation for DIA-LC-MS analysis of host cell proteins: A case study of a bacterially expressed recombinant biopharmaceutical protein.

    Science.gov (United States)

    Heissel, Søren; Bunkenborg, Jakob; Kristiansen, Max Per; Holmbjerg, Anne Fich; Grimstrup, Marie; Mørtz, Ejvind; Kofoed, Thomas; Højrup, Peter

    2018-07-01

    Recombinantly expressed biopharmaceutical proteins often undergo a series of purification steps with the aim of removing contaminating material. Depending on the application of the protein, there are various requirements for the degree of purity, but host cell proteins (HCPs) will in general remain in small amounts. LC-MS has emerged as an orthogonal technique, capable of providing detailed information regarding the individual proteins. The aim of this case study was to characterize the HCPs associated with a biopharmaceutical protein, provided by Statens Serum Institut (DK), which is used in the field of tuberculosis and has not previously been studied by LC-MS. The developed method and acquired experiences served to develop a generalized strategy for HCP-characterization in our laboratory. We evaluated the use of different spectral libraries, recorded in data-dependent mode for obtaining the highest HCP coverage, combined with SWATH-based absolute quantification. The accuracy of two label-free absolute quantification strategies was evaluated using stable isotope peptides. Two different sample preparation workflows were evaluated for optimal HCP yield. . The label-free strategy produced accurate quantification across several orders of magnitude, and the calculated purity was found to be in agreement with previously obtained ELISA data. Copyright © 2018 Elsevier Inc. All rights reserved.

  20. Comparison between medium-chain acyl-CoA dehydrogenase mutant proteins overexpressed in bacterial and mammalian cells

    DEFF Research Database (Denmark)

    Jensen, T G; Bross, P; Andresen, B S

    1995-01-01

    ." Upon expression in E. coli, these mutant proteins produce activity levels in the range of the wild-type enzyme only if the chaperonins GroESL are co-overproduced. When overexpressed in COS cells, the pure folding mutants display enzyme activities comparable to the wild-type enzyme. The results suggest...

  1. Bacterial expression and one-step purification of an isotope-labeled heterotrimeric G-protein {alpha}-subunit

    Energy Technology Data Exchange (ETDEWEB)

    Abdulaev, Najmoutin G. [University of Maryland Biotechnology Institute, Center for Advanced Research in Biotechnology (United States); Zhang Cheng; Dinh, Andy [University of Texas Health Science Center, Center for Membrane Biology, Department of Biochemistry and Molecular Biology (United States); Ngo, Tony; Bryan, Philip N. [University of Maryland Biotechnology Institute, Center for Advanced Research in Biotechnology (United States); Brabazon, Danielle M. [Loyola College in Maryland, Department of Chemistry (United States); Marino, John P. [University of Maryland Biotechnology Institute, Center for Advanced Research in Biotechnology (United States)], E-mail: marino@carb.nist.gov; Ridge, Kevin D. [University of Texas Health Science Center, Center for Membrane Biology, Department of Biochemistry and Molecular Biology (United States)

    2005-05-15

    Heterologous expression systems are often employed to generate sufficient quantities of isotope-labeled proteins for high-resolution NMR studies. Recently, the interaction between the prodomain region of subtilisin and an active, mutant form of the mature enzyme has been exploited to develop a cleavable affinity tag fusion system for one-step generation and purification of full-length soluble proteins obtained by inducible prokaryotic expression. As a first step towards applying high-resolution NMR methods to study heterotrimeric G-protein {alpha}-subunit (G{sub {alpha}}) conformation and dynamics, the utility of this subtilisin prodomain fusion system for expressing and purifying an isotope-labeled G{sub {alpha}} chimera ({approx}40 kDa polypeptide) has been tested. The results show that a prodomain fused G{sub {alpha}} chimera can be expressed to levels approaching 6-8 mg/l in minimal media and that the processed, mature protein exhibits properties similar to those of G{sub {alpha}} isolated from natural sources. To assay for the functional integrity of the purified G{sub {alpha}} chimera at NMR concentrations and probe for changes in the structure and dynamics of G{sub {alpha}} that result from activation, {sup 15}N-HSQC spectra of the GDP/Mg{sup 2+} bound form of G{sub {alpha}} obtained in the absence and presence of aluminum fluoride, a well known activator of the GDP bound state, have been acquired. Comparisons of the {sup 15}N-HSQC spectra reveals a number of changes in chemical shifts of the {sup 1}HN, {sup 15}N crosspeaks that are discussed with respect to expected changes in the protein conformation associated with G{sub {alpha}} activation.

  2. Bacterial expression and one-step purification of an isotope-labeled heterotrimeric G-protein α-subunit

    International Nuclear Information System (INIS)

    Abdulaev, Najmoutin G.; Zhang Cheng; Dinh, Andy; Ngo, Tony; Bryan, Philip N.; Brabazon, Danielle M.; Marino, John P.; Ridge, Kevin D.

    2005-01-01

    Heterologous expression systems are often employed to generate sufficient quantities of isotope-labeled proteins for high-resolution NMR studies. Recently, the interaction between the prodomain region of subtilisin and an active, mutant form of the mature enzyme has been exploited to develop a cleavable affinity tag fusion system for one-step generation and purification of full-length soluble proteins obtained by inducible prokaryotic expression. As a first step towards applying high-resolution NMR methods to study heterotrimeric G-protein α-subunit (G α ) conformation and dynamics, the utility of this subtilisin prodomain fusion system for expressing and purifying an isotope-labeled G α chimera (∼40 kDa polypeptide) has been tested. The results show that a prodomain fused G α chimera can be expressed to levels approaching 6-8 mg/l in minimal media and that the processed, mature protein exhibits properties similar to those of G α isolated from natural sources. To assay for the functional integrity of the purified G α chimera at NMR concentrations and probe for changes in the structure and dynamics of G α that result from activation, 15 N-HSQC spectra of the GDP/Mg 2+ bound form of G α obtained in the absence and presence of aluminum fluoride, a well known activator of the GDP bound state, have been acquired. Comparisons of the 15 N-HSQC spectra reveals a number of changes in chemical shifts of the 1 HN, 15 N crosspeaks that are discussed with respect to expected changes in the protein conformation associated with G α activation

  3. Regulatory Phosphorylation of Bacterial-Type PEP Carboxylase by the Ca2+-Dependent Protein Kinase RcCDPK1 in Developing Castor Oil Seeds.

    Science.gov (United States)

    Ying, Sheng; Hill, Allyson T; Pyc, Michal; Anderson, Erin M; Snedden, Wayne A; Mullen, Robert T; She, Yi-Min; Plaxton, William C

    2017-06-01

    Phosphoenolpyruvate carboxylase (PEPC) is a tightly controlled cytosolic enzyme situated at a crucial branch point of central plant metabolism. In developing castor oil seeds ( Ricinus communis ) a novel, allosterically desensitized 910-kD Class-2 PEPC hetero-octameric complex, arises from a tight interaction between 107-kD plant-type PEPC and 118-kD bacterial-type (BTPC) subunits. The native Ca 2+ -dependent protein kinase (CDPK) responsible for in vivo inhibitory phosphorylation of Class-2 PEPC's BTPC subunit's at Ser-451 was highly purified from COS and identified as RcCDPK1 (XP_002526815) by mass spectrometry. Heterologously expressed RcCDPK1 catalyzed Ca 2+ -dependent, inhibitory phosphorylation of BTPC at Ser-451 while exhibiting: ( i ) a pair of Ca 2+ binding sites with identical dissociation constants of 5.03 μM, ( ii ) a Ca 2+ -dependent electrophoretic mobility shift, and ( iii ) a marked Ca 2+ -independent hydrophobicity. Pull-down experiments established the Ca 2+ -dependent interaction of N-terminal GST-tagged RcCDPK1 with BTPC. RcCDPK1-Cherry localized to the cytosol and nucleus of tobacco bright yellow-2 cells, but colocalized with mitochondrial-surface associated BTPC-enhanced yellow fluorescent protein when both fusion proteins were coexpressed. Deletion analyses demonstrated that although its N-terminal variable domain plays an essential role in optimizing Ca 2+ -dependent RcCDPK1 autophosphorylation and BTPC transphosphorylation activity, it is not critical for in vitro or in vivo target recognition. Arabidopsis ( Arabidopsis thaliana ) CPK4 and soybean ( Glycine max ) CDPKβ are RcCDPK1 orthologs that effectively phosphorylated castor BTPC at Ser-451. Overall, the results highlight a potential link between cytosolic Ca 2+ signaling and the posttranslational control of respiratory CO 2 refixation and anaplerotic photosynthate partitioning in support of storage oil and protein biosynthesis in developing COS. © 2017 American Society of Plant

  4. Regulatory Phosphorylation of Bacterial-Type PEP Carboxylase by the Ca2+-Dependent Protein Kinase RcCDPK1 in Developing Castor Oil Seeds1[OPEN

    Science.gov (United States)

    Hill, Allyson T.; Anderson, Erin M.; She, Yi-Min

    2017-01-01

    Phosphoenolpyruvate carboxylase (PEPC) is a tightly controlled cytosolic enzyme situated at a crucial branch point of central plant metabolism. In developing castor oil seeds (Ricinus communis) a novel, allosterically desensitized 910-kD Class-2 PEPC hetero-octameric complex, arises from a tight interaction between 107-kD plant-type PEPC and 118-kD bacterial-type (BTPC) subunits. The native Ca2+-dependent protein kinase (CDPK) responsible for in vivo inhibitory phosphorylation of Class-2 PEPC’s BTPC subunit’s at Ser-451 was highly purified from COS and identified as RcCDPK1 (XP_002526815) by mass spectrometry. Heterologously expressed RcCDPK1 catalyzed Ca2+-dependent, inhibitory phosphorylation of BTPC at Ser-451 while exhibiting: (i) a pair of Ca2+ binding sites with identical dissociation constants of 5.03 μM, (ii) a Ca2+-dependent electrophoretic mobility shift, and (iii) a marked Ca2+-independent hydrophobicity. Pull-down experiments established the Ca2+-dependent interaction of N-terminal GST-tagged RcCDPK1 with BTPC. RcCDPK1-Cherry localized to the cytosol and nucleus of tobacco bright yellow-2 cells, but colocalized with mitochondrial-surface associated BTPC-enhanced yellow fluorescent protein when both fusion proteins were coexpressed. Deletion analyses demonstrated that although its N-terminal variable domain plays an essential role in optimizing Ca2+-dependent RcCDPK1 autophosphorylation and BTPC transphosphorylation activity, it is not critical for in vitro or in vivo target recognition. Arabidopsis (Arabidopsis thaliana) CPK4 and soybean (Glycine max) CDPKβ are RcCDPK1 orthologs that effectively phosphorylated castor BTPC at Ser-451. Overall, the results highlight a potential link between cytosolic Ca2+ signaling and the posttranslational control of respiratory CO2 refixation and anaplerotic photosynthate partitioning in support of storage oil and protein biosynthesis in developing COS. PMID:28363991

  5. Targets of DNA-binding proteins in bacterial promoter regions present enhanced probabilities for spontaneous thermal openings

    International Nuclear Information System (INIS)

    Apostolaki, Angeliki; Kalosakas, George

    2011-01-01

    We mapped promoter regions of double-stranded DNA with respect to the probabilities of appearance of relatively large bubble openings exclusively due to thermal fluctuations at physiological temperatures. We analyzed five well-studied promoter regions of procaryotic type and found a spatial correlation between the binding sites of transcription factors and the position of peaks in the probability pattern of large thermal openings. Other distinct peaks of the calculated patterns correlate with potential binding sites of DNA-binding proteins. These results suggest that a DNA molecule would more frequently expose the bases that participate in contacts with proteins, which would probably enhance the probability of the latter to reach their targets. It also stands for using this method as a means to analyze DNA sequences based on their intrinsic thermal properties

  6. The small G-protein MglA connects to the MreB actin cytoskeleton at bacterial focal adhesions.

    Science.gov (United States)

    Treuner-Lange, Anke; Macia, Eric; Guzzo, Mathilde; Hot, Edina; Faure, Laura M; Jakobczak, Beata; Espinosa, Leon; Alcor, Damien; Ducret, Adrien; Keilberg, Daniela; Castaing, Jean Philippe; Lacas Gervais, Sandra; Franco, Michel; Søgaard-Andersen, Lotte; Mignot, Tâm

    2015-07-20

    In Myxococcus xanthus the gliding motility machinery is assembled at the leading cell pole to form focal adhesions, translocated rearward to propel the cell, and disassembled at the lagging pole. We show that MglA, a Ras-like small G-protein, is an integral part of this machinery. In this function, MglA stimulates the assembly of the motility complex by directly connecting it to the MreB actin cytoskeleton. Because the nucleotide state of MglA is regulated spatially and MglA only binds MreB in the guanosine triphosphate-bound form, the motility complexes are assembled at the leading pole and dispersed at the lagging pole where the guanosine triphosphatase activating protein MglB disrupts the MglA-MreB interaction. Thus, MglA acts as a nucleotide-dependent molecular switch to regulate the motility machinery spatially. The function of MreB in motility is independent of its function in peptidoglycan synthesis, representing a coopted function. Our findings highlight a new function for the MreB cytoskeleton and suggest that G-protein-cytoskeleton interactions are a universally conserved feature. © 2015 Treuner-Lange et al.

  7. Protective effects of a bacterially expressed NIF-KGF fusion protein against bleomycin-induced acute lung injury in mice.

    Science.gov (United States)

    Li, Xinping; Li, Shengli; Zhang, Miaotao; Li, Xiukun; Zhang, Xiaoming; Zhang, Wenlong; Li, Chuanghong

    2010-08-01

    Current evidence suggests that the keratinocyte growth factor (KGF) and the polymorphonuclear leukocyte may play key roles in the development of lung fibrosis. Here we describe the construction, expression, purification, and identification of a novel NIF (neutrophil inhibitory factor)-KGF mutant fusion protein (NKM). The fusion gene was ligated via a flexible octapeptide hinge and expressed as an insoluble protein in Escherichia coli BL21 (DE3). The fusion protein retained the activities of KGF and NIF, as it inhibited both fibroblast proliferation and leukocyte adhesion. Next, the effects of NKM on bleomycin-induced lung fibrosis in mice were examined. The mice were divided into the following four groups: (i) saline group; (ii) bleomycin group (instilled with 5 mg/kg bleomycin intratracheally); (iii) bleomycin plus dexamethasone (Dex) group (Dex was given intraperitoneally (i.p.) at 1 mg/kg/day 2 days prior to bleomycin instillation and daily after bleomycin instillation until the end of the treatment); and (iv) bleomycin plus NKM group (NKM was given i.p. at 2 mg/kg/day using the same protocol as the Dex group). NKM significantly improved the survival rates of mice exposed to bleomycin. The marked morphological changes and increased hydroxyproline levels resulted from the instillation of bleomycin (on Day 17) in the lungs were significantly inhibited by NKM. These results revealed that NKM can attenuate bleomycin-induced lung fibrosis, suggesting that NKM could be used to prevent bleomycin-induced lung damage or other interstitial pulmonary fibrosis.

  8. The Hyr1 protein from the fungus Candida albicans is a cross kingdom immunotherapeutic target for Acinetobacter bacterial infection.

    Directory of Open Access Journals (Sweden)

    Priya Uppuluri

    2018-05-01

    Full Text Available Different pathogens share similar medical settings and rely on similar virulence strategies to cause infections. We have previously applied 3-D computational modeling and bioinformatics to discover novel antigens that target more than one human pathogen. Active and passive immunization with the recombinant N-terminus of Candida albicans Hyr1 (rHyr1p-N protect mice against lethal candidemia. Here we determine that Hyr1p shares homology with cell surface proteins of the multidrug resistant Gram negative bacterium, Acinetobacter baumannii including hemagglutinin (FhaB and outer membrane protein A (OmpA. The A. baumannii OmpA binds to C. albicans Hyr1p, leading to a mixed species biofilm. Deletion of HYR1, or blocking of Hyr1p using polyclonal antibodies, significantly reduce A. baumannii binding to C. albicans hyphae. Furthermore, active vaccination with rHyr1p-N or passive immunization with polyclonal antibodies raised against specific peptide motifs of rHyr1p-N markedly improve survival of diabetic or neutropenic mice infected with A. baumannii bacteremia or pneumonia. Antibody raised against one particular peptide of the rHyr1p-N sequence (peptide 5 confers majority of the protection through blocking A. baumannii invasion of host cells and inducing death of the bacterium by a putative iron starvation mechanism. Anti-Hyr1 peptide 5 antibodies also mitigate A. baumannii /C. albicans mixed biofilm formation in vitro. Consistent with our bioinformatic analysis and structural modeling of Hyr1p, anti-Hyr1p peptide 5 antibodies bound to A. baumannii FhaB, OmpA, and an outer membrane siderophore binding protein. Our studies highlight the concept of cross-kingdom vaccine protection against high priority human pathogens such as A. baumannii and C. albicans that share similar ecological niches in immunocompromised patients.

  9. Studies of the structure and function of Mms6, a bacterial protein that promotes the formation of magnetic nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Lijun [Iowa State Univ., Ames, IA (United States)

    2011-01-01

    Here we report structural and functional studies of Mms6, a biomineralization protein that can promote the formation in vitro of magnetic nanoparticles with sizes and morphologies similar to the magnetites synthesized by magnetotactic bacteria. We found the binding pattern of Mms6 to ferric ion to be two-phase and multivalent. We quantatively determined that Mms6 binds one Fe3+ with a very high affinity (Kd = 1016 M). The second phase of iron binding is multivalent and cooperative with respect to iron with a Kd in the μM range and a stoichiometry of about 20 ferric ion per protein molecule. We found that Mms6 exists in large particles of two sizes, one consisting of 20-40 monomeric units and the other of 200 units. From proteolytic digestion, ultracentrifugation and liposome fusion studies, we found that Mms6 forms a large micellar quaternary structure with the N-terminal domain self-assembling into a uniformly sized micelle and the C-terminal domain on the surface. The two-phase iron-binding pattern may be relevant to iron crystal formation. We propose that the first high affinity phase may stabilize a new conformation of the C-terminal domain that allows interaction with other C-terminal domains leading to a structural change in the multimeric protein complex that enables the second low affinity iron binding phase to organize iron and initiate crystal formation. We also observed a dimeric apparent molecular mass of the Mms6 C-terminal peptide (C21Mms6). We speculate that the C-terminal domain may form higher order quaternary arrangements on the surface of the micelle or when anchored to a membrane by the N-terminal domain. The change in fluorescence quenching in the N-terminal domain with iron binding suggests a structural integrity between the C- and N-terminal domains. The slow change in trp fluorescence as a function of time after adding iron suggests a very slow conformational change in the protein that involves

  10. TaCPK2-A, a calcium-dependent protein kinase gene that is required for wheat powdery mildew resistance enhances bacterial blight resistance in transgenic rice.

    Science.gov (United States)

    Geng, Shuaifeng; Li, Aili; Tang, Lichuan; Yin, Lingjie; Wu, Liang; Lei, Cailin; Guo, Xiuping; Zhang, Xin; Jiang, Guanghuai; Zhai, Wenxue; Wei, Yuming; Zheng, Youliang; Lan, Xiujin; Mao, Long

    2013-08-01

    Calcium-dependent protein kinases (CPKs) are important Ca2+ signalling components involved in complex immune and stress signalling networks; but the knowledge of CPK gene functions in the hexaploid wheat is limited. Previously, TaCPK2 was shown to be inducible by powdery mildew (Blumeria graminis tritici, Bgt) infection in wheat. Here, its functions in disease resistance are characterized further. This study shows the presence of defence-response and cold-response cis-elements on the promoters of the A subgenome homoeologue (TaCPK2-A) and D subgenome homoeologue (TaCPK2-D), respectively. Their expression patterns were then confirmed by quantitative real-time PCR (qRT-PCR) using genome-specific primers, where TaCPK2-A was induced by Bgt treatment while TaCPK2-D mainly responded to cold treatment. Downregulation of TaCPK2-A by virus-induced gene silencing (VIGS) causes loss of resistance to Bgt in resistant wheat lines, indicating that TaCPK2-A is required for powdery mildew resistance. Furthermore, overexpression of TaCPK2-A in rice enhanced bacterial blight (Xanthomonas oryzae pv. oryzae, Xoo) resistance. qRT-PCR analysis showed that overexpression of TaCPK2-A in rice promoted the expression of OsWRKY45-1, a transcription factor involved in both fungal and bacterial resistance by regulating jasmonic acid and salicylic acid signalling genes. The opposite effect was found in wheat TaCPK2-A VIGS plants, where the homologue of OsWRKY45-1 was significantly repressed. These data suggest that modulation of WRKY45-1 and associated defence-response genes by CPK2 genes may be the common mechanism for multiple disease resistance in grass species, which may have undergone subfunctionalization in promoters before the formation of hexaploid wheat.

  11. Structural and biochemical analyses indicate that a bacterial persulfide dioxygenase–rhodanese fusion protein functions in sulfur assimilation

    Energy Technology Data Exchange (ETDEWEB)

    Motl, Nicole; Skiba, Meredith A.; Kabil, Omer; Smith, Janet L.; Banerjee, Ruma

    2017-07-06

    Hydrogen sulfide (H2S) is a signaling molecule that is toxic at elevated concentrations. In eukaryotes, it is cleared via a mitochondrial sulfide oxidation pathway, which comprises sulfide quinone oxidoreductase, persulfide dioxygenase (PDO), rhodanese, and sulfite oxidase and converts H2S to thiosulfate and sulfate. Natural fusions between the non-heme iron containing PDO and rhodanese, a thiol sulfurtransferase, exist in some bacteria. However, little is known about the role of the PDO–rhodanese fusion (PRF) proteins in sulfur metabolism. Herein, we report the kinetic properties and the crystal structure of a PRF from the Gram-negative endophytic bacterium Burkholderia phytofirmans. The crystal structures of wild-type PRF and a sulfurtransferase-inactivated C314S mutant with and without glutathione were determined at 1.8, 2.4, and 2.7 Å resolution, respectively. We found that the two active sites are distant and do not show evidence of direct communication. The B. phytofirmans PRF exhibited robust PDO activity and preferentially catalyzed sulfur transfer in the direction of thiosulfate to sulfite and glutathione persulfide; sulfur transfer in the reverse direction was detectable only under limited turnover conditions. Together with the kinetic data, our bioinformatics analysis reveals that B. phytofirmans PRF is poised to metabolize thiosulfate to sulfite in a sulfur assimilation pathway rather than in sulfide stress response as seen, for example, with the Staphylococcus aureus PRF or sulfide oxidation and disposal as observed with the homologous mammalian proteins.

  12. The C-terminal domain of the bacterial SSB protein acts as a DNA maintenance hub at active chromosome replication forks.

    Directory of Open Access Journals (Sweden)

    Audrey Costes

    2010-12-01

    Full Text Available We have investigated in vivo the role of the carboxy-terminal domain of the Bacillus subtilis Single-Stranded DNA Binding protein (SSB(Cter as a recruitment platform at active chromosomal forks for many proteins of the genome maintenance machineries. We probed this SSB(Cter interactome using GFP fusions and by Tap-tag and biochemical analysis. It includes at least 12 proteins. The interactome was previously shown to include PriA, RecG, and RecQ and extended in this study by addition of DnaE, SbcC, RarA, RecJ, RecO, XseA, Ung, YpbB, and YrrC. Targeting of YpbB to active forks appears to depend on RecS, a RecQ paralogue, with which it forms a stable complex. Most of these SSB partners are conserved in bacteria, while others, such as the essential DNA polymerase DnaE, YrrC, and the YpbB/RecS complex, appear to be specific to B. subtilis. SSB(Cter deletion has a moderate impact on B. subtilis cell growth. However, it markedly affects the efficiency of repair of damaged genomic DNA and arrested replication forks. ssbΔCter mutant cells appear deficient in RecA loading on ssDNA, explaining their inefficiency in triggering the SOS response upon exposure to genotoxic agents. Together, our findings show that the bacterial SSB(Cter acts as a DNA maintenance hub at active chromosomal forks that secures their propagation along the genome.

  13. The pepper Bs4C proteins are localized to the endoplasmic reticulum (ER) membrane and confer disease resistance to bacterial blight in transgenic rice.

    Science.gov (United States)

    Wang, Jun; Zeng, Xuan; Tian, Dongsheng; Yang, Xiaobei; Wang, Lanlan; Yin, Zhongchao

    2018-03-30

    Transcription activator-like effector (TALE)-dependent dominant disease resistance (R) genes in plants, also referred to as executor R genes, are induced on infection by phytopathogenic bacteria of the genus Xanthomonas harbouring the corresponding TALE genes. Unlike the traditional R proteins, the executor R proteins do not determine the resistance specificity and may function broadly in different plant species. The executor R gene Bs4C-R in the resistant genotype PI 235047 of the pepper species Capsicum pubescens (CpBs4C-R) confers disease resistance to Xanthomonas campestris pv. vesicatoria (Xcv) harbouring the TALE genes avrBsP/avrBs4. In this study, the synthetic genes of CpBs4C-R and two other Bs4C-like genes, the susceptible allele in the genotype PI585270 of C. pubescens (CpBs4C-S) and the CaBs4C-R homologue gene in the cultivar 'CM334' of Capsicum annum (CaBs4C), were characterized in tobacco (Nicotiana benthamiana) and rice (Oryza sativa). The Bs4C genes induced cell death in N. benthamiana. The functional Bs4C-eCFP fusion proteins were localized to the endoplasmic reticulum (ER) membrane in the leaf epidermal cells of N. benthamiana. The Xa10 promoter-Bs4C fusion genes in transgenic rice conferred strain-specific disease resistance to Xanthomonas oryzae pv. oryzae (Xoo), the causal agent of bacterial blight in rice, and were specifically induced by the Xa10-incompatible Xoo strain PXO99 A (pHM1avrXa10). The results indicate that the Bs4C proteins from pepper species function broadly in rice and the Bs4C protein-mediated cell death from the ER is conserved between dicotyledonous and monocotyledonous plants, which can be utilized to engineer novel and enhanced disease resistance in heterologous plants. © 2018 TEMASEK LIFE SCIENCES LABORATORY. MOLECULAR PLANT PATHOLOGY © 2018 JOHN WILEY & SONS LTD.

  14. Contribution of the Collagen-Binding Proteins of Streptococcus mutans to Bacterial Colonization of Inflamed Dental Pulp.

    Science.gov (United States)

    Nomura, Ryota; Ogaya, Yuko; Nakano, Kazuhiko

    2016-01-01

    Streptococcus mutans is a major pathogen of dental caries. Collagen-binding proteins (CBPs) (approximately 120 kDa), termed Cnm and Cbm, are regarded as important cell surface antigens related to the adherence of S. mutans to collagenous tissue. Furthermore, CBP-positive S. mutans strains are associated with various systemic diseases involving bacteremia, such as infective endocarditis. Endodontic infection is considered to be an important cause of bacteremia, but little is known regarding the presence of S. mutans in dental pulp tissue. In the present study, the distribution and virulence of S. mutans in dental pulp tissues were investigated by focusing on CBPs. Adhesion and invasion properties of various S. mutans strains were analyzed using human dental pulp fibroblasts (HDPFs). CBP-positive strains had a significantly higher rate of adhesion to HDPFs compared with CBP-defective isogenic mutant strains (Ppulp. This could be a possible virulence factor for various systemic diseases.

  15. Pigments and proteins in green bacterial chlorosomes studied by matrix-assisted laser desorption ionization mass spectrometry

    DEFF Research Database (Denmark)

    Persson, S; Sönksen, C P; Frigaard, N U

    2000-01-01

    We have used matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) for mass determination of pigments and proteins in chlorosomes, the light-harvesting organelles from the photosynthetic green sulfur bacterium Chlorobium tepidum. By applying a small volume (1...... microL) of a concentrated suspension of isolated chlorosomes directly to the target of the mass spectrometer we have been able to detect bacteriochlorophyll a and all the major homologs of bacteriochlorophyll c. The peak heights of the different bacteriochlorophyll c homologs in the MALDI spectra were...... proportional to peak areas obtained from HPLC analysis of the same sample. The same result was also obtained when whole cells of Chl. tepidum were applied to the target, indicating that MALDI-MS can provide a rapid method for obtaining a semiquantitative determination or finger-print of the bacteriochlorophyll...

  16. Biosynthesis of Chlorophyll a in a Purple Bacterial Phototroph and Assembly into a Plant Chlorophyll-Protein Complex.

    Science.gov (United States)

    Hitchcock, Andrew; Jackson, Philip J; Chidgey, Jack W; Dickman, Mark J; Hunter, C Neil; Canniffe, Daniel P

    2016-09-16

    Improvements to photosynthetic efficiency could be achieved by manipulating pigment biosynthetic pathways of photosynthetic organisms in order to increase the spectral coverage for light absorption. The development of organisms that can produce both bacteriochlorophylls and chlorophylls is one way to achieve this aim, and accordingly we have engineered the bacteriochlorophyll-utilizing anoxygenic phototroph Rhodobacter sphaeroides to make chlorophyll a. Bacteriochlorophyll and chlorophyll share a common biosynthetic pathway up to the precursor chlorophyllide. Deletion of genes responsible for the bacteriochlorophyll-specific modifications of chlorophyllide and replacement of the native bacteriochlorophyll synthase with a cyanobacterial chlorophyll synthase resulted in the production of chlorophyll a. This pigment could be assembled in vivo into the plant water-soluble chlorophyll protein, heterologously produced in Rhodobacter sphaeroides, which represents a proof-of-principle for the engineering of novel antenna complexes that enhance the spectral range of photosynthesis.

  17. Characterization of foot-and-mouth disease virus gene products with antisera against bacterially synthesized fusion proteins

    International Nuclear Information System (INIS)

    Strebel, K.; Beck, E.; Strohmaier, K.; Schaller, H.

    1986-01-01

    Defined segments of the cloned foot-and-mouth disease virus genome corresponding to all parts of the coding region were expressed in Escherichia coli as fusions to the N-terminal part of the MS2-polymerase gene under the control of the inducible λPL promoter. All constructs yielded large amounts of proteins, which were purified and used to raise sequence-specific antisera in rabbits. These antisera were used to identify the corresponding viral gene products in 35 S-labeled extracts from foot-and-mouth disease virus-infected BHK cells. This allowed us to locate unequivocally all mature foot-and-mouth disease virus gene products in the nucleotide sequence, to identify precursor-product relationships, and to detect several foot-and mouth disease virus gene products not previously identified in vivo or in vitro

  18. Effects of corn-based diet starch content and neutral detergent fiber source on lactation performance, digestibility, and bacterial protein flow in dairy cows.

    Science.gov (United States)

    Fredin, S M; Akins, M S; Ferraretto, L F; Shaver, R D

    2015-01-01

    An experiment was conducted to evaluate the effects of corn-based dietary starch content and source of neutral detergent fiber (NDF) on lactation performance, nutrient digestion, bacterial protein flow, and ruminal parameters in lactating dairy cows. Eight ruminally cannulated multiparous Holstein cows averaging 193±11d in milk were randomly assigned to treatments in a replicated 4×4 Latin square design with 21-d periods. Treatment diets were high corn grain (HCG; 38% corn silage, 19% dry ground corn, and 4% soy hulls), high soy hulls (HSH; 38% corn silage, 11% dry ground corn, and 13% soy hulls), high corn silage (HCS; 50% corn silage, 6% dry ground corn, and 4% soy hulls), and low corn silage (LCS; 29% corn silage, 15% corn, and 19% soy hulls). The HCG, HSH, HCS, and LCS diets contained 29, 23, 24, and 22% starch; 27, 32, 30, and 32% total NDF; and 21, 21, 25, and 17% forage NDF (dry matter basis), respectively. Mean dry matter intake and milk yield were unaffected by treatment. Cows fed LCS had reduced milk fat content compared with HSH and HCS. The concentration of milk urea nitrogen was greater for cows fed HCS compared with the other treatments. Total-tract digestion of NDF was reduced for cows fed the HCG diet. Total-tract starch digestion was increased for cows fed the HSH and HCS compared with HCG and LCS diets. Bacterial protein flow was unaffected by treatment. Ruminal ammonia concentration was reduced in cows fed the HCG and LCS diets compared with the HCS diet. Ruminal propionate increased and the acetate:propionate ratio decreased in cows fed the LCS diet compared with the HCS diet. Ruminal pH was greater for cows fed the HCS diet compared with cows fed the LCS diet. Diet digestibility and performance of mid- to late-lactation cows fed reduced-starch diets by partially replacing corn grain with soy hulls or corn silage was similar to or improved compared with cows fed a normal-starch diet. Copyright © 2015 American Dairy Science Association

  19. Bacterial Cell Mechanics.

    Science.gov (United States)

    Auer, George K; Weibel, Douglas B

    2017-07-25

    Cellular mechanical properties play an integral role in bacterial survival and adaptation. Historically, the bacterial cell wall and, in particular, the layer of polymeric material called the peptidoglycan were the elements to which cell mechanics could be primarily attributed. Disrupting the biochemical machinery that assembles the peptidoglycan (e.g., using the β-lactam family of antibiotics) alters the structure of this material, leads to mechanical defects, and results in cell lysis. Decades after the discovery of peptidoglycan-synthesizing enzymes, the mechanisms that underlie their positioning and regulation are still not entirely understood. In addition, recent evidence suggests a diverse group of other biochemical elements influence bacterial cell mechanics, may be regulated by new cellular mechanisms, and may be triggered in different environmental contexts to enable cell adaptation and survival. This review summarizes the contributions that different biomolecular components of the cell wall (e.g., lipopolysaccharides, wall and lipoteichoic acids, lipid bilayers, peptidoglycan, and proteins) make to Gram-negative and Gram-positive bacterial cell mechanics. We discuss the contribution of individual proteins and macromolecular complexes in cell mechanics and the tools that make it possible to quantitatively decipher the biochemical machinery that contributes to bacterial cell mechanics. Advances in this area may provide insight into new biology and influence the development of antibacterial chemotherapies.

  20. Receptor density balances signal stimulation and attenuation in membrane-assembled complexes of bacterial chemotaxis signaling proteins

    Science.gov (United States)

    Besschetnova, Tatiana Y.; Montefusco, David J.; Asinas, Abdalin E.; Shrout, Anthony L.; Antommattei, Frances M.; Weis, Robert M.

    2008-01-01

    All cells possess transmembrane signaling systems that function in the environment of the lipid bilayer. In the Escherichia coli chemotaxis pathway, the binding of attractants to a two-dimensional array of receptors and signaling proteins simultaneously inhibits an associated kinase and stimulates receptor methylation—a slower process that restores kinase activity. These two opposing effects lead to robust adaptation toward stimuli through a physical mechanism that is not understood. Here, we provide evidence of a counterbalancing influence exerted by receptor density on kinase stimulation and receptor methylation. Receptor signaling complexes were reconstituted over a range of defined surface concentrations by using a template-directed assembly method, and the kinase and receptor methylation activities were measured. Kinase activity and methylation rates were both found to vary significantly with surface concentration—yet in opposite ways: samples prepared at high surface densities stimulated kinase activity more effectively than low-density samples, whereas lower surface densities produced greater methylation rates than higher densities. FRET experiments demonstrated that the cooperative change in kinase activity coincided with a change in the arrangement of the membrane-associated receptor domains. The counterbalancing influence of density on receptor methylation and kinase stimulation leads naturally to a model for signal regulation that is compatible with the known logic of the E. coli pathway. Density-dependent mechanisms are likely to be general and may operate when two or more membrane-related processes are influenced differently by the two-dimensional concentration of pathway elements. PMID:18711126

  1. Dimerization site 2 of the bacterial DNA-binding protein H-NS is required for gene silencing and stiffened nucleoprotein filament formation.

    Science.gov (United States)

    Yamanaka, Yuki; Winardhi, Ricksen S; Yamauchi, Erika; Nishiyama, So-Ichiro; Sowa, Yoshiyuki; Yan, Jie; Kawagishi, Ikuro; Ishihama, Akira; Yamamoto, Kaneyoshi

    2018-06-15

    The bacterial nucleoid-associated protein H-NS is a DNA-binding protein, playing a major role in gene regulation. To regulate transcription, H-NS silences genes, including horizontally acquired foreign genes. Escherichia coli H-NS is 137 residues long and consists of two discrete and independent structural domains: an N-terminal oligomerization domain and a C-terminal DNA-binding domain, joined by a flexible linker. The N-terminal oligomerization domain is composed of two dimerization sites, dimerization sites 1 and 2, which are both required for H-NS oligomerization, but the exact role of dimerization site 2 in gene silencing is unclear. To this end, we constructed a whole set of single amino acid substitution variants spanning residues 2 to 137. Using a well-characterized H-NS target, the slp promoter of the glutamic acid-dependent acid resistance (GAD) cluster promoters, we screened for any variants defective in gene silencing. Focusing on the function of dimerization site 2, we analyzed four variants, I70C/I70A and L75C/L75A, which all could actively bind DNA but are defective in gene silencing. Atomic force microscopy analysis of DNA-H-NS complexes revealed that all of these four variants formed condensed complexes on DNA, whereas WT H-NS formed rigid and extended nucleoprotein filaments, a conformation required for gene silencing. Single-molecule stretching experiments confirmed that the four variants had lost the ability to form stiffened filaments. We conclude that dimerization site 2 of H-NS plays a key role in the formation of rigid H-NS nucleoprotein filament structures required for gene silencing. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

  2. Bacterial prostatitis.

    Science.gov (United States)

    Gill, Bradley C; Shoskes, Daniel A

    2016-02-01

    The review provides the infectious disease community with a urologic perspective on bacterial prostatitis. Specifically, the article briefly reviews the categorization of prostatitis by type and provides a distillation of new findings published on bacterial prostatitis over the past year. It also highlights key points from the established literature. Cross-sectional prostate imaging is becoming more common and may lead to more incidental diagnoses of acute bacterial prostatitis. As drug resistance remains problematic in this condition, the reemergence of older antibiotics such as fosfomycin, has proven beneficial. With regard to chronic bacterial prostatitis, no clear clinical risk factors emerged in a large epidemiological study. However, bacterial biofilm formation has been associated with more severe cases. Surgery has a limited role in bacterial prostatitis and should be reserved for draining of a prostatic abscess or the removal of infected prostatic stones. Prostatitis remains a common and bothersome clinical condition. Antibiotic therapy remains the basis of treatment for both acute and chronic bacterial prostatitis. Further research into improving prostatitis treatment is indicated.

  3. The XylS/Pm regulator/promoter system and its use in fundamental studies of bacterial gene expression, recombinant protein production and metabolic engineering.

    Science.gov (United States)

    Gawin, Agnieszka; Valla, Svein; Brautaset, Trygve

    2017-07-01

    The XylS/Pm regulator/promoter system originating from the Pseudomonas putida TOL plasmid pWW0 is widely used for regulated low- and high-level recombinant expression of genes and gene clusters in Escherichia coli and other bacteria. Induction of this system can be graded by using different cheap benzoic acid derivatives, which enter cells by passive diffusion, operate in a dose-dependent manner and are typically not metabolized by the host cells. Combinatorial mutagenesis and selection using the bla gene encoding β-lactamase as a reporter have demonstrated that the Pm promoter, the DNA sequence corresponding to the 5' untranslated end of its cognate mRNA and the xylS coding region can be modified and improved relative to various types of applications. By combining such mutant genetic elements, altered and extended expression profiles were achieved. Due to their unique properties, obtained systems serve as a genetic toolbox valuable for heterologous protein production and metabolic engineering, as well as for basic studies aiming at understanding fundamental parameters affecting bacterial gene expression. The approaches used to modify XylS/Pm should be adaptable for similar improvements also of other microbial expression systems. In this review, we summarize constructions, characteristics, refinements and applications of expression tools using the XylS/Pm system. © 2017 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.

  4. Inhibition of bacterial DD-peptidases (penicillin-binding proteins) in membranes and in vivo by peptidoglycan-mimetic boronic acids.

    Science.gov (United States)

    Dzhekieva, Liudmila; Kumar, Ish; Pratt, R F

    2012-04-03

    The DD-peptidases or penicillin-binding proteins (PBPs) catalyze the final steps of bacterial peptidoglycan biosynthesis and are inhibited by the β-lactam antibiotics. There is at present a question of whether the active site structure and activity of these enzymes is the same in the solubilized (truncated) DD-peptidase constructs employed in crystallographic and kinetics studies as in membrane-bound holoenzymes. Recent experiments with peptidoglycan-mimetic boronic acids have suggested that these transition state analogue-generating inhibitors may be able to induce reactive conformations of these enzymes and thus inhibit strongly. We have now, therefore, measured the dissociation constants of peptidoglycan-mimetic boronic acids from Escherichia coli and Bacillus subtilis PBPs in membrane preparations and, in the former case, in vivo, by means of competition experiments with the fluorescent penicillin Bocillin Fl. The experiments showed that the boronic acids bound measurably (K(i) DD-peptidase inhibitors are more or less effective in vivo than in homogeneous solution.

  5. Immobilization of bacterial S-layer proteins from Caulobacter crescentus on iron oxide-based nanocomposite: synthesis and spectroscopic characterization of zincite-coated Fe₂O₃ nanoparticles.

    Science.gov (United States)

    Habibi, Neda

    2014-05-05

    Zinc oxide was coated on Fe2O3 nanoparticles using sol-gel spin-coating. Caulobacter crescentus have a crystalline surface layer (S-layer), which consist of one protein or glycoprotein species. The immobilization of bacterial S-layers obtained from C. crescentus on zincite-coated nanoparticles of iron oxide was investigated. The SDS PAGE results of S-layers isolated from C. crescentus showed the weight of 50 KDa. Nanoparticles of the Fe2O3 and zinc oxide were synthesized by a sol-gel technique. Fe2O3 nanoparticles with an average size of 50 nm were successfully prepared by the proper deposition of zinc oxide onto iron oxide nanoparticles surface annealed at 450 °C. The samples were characterized by field-emission scanning electron microscope (FESEM), atomic force microscopy (AFM), powder X-ray diffraction (XRD) and Fourier-transform infrared spectroscopy (FT-IR). Copyright © 2014 Elsevier B.V. All rights reserved.

  6. A Bioinformatics Analysis Reveals a Group of MocR Bacterial Transcriptional Regulators Linked to a Family of Genes Coding for Membrane Proteins

    Directory of Open Access Journals (Sweden)

    Teresa Milano

    2016-01-01

    Full Text Available The MocR bacterial transcriptional regulators are characterized by an N-terminal domain, 60 residues long on average, possessing the winged-helix-turn-helix (wHTH architecture responsible for DNA recognition and binding, linked to a large C-terminal domain (350 residues on average that is homologous to fold type-I pyridoxal 5′-phosphate (PLP dependent enzymes like aspartate aminotransferase (AAT. These regulators are involved in the expression of genes taking part in several metabolic pathways directly or indirectly connected to PLP chemistry, many of which are still uncharacterized. A bioinformatics analysis is here reported that studied the features of a distinct group of MocR regulators predicted to be functionally linked to a family of homologous genes coding for integral membrane proteins of unknown function. This group occurs mainly in the Actinobacteria and Gammaproteobacteria phyla. An analysis of the multiple sequence alignments of their wHTH and AAT domains suggested the presence of specificity-determining positions (SDPs. Mapping of SDPs onto a homology model of the AAT domain hinted at possible structural/functional roles in effector recognition. Likewise, SDPs in wHTH domain suggested the basis of specificity of Transcription Factor Binding Site recognition. The results reported represent a framework for rational design of experiments and for bioinformatics analysis of other MocR subgroups.

  7. Bacterial Vaginosis

    Science.gov (United States)

    ... Archive STDs Home Page Bacterial Vaginosis (BV) Chlamydia Gonorrhea Genital Herpes Hepatitis HIV/AIDS & STDs Human Papillomavirus ( ... of getting other STDs, such as chlamydia and gonorrhea . These bacteria can sometimes cause pelvic inflammatory disease ( ...

  8. The TLR2 Antagonist Staphylococcal Superantigen-Like Protein 3 Acts as a Virulence Factor to Promote Bacterial Pathogenicity in vivo

    NARCIS (Netherlands)

    Koymans, Kirsten J.; Goldmann, Oliver; Karlsson, Christofer A.Q.; Sital, Wiedjai; Thänert, Robert; Bisschop, Adinda; Vrieling, Manouk; Malmström, Johan; van Kessel, Kok P M; De Haas, Carla J.C.; Van Strijp, Jos A.G.; Medina, Eva

    2017-01-01

    Toll-like receptor (TLR) signaling is important in the initiation of immune responses and subsequent instigation of adaptive immunity. TLR2 recognizes bacterial lipoproteins and plays a central role in the host defense against bacterial infections, including those caused by Staphylococcus aureus.

  9. Xanthomonas oryzae pv. oryzae TALE proteins recruit OsTFIIAγ1 to compensate for the absence of OsTFIIAγ5 in bacterial blight in rice.

    Science.gov (United States)

    Ma, Wenxiu; Zou, Lifang; Ji, Zhiyuan; Xu, Xiameng; Xu, Zhengyin; Yang, Yangyang; Alfano, James R; Chen, Gongyou

    2018-04-28

    Xanthomonas oryzae pv. oryzae (Xoo), causal agent of bacterial blight (BB) of rice, uses transcription activator-like effectors (TALEs) to interact with the basal transcription factor gama subunit OsTFIIAγ5 (Xa5) and activates transcription of host genes. However, how OsTFIIAγ1, the other OsTFIIAγ protein, functions in the presence of TALEs remains unclear. In this study, we show that OsTFIIAγ1 plays a compensatory role in the absence of Xa5. The expression of OsTFIIAγ1, which is activated by TALE PthXo7, increased the expression of host genes targeted by avirulent and virulent TALEs. Defective OsTFIIAγ1 rice lines showed reduced expression of the TALE-targeted susceptibility (S) genes, OsSWEET11 and OsSWEET14, which resulted in increased BB resistance. Selected TALEs (PthXo1, AvrXa7, and AvrXa27) were evaluated for interactions with OsTFIIAγ1, Xa5 and xa5 (naturally-occurring mutant form of Xa5) using biomolecular fluorescence complementation (BiFC) and microscale thermophoresis (MST). BiFC and MST demonstrated that the three TALEs bind Xa5 and OsTFIIAγ1 with a stronger affinity than xa5. These results provide insight into the complex roles of OsTFIIAγ1 and OsTFIIAγ5 in TALE-mediated host gene transcription. This article is protected by copyright. All rights reserved. © 2018 BSPP and John Wiley & Sons Ltd.

  10. Platelet activation by bacterial phospholipase C involves phosphoinositide turnover and phosphorylation of 47,000 dalton but not 20,000 dalton protein

    International Nuclear Information System (INIS)

    Huzoor-Akbar; Anwer, K.

    1986-01-01

    This study was conducted to examine the role of phosphoinositides (PIns) and phosphorylation of 47,000 dalton (P47) and 20,000 dalton (P20) proteins in platelet activation by bacterial phospholipase C (PLC). PLC induced serotonin secretion (SS) and platelet aggregation (PA) in a concentration dependent manner. PLC (0.02 U/ml) caused phosphorylation of P47 in a time dependent manner (27% at 0.5 min to 378% at 7 min). PLC did not induce more than 15% phosphorylation of P20 by 7 min. Aspirin (500 μM) blocked phosphorylation of P20 but did not inhibit SS, PA or phosphorylation of P47. PLC (0.04 U/ml) decreased radioactivity (cpm) in 32 P labeled phosphatidylinositol (PI), PI-4,5-bis-PO4 (PIP2) and PI-4-PO4 (PIP) by 20%, 12% and 7.5% respectively at 15 sec. The level of PI but not that of PIP2 returned to base line in 3 min. PIP level increased above control values within one min. PLC increased phosphatidic acid level (75% at 0.5 min. to 1545% at 3 min). In other experiments PLC produced diacylglycerol (DAG) in a time and concentration dependent manner. However, no DAG was detectable in the first 60 sec. These data suggest that: (a) PIns turnover and phosphorylation of P47 but not that of P20 is involved in platelet activation by PLC; and (b) DAG production from outer membrane phospholipids is not a prerequisite for platelet activation by PLC

  11. BACTERIAL CONSORTIUM

    Directory of Open Access Journals (Sweden)

    Payel Sarkar

    2013-01-01

    Full Text Available Petroleum aromatic hydrocarbons like benzen e, toluene, ethyl benzene and xylene, together known as BTEX, has almost the same chemical structure. These aromatic hydrocarbons are released as pollutants in th e environment. This work was taken up to develop a solvent tolerant bacterial cons ortium that could degrade BTEX compounds as they all share a common chemical structure. We have isolated almost 60 different types of bacterial strains from different petroleum contaminated sites. Of these 60 bacterial strains almost 20 microorganisms were screene d on the basis of capability to tolerate high concentration of BTEX. Ten differe nt consortia were prepared and the compatibility of the bacterial strains within the consortia was checked by gram staining and BTEX tolerance level. Four successful mi crobial consortia were selected in which all the bacterial strains concomitantly grew in presence of high concentration of BTEX (10% of toluene, 10% of benzene 5% ethyl benzene and 1% xylene. Consortium #2 showed the highest growth rate in pr esence of BTEX. Degradation of BTEX by consortium #2 was monitored for 5 days by gradual decrease in the volume of the solvents. The maximum reduction observed wa s 85% in 5 days. Gas chromatography results also reveal that could completely degrade benzene and ethyl benzene within 48 hours. Almost 90% degradation of toluene and xylene in 48 hours was exhibited by consortium #2. It could also tolerate and degrade many industrial solvents such as chloroform, DMSO, acetonitrile having a wide range of log P values (0.03–3.1. Degradation of aromatic hydrocarbon like BTEX by a solvent tolerant bacterial consortium is greatly significant as it could degrade high concentration of pollutants compared to a bacterium and also reduces the time span of degradation.

  12. Molecular approaches for bacterial azoreductases

    Directory of Open Access Journals (Sweden)

    Montira Leelakriangsak

    2013-12-01

    Full Text Available Azo dyes are the dominant types of synthetic dyes, widely used in textiles, foods, leather, printing, tattooing, cosmetics, and pharmaceutical industries. Many microorganisms are able to decolorize azo dyes, and there is increasing interest in biological waste treatment methods. Bacterial azoreductases can cleave azo linkages (-N=N- in azo dyes, forming aromatic amines. This review mainly focuses on employing molecular approaches, including gene manipulation and recombinant strains, to study bacterial azoreductases. The construction of the recombinant protein by cloning and the overexpression of azoreductase is described. The mechanisms and function of bacterial azoreductases can be studied by other molecular techniques discussed in this review, such as RT-PCR, southern blot analysis, western blot analysis, zymography, and muta-genesis in order to understand bacterial azoreductase properties, function and application. In addition, understanding the regulation of azoreductase gene expression will lead to the systematic use of gene manipulation in bacterial strains for new strategies in future waste remediation technologies.

  13. Colonization of Vitis vinifera by a Green Fluorescence Protein-Labeled, gfp-Marked Strain of Xylophilus ampelinus, the Causal Agent of Bacterial Necrosis of Grapevine

    OpenAIRE

    Grall, Sophie; Manceau, Charles

    2003-01-01

    The dynamics of Xylophilus ampelinus were studied in Vitis vinifera cv. Ugni blanc using gfp-marked bacterial strains to evaluate the relative importance of epiphytic and endophytic phases of plant colonization in disease development. Currently, bacterial necrosis of grapevine is of economic importance in vineyards in three regions in France: the Cognac, Armagnac, and Die areas. This disease is responsible for progressive destruction of vine shoots, leading to their death. We constructed gfp-...

  14. Bacterial meningitis

    NARCIS (Netherlands)

    Heckenberg, Sebastiaan G. B.; Brouwer, Matthijs C.; van de Beek, Diederik

    2014-01-01

    Bacterial meningitis is a neurologic emergency. Vaccination against common pathogens has decreased the burden of disease. Early diagnosis and rapid initiation of empiric antimicrobial and adjunctive therapy are vital. Therapy should be initiated as soon as blood cultures have been obtained,

  15. Bacterial lipases

    NARCIS (Netherlands)

    Jaeger, Karl-Erich; Ransac, Stéphane; Dijkstra, Bauke W.; Colson, Charles; Heuvel, Margreet van; Misset, Onno

    Many different bacterial species produce lipases which hydrolyze esters of glycerol with preferably long-chain fatty acids. They act at the interface generated by a hydrophobic lipid substrate in a hydrophilic aqueous medium. A characteristic property of lipases is called interfacial activation,

  16. Bacterial stress

    Indian Academy of Sciences (India)

    First page Back Continue Last page Graphics. Bacterial stress. Physicochemical and chemical parameters: temperature, pressure, pH, salt concentration, oxygen, irradiation. Nutritional depravation: nutrient starvation, water shortage. Toxic compounds: Antibiotics, heavy metals, toxins, mutagens. Interactions with other cells: ...

  17. Novel Accurate Bacterial Discrimination by MALDI-Time-of-Flight MS Based on Ribosomal Proteins Coding in S10-spc-alpha Operon at Strain Level S10-GERMS

    Science.gov (United States)

    Tamura, Hiroto; Hotta, Yudai; Sato, Hiroaki

    2013-08-01

    Matrix-assisted laser-desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is one of the most widely used mass-based approaches for bacterial identification and classification because of the simple sample preparation and extremely rapid analysis within a few minutes. To establish the accurate MALDI-TOF MS bacterial discrimination method at strain level, the ribosomal subunit proteins coded in the S 10-spc-alpha operon, which encodes half of the ribosomal subunit protein and is highly conserved in eubacterial genomes, were selected as reliable biomarkers. This method, named the S10-GERMS method, revealed that the strains of genus Pseudomonas were successfully identified and discriminated at species and strain levels, respectively; therefore, the S10-GERMS method was further applied to discriminate the pathovar of P. syringae. The eight selected biomarkers (L24, L30, S10, S12, S14, S16, S17, and S19) suggested the rapid discrimination of P. syringae at the strain (pathovar) level. The S10-GERMS method appears to be a powerful tool for rapid and reliable bacterial discrimination and successful phylogenetic characterization. In this article, an overview of the utilization of results from the S10-GERMS method is presented, highlighting the characterization of the Lactobacillus casei group and discrimination of the bacteria of genera Bacillus and Sphingopyxis despite only two and one base difference in the 16S rRNA gene sequence, respectively.

  18. [Spontaneous bacterial peritonitis].

    Science.gov (United States)

    Velkey, Bálint; Vitális, Eszter; Vitális, Zsuzsanna

    2017-01-01

    Spontaneous bacterial peritonitis occurs most commonly in cirrhotic patients with ascites. Pathogens get into the circulation by intestinal translocation and colonize in peritoneal fluid. Diagnosis of spontaneous bacterial peritonitis is based on elevated polymorphonuclear leukocyte count in the ascites (>0,25 G/L). Ascites culture is often negative but aids to get information about antibiotic sensitivity in positive cases. Treatment in stable patient can be intravenous then orally administrated ciprofloxacin or amoxicillin/clavulanic acid, while in severe cases intravenous III. generation cephalosporin. Nosocomial spontaneous bacterial peritonitis often caused by Gram-positive bacteria and multi-resistant pathogens can also be expected thus carbapenem should be the choice of the empiric treatment. Antibiotic prophylaxis should be considered. Norfloxacin is used most commonly, but changes are expected due to increase in quinolone resistance. As a primary prophylaxis, a short-term antibiotic treatment is recommended after gastrointestinal bleeding for 5 days, while long-term prophylaxis is for patients with low ascites protein, and advanced disease (400 mg/day). Secondary prophylaxis is recommended for all patients recovered from spontaneous bacterial peritonitis. Due to increasing antibiotic use of antibiotics prophylaxis is debated to some degree. Orv. Hetil., 2017, 158(2), 50-57.

  19. [Bacterial vaginosis].

    Science.gov (United States)

    Romero Herrero, Daniel; Andreu Domingo, Antonia

    2016-07-01

    Bacterial vaginosis (BV) is the main cause of vaginal dysbacteriosis in the women during the reproductive age. It is an entity in which many studies have focused for years and which is still open for discussion topics. This is due to the diversity of microorganisms that cause it and therefore, its difficult treatment. Bacterial vaginosis is probably the result of vaginal colonization by complex bacterial communities, many of them non-cultivable and with interdependent metabolism where anaerobic populations most likely play an important role in its pathogenesis. The main symptoms are an increase of vaginal discharge and the unpleasant smell of it. It can lead to serious consequences for women, such as an increased risk of contracting sexually transmitted infections including human immunodeficiency virus and upper genital tract and pregnancy complications. Gram stain is the gold standard for microbiological diagnosis of BV, but can also be diagnosed using the Amsel clinical criteria. It should not be considered a sexually transmitted disease but it is highly related to sex. Recurrence is the main problem of medical treatment. Apart from BV, there are other dysbacteriosis less characterized like aerobic vaginitis of which further studies are coming slowly but are achieving more attention and consensus among specialists. Copyright © 2016 Elsevier España, S.L.U. All rights reserved.

  20. Expression of Human papillomavirus 16 E7ggg oncoprotein on N- and C-terminus of Potato virus X coat protein in bacterial and plant cells

    Czech Academy of Sciences Publication Activity Database

    Plchová, Helena; Moravec, Tomáš; Hoffmeisterová, Hana; Folwarczna, Jitka; Čeřovská, Noemi

    2011-01-01

    Roč. 77, č. 2 (2011), s. 146-152 ISSN 1046-5928 R&D Projects: GA ČR GA521/09/1525 Institutional research plan: CEZ:AV0Z50380511 Keywords : Bacterial expression * Human papillomavirus * Oncoprotein E7 Subject RIV: EI - Biotechnology ; Bionics Impact factor: 1.587, year: 2011

  1. Functional microdomains in bacterial membranes.

    Science.gov (United States)

    López, Daniel; Kolter, Roberto

    2010-09-01

    The membranes of eukaryotic cells harbor microdomains known as lipid rafts that contain a variety of signaling and transport proteins. Here we show that bacterial membranes contain microdomains functionally similar to those of eukaryotic cells. These membrane microdomains from diverse bacteria harbor homologs of Flotillin-1, a eukaryotic protein found exclusively in lipid rafts, along with proteins involved in signaling and transport. Inhibition of lipid raft formation through the action of zaragozic acid--a known inhibitor of squalene synthases--impaired biofilm formation and protein secretion but not cell viability. The orchestration of physiological processes in microdomains may be a more widespread feature of membranes than previously appreciated.

  2. Effect of diet and absence of protozoa on the rumen microbial community and on the representativeness of bacterial fractions used in the determination of microbial protein synthesis.

    Science.gov (United States)

    Belanche, A; de la Fuente, G; Pinloche, E; Newbold, C J; Balcells, J

    2012-11-01

    Accurate estimates of microbial synthesis in the rumen are vital to optimize ruminant nutrition. Liquid- (LAB) and solid-associated bacterial fractions (SAB) harvested from the rumen are generally considered as microbial references when microbial yield is calculated; however, factors that determine their composition are not completely understood. The aim of this study was to evaluate the effect of diet and absence or presence of rumen protozoa on the rumen microbial community. It was hypothesized that these treatments could modify the composition and representativeness of LAB and SAB. Twenty twin lambs (Ovis aries) were used; one-half of the twins were kept protozoa-free, and each respective twin sibling was faunated. At 6 mo of age, 5 animals from each group were randomly allocated to the experimental diets consisting of either alfalfa hay as the sole diet, or 50:50 mixed with ground barley grain. After 15 d of adaptation to the diet, animals were euthanized, rumen and abomasum contents were sampled, and LAB and SAB isolated. The presence of protozoa buffered the effect of diet on the rumen bacterial population. Faunated animals fed alfalfa hay had a greater abundance of F. succinogenes, anaerobic fungi and methanogens, as well as an enhanced rumen bacterial diversity. Cellulolytic bacteria were more abundant in SAB, whereas the abomasal abundance of most of the microorganisms studied was closer to those values observed in LAB. Rumen and abomasal samples showed similar bacterial DNA concentrations, but the fungal and protozoal DNA concentration in the abomasum was only 69% and 13% of that observed in the rumen, respectively, suggesting fungal and protozoal sequestration in the rumen or possible preferential degradation of fungal and protozoal DNA in the abomasum, or both. In conclusion, absence of protozoa and type of diet extensively modified the chemical composition of LAB and SAB as a consequence of changes in the microbial composition of these fractions.

  3. Structure of Stenotrophomonas maltophilia FeoA complexed with zinc: a unique prokaryotic SH3-domain protein that possibly acts as a bacterial ferrous iron-transport activating factor

    International Nuclear Information System (INIS)

    Su, Yi-Che; Chin, Ko-Hsin; Hung, Hui-Chih; Shen, Gwan-Han; Wang, Andrew H.-J.; Chou, Shan-Ho

    2010-01-01

    The crystal structure of FeoA from Stenotrophomonas maltophilia has been determined to a resolution of 1.7 Å using an Se single-wavelength anomalous dispersion (Se-SAD) approach and revealed a unique dimer cross-linked by two zinc ions and six chloride ions. Iron is vital to the majority of prokaryotes, with ferrous iron believed to be the preferred form for iron uptake owing to its much better solubility. The major route for bacterial ferrous iron uptake is found to be via an Feo (ferrous iron-transport) system comprising the three proteins FeoA, FeoB and FeoC. Although the structure and function of FeoB have received much attention recently, the roles played by FeoA and FeoC have been little investigated to date. Here, the tertiary structure of FeoA from Stenotrophomonas maltophilia (Sm), a vital opportunistic pathogen in immunodepressed hosts, is reported. The crystal structure of SmFeoA has been determined to a resolution of 1.7 Å using an Se single-wavelength anomalous dispersion (Se-SAD) approach. Although SmFeoA bears low sequence identity to eukaryotic proteins, its structure is found to adopt a eukaryotic SH3-domain-like fold. It also bears weak similarity to the C-terminal SH3 domain of bacterial DtxR (diphtheria toxin regulator), with some unique characteristics. Intriguingly, SmFeoA is found to adopt a unique dimer cross-linked by two zinc ions and six anions (chloride ions). Since FeoB has been found to contain a G-protein-like domain with low GTPase activity, FeoA may interact with FeoB through the SH3–G-protein domain interaction to act as a ferrous iron-transport activating factor

  4. Colonization of Vitis vinifera by a green fluorescence protein-labeled, gfp-marked strain of Xylophilus ampelinus, the causal agent of bacterial necrosis of grapevine.

    Science.gov (United States)

    Grall, Sophie; Manceau, Charles

    2003-04-01

    The dynamics of Xylophilus ampelinus were studied in Vitis vinifera cv. Ugni blanc using gfp-marked bacterial strains to evaluate the relative importance of epiphytic and endophytic phases of plant colonization in disease development. Currently, bacterial necrosis of grapevine is of economic importance in vineyards in three regions in France: the Cognac, Armagnac, and Die areas. This disease is responsible for progressive destruction of vine shoots, leading to their death. We constructed gfp-marked strains of the CFBP2098 strain of X. ampelinus for histological studies. We studied the colonization of young plants of V. vinifera cv. Ugni blanc by X. ampelinus after three types of artificial contamination in a growth chamber and in a greenhouse. (i) After wounding of the stem and inoculation, the bacteria progressed down to the crown through the xylem vessels, where they organized into biofilms. (ii) When the bacteria were forced into woody cuttings, they rarely colonized the emerging plantlets. Xylem vessels could play a key role in the multiplication and conservation of the bacteria, rather than being a route for plant colonization. (iii) When bacterial suspensions were sprayed onto the plants, bacteria progressed in two directions: both in emerging organs and down to the crown, thus displaying the importance of epiphytic colonization in disease development.

  5. Bacterial Ecology

    DEFF Research Database (Denmark)

    Fenchel, Tom

    2011-01-01

    compounds these must first be undergo extracellular hydrolysis. Bacteria have a great diversity with respect to types of metabolism that far exceeds the metabolic repertoire of eukaryotic organisms. Bacteria play a fundamental role in the biosphere and certain key processes such as, for example......, the production and oxidation of methane, nitrate reduction and fixation of atmospheric nitrogen are exclusively carried out by different groups of bacteria. Some bacterial species – ‘extremophiles’ – thrive in extreme environments in which no eukaryotic organisms can survive with respect to temperature, salinity...... biogeochemical processes are carried exclusively by bacteria. * Bacteria play an important role in all types of habitats including some that cannot support eukaryotic life....

  6. Diagnostic challenges with acellular bacterial meningitis

    African Journals Online (AJOL)

    tests performed included a non-reactive HIV ELISA and syphilis serology. ... Despite our patient's reduced CSF glucose and raised protein, the inconsistent .... Suzuki W, et al. Cerebrospinal fluid/blood glucose ratio as an indicator for bacterial ...

  7. Proteomic Analysis of Bacterial Expression Profiles Following ...

    African Journals Online (AJOL)

    mass spectrometry (GC-MS) were performed to determine the phytochemicals in the active fraction. Results: Five differentially expressed bacterial proteins (four from Escherichia coli and one from Staphylococcus aureus), were identified via ...

  8. Bacterial mitotic machineries

    DEFF Research Database (Denmark)

    Gerdes, Kenn; Møller-Jensen, Jakob; Ebersbach, Gitte

    2004-01-01

    Here, we review recent progress that yields fundamental new insight into the molecular mechanisms behind plasmid and chromosome segregation in prokaryotic cells. In particular, we describe how prokaryotic actin homologs form mitotic machineries that segregate DNA before cell division. Thus, the P......M protein of plasmid R1 forms F actin-like filaments that separate and move plasmid DNA from mid-cell to the cell poles. Evidence from three different laboratories indicate that the morphogenetic MreB protein may be involved in segregation of the bacterial chromosome.......Here, we review recent progress that yields fundamental new insight into the molecular mechanisms behind plasmid and chromosome segregation in prokaryotic cells. In particular, we describe how prokaryotic actin homologs form mitotic machineries that segregate DNA before cell division. Thus, the Par...

  9. Bacterial surface-displayed GII.4 human norovirus capsid proteins bound to surface of Romaine lettuce through HBGA-like molecules

    Science.gov (United States)

    Human Noroviruses (HuNoVs) are the main cause of nonbacterial gastroenteritis. Contaminated produce is a main vehicle for dissemination of HuNoVs. In this study, we used an ice nucleation protein (INP) mediated surface display system to present the protruding domain of GII.4 HuNoV capsid protein (G...

  10. Handling Gene and Protein Names in the Age of Bioinformatics: The Special Challenge of Secreted Multimodular Bacterial Enzymes such as the cbhA/cbh9A Gene of Clostridium thermocellum

    Energy Technology Data Exchange (ETDEWEB)

    Brunecky, Roman [National Renewable Energy Laboratory (NREL), Golden, CO (United States); Schwarz, Wolfgang H. [Technical University of Munich; Broeker, Jannis [Technical University of Munich; Liebl, Wolfgang [Technical University of Munich; Zverlov, Vladimir V. [Technical University of Munich; Russian Academy of Science

    2018-02-26

    An increasing number of researchers working in biology, biochemistry, biotechnology, bioengineering, bioinformatics and other related fields of science are using biological molecules. As the scientific background of the members of different scientific communities is more diverse than ever before, the number of scientists not familiar with the rules for non-ambiguous designation of genetic elements is increasing. However, with biological molecules gaining importance through biotechnology, their functional and unambiguous designation is vital. Unfortunately, naming genes and proteins is not an easy task. In addition, the traditional concepts of bioinformatics are challenged with the appearance of proteins comprising different modules with a respective function in each module. This article highlights basic rules and novel solutions in designation recently used within the community of bacterial geneticists, and we discuss the present-day handling of gene and protein designations. As an example we will utilize a recent mischaracterization of gene nomenclature. We make suggestions for better handling of names in future literature as well as in databases and annotation projects. Our methodology emphasizes the hydrolytic function of multi-modular genes and extracellular proteins from bacteria.

  11. Measurement of the rate of bacterial protein synthesis in rumen. Part of a coordinated programme on tracer techniques in studies on the use of non-protein nitrogen in ruminants

    International Nuclear Information System (INIS)

    Leng, R.A.

    1975-10-01

    A technique has been developed to differentiate between rumen microbial and dietary protein at the duodenum. Mature whethers fitted with rumen, duodenal and ileal cannulas were fed protein poor diets with added urea, casein and formaldehyde treated casein. These contained 0.30, 1.54, 3.26 and 3.26 % N respectively. The isotopes 35 S or 15 N were infused into the rumen to label microbial protein so that microbial protein could be differentiated from dietary protein. By using the 35 S or 15 N system to mark microbes, it was found that 40% of the formaldehyde treated protein is fermented in the rumen and about 10% is lost in the feces. This indicates that approximately 50% of the protected protein was utilized by the sheep

  12. Antibiotic resistance of bacterial biofilms

    DEFF Research Database (Denmark)

    Hoiby, N.; Bjarnsholt, T.; Givskov, M.

    2010-01-01

    A biofilm is a structured consortium of bacteria embedded in a self-produced polymer matrix consisting of polysaccharide, protein and DNA. Bacterial biofilms cause chronic infections because they show increased tolerance to antibiotics and disinfectant chemicals as well as resisting phagocytosis...... and other components of the body's defence system. The persistence of, for example, staphylococcal infections related to foreign bodies is due to biofilm formation. Likewise, chronic Pseudomonas aeruginosa lung infection in cystic fibrosis patients is caused by biofilm-growing mucoid strains....... Characteristically, gradients of nutrients and oxygen exist from the top to the bottom of biofilms and these gradients are associated with decreased bacterial metabolic activity and increased doubling times of the bacterial cells; it is these more or less dormant cells that are responsible for some of the tolerance...

  13. Application of a loading dose of colistin methanesulfonate in critically ill patients: population pharmacokinetics, protein binding, and prediction of bacterial kill.

    Science.gov (United States)

    Mohamed, Ami F; Karaiskos, Ilias; Plachouras, Diamantis; Karvanen, Matti; Pontikis, Konstantinos; Jansson, Britt; Papadomichelakis, Evangelos; Antoniadou, Anastasia; Giamarellou, Helen; Armaganidis, Apostolos; Cars, Otto; Friberg, Lena E

    2012-08-01

    A previous pharmacokinetic study on dosing of colistin methanesulfonate (CMS) at 240 mg (3 million units [MU]) every 8 h indicated that colistin has a long half-life, resulting in insufficient concentrations for the first 12 to 48 h after initiation of treatment. A loading dose would therefore be beneficial. The aim of this study was to evaluate CMS and colistin pharmacokinetics following a 480-mg (6-MU) loading dose in critically ill patients and to explore the bacterial kill following the use of different dosing regimens obtained by predictions from a pharmacokinetic-pharmacodynamic model developed from an in vitro study on Pseudomonas aeruginosa. The unbound fractions of colistin A and colistin B were determined using equilibrium dialysis and considered in the predictions. Ten critically ill patients (6 males; mean age, 54 years; mean creatinine clearance, 82 ml/min) with infections caused by multidrug-resistant Gram-negative bacteria were enrolled in the study. The pharmacokinetic data collected after the first and eighth doses were analyzed simultaneously with the data from the previous study (total, 28 patients) in the NONMEM program. For CMS, a two-compartment model best described the pharmacokinetics, and the half-lives of the two phases were estimated to be 0.026 and 2.2 h, respectively. For colistin, a one-compartment model was sufficient and the estimated half-life was 18.5 h. The unbound fractions of colistin in the patients were 26 to 41% at clinical concentrations. Colistin A, but not colistin B, had a concentration-dependent binding. The predictions suggested that the time to 3-log-unit bacterial kill for a 480-mg loading dose was reduced to half of that for the dose of 240 mg.

  14. Increase in Bacterial Colony Formation from a Permafrost Ice Wedge Dosed with a Tomitella biformata Recombinant Resuscitation-Promoting Factor Protein.

    Science.gov (United States)

    Puspita, Indun Dewi; Kitagawa, Wataru; Kamagata, Yoichi; Tanaka, Michiko; Nakatsu, Cindy H

    2015-01-01

    Resuscitation-promoting factor (Rpf) is a protein that has been found in a number of different Actinobacteria species and has been shown to promote the growth of active cells and resuscitate dormant (non-dividing) cells. We previously reported the biological activity of an Rpf protein in Tomitella biformata AHU 1821(T), an Actinobacteria isolated from a permafrost ice wedge. This protein is excreted outside the cell; however, few studies have investigated its contribution in environmental samples to the growth or resuscitation of bacteria other than the original host. Therefore, the aim of the present study was to determine whether Rpf from T. biformata impacted the cultivation of other bacteria from the permafrost ice wedge from which it was originally isolated. All experiments used recombinant Rpf proteins produced using a Rhodococcus erythropolis expression system. Dilutions of melted surface sterilized ice wedge samples mixed with different doses of the purified recombinant Rpf (rRpf) protein indicated that the highest concentration tested, 1250 pM, had a significantly (p permafrost sediments. The results of the present study demonstrated that rRpf not only promoted the growth of T. biformata from which it was isolated, but also enhanced colony formation by another Actinobacteria in an environmental sample.

  15. Bacterial Microcompartments

    Energy Technology Data Exchange (ETDEWEB)

    Kerfeld, Cheryl A.; Heinhorst, Sabine; Cannon, Gordon C.

    2010-06-05

    Bacterialmicrocompartments (BMCs) are organelles composed entirely of protein. They promote specific metabolic processes by encapsulatingand colocalizing enzymes with their substrates and cofactors, by protecting vulnerable enzymes in a defined microenvironment, and bysequestering toxic or volatile intermediates. Prototypes of the BMCsare the carboxysomes of autotrophic bacteria. However, structures of similarpolyhedral shape are being discovered in an ever-increasing number of heterotrophic bacteria, where they participate in the utilization ofspecialty carbon and energy sources.Comparative genomics reveals that the potential for this type of compartmentalization is widespread acrossbacterial phyla and suggests that genetic modules encoding BMCs are frequently laterally transferred among bacteria. The diverse functionsof these BMCs suggest that they contribute to metabolic innovation in bacteria in a broad range of environments.

  16. New players in the same old game: a system level in silico study to predict type III secretion system and effector proteins in bacterial genomes reveals common themes in T3SS mediated pathogenesis.

    Science.gov (United States)

    Sadarangani, Vineet; Datta, Sunando; Arunachalam, Manonmani

    2013-07-26

    Type III secretion system (T3SS) plays an important role in virulence or symbiosis of many pathogenic or symbiotic bacteria [CHM 2:291-294, 2007; Physiology (Bethesda) 20:326-339, 2005]. T3SS acts like a tunnel between a bacterium and its host through which the bacterium injects 'effector' proteins into the latter [Nature 444:567-573, 2006; COSB 18:258-266, 2008]. The effectors spatially and temporally modify the host signalling pathways [FEMS Microbiol Rev 35:1100-1125, 2011; Cell Host Microbe5:571-579, 2009]. In spite its crucial role in host-pathogen interaction, the study of T3SS and the associated effectors has been limited to a few bacteria [Cell Microbiol 13:1858-1869, 2011; Nat Rev Microbiol 6:11-16, 2008; Mol Microbiol 80:1420-1438, 2011]. Before one set out to perform systematic experimental studies on an unknown set of bacteria it would be beneficial to identify the potential candidates by developing an in silico screening algorithm. A system level study would also be advantageous over traditional laboratory methods to extract an overriding theme for host-pathogen interaction, if any, from the vast resources of data generated by sequencing multiple bacterial genomes. We have developed an in silico protocol in which the most conserved set of T3SS proteins was used as the query against the entire bacterial database with increasingly stringent search parameters. It enabled us to identify several uncharacterized T3SS positive bacteria. We adopted a similar strategy to predict the presence of the already known effectors in the newly identified T3SS positive bacteria. The huge resources of biochemical data [FEMS Microbiol Rev 35:1100-1125, 2011; Cell Host Microbe 5:571-579, 2009; BMC Bioinformatics 7(11):S4, 2010] on the T3SS effectors enabled us to search for the common theme in T3SS mediated pathogenesis. We identified few cellular signalling networks in the host, which are manipulated by most of the T3SS containing pathogens. We went on to look for

  17. Efficient bacterial expression of recombinant potato mop-top virus non-structural triple gene block protein 1 modified by progressive deletion of its N-terminus

    Czech Academy of Sciences Publication Activity Database

    Pečenková, Tamara; Filigarová, Marie; Čeřovská, Noemi

    2005-01-01

    Roč. 41, - (2005), s. 128-135 ISSN 1046-5928 R&D Projects: GA ČR GA522/04/1329 Institutional research plan: CEZ:AV0Z50380511 Keywords : Protein expression * Potato mop-top virus * Triple gene block Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.553, year: 2005

  18. The Bacterial Effector AvrPto Targets the Regulatory Coreceptor SOBIR1 and Suppresses Defense Signaling Mediated by the Receptor-Like Protein Cf-4

    NARCIS (Netherlands)

    Wu, Jinbin; Burgh, Van Der Aranka M.; Bi, Guozhi; Zhang, Lisha; Alfano, James R.; Martin, Gregory B.; Joosten, Matthieu H.A.J.

    2018-01-01

    Receptor-like proteins (RLPs) and receptor-like kinases (RLKs) are cell-surface receptors that are essential for detecting invading pathogens and subsequent activation of plant defense responses. RLPs lack a cytoplasmic kinase domain to trigger downstream signaling leading to host resistance. The

  19. 2-DE and MS analysis of key proteins in the adhesion of Lactobacillus plantarum, a first step toward early selection of probiotics based on bacterial biomarkers

    NARCIS (Netherlands)

    Izquierdo, Esther; Horvatovich, Peter; Marchioni, Eric; Aoude-Werner, Dalal; Sanz, Yolanda; Ennahar, Saied

    The identification of cell components involved in probiotic activities is a challenge in current probiotic research. In this work, a new approach based on proteomics as an analytical tool for the identification of characteristic protein profiles related to adhesion to mucin as a model probiotic

  20. Structure of thrombospondin type 3 repeats in bacterial outer membrane protein A reveals its intra-repeat disulfide bond-dependent calcium-binding capability

    Energy Technology Data Exchange (ETDEWEB)

    Dai, Shuyan; Sun, Cancan; Tan, Kemin; Ye, Sheng; Zhang, Rongguang

    2017-09-01

    Eukaryotic thrombospondin type 3 repeat (TT3R) is an efficient calcium ion (Ca2+) binding motif only found in mammalian thrombospondin family. TT3R has also been found in prokaryotic cellulase Cel5G, which was thought to forfeit the Ca2+-binding capability due to the formation of intra-repeat disulfide bonds, instead of the inter-repeat ones possessed by eukaryotic TT3Rs. In this study, we have identified an enormous number of prokaryotic TT3R-containing proteins belonging to several different protein families, including outer membrane protein A (OmpA), an important structural protein connecting the outer membrane and the periplasmic peptidoglycan layer in gram-negative bacteria. Here, we report the crystal structure of the periplasmic region of OmpA from Capnocytophaga gingivalis, which contains a linker region comprising five consecutive TT3Rs. The structure of OmpA-TT3R exhibits a well-ordered architecture organized around two tightly-coordinated Ca2+ and confirms the presence of abnormal intra-repeat disulfide bonds. Further mutagenesis studies showed that the Ca2+-binding capability of OmpA-TT3R is indeed dependent on the proper formation of intra-repeat disulfide bonds, which help to fix a conserved glycine residue at its proper position for Ca2+ coordination. Additionally, despite lacking inter repeat disulfide bonds, the interfaces between adjacent OmpA-TT3Rs are enhanced by both hydrophobic and conserved aromatic-proline interactions.

  1. Potential costs of bacterial infection on storage protein gene expression and reproduction in queenless Apis mellifera worker bees on distinct dietary regimes.

    Science.gov (United States)

    Lourenço, Anete Pedro; Martins, Juliana Ramos; Guidugli-Lazzarini, Karina Rosa; Macedo, Liliane Maria Fróes; Bitondi, Márcia Maria Gentile; Simões, Zilá Luz Paulino

    2012-09-01

    Insects are able to combat infection by initiating an efficient immune response that involves synthesizing antimicrobial peptides and a range of other defense molecules. These responses may be costly to the organism, resulting in it exploiting endogenous resources to maintain homeostasis or support defense to the detriment of other physiological needs. We used queenless worker bees on distinct dietary regimes that may alter hemolymph protein storage and ovary activation to investigate the physiological costs of infection with Serratia marcescens. The expression of the genes encoding the storage proteins vitellogenin and hexamerin 70a, the vitellogenin receptor, and vasa (which has a putative role in reproduction), was impaired in the infected bees. This impairment was mainly evident in the bees fed beebread, which caused significantly higher expression of these genes than did royal jelly or syrup, and this was confirmed at the vitellogenin and hexamerin 70a protein levels. Beebread was also the only diet that promoted ovary activation in the queenless bees, but this activation was significantly impaired by the infection. The expression of the genes encoding the storage proteins apolipophorins-I and -III and the lipophorin receptor was not altered by infection regardless the diet provided to the bees. Similarly, the storage of apolipophorin-I in the hemolymph was only slightly impaired by the infection, independently of the supplied diet. Taken together these results indicate that, infection demands a physiological cost from the transcription of specific protein storage-related genes and from the reproductive capacity. Copyright © 2012 Elsevier Ltd. All rights reserved.

  2. Structural mapping of the coiled-coil domain of a bacterial condensin and comparative analyses across all domains of life suggest conserved features of SMC proteins.

    Science.gov (United States)

    Waldman, Vincent M; Stanage, Tyler H; Mims, Alexandra; Norden, Ian S; Oakley, Martha G

    2015-06-01

    The structural maintenance of chromosomes (SMC) proteins form the cores of multisubunit complexes that are required for the segregation and global organization of chromosomes in all domains of life. These proteins share a common domain structure in which N- and C- terminal regions pack against one another to form a globular ATPase domain. This "head" domain is connected to a central, globular, "hinge" or dimerization domain by a long, antiparallel coiled coil. To date, most efforts for structural characterization of SMC proteins have focused on the globular domains. Recently, however, we developed a method to map interstrand interactions in the 50-nm coiled-coil domain of MukB, the divergent SMC protein found in γ-proteobacteria. Here, we apply that technique to map the structure of the Bacillus subtilis SMC (BsSMC) coiled-coil domain. We find that, in contrast to the relatively complicated coiled-coil domain of MukB, the BsSMC domain is nearly continuous, with only two detectable coiled-coil interruptions. Near the middle of the domain is a break in coiled-coil structure in which there are three more residues on the C-terminal strand than on the N-terminal strand. Close to the head domain, there is a second break with a significantly longer insertion on the same strand. These results provide an experience base that allows an informed interpretation of the output of coiled-coil prediction algorithms for this family of proteins. A comparison of such predictions suggests that these coiled-coil deviations are highly conserved across SMC types in a wide variety of organisms, including humans. © 2015 Wiley Periodicals, Inc.

  3. A study of bacterial gene regulatory mechanisms

    DEFF Research Database (Denmark)

    Hansen, Sabine

    Bacterial cells are capable of rapidly changing their protein expression in response to ever-changing environments and physiological conditions. The cells are able to switch on the expression of proteins that due to changing environmental conditions have become vital to sustain life and likewise ...

  4. Addressing the Immunogenicity of the Cargo and of the Targeting Antibodies with a Focus on Deimmunized Bacterial Toxins and on Antibody-Targeted Human Effector Proteins

    OpenAIRE

    Grinberg, Yehudit; Benhar, Itai

    2017-01-01

    Third-generation immunotoxins are composed of a human, or humanized, targeting moiety, usually a monoclonal antibody or an antibody fragment, and a non-human effector molecule. Due to the non-human origin of the cytotoxic domain, these molecules stimulate potent anti-drug immune responses, which limit treatment options. Efforts are made to deimmunize such immunotoxins or to combine treatment with immunosuppression. An alternative approach is using the so-called ?human cytotoxic fusion protein...

  5. Connecting the dots between bacterial biofilms and ice cream

    Science.gov (United States)

    Stanley-Wall, Nicola R.; MacPhee, Cait E.

    2015-12-01

    Emerging research is revealing a diverse array of interfacially-active proteins that are involved in varied biological process from foaming horse sweat to bacterial raincoat formation. We describe an interdisciplinary approach to study the molecular and biophysical mechanisms controlling the activity of an unusual bacterial protein called BslA. This protein is needed for biofilm formation and forms a protective layer or raincoat over the bacterial community, but also has a multitude of potential applications in multiphase formulations. Here we document our journey from fundamental research to an examination of the applications for this surface-active protein in ice cream.

  6. [Expression and purification of a novel thermophilic bacterial single-stranded DNA-binding protein and enhancement the synthesis of DNA and cDNA].

    Science.gov (United States)

    Jia, Xiao-Wei; Zhang, Guo-Hui; Shi, Hai-Yan

    2012-12-01

    Express a novel species of single-stranded DNA-binding protein (SSB) derived from Thermococcus kodakarensis KOD1, abbreviated kod-ssb. And evaluate the effect of kod-ssb on PCR-based DNA amplification and reverse transcription. We express kod-ssb with the Transrtta (DE3), and kod-ssb was purified by affinity chromatography on a Ni2+ Sepharose column, detected by SDS-PAGE. To evaluate the effect of kod-ssb on PCR-based DNA amplification, the human beta globin gene was used as template to amplify a 5-kb, 9-kb and 13-kb. And to detect the effect of kod-ssb on reverse transcription, we used RNA from flu cell culture supernatant extraction as templates to implement qRT-PCR reaction. The plasmid pET11a-kod was transformed into Transetta (DE3) and the recombinant strain Transetta (pET11 a-kod) was obtained. The kod-ssb was highly expressed when the recombinant strain Transetta(pET11a-kod) was induced by IPTG. The specific protein was detected by SDS-PAGE. To confirm that kod-ssb can enhance target DNA synthesis and reduce PCR by-products, 5-, 9-, and 13-kb human beta globin gene fragments were used as templates for PCR. When PCR reactions did not include SSB proteins, the specific PCR product was contaminated with non-specific products. When kod -ssb was added, kod-ssb significantly enhanced amplification of the 5-, 9-and 13-kb target product and minimised the non-specific PCR products. To confirm that kod-ssb can enhance target cDNA synthesis, RNA from flu cell culture supernatant extraction was used as templates for qRT-PCR reaction. The results was that when kod-ssb was added, kod-ssb significantly enhanced the synthesis of cDNA, average Ct value is 19.42, and the average Ct value without kod-ssb is 22.15. kod-ssb may in future be used to enhance DNA and cDNA amplification.

  7. Bacterial Carriers for Glioblastoma Therapy

    Directory of Open Access Journals (Sweden)

    Nalini Mehta

    2017-03-01

    Full Text Available Treatment of aggressive glioblastoma brain tumors is challenging, largely due to diffusion barriers preventing efficient drug dosing to tumors. To overcome these barriers, bacterial carriers that are actively motile and programmed to migrate and localize to tumor zones were designed. These carriers can induce apoptosis via hypoxia-controlled expression of a tumor suppressor protein p53 and a pro-apoptotic drug, Azurin. In a xenograft model of human glioblastoma in rats, bacterial carrier therapy conferred a significant survival benefit with 19% overall long-term survival of >100 days in treated animals relative to a median survival of 26 days in control untreated animals. Histological and proteomic analyses were performed to elucidate the safety and efficacy of these carriers, showing an absence of systemic toxicity and a restored neural environment in treated responders. In the treated non-responders, proteomic analysis revealed competing mechanisms of pro-apoptotic and drug-resistant activity. This bacterial carrier opens a versatile avenue to overcome diffusion barriers in glioblastoma by virtue of its active motility in extracellular space and can lead to tailored therapies via tumor-specific expression of tumoricidal proteins.

  8. Roles of the Protruding Loop of Factor B Essential for the Localization of Lipoproteins (LolB) in the Anchoring of Bacterial Triacylated Proteins to the Outer Membrane*

    Science.gov (United States)

    Hayashi, Yumi; Tsurumizu, Ryoji; Tsukahara, Jun; Takeda, Kazuki; Narita, Shin-ichiro; Mori, Makiko; Miki, Kunio; Tokuda, Hajime

    2014-01-01

    The Lol system comprising five Lol proteins, LolA through LolE, sorts Escherichia coli lipoproteins to outer membranes. The LolCDE complex, an ATP binding cassette transporter in inner membranes, releases outer membrane-specific lipoproteins in an ATP-dependent manner, causing formation of the LolA-lipoprotein complex in the periplasm. LolA transports lipoproteins through the periplasm to LolB on outer membranes. LolB is itself a lipoprotein anchored to outer membranes, although the membrane anchor is functionally dispensable. LolB then localizes lipoproteins to outer membranes through largely unknown mechanisms. The crystal structure of LolB is similar to that of LolA, and it possesses a hydrophobic cavity that accommodates acyl chains of lipoproteins. To elucidate the molecular function of LolB, a periplasmic version of LolB, mLolB, was mutagenized at various conserved residues. Despite the lack of acyl chains, most defective mutants were insoluble. However, a derivative with glutamate in place of leucine 68 was soluble and unable to localize lipoproteins to outer membranes. This leucine is present in a loop protruding from mLolB into an aqueous environment, and no analogous loop is present in LolA. Thus, leucine 68 was replaced with other residues. Replacement by acidic, but not hydrophobic, residues generated for the first time mLolB derivatives that can accept but cannot localize lipoproteins to outer membranes. Moreover, deletion of the leucine with neighboring residues impaired the lipoprotein receptor activity. Based on these observations, the roles of the protruding loop of LolB in the last step of lipoprotein sorting are discussed. PMID:24569999

  9. Roles of the protruding loop of factor B essential for the localization of lipoproteins (LolB) in the anchoring of bacterial triacylated proteins to the outer membrane.

    Science.gov (United States)

    Hayashi, Yumi; Tsurumizu, Ryoji; Tsukahara, Jun; Takeda, Kazuki; Narita, Shin-ichiro; Mori, Makiko; Miki, Kunio; Tokuda, Hajime

    2014-04-11

    The Lol system comprising five Lol proteins, LolA through LolE, sorts Escherichia coli lipoproteins to outer membranes. The LolCDE complex, an ATP binding cassette transporter in inner membranes, releases outer membrane-specific lipoproteins in an ATP-dependent manner, causing formation of the LolA-lipoprotein complex in the periplasm. LolA transports lipoproteins through the periplasm to LolB on outer membranes. LolB is itself a lipoprotein anchored to outer membranes, although the membrane anchor is functionally dispensable. LolB then localizes lipoproteins to outer membranes through largely unknown mechanisms. The crystal structure of LolB is similar to that of LolA, and it possesses a hydrophobic cavity that accommodates acyl chains of lipoproteins. To elucidate the molecular function of LolB, a periplasmic version of LolB, mLolB, was mutagenized at various conserved residues. Despite the lack of acyl chains, most defective mutants were insoluble. However, a derivative with glutamate in place of leucine 68 was soluble and unable to localize lipoproteins to outer membranes. This leucine is present in a loop protruding from mLolB into an aqueous environment, and no analogous loop is present in LolA. Thus, leucine 68 was replaced with other residues. Replacement by acidic, but not hydrophobic, residues generated for the first time mLolB derivatives that can accept but cannot localize lipoproteins to outer membranes. Moreover, deletion of the leucine with neighboring residues impaired the lipoprotein receptor activity. Based on these observations, the roles of the protruding loop of LolB in the last step of lipoprotein sorting are discussed.

  10. Structural studies of a bacterial tRNA(HIS guanylyltransferase (Thg1-like protein, with nucleotide in the activation and nucleotidyl transfer sites.

    Directory of Open Access Journals (Sweden)

    Samantha J Hyde

    Full Text Available All nucleotide polymerases and transferases catalyze nucleotide addition in a 5' to 3' direction. In contrast, tRNA(His guanylyltransferase (Thg1 enzymes catalyze the unusual reverse addition (3' to 5' of nucleotides to polynucleotide substrates. In eukaryotes, Thg1 enzymes use the 3'-5' addition activity to add G-1 to the 5'-end of tRNA(His, a modification required for efficient aminoacylation of the tRNA by the histidyl-tRNA synthetase. Thg1-like proteins (TLPs are found in Archaea, Bacteria, and mitochondria and are biochemically distinct from their eukaryotic Thg1 counterparts TLPs catalyze 5'-end repair of truncated tRNAs and act on a broad range of tRNA substrates instead of exhibiting strict specificity for tRNA(His. Taken together, these data suggest that TLPs function in distinct biological pathways from the tRNA(His maturation pathway, perhaps in tRNA quality control. Here we present the first crystal structure of a TLP, from the gram-positive soil bacterium Bacillus thuringiensis (BtTLP. The enzyme is a tetramer like human THG1, with which it shares substantial structural similarity. Catalysis of the 3'-5' reaction with 5'-monophosphorylated tRNA necessitates first an activation step, generating a 5'-adenylylated intermediate prior to a second nucleotidyl transfer step, in which a nucleotide is transferred to the tRNA 5'-end. Consistent with earlier characterization of human THG1, we observed distinct binding sites for the nucleotides involved in these two steps of activation and nucleotidyl transfer. A BtTLP complex with GTP reveals new interactions with the GTP nucleotide in the activation site that were not evident from the previously solved structure. Moreover, the BtTLP-ATP structure allows direct observation of ATP in the activation site for the first time. The BtTLP structural data, combined with kinetic analysis of selected variants, provide new insight into the role of key residues in the activation step.

  11. Bacterial lung abscess

    International Nuclear Information System (INIS)

    Groskin, S.A.; Panicek, D.M.; Ewing, D.K.; Rivera, F.; Math, K.; Teixeira, J.; Heitzman, E.R.

    1987-01-01

    A retrospective review of patients with bacterial lung abscess was carried out. Demographic, clinical, and radiographical features of this patient group are compared with similar data from patients with empyema and/or cavitated lung carcinoma; differential diagnostic points are stressed. The entity of radiographically occult lung abscess is discussed. Complications associated with bacterial lung abscess are discussed. Current therapeutic options and treatment philosophy for patients with bacterial lung abscess are noted

  12. Peritonitis - spontaneous bacterial

    Science.gov (United States)

    Spontaneous bacterial peritonitis (SBP); Ascites - peritonitis; Cirrhosis - peritonitis ... who are on peritoneal dialysis for kidney failure. Peritonitis may have other causes . These include infection from ...

  13. MIPS bacterial genomes functional annotation benchmark dataset.

    Science.gov (United States)

    Tetko, Igor V; Brauner, Barbara; Dunger-Kaltenbach, Irmtraud; Frishman, Goar; Montrone, Corinna; Fobo, Gisela; Ruepp, Andreas; Antonov, Alexey V; Surmeli, Dimitrij; Mewes, Hans-Wernen

    2005-05-15

    Any development of new methods for automatic functional annotation of proteins according to their sequences requires high-quality data (as benchmark) as well as tedious preparatory work to generate sequence parameters required as input data for the machine learning methods. Different program settings and incompatible protocols make a comparison of the analyzed methods difficult. The MIPS Bacterial Functional Annotation Benchmark dataset (MIPS-BFAB) is a new, high-quality resource comprising four bacterial genomes manually annotated according to the MIPS functional catalogue (FunCat). These resources include precalculated sequence parameters, such as sequence similarity scores, InterPro domain composition and other parameters that could be used to develop and benchmark methods for functional annotation of bacterial protein sequences. These data are provided in XML format and can be used by scientists who are not necessarily experts in genome annotation. BFAB is available at http://mips.gsf.de/proj/bfab

  14. Effect of vaccines on bacterial meningitis worldwide

    NARCIS (Netherlands)

    McIntyre, Peter B.; O'Brien, Katherine L.; Greenwood, Brian; van de Beek, Diederik

    2012-01-01

    Three bacteria-Haemophilus influenzae, Streptococcus pneumoniae, and Neisseria meningitidis-account for most acute bacterial meningitis. Measurement of the effect of protein-polysaccharide conjugate vaccines is most reliable for H influenzae meningitis because one serotype and one age group account

  15. SnapShot: The Bacterial Cytoskeleton.

    Science.gov (United States)

    Fink, Gero; Szewczak-Harris, Andrzej; Löwe, Jan

    2016-07-14

    Most bacteria and archaea contain filamentous proteins and filament systems that are collectively known as the bacterial cytoskeleton, though not all of them are cytoskeletal, affect cell shape, or maintain intracellular organization. To view this SnapShot, open or download the PDF. Copyright © 2016. Published by Elsevier Inc.

  16. The normal bacterial flora prevents GI disease

    Indian Academy of Sciences (India)

    First page Back Continue Last page Overview Graphics. The normal bacterial flora prevents GI disease. Inhibits pathogenic enteric bacteria. Decrease luminal pH; Secrete bacteriocidal proteins; Colonization resistance; Block epithelial binding – induce MUC2. Improves epithelial and mucosal barrier integrity. Produce ...

  17. Structural Genomics of Bacterial Virulence Factors

    Science.gov (United States)

    2006-05-01

    Moller, T., T. Franch , P. Hojrup, D.R. Keene, H.P. Bachinger, R.G. Brennan, and P. Valentin-Hansen. 2002. Hfq: a bacterial Sm-like protein that...level, an unre- sponsiveness to external stimuli, or an inability to obtain readily available food or water, along with any of the following accompa

  18. Prevention of bacterial adhesion

    DEFF Research Database (Denmark)

    Klemm, Per; Vejborg, Rebecca Munk; Hancock, Viktoria

    2010-01-01

    . As such, adhesion represents the Achilles heel of crucial pathogenic functions. It follows that interference with adhesion can reduce bacterial virulence. Here, we illustrate this important topic with examples of techniques being developed that can inhibit bacterial adhesion. Some of these will become...

  19. Protein malnutrition and metronidazole induced intestinal bacterial ...

    African Journals Online (AJOL)

    STORAGESEVER

    2008-09-17

    Sep 17, 2008 ... Figure 1. Weight loss in different situation of diets. Each point represents ... using sterile ground-glass stoppers (Heimo et al., 2001). To .... In fact, selective elimination ... attention in recent years, because the ability of bacteria.

  20. Bacterial proteins pinpoint a single eukaryotic root

    Czech Academy of Sciences Publication Activity Database

    Derelle, R.; Torruella, G.; Klimeš, V.; Brinkmann, H.; Kim, E.; Vlček, Čestmír; Lang, B.F.; Eliáš, M.

    2015-01-01

    Roč. 112, č. 7 (2015), E693-E699 ISSN 0027-8424 R&D Projects: GA ČR GA13-24983S Grant - others:GA MŠk(CZ) ED2.1.00/03.0100; Howard Hughes Medical Institute International Early Career Scientist Program(US) 55007424; Spanish Ministry of Economy and Competitiveness, European Molecular Biology Organization Young Investigator Program(ES) BFU2012-31329; Spanish Ministry of Economy and Competitiveness, "Centro de Excelencia Severo Ochoa" - European Regional Development Fund(ES) Sev-2012-0208, BES-2013-064004 Institutional support: RVO:68378050 Keywords : eukaryote phylogeny * phylogenomics * Opimoda * Diphoda * LECA Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 9.423, year: 2015

  1. Bacterial Actins? An Evolutionary Perspective

    Science.gov (United States)

    Doolittle, Russell F.; York, Amanda L.

    2003-01-01

    According to the conventional wisdom, the existence of a cytoskeleton in eukaryotes and its absence in prokaryotes constitute a fundamental divide between the two domains of life. An integral part of the dogma is that a cytoskeleton enabled an early eukaryote to feed upon prokaryotes, a consequence of which was the occasional endosymbiosis and the eventual evolution of organelles. Two recent papers present compelling evidence that actin, one of the principal components of a cytoskeleton, has a homolog in Bacteria that behaves in many ways like eukaryotic actin. Sequence comparisons reveml that eukaryotic actin and the bacterial homolog (mreB protein), unlike many other proteins common to eukaryotes and Bacteria, have very different and more highly extended evolutionary histories.

  2. The papain inhibitor (SPI) of Streptomyces mobaraensis inhibits bacterial cysteine proteases and is an antagonist of bacterial growth

    NARCIS (Netherlands)

    S. Zindel (Stephan); W.E. Kaman (Wendy); S. Fröls (Sabrina); F. Pfeifer (Felicitas); A. Peters (Annette); J.P. Hays (John); H.-L. Fuchsbauer (Hans-Lothar)

    2013-01-01

    textabstractA novel papain inhibitory protein (SPI) from Streptomyces mobaraensis was studied to measure its inhibitory effect on bacterial cysteine protease activity (Staphylococcus aureus SspB) and culture supernatants (Porphyromonas gingivalis, Bacillus anthracis). Further, growth of Bacillus

  3. Bacterial Inclusion Bodies: Discovering Their Better Half.

    Science.gov (United States)

    Rinas, Ursula; Garcia-Fruitós, Elena; Corchero, José Luis; Vázquez, Esther; Seras-Franzoso, Joaquin; Villaverde, Antonio

    2017-09-01

    Bacterial inclusion bodies (IBs) are functional, non-toxic amyloids occurring in recombinant bacteria showing analogies with secretory granules of the mammalian endocrine system. The scientific interest in these mesoscale protein aggregates has been historically masked by their status as a hurdle in recombinant protein production. However, progressive understanding of how the cell handles the quality of recombinant polypeptides and the main features of their intriguing molecular organization has stimulated the interest in inclusion bodies and spurred their use in diverse technological fields. The engineering and tailoring of IBs as functional protein particles for materials science and biomedicine is a good example of how formerly undesired bacterial byproducts can be rediscovered as promising functional materials for a broad spectrum of applications. Copyright © 2017 Elsevier Ltd. All rights reserved.

  4. Molecular mechanisms of action of bacterial exotoxins.

    Science.gov (United States)

    Balfanz, J; Rautenberg, P; Ullmann, U

    1996-07-01

    Toxins are one of the inventive strategies that bacteria have developed in order to survive. As virulence factors, they play a major role in the pathogenesis of infectious diseases. Recent discoveries have once more highlighted the effectiveness of these precisely adjusted bacterial weapons. Furthermore, toxins have become an invaluable tool in the investigation of fundamental cell processes, including regulation of cellular functions by various G proteins, cytoskeletal dynamics and neural transmission. In this review, the bacterial toxins are presented in a rational classification based on the molecular mechanisms of action.

  5. Postviral Complications: Bacterial Pneumonia.

    Science.gov (United States)

    Prasso, Jason E; Deng, Jane C

    2017-03-01

    Secondary bacterial pneumonia after viral respiratory infection remains a significant source of morbidity and mortality. Susceptibility is mediated by a variety of viral and bacterial factors, and complex interactions with the host immune system. Prevention and treatment strategies are limited to influenza vaccination and antibiotics/antivirals respectively. Novel approaches to identifying the individuals with influenza who are at increased risk for secondary bacterial pneumonias are urgently needed. Given the threat of further pandemics and the heightened prevalence of these viruses, more research into the immunologic mechanisms of this disease is warranted with the hope of discovering new potential therapies. Published by Elsevier Inc.

  6. Bacterial vaginosis - aftercare

    Science.gov (United States)

    Bacterial vaginosis (BV) is a type of vaginal infection. The vagina normally contains both healthy bacteria and unhealthy bacteria. BV occurs when more unhealthy bacteria grow than healthy bacteria. No one knows ...

  7. Bacterial surface adaptation

    Science.gov (United States)

    Utada, Andrew

    2014-03-01

    Biofilms are structured multi-cellular communities that are fundamental to the biology and ecology of bacteria. Parasitic bacterial biofilms can cause lethal infections and biofouling, but commensal bacterial biofilms, such as those found in the gut, can break down otherwise indigestible plant polysaccharides and allow us to enjoy vegetables. The first step in biofilm formation, adaptation to life on a surface, requires a working knowledge of low Reynolds number fluid physics, and the coordination of biochemical signaling, polysaccharide production, and molecular motility motors. These crucial early stages of biofilm formation are at present poorly understood. By adapting methods from soft matter physics, we dissect bacterial social behavior at the single cell level for several prototypical bacterial species, including Pseudomonas aeruginosa and Vibrio cholerae.

  8. Diagnosis of bacterial infection

    African Journals Online (AJOL)

    direct or indirect evidence of a compatible bacterial pathogen. Inflammation may be .... cardinal features (fever, confusion, headache and neck stiffness). .... specificity, inappropriate indications or poor sampling technique may diminish this ...

  9. Biodegradability of bacterial surfactants.

    Science.gov (United States)

    Lima, Tânia M S; Procópio, Lorena C; Brandão, Felipe D; Carvalho, André M X; Tótola, Marcos R; Borges, Arnaldo C

    2011-06-01

    This work aimed at evaluating the biodegradability of different bacterial surfactants in liquid medium and in soil microcosms. The biodegradability of biosurfactants by pure and mixed bacterial cultures was evaluated through CO(2) evolution. Three bacterial strains, Acinetobacter baumanni LBBMA ES11, Acinetobacter haemolyticus LBBMA 53 and Pseudomonas sp. LBBMA 101B, used the biosurfactants produced by Bacillus sp. LBBMA 111A (mixed lipopeptide), Bacillus subtilis LBBMA 155 (lipopeptide), Flavobacterium sp. LBBMA 168 (mixture of flavolipids), Dietzia Maris LBBMA 191(glycolipid) and Arthrobacter oxydans LBBMA 201(lipopeptide) as carbon sources in minimal medium. The synthetic surfactant sodium dodecyl sulfate (SDS) was also mineralized by these microorganisms, but at a lower rate. CO(2) emitted by a mixed bacterial culture in soil microcosms with biosurfactants was higher than in the microcosm containing SDS. Biosurfactant mineralization in soil was confirmed by the increase in surface tension of the soil aqueous extracts after incubation with the mixed bacterial culture. It can be concluded that, in terms of biodegradability and environmental security, these compounds are more suitable for applications in remediation technologies in comparison to synthetic surfactants. However, more information is needed on structure of biosurfactants, their interaction with soil and contaminants and scale up and cost for biosurfactant production.

  10. Structure and operation of bacterial tripartite pumps.

    Science.gov (United States)

    Hinchliffe, Philip; Symmons, Martyn F; Hughes, Colin; Koronakis, Vassilis

    2013-01-01

    In bacteria such as Pseudomonas aeruginosa and Escherichia coli, tripartite membrane machineries, or pumps, determine the efflux of small noxious molecules, such as detergents, heavy metals, and antibiotics, and the export of large proteins including toxins. They are therefore influential in bacterial survival, particularly during infections caused by multidrug-resistant pathogens. In these tripartite pumps an inner membrane transporter, typically an ATPase or proton antiporter, binds and translocates export or efflux substrates. In cooperation with a periplasmic adaptor protein it recruits and opens a TolC family cell exit duct, which is anchored in the outer membrane and projects across the periplasmic space between inner and outer membranes. Assembled tripartite pumps thus span the entire bacterial cell envelope. We review the atomic structures of each of the three pump components and discuss how these have allowed high-resolution views of tripartite pump assembly, operation, and possible inhibition.

  11. Method of inactivation of viral and bacterial blood contaminants

    International Nuclear Information System (INIS)

    Hackett, R.; Goodrich, R.P.; Van Borssum Waalkes, M.; Wong, V.A.

    1992-01-01

    A method is provided for inactivating viral and/or bacterial contamination in blood cellular matter, such as erythrocytes and platelets, or protein fractions. The cells or protein fractions are mixed with chemical sensitizers and irradiated with, for example, gamma or X-ray radiation

  12. Bithionol blocks pathogenicity of bacterial toxins, ricin, and Zika virus

    Science.gov (United States)

    Disease pathways form overlapping networks, and hub proteins represent attractive targets for broad-spectrum drugs. Using bacterial toxins as a proof of concept, we describe a new approach of discovering broad-spectrum therapies capable of inhibiting host proteins that mediate multiple pathogenic pa...

  13. Bacterial meningitis in children

    International Nuclear Information System (INIS)

    Marji, S.

    2007-01-01

    To demonstrate the epidemiology, clinical manifestations and bacteriological profile of bacterial meningitis in children beyond the neonatal period in our hospital. This was a retrospective descriptive study conducted at Prince Rashid Hospital in Irbid, Jordan. The medical records of 50 children with the diagnosis of bacterial meningitis during 4 years period, were reviewed. The main cause of infection was streptococcus pneumoniae, followed by Haemophilus influenza and Niesseria meningitides. Mortality was higher in infants and meningococcal infection, while complications were more encountered in cases of streptococcus pneumoniae. Cerebrospinal fluid culture was positive in 11 cases and Latex agglutination test in 39. There is a significant reduction of the numbers of bacterial meningitis caused by Haemophilus influenza type B species. (author)

  14. Rapid Identification of Bacterial Virulence Factors

    Science.gov (United States)

    2014-04-15

    protein sorting and transport. F/’/wyi-deletion mutants had decreased invasiveness of HeLa cells when compared to their parental strain, and it has...mileux. Bacteria with intracellular life styles and have reductive genomes often have many different ABC transporters. This is certainly the case in...34 Microbiology 151:2975-2986. Newman , R.M., P. Salunkhe, A. Godzik, J.C. Reed. 2006. Identification and Characterization of a Novel Bacterial

  15. STRUCTURAL ORGANIZATION OF BACTERIAL UREASES

    Directory of Open Access Journals (Sweden)

    Lisnyak YuV

    2016-09-01

    Full Text Available This brief review concerns the basic principles of structural organization of multi-subunit bacterial ureases and formation of their quaternary structure. Urease is a nickel-containing enzyme (urea amidohydrolase, ЕС 3.5.1.5 that catalyses the hydrolysis of urea to get ammonia and carbamate which then decomposes with water to get ammonia and carbon dioxide. Urease is produced by bacteria, fungi, yeast and plants. On the basis of similarities in amino acid sequences, ureases assumed to have a similar structure and conservative catalytic mechanism. Within past two decades bacterial ureases have gained much attention in research field as a virulence factor in human and animal infections. The first crystal structure of urease has been determined for that from Klebsiella aerogenes. The native enzyme consists of three subunits, UreA (α-chain, UreB (β-chain and UreC (γ-chain, and contains four structural domains: two in α-chain (α-domain 1 and α-domain-2, one in β- and one in γ-chain. These three chains form a T-shaped heterotrimer αβγ. Three αβγ heterotrimers form quaternary complex (αβγ3. In case of Helicobacter pilori, the analogous trimers of corresponding dimeric subunits (αβ3 form tetrameric structure ((αβ34 in which four trimers are situated at the vertexes of the regular triangle pyramid. Active center is located in α-domain 1 and contains two atoms of nickel coordinated by residues His134, His136, carboxylated Lys217, His 246, His272 and Asp360, as well as residues involved in binding (His219 and catalysis (His320. Active site is capped by a flap that controls substrate ingress to and product egress from the dinickel center. Urease requires accessory proteins (UreD, UreF, UreG and UreE for the correct assembly of their Ni-containing metallocenters. The accessory proteins UreD, UreF, and UreG sequentially bind to the apoprotein (UreABC3 to finally form (UreABC-UreDFG3 activation complex. UreE metallochaperone delivers

  16. Adult bacterial meningitis

    DEFF Research Database (Denmark)

    Meyer, C N; Samuelsson, I S; Galle, M

    2004-01-01

    Episodes of adult bacterial meningitis (ABM) at a Danish hospital in 1991-2000 were identified from the databases of the Department of Clinical Microbiology, and compared with data from the Danish National Patient Register and the Danish National Notification System. Reduced penicillin susceptibi......Episodes of adult bacterial meningitis (ABM) at a Danish hospital in 1991-2000 were identified from the databases of the Department of Clinical Microbiology, and compared with data from the Danish National Patient Register and the Danish National Notification System. Reduced penicillin...

  17. The Carboxysome and Other Bacterial Microcompartments

    Energy Technology Data Exchange (ETDEWEB)

    Kerfeld, Cheryl A.; Greenleaf, William B.; Kinney, James N.

    2010-06-23

    - Carboxysomes are part of the carbon concentrating mechanism in cyanobacteria and chemoautotrophs. - Carboxysomes are a subclass of bacterial microcompartments (BMCs); BMCs can encapsulate a range of metabolic processes. - Like some viral particles, the carboxysome can be modeled as an icosahedron-in its case, having 4,000-5,000 hexameric shell subunits and 12 surface pentamers to generate curvature. - The threefold axis of symmetry of the CsoS1D protein in carboxysomes forms a pore that can open and close, allowing for selective diffusion. - Genetic modules encoding BMC shell proteins and the enzymes that they encapsulate are horizontally transferable, suggesting they enable bacteria to adapt to diverse environments.

  18. [Bacterial biofilms and infection].

    Science.gov (United States)

    Lasa, I; Del Pozo, J L; Penadés, J R; Leiva, J

    2005-01-01

    In developed countries we tend to think of heart disease and the numerous forms of cancer as the main causes of mortality, but on a global scale infectious diseases come close, or may even be ahead: 14.9 million deaths in 2002 compared to cardiovascular diseases (16.9 million deaths) and cancer (7.1 million deaths) (WHO report 2004). The infectious agents responsible for human mortality have evolved as medical techniques and hygienic measures have changed. Modern-day acute infectious diseases caused by specialized bacterial pathogens such as diphtheria, tetanus, cholera, plague, which represented the main causes of death at the beginning of XX century, have been effectively controlled with antibiotics and vaccines. In their place, more than half of the infectious diseases that affect mildly immunocompromised patients involve bacterial species that are commensal with the human body; these can produce chronic infections, are resistant to antimicrobial agents and there is no effective vaccine against them. Examples of these infections are the otitis media, native valve endocarditis, chronic urinary infections, bacterial prostatitis, osteomyelitis and all the infections related to medical devices. Direct analysis of the surface of medical devices or of tissues that have been foci of chronic infections shows the presence of large numbers of bacteria surrounded by an exopolysaccharide matrix, which has been named the "biofilm". Inside the biofilm, bacteria grow protected from the action of the antibodies, phagocytic cells and antimicrobial treatments. In this article, we describe the role of bacterial biofilms in human persistent infections.

  19. Interfering with bacterial gossip

    DEFF Research Database (Denmark)

    Bjarnsholt, Thomas; Tolker-Nielsen, Tim; Givskov, Michael

    2011-01-01

    that appropriately target bacteria in their relevant habitat with the aim of mitigating their destructive impact on patients. In this review we describe molecular mechanisms involved in “bacterial gossip” (more scientifically referred to as quorum sensing (QS) and c-di-GMP signaling), virulence, biofilm formation...

  20. Bacterial fingerprints across Europe

    NARCIS (Netherlands)

    Glasner, Corinna

    2014-01-01

    Bacterial pathogens, such as Staphylococcus aureus and carbapenemase-producing Enterobacteriaceae (CPE), impose major threats to human health worldwide. Both have a ‘Jekyll & Hyde’ character, since they can be present as human commensals, but can also become harmful invasive pathogens especially

  1. Towards understanding the molecular basis of bacterial DNA segregation

    DEFF Research Database (Denmark)

    Leonard, Thomas A.; Møller-Jensen, Jakob; Löwe, Jan

    2005-01-01

    Bacteria ensure the fidelity of genetic inheritance by the coordinated control of chromosome segregation and cell division. Here, we review the molecules and mechanisms that govern the correct subcellular positioning and rapid separation of newly replicated chromosomes and plasmids towards the ce...... common to the two processes. Finally, we discuss the role that the bacterial cytoskeleton plays in DNA partitioning and the missing link between prokaryotes and eukaryotes that is bacterial mechano-chemical motor proteins. Udgivelsesdato: Mar 29...

  2. Corticosteroids for Bacterial Keratitis

    Science.gov (United States)

    Srinivasan, Muthiah; Mascarenhas, Jeena; Rajaraman, Revathi; Ravindran, Meenakshi; Lalitha, Prajna; Glidden, David V.; Ray, Kathryn J.; Hong, Kevin C.; Oldenburg, Catherine E.; Lee, Salena M.; Zegans, Michael E.; McLeod, Stephen D.; Lietman, Thomas M.; Acharya, Nisha R.

    2013-01-01

    Objective To determine whether there is a benefit in clinical outcomes with the use of topical corticosteroids as adjunctive therapy in the treatment of bacterial corneal ulcers. Methods Randomized, placebo-controlled, double-masked, multicenter clinical trial comparing prednisolone sodium phosphate, 1.0%, to placebo as adjunctive therapy for the treatment of bacterial corneal ulcers. Eligible patients had a culture-positive bacterial corneal ulcer and received topical moxifloxacin for at least 48 hours before randomization. Main Outcome Measures The primary outcome was best spectacle-corrected visual acuity (BSCVA) at 3 months from enrollment. Secondary outcomes included infiltrate/scar size, reepithelialization, and corneal perforation. Results Between September 1, 2006, and February 22, 2010, 1769 patients were screened for the trial and 500 patients were enrolled. No significant difference was observed in the 3-month BSCVA (−0.009 logarithm of the minimum angle of resolution [logMAR]; 95% CI, −0.085 to 0.068; P = .82), infiltrate/scar size (P = .40), time to reepithelialization (P = .44), or corneal perforation (P > .99). A significant effect of corticosteroids was observed in subgroups of baseline BSCVA (P = .03) and ulcer location (P = .04). At 3 months, patients with vision of counting fingers or worse at baseline had 0.17 logMAR better visual acuity with corticosteroids (95% CI, −0.31 to −0.02; P = .03) compared with placebo, and patients with ulcers that were completely central at baseline had 0.20 logMAR better visual acuity with corticosteroids (−0.37 to −0.04; P = .02). Conclusions We found no overall difference in 3-month BSCVA and no safety concerns with adjunctive corticosteroid therapy for bacterial corneal ulcers. Application to Clinical Practice Adjunctive topical corticosteroid use does not improve 3-month vision in patients with bacterial corneal ulcers. PMID:21987582

  3. LATERAL GENE TRANSFER AND THE HISTORY OF BACTERIAL GENOMES

    Energy Technology Data Exchange (ETDEWEB)

    Howard Ochman

    2006-02-22

    The aims of this research were to elucidate the role and extent of lateral transfer in the differentiation of bacterial strains and species, and to assess the impact of gene transfer on the evolution of bacterial genomes. The ultimate goal of the project is to examine the dynamics of a core set of protein-coding genes (i.e., those that are distributed universally among Bacteria) by developing conserved primers that would allow their amplification and sequencing in any bacterial taxa. In addition, we adopted a bioinformatic approach to elucidate the extent of lateral gene transfer in sequenced genome.

  4. The type III protein secretion system contributes to Xanthomonas citri subsp. citri biofilm formation

    KAUST Repository

    Zimaro, Tamara; Thomas, Ludivine; Marondedze, Claudius; Sgro, Germá n G; Garofalo, Cecilia G; Ficarra, Florencia A; Gehring, Christoph A; Ottado, Jorgelina; Gottig, Natalia

    2014-01-01

    Background: Several bacterial plant pathogens colonize their hosts through the secretion of effector proteins by a Type III protein secretion system (T3SS). The role of T3SS in bacterial pathogenesis is well established but whether this system

  5. Copper effects on bacterial activity of estuarine silty sediments

    Science.gov (United States)

    Almeida, Adelaide; Cunha, Ângela; Fernandes, Sandra; Sobral, Paula; Alcântara, Fernanda

    2007-07-01

    , mainly, by the great intensification of bacterial biomass production and leucine turnover rate. We conclude that the bacterial community of silty estuarine sediments seems to withstand considerable concentrations of copper at the cost of reduced bacterial organic matter degradation and of the almost halting of bacterial production. The toxic effects elicited by copper on protein and carbohydrate degradation were not rapidly repaired by erosion and oxygenation of the sediment cells but, in contrast, bacterial biomass production and leucine turnover were rapidly and efficiently reactivated.

  6. Bacterial infec tions in travellers

    African Journals Online (AJOL)

    namely bacterial causes of travellers' diarrhoea and skin infections, as well as .... Vaccination: protective efficacy against typhoid may be overcome by ingesting a high bacterial load. Vaccine ..... preparation such as cream sauce. Only after ...

  7. Discovery of novel bacterial toxins by genomics and computational biology.

    Science.gov (United States)

    Doxey, Andrew C; Mansfield, Michael J; Montecucco, Cesare

    2018-06-01

    Hundreds and hundreds of bacterial protein toxins are presently known. Traditionally, toxin identification begins with pathological studies of bacterial infectious disease. Following identification and cultivation of a bacterial pathogen, the protein toxin is purified from the culture medium and its pathogenic activity is studied using the methods of biochemistry and structural biology, cell biology, tissue and organ biology, and appropriate animal models, supplemented by bioimaging techniques. The ongoing and explosive development of high-throughput DNA sequencing and bioinformatic approaches have set in motion a revolution in many fields of biology, including microbiology. One consequence is that genes encoding novel bacterial toxins can be identified by bioinformatic and computational methods based on previous knowledge accumulated from studies of the biology and pathology of thousands of known bacterial protein toxins. Starting from the paradigmatic cases of diphtheria toxin, tetanus and botulinum neurotoxins, this review discusses traditional experimental approaches as well as bioinformatics and genomics-driven approaches that facilitate the discovery of novel bacterial toxins. We discuss recent work on the identification of novel botulinum-like toxins from genera such as Weissella, Chryseobacterium, and Enteroccocus, and the implications of these computationally identified toxins in the field. Finally, we discuss the promise of metagenomics in the discovery of novel toxins and their ecological niches, and present data suggesting the existence of uncharacterized, botulinum-like toxin genes in insect gut metagenomes. Copyright © 2018. Published by Elsevier Ltd.

  8. Factors influencing bacterial adhesion to contact lenses.

    Science.gov (United States)

    Dutta, Debarun; Cole, Nerida; Willcox, Mark

    2012-01-01

    The process of any contact lens related keratitis generally starts with the adhesion of opportunistic pathogens to contact lens surface. This article focuses on identifying the factors which have been reported to affect bacterial adhesion to contact lenses. Adhesion to lenses differs between various genera/species/strains of bacteria. Pseudomonas aeruginosa, which is the predominant causative organism, adheres in the highest numbers to both hydrogel and silicone hydrogel lenses in vitro. The adhesion of this strain reaches maximum numbers within 1h in most in vitro studies and a biofilm has generally formed within 24 h of cells adhering to the lens surface. Physical and chemical properties of contact lens material affect bacterial adhesion. The water content of hydroxyethylmethacrylate (HEMA)-based lenses and their iconicity affect the ability of bacteria to adhere. The higher hydrophobicity of silicone hydrogel lenses compared to HEMA-based lenses has been implicated in the higher numbers of bacteria that can adhere to their surfaces. Lens wear has different effects on bacterial adhesion, partly due to differences between wearers, responses of bacterial strains and the ability of certain tear film proteins when bound to a lens surface to kill certain types of bacteria.

  9. Structure of bacterial lipopolysaccharides.

    Science.gov (United States)

    Caroff, Martine; Karibian, Doris

    2003-11-14

    Bacterial lipopolysaccharides are the major components of the outer surface of Gram-negative bacteria They are often of interest in medicine for their immunomodulatory properties. In small amounts they can be beneficial, but in larger amounts they may cause endotoxic shock. Although they share a common architecture, their structural details exert a strong influence on their activity. These molecules comprise: a lipid moiety, called lipid A, which is considered to be the endotoxic component, a glycosidic part consisting of a core of approximately 10 monosaccharides and, in "smooth-type" lipopolysaccharides, a third region, named O-chain, consisting of repetitive subunits of one to eight monosaccharides responsible for much of the immunospecificity of the bacterial cell.

  10. Cooperative Bacterial Foraging Optimization

    Directory of Open Access Journals (Sweden)

    Hanning Chen

    2009-01-01

    Full Text Available Bacterial Foraging Optimization (BFO is a novel optimization algorithm based on the social foraging behavior of E. coli bacteria. This paper presents a variation on the original BFO algorithm, namely, the Cooperative Bacterial Foraging Optimization (CBFO, which significantly improve the original BFO in solving complex optimization problems. This significant improvement is achieved by applying two cooperative approaches to the original BFO, namely, the serial heterogeneous cooperation on the implicit space decomposition level and the serial heterogeneous cooperation on the hybrid space decomposition level. The experiments compare the performance of two CBFO variants with the original BFO, the standard PSO and a real-coded GA on four widely used benchmark functions. The new method shows a marked improvement in performance over the original BFO and appears to be comparable with the PSO and GA.

  11. Bacterial control of cyanobacteria

    CSIR Research Space (South Africa)

    Ndlela, Luyanda L

    2017-08-01

    Full Text Available of biological control appears to be direct contact. • Ndlela, L. L. et al. (2016) ‘An overview of cyanobacterial bloom occurrences and research in Africa over the last decade’, Harmful Algae, 60 • Gumbo, J.R. et al. (2010) The Isolation and identification... of Predatory Bacteria from a Microcystis algal Bloom.. African Journal of Biotechnology, 9. *Special acknowledgement goes to the National Research foundation for funding this presentation Bacterial control of cyanobacteria Luyanda...

  12. Bacterial lipoproteins; biogenesis, sorting and quality control.

    Science.gov (United States)

    Narita, Shin-Ichiro; Tokuda, Hajime

    2017-11-01

    Bacterial lipoproteins are a subset of membrane proteins localized on either leaflet of the lipid bilayer. These proteins are anchored to membranes through their N-terminal lipid moiety attached to a conserved Cys. Since the protein moiety of most lipoproteins is hydrophilic, they are expected to play various roles in a hydrophilic environment outside the cytoplasmic membrane. Gram-negative bacteria such as Escherichia coli possess an outer membrane, to which most lipoproteins are sorted. The Lol pathway plays a central role in the sorting of lipoproteins to the outer membrane after lipoprotein precursors are processed to mature forms in the cytoplasmic membrane. Most lipoproteins are anchored to the inner leaflet of the outer membrane with their protein moiety in the periplasm. However, recent studies indicated that some lipoproteins further undergo topology change in the outer membrane, and play critical roles in the biogenesis and quality control of the outer membrane. This article is part of a Special Issue entitled: Bacterial Lipids edited by Russell E. Bishop. Copyright © 2016 Elsevier B.V. All rights reserved.

  13. Serum amyloid A and C-reactive protein combined determination for diagnosis of bacterial infection in neonates%血清淀粉样蛋白A与C反应蛋白联合检测在新生儿细菌感染诊断中的应用

    Institute of Scientific and Technical Information of China (English)

    沈伟虹; 岳朝艳; 孙振东; 应春妹

    2016-01-01

    目的:研究血清淀粉样蛋白 A(SAA)和 C 反应蛋白(CRP)以及白细胞总数在新生儿细菌感染诊断中的应用价值。方法分别检测80例细菌感染新生儿及58名体检正常的新生儿(非感染组)SAA、CRP 和白细胞总数并做比较。结果新生儿细菌感染组 SAA 与 CRP 水平分别为(67.3±38.4)、(19.2±11.2)mg/L,明显高于非感染组[(8.7±2.1)、(5.6±2.5)mg/L](P15.00×109/L,明显高于非感染组[24例(41%)]。结论 SAA 和 CRP 可作为新生儿细菌感染诊断的重要指标,SAA 较 CRP 更为敏感,SAA 与 CRP 的联合检测能够提高新生儿细菌感染的检出率,为疾病的早期诊断和监测提供实验依据。%Objective To study the significance of serum amyloid A(SAA),C-reactive protein(CRP)and white blood cell count determinations for diagnosis of bacterial infection in neonates. Methods SAA,CRP and white blood cell count were determined in 80 neonates with bacterial infection and 58 healthy neonates(non-bacterial infection group),and the results were analyzed. Results The concentrations of SAA and CRP in infection group [(67.3±38.4)and(19.2±11.2) mg/L] were significantly higher than those in non-bacterial infection group [(8.7±2.1)and(5.6±2.5)mg/L](P15.00×109/L in 64 neonates of 80 cases of infection group(80%),which was higher than that in non-bacterial infection group [24 cases(41%)]. Conclusions SAA and CRP could be used as important markers for monitoring bacterial infection in neonates,and SAA is more sensitive than CRP. The combined determination of SAA and CRP could improve the determination rate of bacterial infection,which provides an experimental reference for early diagnosis and monitoring of bacterial infection.

  14. Bacterial growth kinetics

    International Nuclear Information System (INIS)

    Boonkitticharoen, V.; Ehrhardt, J.C.; Kirchner, P.T.

    1989-01-01

    Quantitative measurement of bacterial growth may be made using a radioassay technique. This method measures, by scintillation counting, the 14 CO 2 derived from the bacterial metabolism of a 14 C-labeled substrate. Mathematical growth models may serve as reliable tools for estimation of the generation rate constant (or slope of the growth curve) and provide a basis for evaluating assay performance. Two models, i.e., exponential and logistic, are proposed. Both models yielded an accurate fit to the data from radioactive measurement of bacterial growth. The exponential model yielded high precision values of the generation rate constant, with an average relative standard deviation of 1.2%. Under most conditions the assay demonstrated no changes in the slopes of growth curves when the number of bacteria per inoculation was changed. However, the radiometric assay by scintillation method had a growth-inhibiting effect on a few strains of bacteria. The source of this problem was thought to be hypersensitivity to trace amounts of toluene remaining on the detector

  15. Adaptive Bacterial Foraging Optimization

    Directory of Open Access Journals (Sweden)

    Hanning Chen

    2011-01-01

    Full Text Available Bacterial Foraging Optimization (BFO is a recently developed nature-inspired optimization algorithm, which is based on the foraging behavior of E. coli bacteria. Up to now, BFO has been applied successfully to some engineering problems due to its simplicity and ease of implementation. However, BFO possesses a poor convergence behavior over complex optimization problems as compared to other nature-inspired optimization techniques. This paper first analyzes how the run-length unit parameter of BFO controls the exploration of the whole search space and the exploitation of the promising areas. Then it presents a variation on the original BFO, called the adaptive bacterial foraging optimization (ABFO, employing the adaptive foraging strategies to improve the performance of the original BFO. This improvement is achieved by enabling the bacterial foraging algorithm to adjust the run-length unit parameter dynamically during algorithm execution in order to balance the exploration/exploitation tradeoff. The experiments compare the performance of two versions of ABFO with the original BFO, the standard particle swarm optimization (PSO and a real-coded genetic algorithm (GA on four widely-used benchmark functions. The proposed ABFO shows a marked improvement in performance over the original BFO and appears to be comparable with the PSO and GA.

  16. A growing family: the expanding universe of the bacterial cytoskeleton.

    Science.gov (United States)

    Ingerson-Mahar, Michael; Gitai, Zemer

    2012-01-01

    Cytoskeletal proteins are important mediators of cellular organization in both eukaryotes and bacteria. In the past, cytoskeletal studies have largely focused on three major cytoskeletal families, namely the eukaryotic actin, tubulin, and intermediate filament (IF) proteins and their bacterial homologs MreB, FtsZ, and crescentin. However, mounting evidence suggests that these proteins represent only the tip of the iceberg, as the cellular cytoskeletal network is far more complex. In bacteria, each of MreB, FtsZ, and crescentin represents only one member of large families of diverse homologs. There are also newly identified bacterial cytoskeletal proteins with no eukaryotic homologs, such as WACA proteins and bactofilins. Furthermore, there are universally conserved proteins, such as the metabolic enzyme CtpS, that assemble into filamentous structures that can be repurposed for structural cytoskeletal functions. Recent studies have also identified an increasing number of eukaryotic cytoskeletal proteins that are unrelated to actin, tubulin, and IFs, such that expanding our understanding of cytoskeletal proteins is advancing the understanding of the cell biology of all organisms. Here, we summarize the recent explosion in the identification of new members of the bacterial cytoskeleton and describe a hypothesis for the evolution of the cytoskeleton from self-assembling enzymes. © 2011 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

  17. The clinical impact of bacterial biofilms

    DEFF Research Database (Denmark)

    Høiby, Niels; Ciofu, Oana; Johansen, Helle Krogh

    2011-01-01

    Bacteria survive in nature by forming biofilms on surfaces and probably most, if not all, bacteria (and fungi) are capable of forming biofilms. A biofilm is a structured consortium of bacteria embedded in a self-produced polymer matrix consisting of polysaccharide, protein and extracellular DNA....... Bacterial biofilms are resistant to antibiotics, disinfectant chemicals and to phagocytosis and other components of the innate and adaptive inflammatory defense system of the body. It is known, for example, that persistence of staphylococcal infections related to foreign bodies is due to biofilm formation....... Likewise, chronic Pseudomonas aeruginosa lung infections in cystic fibrosis patients are caused by biofilm growing mucoid strains. Gradients of nutrients and oxygen exist from the top to the bottom of biofilms and the bacterial cells located in nutrient poor areas have decreased metabolic activity...

  18. Gene calling and bacterial genome annotation with BG7.

    Science.gov (United States)

    Tobes, Raquel; Pareja-Tobes, Pablo; Manrique, Marina; Pareja-Tobes, Eduardo; Kovach, Evdokim; Alekhin, Alexey; Pareja, Eduardo

    2015-01-01

    New massive sequencing technologies are providing many bacterial genome sequences from diverse taxa but a refined annotation of these genomes is crucial for obtaining scientific findings and new knowledge. Thus, bacterial genome annotation has emerged as a key point to investigate in bacteria. Any efficient tool designed specifically to annotate bacterial genomes sequenced with massively parallel technologies has to consider the specific features of bacterial genomes (absence of introns and scarcity of nonprotein-coding sequence) and of next-generation sequencing (NGS) technologies (presence of errors and not perfectly assembled genomes). These features make it convenient to focus on coding regions and, hence, on protein sequences that are the elements directly related with biological functions. In this chapter we describe how to annotate bacterial genomes with BG7, an open-source tool based on a protein-centered gene calling/annotation paradigm. BG7 is specifically designed for the annotation of bacterial genomes sequenced with NGS. This tool is sequence error tolerant maintaining their capabilities for the annotation of highly fragmented genomes or for annotating mixed sequences coming from several genomes (as those obtained through metagenomics samples). BG7 has been designed with scalability as a requirement, with a computing infrastructure completely based on cloud computing (Amazon Web Services).

  19. Radiology of bacterial pneumonia

    International Nuclear Information System (INIS)

    Vilar, Jose; Domingo, Maria Luisa; Soto, Cristina; Cogollos, Jonathan

    2004-01-01

    Bacterial pneumonia is commonly encountered in clinical practice. Radiology plays a prominent role in the evaluation of pneumonia. Chest radiography is the most commonly used imaging tool in pneumonias due to its availability and excellent cost benefit ratio. CT should be used in unresolved cases or when complications of pneumonia are suspected. The main applications of radiology in pneumonia are oriented to detection, characterisation and follow-up, especially regarding complications. The classical classification of pneumonias into lobar and bronchial pneumonia has been abandoned for a more clinical classification. Thus, bacterial pneumonias are typified into three main groups: Community acquired pneumonia (CAD), Aspiration pneumonia and Nosocomial pneumonia (NP).The usual pattern of CAD is that of the previously called lobar pneumonia; an air-space consolidation limited to one lobe or segment. Nevertheless, the radiographic patterns of CAD may be variable and are often related to the causative agent. Aspiration pneumonia generally involves the lower lobes with bilateral multicentric opacities. Nosocomial Pneumonia (NP) occurs in hospitalised patients. The importance of NP is related to its high mortality and, thus, the need to obtain a prompt diagnosis. The role of imaging in NP is limited but decisive. The most valuable information is when the chest radiographs are negative and rule out pneumonia. The radiographic patterns of NP are very variable, most commonly showing diffuse multifocal involvement and pleural effusion. Imaging plays also an important role in the detection and evaluation of complications of bacterial pneumonias. In many of these cases, especially in hospitalised patients, chest CT must be obtained in order to better depict these associate findings

  20. Radiology of bacterial pneumonia

    Energy Technology Data Exchange (ETDEWEB)

    Vilar, Jose E-mail: vilar_jlu@gva.es; Domingo, Maria Luisa; Soto, Cristina; Cogollos, Jonathan

    2004-08-01

    Bacterial pneumonia is commonly encountered in clinical practice. Radiology plays a prominent role in the evaluation of pneumonia. Chest radiography is the most commonly used imaging tool in pneumonias due to its availability and excellent cost benefit ratio. CT should be used in unresolved cases or when complications of pneumonia are suspected. The main applications of radiology in pneumonia are oriented to detection, characterisation and follow-up, especially regarding complications. The classical classification of pneumonias into lobar and bronchial pneumonia has been abandoned for a more clinical classification. Thus, bacterial pneumonias are typified into three main groups: Community acquired pneumonia (CAD), Aspiration pneumonia and Nosocomial pneumonia (NP).The usual pattern of CAD is that of the previously called lobar pneumonia; an air-space consolidation limited to one lobe or segment. Nevertheless, the radiographic patterns of CAD may be variable and are often related to the causative agent. Aspiration pneumonia generally involves the lower lobes with bilateral multicentric opacities. Nosocomial Pneumonia (NP) occurs in hospitalised patients. The importance of NP is related to its high mortality and, thus, the need to obtain a prompt diagnosis. The role of imaging in NP is limited but decisive. The most valuable information is when the chest radiographs are negative and rule out pneumonia. The radiographic patterns of NP are very variable, most commonly showing diffuse multifocal involvement and pleural effusion. Imaging plays also an important role in the detection and evaluation of complications of bacterial pneumonias. In many of these cases, especially in hospitalised patients, chest CT must be obtained in order to better depict these associate findings.

  1. Bacterial Multidrug Efflux Pumps: Much More Than Antibiotic Resistance Determinants.

    Science.gov (United States)

    Blanco, Paula; Hernando-Amado, Sara; Reales-Calderon, Jose Antonio; Corona, Fernando; Lira, Felipe; Alcalde-Rico, Manuel; Bernardini, Alejandra; Sanchez, Maria Blanca; Martinez, Jose Luis

    2016-02-16

    Bacterial multidrug efflux pumps are antibiotic resistance determinants present in all microorganisms. With few exceptions, they are chromosomally encoded and present a conserved organization both at the genetic and at the protein levels. In addition, most, if not all, strains of a given bacterial species present the same chromosomally-encoded efflux pumps. Altogether this indicates that multidrug efflux pumps are ancient elements encoded in bacterial genomes long before the recent use of antibiotics for human and animal therapy. In this regard, it is worth mentioning that efflux pumps can extrude a wide range of substrates that include, besides antibiotics, heavy metals, organic pollutants, plant-produced compounds, quorum sensing signals or bacterial metabolites, among others. In the current review, we present information on the different functions that multidrug efflux pumps may have for the bacterial behaviour in different habitats as well as on their regulation by specific signals. Since, in addition to their function in non-clinical ecosystems, multidrug efflux pumps contribute to intrinsic, acquired, and phenotypic resistance of bacterial pathogens, the review also presents information on the search for inhibitors of multidrug efflux pumps, which are currently under development, in the aim of increasing the susceptibility of bacterial pathogens to antibiotics.

  2. Cytosolic Access of Intracellular Bacterial Pathogens: The Shigella Paradigm.

    Science.gov (United States)

    Mellouk, Nora; Enninga, Jost

    2016-01-01

    Shigella is a Gram-negative bacterial pathogen, which causes bacillary dysentery in humans. A crucial step of Shigella infection is its invasion of epithelial cells. Using a type III secretion system, Shigella injects several bacterial effectors ultimately leading to bacterial internalization within a vacuole. Then, Shigella escapes rapidly from the vacuole, it replicates within the cytosol and spreads from cell-to-cell. The molecular mechanism of vacuolar rupture used by Shigella has been studied in some detail during the recent years and new paradigms are emerging about the underlying molecular events. For decades, bacterial effector proteins were portrayed as main actors inducing vacuolar rupture. This includes the effector/translocators IpaB and IpaC. More recently, this has been challenged and an implication of the host cell in the process of vacuolar rupture has been put forward. This includes the bacterial subversion of host trafficking regulators, such as the Rab GTPase Rab11. The involvement of the host in determining bacterial vacuolar integrity has also been found for other bacterial pathogens, particularly for Salmonella. Here, we will discuss our current view of host factor and pathogen effector implications during Shigella vacuolar rupture and the steps leading to it.

  3. Cytosolic access of intracellular bacterial pathogens: the Shigella paradigm

    Directory of Open Access Journals (Sweden)

    Nora eMellouk

    2016-04-01

    Full Text Available Shigella is a Gram-negative bacterial pathogen, which causes bacillary dysentery in humans. A crucial step of Shigella infection is its invasion of epithelial cells. Using a type III secretion system, Shigella injects several bacterial effectors ultimately leading to bacterial internalization within a vacuole. Then, Shigella escapes rapidly from the vacuole, it replicates within the cytosol and spreads from cell-to-cell. The molecular mechanism of vacuolar rupture used by Shigella has been studied in some detail during the recent years and new paradigms are emerging about the underlying molecular events. For decades, bacterial effector proteins were portrayed as main actors inducing vacuolar rupture. This includes the effector/translocators IpaB and IpaC. More recently, this has been challenged and an implication of the host cell in the process of vacuolar rupture has been put forward. This includes the bacterial subversion of host trafficking regulators, such as the Rab GTPase Rab11. The involvement of the host in determining bacterial vacuolar integrity has also been found for other bacterial pathogens, particularly for Salmonella. Here, we will discuss our current view of host factor and pathogen effector implications during Shigella vacuolar rupture and the steps leading to it.

  4. Rheumatoid arthritis and bacterial infections

    Directory of Open Access Journals (Sweden)

    N L Prokopjeva

    2008-01-01

    Full Text Available To study features of bacterial infections course in pts with rheumatoid arthritis (RA and changes of laboratory measures after focus of infection sanation. Material and methods. 46 pts with definite rheumatoid arthritis were examined at the time of comorbid infection (Cl detection and after infection focus sanation. Bacteriological test with evaluation of flora sensitivity to antibiotics by disco-diffusion method was performed at baseline and after the course of antibacterial therapy to assess its efficacy. Hemogram, serum fibrinogen, rheumatoid factor, circulating immune complexes (CIC, C-reactive protein levels were assessed. Serum interleukin (IL 1(3, IL6 and neopterin concentrations were examined by immune-enzyme assay in a part of pts. Typical clinical features of Cl were present in only 28 (60,9% pts. 13 (28,3% pts had fever, 12 (26,0% — leukocytosis, 15 (32,6% — changes of leucocyte populations. Some laboratory measures (thrombocytes, fibrinogen, CIC, neopterin levels significantly decreased (p<0,05 after infection focus sanation without correction of disease modifying therapy. Cl quite often develop as asymptomatic processes most often in pts with high activity and can induce disturbances promoting appearance of endothelial dysfunction, atherothrombosis and reduction of life duration. So timely detection and proper sanation of infection focuses should be performed in pts with RA

  5. Conserved water molecules in bacterial serine hydroxymethyltransferases.

    Science.gov (United States)

    Milano, Teresa; Di Salvo, Martino Luigi; Angelaccio, Sebastiana; Pascarella, Stefano

    2015-10-01

    Water molecules occurring in the interior of protein structures often are endowed with key structural and functional roles. We report the results of a systematic analysis of conserved water molecules in bacterial serine hydroxymethyltransferases (SHMTs). SHMTs are an important group of pyridoxal-5'-phosphate-dependent enzymes that catalyze the reversible conversion of l-serine and tetrahydropteroylglutamate to glycine and 5,10-methylenetetrahydropteroylglutamate. The approach utilized in this study relies on two programs, ProACT2 and WatCH. The first software is able to categorize water molecules in a protein crystallographic structure as buried, positioned in clefts or at the surface. The other program finds, in a set of superposed homologous proteins, water molecules that occur approximately in equivalent position in each of the considered structures. These groups of molecules are referred to as 'clusters' and represent structurally conserved water molecules. Several conserved clusters of buried or cleft water molecules were found in the set of 11 bacterial SHMTs we took into account for this work. The majority of these clusters were not described previously. Possible structural and functional roles for the conserved water molecules are envisaged. This work provides a map of the conserved water molecules helpful for deciphering SHMT mechanism and for rational design of molecular engineering experiments. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  6. S1PR3 Signaling Drives Bacterial Killing and Is Required for Survival in Bacterial Sepsis.

    Science.gov (United States)

    Hou, JinChao; Chen, QiXing; Wu, XiaoLiang; Zhao, DongYan; Reuveni, Hadas; Licht, Tamar; Xu, MengLong; Hu, Hu; Hoeft, Andreas; Ben-Sasson, Shmuel A; Shu, Qiang; Fang, XiangMing

    2017-12-15

    Efficient elimination of pathogenic bacteria is a critical determinant in the outcome of sepsis. Sphingosine-1-phosphate receptor 3 (S1PR3) mediates multiple aspects of the inflammatory response during sepsis, but whether S1PR3 signaling is necessary for eliminating the invading pathogens remains unknown. To investigate the role of S1PR3 in antibacterial immunity during sepsis. Loss- and gain-of-function experiments were performed using cell and murine models. S1PR3 levels were determined in patients with sepsis and healthy volunteers. S1PR3 protein levels were up-regulated in macrophages upon bacterial stimulation. S1pr3 -/- mice showed increased mortality and increased bacterial burden in multiple models of sepsis. The transfer of wild-type bone marrow-derived macrophages rescued S1pr3 -/- mice from lethal sepsis. S1PR3-overexpressing macrophages further ameliorated the mortality rate of sepsis. Loss of S1PR3 led to markedly decreased bacterial killing in macrophages. Enhancing endogenous S1PR3 activity using a peptide agonist potentiated the macrophage bactericidal function and improved survival rates in multiple models of sepsis. Mechanically, the reactive oxygen species levels were decreased and phagosome maturation was delayed in S1pr3 -/- macrophages due to impaired recruitment of vacuolar protein-sorting 34 to the phagosomes. In addition, S1RP3 expression levels were elevated in monocytes from patients with sepsis. Higher levels of monocytic S1PR3 were associated with efficient intracellular bactericidal activity, better immune status, and preferable outcomes. S1PR3 signaling drives bacterial killing and is essential for survival in bacterial sepsis. Interventions targeting S1PR3 signaling could have translational implications for manipulating the innate immune response to combat pathogens.

  7. Enhancing Nanos expression via the bacterial TomO protein is a conserved strategy used by the symbiont Wolbachia to fuel germ stem cell maintenance in infected Drosophila females.

    Science.gov (United States)

    Ote, Manabu; Yamamoto, Daisuke

    2018-04-27

    The toxic manipulator of oogenesis (TomO) protein has been identified in the wMel strain of Wolbachia that symbioses with the vinegar fly Drosophila melanogaster, as a protein that affects host reproduction. TomO protects germ stem cells (GSCs) from degeneration, which otherwise occurs in ovaries of host females that are mutant for the gene Sex-lethal (Sxl). We isolated the TomO homologs from wPip, a Wolbachia strain from the mosquito Culex quinquefasciatus. One of the homologs, TomO w Pip 1, exerted the GSC rescue activity in fly Sxl mutants when lacking its hydrophobic stretches. The GSC-rescuing action of the TomO w Pip 1 variant was ascribable to its abilities to associate with Nanos (nos) mRNA and to enhance Nos protein expression. The analysis of structure-activity relationships with TomO homologs and TomO deletion variants revealed distinct modules in the protein that are each dedicated to different functions, i.e., subcellular localization, nos mRNA binding or Nos expression enhancement. We propose that modular reshuffling is the basis for structural and functional diversification of TomO protein members. © 2018 Wiley Periodicals, Inc.

  8. Evolvability of thermophilic proteins from archaea and bacteria.

    Science.gov (United States)

    Takano, Kazufumi; Aoi, Atsushi; Koga, Yuichi; Kanaya, Shigenori

    2013-07-16

    Proteins from thermophiles possess high thermostability. The stabilization mechanisms differ between archaeal and bacterial proteins, whereby archaeal proteins are mainly stabilized via hydrophobic interactions and bacterial proteins by ion pairs. High stability is an important factor in promoting protein evolution, but the precise means by which different stabilization mechanisms affect the evolution process remain unclear. In this study, we investigated a random mutational drift of esterases from thermophilic archaea and bacteria at high temperatures. Our results indicate that mutations in archaeal proteins lead to improved function with no loss of stability, while mutant bacterial proteins are largely destabilized with decreased activity at high temperatures. On the basis of these findings, we suggest that archaeal proteins possess higher "evolvability" than bacterial proteins under temperature selection and are additionally able to evolve into eukaryotic proteins.

  9. Evaluating bacterial gene-finding HMM structures as probabilistic logic programs

    DEFF Research Database (Denmark)

    Mørk, Søren; Holmes, Ian

    2012-01-01

    , a probabilistic dialect of Prolog. Results: We evaluate Hidden Markov Model structures for bacterial protein-coding gene potential, including a simple null model structure, three structures based on existing bacterial gene finders and two novel model structures. We test standard versions as well as ADPH length...

  10. Classification of EC 3.1.1.3 bacterial true lipases using phylogenetic ...

    African Journals Online (AJOL)

    To obtain an overview of this industrially and very important class of enzymes and their characteristics, we collected and classified bacterial lipases sequences available from protein databases. Here we proposed an updated and revised classification of family I bacterial "true" lipases based mainly on a comparison of their ...

  11. Capsule Shields the Function of Short Bacterial Adhesins

    OpenAIRE

    Schembri, Mark A.; Dalsgaard, Dorte; Klemm, Per

    2004-01-01

    Bacterial surface structures such as capsules and adhesins are generally regarded as important virulence factors. Here we demonstrate that capsules block the function of the self-recognizing protein antigen 43 through physical shielding. The phenomenon is not restricted to Escherichia coli but can occur in other gram-negative bacteria. Likewise, we show that other short adhesins exemplified by the AIDA-I protein are blocked by the presence of a capsule. The results support the notion that cap...

  12. Corruption of innate immunity by bacterial proteases.

    Science.gov (United States)

    Potempa, Jan; Pike, Robert N

    2009-01-01

    The innate immune system of the human body has developed numerous mechanisms to control endogenous and exogenous bacteria and thus prevent infections by these microorganisms. These mechanisms range from physical barriers such as the skin or mucosal epithelium to a sophisticated array of molecules and cells that function to suppress or prevent bacterial infection. Many bacteria express a variety of proteases, ranging from non-specific and powerful enzymes that degrade many proteins involved in innate immunity to proteases that are extremely precise and specific in their mode of action. Here we have assembled a comprehensive picture of how bacterial proteases affect the host's innate immune system to gain advantage and cause infection. This picture is far from being complete since the numbers of mechanisms utilized are as astonishing as they are diverse, ranging from degradation of molecules vital to innate immune mechanisms to subversion of the mechanisms to allow the bacterium to hide from the system or take advantage of it. It is vital that such mechanisms are elucidated to allow strategies to be developed to aid the innate immune system in controlling bacterial infections.

  13. Capsule shields the function of short bacterial adhesins

    DEFF Research Database (Denmark)

    Schembri, Mark; Dalsgaard, D.; Klemm, Per

    2004-01-01

    Bacterial surface structures such as capsules and adhesins are generally regarded as important virulence factors. Here we demonstrate that capsules block the function of the self-recognizing protein antigen 43 through physical shielding. The phenomenon is not restricted to Escherichia coli but can...

  14. Molecular analysis of cross-bacterial contamination detected in ...

    African Journals Online (AJOL)

    ... the isolate Delftia acidovorans BP(R2) and it is also coupled to protein with molecular weight 25-26 KDa. As well as, this bacterial contamination was the reason for the false positive results observed during the detection of HCV infections. Journal of Applied Sciences and Environmental Management Vol. 9(1) 2005: 5-10.

  15. Recombinant bacterial hemoglobin alters metabolism of Aspergillus niger

    DEFF Research Database (Denmark)

    Hofmann, Gerald; Diano, Audrey; Nielsen, Jens

    2009-01-01

    , the fungus will produce various by-products like organic acids and polyols. In order to circumvent this problem we here study the effects of the expression of a bacterial hemoglobin protein on the metabolism of A. niger. We integrated the vgb gene from Vitreoscilla sp. into the genome at the pyrA locus...

  16. Effects of bacterial inoculants and an enzyme on the fermentation ...

    African Journals Online (AJOL)

    ... the effects of bacterial inoculation and cellulase on the fermentation quality of ensiled whole-crop sweet sorghum (WCSS, Sorghum bicolor L. Moench). The WCSS (323 g dry matter (DM)/kg, 251 g water soluble carbohydrates (WSC)/kg DM, 43 g crude protein (CP)/kg DM and 439 g neutral detergent fibre (NDF)/kg DM) ...

  17. Modeling base excision repair in Escherichia coli bacterial cells

    International Nuclear Information System (INIS)

    Belov, O.V.

    2011-01-01

    A model describing the key processes in Escherichia coli bacterial cells during base excision repair is developed. The mechanism is modeled of damaged base elimination involving formamidopyrimidine DNA glycosylase (the Fpg protein), which possesses several types of activities. The modeling of the transitions between DNA states is based on a stochastic approach to the chemical reaction description

  18. A BACTERIAL CYTOKINE Mukamolova et al (1998) PNAS, 95, 8916 ...

    Indian Academy of Sciences (India)

    First page Back Continue Last page Overview Graphics. A BACTERIAL CYTOKINE Mukamolova et al (1998) PNAS, 95, 8916-8921. To grow or not to grow is a social decision. A resuscitation promoting factor (rpf) identified. The rpf protein shares similarity with M. tuberculosis and M. leprae ...

  19. Isolation of cell-free bacterial inclusion bodies.

    Science.gov (United States)

    Rodríguez-Carmona, Escarlata; Cano-Garrido, Olivia; Seras-Franzoso, Joaquin; Villaverde, Antonio; García-Fruitós, Elena

    2010-09-17

    Bacterial inclusion bodies are submicron protein clusters usually found in recombinant bacteria that have been traditionally considered as undesirable products from protein production processes. However, being fully biocompatible, they have been recently characterized as nanoparticulate inert materials useful as scaffolds for tissue engineering, with potentially wider applicability in biomedicine and material sciences. Current protocols for inclusion body isolation from Escherichia coli usually offer between 95 to 99% of protein recovery, what in practical terms, might imply extensive bacterial cell contamination, not compatible with the use of inclusion bodies in biological interfaces. Using an appropriate combination of chemical and mechanical cell disruption methods we have established a convenient procedure for the recovery of bacterial inclusion bodies with undetectable levels of viable cell contamination, below 10⁻¹ cfu/ml, keeping the particulate organization of these aggregates regarding size and protein folding features. The application of the developed protocol allows obtaining bacterial free inclusion bodies suitable for use in mammalian cell cultures and other biological interfaces.

  20. Reaction of Musa balbisiana to Banana bacterial wilt infection ...

    African Journals Online (AJOL)

    The expression of NPR1, a marker gene of the systemic acquired resistance plant defence system provides preliminary evidence that this may be the major form of resistance in Musa balbisiana to bacterial wilt infection. Keywords: NPR1, PR proteins, Uganda, Xanthomonas campestris. African Crop Science Journal, Vol.

  1. Exploring bacterial outer membrane barrier to combat bad bugs

    Directory of Open Access Journals (Sweden)

    Ghai I

    2017-08-01

    Full Text Available Ishan Ghai,1 Shashank Ghai2 1School of Engineering and Life Sciences, Jacobs University, Bremen, 2Leibniz University, Hannover, Germany Abstract: One of the main fundamental mechanisms of antibiotic resistance in Gram-negative bacteria comprises an effective change in the membrane permeability to antibiotics. The Gram-negative bacterial complex cell envelope comprises an outer membrane that delimits the periplasm from the exterior environment. The outer membrane contains numerous protein channels, termed as porins or nanopores, which are mainly involved in the influx of hydrophilic compounds, including antibiotics. Bacterial adaptation to reduce influx through these outer membrane proteins (Omps is one of the crucial mechanisms behind antibiotic resistance. Thus to interpret the molecular basis of the outer membrane permeability is the current challenge. This review attempts to develop a state of knowledge pertinent to Omps and their effective role in antibiotic influx. Further, it aims to study the bacterial response to antibiotic membrane permeability and hopefully provoke a discussion toward understanding and further exploration of prospects to improve our knowledge on physicochemical parameters that direct the translocation of antibiotics through the bacterial membrane protein channels. Keywords: antibiotics, Gram-negative bacteria, cell envelope, protein channels, nanopores, influx, antibiotic resistance

  2. Camel Mastitis, associated Bacterial Pathogens and its impact on ...

    African Journals Online (AJOL)

    The present study was conducted between September 2006 and April 2007 with the aim of assessing the occurrence of camel mastitis and bacterial causes associated with it and evaluating Fat and Protein content of camel milk in Gewane district, Afar Regional State, Northeastern Ethiopia. Lactating camels which are ...

  3. Bacterial polyhydroxyalkanoates: Still fabulous?

    Science.gov (United States)

    Możejko-Ciesielska, Justyna; Kiewisz, Robert

    2016-11-01

    Bacterial polyhydroxyalkanoates (PHA) are polyesters accumulated as carbon and energy storage materials under limited growth conditions in the presence of excess carbon sources. They have been developed as biomaterials with unique properties for the past many years being considered as a potential substitute for conventional non-degradable plastics. Due to the increasing concern towards global climate change, depleting petroleum resource and problems with an utilization of a growing number of synthetic plastics, PHAs have gained much more attention from industry and research. These environmentally friendly microbial polymers have great potential in biomedical, agricultural, and industrial applications. However, their production on a large scale is still limited. This paper describes the backgrounds of PHAs and discussed the current state of knowledge on the polyhydroxyalkanoates. Ability of bacteria to convert different carbon sources to PHAs, the opportunities and challenges of their introduction to global market as valuable renewable products have been also discussed. Copyright © 2016 Elsevier GmbH. All rights reserved.

  4. Energetics of bacterial adhesion

    International Nuclear Information System (INIS)

    Loosdrecht, M.C.M. van; Zehnder, A.J.B.

    1990-01-01

    For the description of bacterial adhesion phenomena two different physico-chemical approaches are available. The first one, based on a surface Gibbs energy balance, assumes intimate contact between the interacting surfaces. The second approach, based on colloid chemical theories (DLVO theory), allows for two types of adhesion: 1) secondary minimum adhesion, which is often weak and reversible, and 2) irreversible primary minimum adhesion. In the secondary minimum adhesion a thin water film remains present between the interacting surface. The merits of both approaches are discussed in this paper. In addition, the methods available to measure the physico-chemical surface characteristics of bacteria and the influence of adsorbing (in)organic compounds, extracellular polymers and cell surface appendages on adhesion are summarized. (author) 2 figs., 1 tab., 50 refs

  5. Biosensors of bacterial cells.

    Science.gov (United States)

    Burlage, Robert S; Tillmann, Joshua

    2017-07-01

    Biosensors are devices which utilize both an electrical component (transducer) and a biological component to study an environment. They are typically used to examine biological structures, organisms and processes. The field of biosensors has now become so large and varied that the technology can often seem impenetrable. Yet the principles which underlie the technology are uncomplicated, even if the details of the mechanisms are elusive. In this review we confine our analysis to relatively current advancements in biosensors for the detection of whole bacterial cells. This includes biosensors which rely on an added labeled component and biosensors which do not have a labeled component and instead detect the binding event or bound structure on the transducer. Methods to concentrate the bacteria prior to biosensor analysis are also described. The variety of biosensor types and their actual and potential uses are described. Copyright © 2016 Elsevier B.V. All rights reserved.

  6. Exploring bacterial lignin degradation.

    Science.gov (United States)

    Brown, Margaret E; Chang, Michelle C Y

    2014-04-01

    Plant biomass represents a renewable carbon feedstock that could potentially be used to replace a significant level of petroleum-derived chemicals. One major challenge in its utilization is that the majority of this carbon is trapped in the recalcitrant structural polymers of the plant cell wall. Deconstruction of lignin is a key step in the processing of biomass to useful monomers but remains challenging. Microbial systems can provide molecular information on lignin depolymerization as they have evolved to break lignin down using metalloenzyme-dependent radical pathways. Both fungi and bacteria have been observed to metabolize lignin; however, their differential reactivity with this substrate indicates that they may utilize different chemical strategies for its breakdown. This review will discuss recent advances in studying bacterial lignin degradation as an approach to exploring greater diversity in the environment. Copyright © 2013 Elsevier Ltd. All rights reserved.

  7. Studying bacterial multispecies biofilms

    DEFF Research Database (Denmark)

    Røder, Henriette Lyng; Sørensen, Søren Johannes; Burmølle, Mette

    2016-01-01

    The high prevalence and significance of multispecies biofilms have now been demonstrated in various bacterial habitats with medical, industrial, and ecological relevance. It is highly evident that several species of bacteria coexist and interact in biofilms, which highlights the need for evaluating...... the approaches used to study these complex communities. This review focuses on the establishment of multispecies biofilms in vitro, interspecies interactions in microhabitats, and how to select communities for evaluation. Studies have used different experimental approaches; here we evaluate the benefits...... and drawbacks of varying the degree of complexity. This review aims to facilitate multispecies biofilm research in order to expand the current limited knowledge on interspecies interactions. Recent technological advances have enabled total diversity analysis of highly complex and diverse microbial communities...

  8. Anaerobes in bacterial vaginosis

    Directory of Open Access Journals (Sweden)

    Aggarwal A

    2003-01-01

    Full Text Available Four hundred high vaginal swabs were taken from patients attending gynaecology and obstetrics department of Govt. medical college, Amritsar. The patients were divided into four groups i.e. women in pregnancy (Group I, in labour/post partum (Group II, with abnormal vaginal discharge or bacterial vaginosis (Group III and asymptomatic women as control (Group IV. Anaerobic culture of vaginal swabs revealed that out of 400 cases, 212(53% were culture positive. Maximum isolation of anaerobes was in group III (84% followed by group II (56%, group I (36% and control group (15%. Gram positive anaerobes (69.2% out numbered gram negatives (30.8%. Among various isolates Peptostreptococcus spp. and Bacteroides spp. were predominant.

  9. Assembly and mechanical properties of the cargo-free and cargo-loaded bacterial nanocompartment encapsulin

    NARCIS (Netherlands)

    Snijder, Joost; Kononova, Olga; Barbu, Ioana M; Uetrecht, Charlotte; Rurup, W Frederik; Burnley, Rebecca J; Koay, Melissa S T; Cornelissen, Jeroen J L M; Roos, Wouter H; Barsegov, Valeri; Wuite, Gijs J L; Heck, Albert J R

    2016-01-01

    Prokaryotes mostly lack membranous compartments that are typical of eukaryotic cells, but instead they have various protein-based organelles. These include bacterial microcompartments like the carboxysome and the virus-like nanocompartment encapsulin. Encapsulins have an adaptable mechanism for

  10. Bacterial meningitis in immunocompromised patients

    NARCIS (Netherlands)

    van Veen, K.E.B.

    2018-01-01

    Bacterial meningitis is an acute infection of the meninges, in The Netherlands most commonly caused by Streptococcus pneumoniae and Neisseria meningitides. Risk factors for acquiring bacterial meningitis include a decreased function of the immune system. The aim of this thesis was to study

  11. Bacteriële meningitis

    NARCIS (Netherlands)

    Brouwer, M. C.; van de Beek, D.

    2012-01-01

    Bacterial meningitis is a severe disease which affects 35.000 Europeans each year and has a mortality rate of about 20%. During the past 25 years the epidemiology of bacterial meningitis has changed significantly due to the implementation of vaccination against Haemophilus influenzae, Neisseria

  12. The antibacterial efficacy of an aceraceous plant [Shantung maple (Acer truncatum Bunge)] may be related to inhibition of bacterial beta-oxoacyl-acyl carrier protein reductase (FabG).

    Science.gov (United States)

    Zhang, Feng; Luo, Shi-Yun; Ye, Yan-Bin; Zhao, Wen-Hua; Sun, Xu-Guang; Wang, Zhi-Qun; Li, Ran; Sun, Ying-Hui; Tian, Wei-Xi; Zhang, Ying-Xia

    2008-10-01

    Polyphenols, including flavonoids, are the major components of the extracts from aceraceous plants. They possess remarkable antibacterial and antitumour activity. Our study focused on whether the inhibition of the bacterial type II fatty acid synthesis system is the mechanism for the antibacterial effect of the related plant polyphenols. Extracts obtained from the fallen leaves of the Shantung maple (Acer truncatum Bunge) using different solvents, and the related pure compound PGG (1,2,3,4,6-penta-O-galloyl-beta-D-glucose), potently inhibited the FabG (beta-oxoacyl-ACP reductase) steps in the fatty-acid-elongation cycle with the IC(50) values between 0.9 and 7.2 microg/ml. An ethyl acetate extract appeared to inhibit FabG reductase in a mixed manner with NADPH, as did PGG with NADPH, demonstrating that they interfered with the binding of the cofactor to the enzyme. Gram-positive and Gram-negative bacteria and some fungi were used to evaluate the antibacterial abilities of different extract samples. The experiments showed that a higher polyphenol content of the extracts led to a more potent inhibitory capacity against FabG, thus enhancing the antibacterial efficacy.

  13. Biochemical composition of the marine conditioning film: Implications for bacterial adhesion

    Digital Repository Service at National Institute of Oceanography (India)

    Jain, A.; Bhosle, N

    1980; Fletcher and Marshall 1982; Abbott et al. 1983; Frolund et al. 1996; Azeredo and Oliveira 2000; Gubner and Beech 2000). Carbohydrates account for *10 to 50% of dissolved organic carbon in marine waters (Pakulski and Benner 1994; Amon and Benner... in bacterial adhesion to surfaces. Conversely, bacterial adhesion to a protein-rich conditioning film was inhibited (Fletcher 1980; Fletcher and Marshall 1982; Abbott et al. 1983; Husmark and Ronner 1993). It appears that the proteins of the conditioning film...

  14. Studying biomolecule localization by engineering bacterial cell wall curvature.

    Directory of Open Access Journals (Sweden)

    Lars D Renner

    Full Text Available In this article we describe two techniques for exploring the relationship between bacterial cell shape and the intracellular organization of proteins. First, we created microchannels in a layer of agarose to reshape live bacterial cells and predictably control their mean cell wall curvature, and quantified the influence of curvature on the localization and distribution of proteins in vivo. Second, we used agarose microchambers to reshape bacteria whose cell wall had been chemically and enzymatically removed. By combining microstructures with different geometries and fluorescence microscopy, we determined the relationship between bacterial shape and the localization for two different membrane-associated proteins: i the cell-shape related protein MreB of Escherichia coli, which is positioned along the long axis of the rod-shaped cell; and ii the negative curvature-sensing cell division protein DivIVA of Bacillus subtilis, which is positioned primarily at cell division sites. Our studies of intracellular organization in live cells of E. coli and B. subtilis demonstrate that MreB is largely excluded from areas of high negative curvature, whereas DivIVA localizes preferentially to regions of high negative curvature. These studies highlight a unique approach for studying the relationship between cell shape and intracellular organization in intact, live bacteria.

  15. Unraveling plant responses to bacterial pathogens through proteomics

    KAUST Repository

    Zimaro, Tamara

    2011-11-03

    Plant pathogenic bacteria cause diseases in important crops and seriously and negatively impact agricultural production. Therefore, an understanding of the mechanisms by which plants resist bacterial infection at the stage of the basal immune response or mount a successful specific R-dependent defense response is crucial since a better understanding of the biochemical and cellular mechanisms underlying these interactions will enable molecular and transgenic approaches to crops with increased biotic resistance. In recent years, proteomics has been used to gain in-depth understanding of many aspects of the host defense against pathogens and has allowed monitoring differences in abundance of proteins as well as posttranscriptional and posttranslational processes, protein activation/inactivation, and turnover. Proteomics also offers a window to study protein trafficking and routes of communication between organelles. Here, we summarize and discuss current progress in proteomics of the basal and specific host defense responses elicited by bacterial pathogens. Copyright 2011 Tamara Zimaro et al.

  16. Unraveling plant responses to bacterial pathogens through proteomics

    KAUST Repository

    Zimaro, Tamara; Gottig, Natalia; Garavaglia, Betiana S.; Gehring, Christoph A; Ottado, Jorgelina

    2011-01-01

    Plant pathogenic bacteria cause diseases in important crops and seriously and negatively impact agricultural production. Therefore, an understanding of the mechanisms by which plants resist bacterial infection at the stage of the basal immune response or mount a successful specific R-dependent defense response is crucial since a better understanding of the biochemical and cellular mechanisms underlying these interactions will enable molecular and transgenic approaches to crops with increased biotic resistance. In recent years, proteomics has been used to gain in-depth understanding of many aspects of the host defense against pathogens and has allowed monitoring differences in abundance of proteins as well as posttranscriptional and posttranslational processes, protein activation/inactivation, and turnover. Proteomics also offers a window to study protein trafficking and routes of communication between organelles. Here, we summarize and discuss current progress in proteomics of the basal and specific host defense responses elicited by bacterial pathogens. Copyright 2011 Tamara Zimaro et al.

  17. Electromagnetism of Bacterial Growth

    Science.gov (United States)

    Ainiwaer, Ailiyasi

    2011-10-01

    There has been increasing concern from the public about personal health due to the significant rise in the daily use of electrical devices such as cell phones, radios, computers, GPS, video games and television. All of these devices create electromagnetic (EM) fields, which are simply magnetic and electric fields surrounding the appliances that simultaneously affect the human bio-system. Although these can affect the human system, obstacles can easily shield or weaken the electrical fields; however, magnetic fields cannot be weakened and can pass through walls, human bodies and most other objects. The present study was conducted to examine the possible effects of bacteria when exposed to magnetic fields. The results indicate that a strong causal relationship is not clear, since different magnetic fields affect the bacteria differently, with some causing an increase in bacterial cells, and others causing a decrease in the same cells. This phenomenon has yet to be explained, but the current study attempts to offer a mathematical explanation for this occurrence. The researchers added cultures to the magnetic fields to examine any effects to ion transportation. Researchers discovered ions such as potassium and sodium are affected by the magnetic field. A formula is presented in the analysis section to explain this effect.

  18. Evolution of Bacterial Suicide

    Science.gov (United States)

    Tchernookov, Martin; Nemenman, Ilya

    2013-03-01

    While active, controlled cellular suicide (autolysis) in bacteria is commonly observed, it has been hard to argue that autolysis can be beneficial to an individual who commits it. We propose a theoretical model that predicts that bacterial autolysis is evolutionarily advantageous to an individualand would fixate in physically structured environments for stationary phase colonies. We perform spatially resolved agent-based simulations of the model, which predict that lower mixing in the environment results in fixation of a higher autolysis rate from a single mutated cell, regardless of the colony's genetic diversity. We argue that quorum sensing will fixate as well, even if initially rare, if it is coupled to controlling the autolysis rate. The model does not predict a strong additional competitive advantage for cells where autolysis is controlled by quorum sensing systems that distinguish self from nonself. These predictions are broadly supported by recent experimental results in B. subtilisand S. pneumoniae. Research partially supported by the James S McDonnell Foundation grant No. 220020321 and by HFSP grant No. RGY0084/2011.

  19. Exploring bacterial outer membrane barrier to combat bad bugs.

    Science.gov (United States)

    Ghai, Ishan; Ghai, Shashank

    2017-01-01

    One of the main fundamental mechanisms of antibiotic resistance in Gram-negative bacteria comprises an effective change in the membrane permeability to antibiotics. The Gram-negative bacterial complex cell envelope comprises an outer membrane that delimits the periplasm from the exterior environment. The outer membrane contains numerous protein channels, termed as porins or nanopores, which are mainly involved in the influx of hydrophilic compounds, including antibiotics. Bacterial adaptation to reduce influx through these outer membrane proteins (Omps) is one of the crucial mechanisms behind antibiotic resistance. Thus to interpret the molecular basis of the outer membrane permeability is the current challenge. This review attempts to develop a state of knowledge pertinent to Omps and their effective role in antibiotic influx. Further, it aims to study the bacterial response to antibiotic membrane permeability and hopefully provoke a discussion toward understanding and further exploration of prospects to improve our knowledge on physicochemical parameters that direct the translocation of antibiotics through the bacterial membrane protein channels.

  20. Towards revealing the structure of bacterial inclusion bodies.

    Science.gov (United States)

    Wang, Lei

    2009-01-01

    Protein aggregation is a widely observed phenomenon in human diseases, biopharmaceutical production, and biological research. Protein aggregates are generally classified as highly ordered, such as amyloid fibrils, or amorphous, such as bacterial inclusion bodies. Amyloid fibrils are elongated filaments with diameters of 6-12 nm, they are comprised of residue-specific cross-beta structure, and display characteristic properties, such as binding with amyloid-specific dyes. Amyloid fibrils are associated with dozens of human pathological conditions, including Alzheimer disease and prion diseases. Distinguished from amyloid fibrils, bacterial inclusion bodies display apparent amorphous morphology. Inclusion bodies are formed during high-level recombinant protein production, and formation of inclusion bodies is a major concern in biotechnology. Despite of the distinctive morphological difference, bacterial inclusion bodies have been found to have some amyloid-like properties, suggesting that they might contain structures similar to amyloid-like fibrils. Recent structural data further support this hypothesis, and this review summarizes the latest progress towards revealing the structural details of bacterial inclusion bodies.

  1. Bacterial growth on macrophyte leachate and fate of bacterial production

    International Nuclear Information System (INIS)

    Findlay, S.; Carlough, L.; Crocker, M.T.; Gill, H.K.; Meyer, J.L.; Smith, P.J.

    1986-01-01

    The role bacteria play in transferring organic carbon to other trophic levels in aquatic ecosystems depends on the efficiency with which they convert dissolved organic [ 14 C]-labelled carbon into bacterial biomass and on the ability of consumers to graze bacteria. The authors have measured the conversion efficiency for bacteria growing on macrophyte-derived dissolved organic carbon and estimated the amount of bacterial production removed by grazing. Bacteria converted this DOC into new tissue with an efficiency of 53%, substantially higher than the apparent conversion efficiency of macrophyte-derived particulate organic carbon or other types of DOC. Two estimates of grazing indicate that the decline in bacterial numbers after the bloom was probably due to grazing by flagellates. These results show the significance of the bacterial link between DOC and other trophic levels

  2. Molecular detection of human bacterial pathogens

    National Research Council Canada - National Science Library

    Liu, Dongyou

    2011-01-01

    .... Molecular Detection of Human Bacterial Pathogens addresses this issue, with international scientists in respective bacterial pathogen research and diagnosis providing expert summaries on current...

  3. Incidence and Predictors of Bacterial infection in Febrile Children with Sickle Cell Disease.

    Science.gov (United States)

    Morrissey, Benita J; Bycroft, Thomas P; Almossawi, Ofran; Wilkey, Olufunke B; Daniels, Justin G

    2015-01-01

    Children with sickle cell disease are at increased risk of developing bacteremia and other serious bacterial infections. Fever is a common symptom in sickle cell disease and can also occur with sickle cell crises and viral infections. We aimed to evaluate the incidence and predictors of bacteremia and bacterial infection in children with sickle cell disease presenting with fever to a district hospital and sickle cell center in London. A retrospective analysis was performed on all attendances of children (aged under 16 years) with sickle cell disease presenting with a fever of 38.5 °C or higher over a 1-year period. Confirmed bacterial infection was defined as bacteremia, bacterial meningitis, urinary tract infection (UTI), pneumonia, osteomyelitis or other bacterial infection with positive identification of organism. Children were defined as having a suspected bacterial infection if a bacterial infection was suspected clinically, but no organism was identified. Over a 1-year period there were 88 episodes analyzed in 59 children. Bacteremia occurred in 3.4% of episodes and confirmed bacterial infection in 7.0%. Suspected bacterial infection occurred in 33.0%. One death occurred from Salmonella typhirium septicemia. C-reactive protein (CRP) level and white blood cell (WBC) count were both significantly associated with bacterial infection (p = 0.004 and 0.02, respectively.) In conclusion, bacterial infections continue to be a significant problem in children with sickle cell disease. C-reactive protein was significantly associated with bacterial infections, and could be included in clinical risk criteria for febrile children with sickle cell disease.

  4. Synthesis and Characterization of Protein-Conjugated Silver Nanoparticles/Silver Salt Loaded Poly (3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) Film for Prevention of Bacterial Infections and Potential Use in Bone Tissue Engineering Applications

    Science.gov (United States)

    Bakare, Rotimi Ayotunde

    Failure of orthopedic implants due to bacterial infection has been a major concern in bone tissue engineering. To this end, we have formulated a potential orthopedic implant made of naturally occurring biodegradable polymer, i.e. poly (3-hydroxylbutyrate-co-3-hydroxylvalerate) (PHBV), modified with BSA conjugated silver nanoparticles and or silver chloride. Upon release of Ag NPs and or Ag+ in the implant region, can promote aseptic environment by inhibition of bacteria growth and also support/maintain bone cell adhesion, growth, and proliferation. For formulating nanoparticles loaded PHBV scaffold, we exploit specific interaction between bovine serum albumin (BSA) of BSA capped silver nanoparticles and collagen of collagen immobilized PHBV scaffold. Therefore, the first part of this study dealt with synthesis and characterization of collagen immobilized PHBV film for loading of BSA stabilized silver (Ag/BSA) nanoparticles. Two different approaches were used to immobilize collagen on macroporous PHBV film. First approach uses thermal radical copolymerization with 2-hydroxyethylmethacrylate (HEMA), while the second approach uses aminolysis to functionalize macroporous PHBV film. Using collagen crosslinker, type I collagen was covalently grafted to formulate collagen immobilized PHEMA-g-PHBV and collagen immobilized NH2-PHBV films, respectively. Spectroscopic (FTIR, XPS), physical (SEM), and thermal (TGA) techniques were used to characterize the functionalized PHBV films. The Ag/BSA nanoparticles were then loaded on collagen immobilized PHBV films and untreated PHBV films. The concentration of nanoparticles loaded on PHBV film was determined by atomic absorption spectrometry and fluorescence spectroscopy. The amount of nanoparticles loaded on collagen immobilized PHBV film was found to be significantly greater than that on untreated PHBV film. The amount of Ag/BSA nanoparticles loaded on collagen immobilized PHBV film was found to depend on the concentration of Ag

  5. Plant pathology: monitoring a pathogen-targeted host protein.

    Science.gov (United States)

    Ellis, Jeff; Dodds, Peter

    2003-05-13

    A plant protein RIN4 is targeted and modified by bacterial pathogens as part of the disease process. At least two host resistance proteins monitor this pathogen interference and trigger the plant's defence responses.

  6. Comparison of different protein sources in enriched grain mixture for ...

    African Journals Online (AJOL)

    -inname voorsien. ..... Animal nutrition. Longman, New York. MERCER, lR., ALLEN, S.A. & MILLER, EL, 1980. Rumen bacterial protein synthesis and the proportion of dietary protein escaping degradation in the rumen of sheep. Br. J. NUlr.

  7. Neurological sequelae of bacterial meningitis

    NARCIS (Netherlands)

    Lucas, Marjolein J.; Brouwer, Matthijs C.; van de Beek, Diederik

    2016-01-01

    We reported on occurrence and impact of neurological sequelae after bacterial meningitis. We reviewed occurrence of neurological sequelae in children and adults after pneumococcal and meningococcal meningitis. Most frequently reported sequelae are focal neurological deficits, hearing loss, cognitive

  8. Bacterial tracheitis in Down's syndrome.

    OpenAIRE

    Cant, A J; Gibson, P J; West, R J

    1987-01-01

    Four children with Down's syndrome and bacterial tracheitis are described. In three the infection was due to Haemophilus influenza. In patients with Down's syndrome presenting with stridor tracheitis should be considered and appropriate treatment started.

  9. Bacterial Communities: Interactions to Scale

    Directory of Open Access Journals (Sweden)

    Reed M. Stubbendieck

    2016-08-01

    Full Text Available In the environment, bacteria live in complex multispecies communities. These communities span in scale from small, multicellular aggregates to billions or trillions of cells within the gastrointestinal tract of animals. The dynamics of bacterial communities are determined by pairwise interactions that occur between different species in the community. Though interactions occur between a few cells at a time, the outcomes of these interchanges have ramifications that ripple through many orders of magnitude, and ultimately affect the macroscopic world including the health of host organisms. In this review we cover how bacterial competition influences the structures of bacterial communities. We also emphasize methods and insights garnered from culture-dependent pairwise interaction studies, metagenomic analyses, and modeling experiments. Finally, we argue that the integration of multiple approaches will be instrumental to future understanding of the underlying dynamics of bacterial communities.

  10. Bacterial flora of sturgeon fingerling

    International Nuclear Information System (INIS)

    Arani, A.S.; Mosahab, R.

    2008-01-01

    The study on microbial populations is a suitable tool to understand and apply control methods to improve the sanitary level of production in fish breeding and rearing centers, ensure health of sturgeon fingerlings at the time of their release into the rivers and also in the conversation and restoration of these valuable stocks in the Caspian Sea, Iran. A laboratory research based on Austin methods (Austin, B., Austin, D.A. 1993) was conducted for bacterial study on 3 sturgeon species naming A. persicus, A. stellatus and A. nudiventris during different growth stages. Bacterial flora of Acinetobacter, Moraxella, Aeromonas, Vibrio, Edwardsiella, Staphylococcus, Proteus, Yersinia, Pseudomonas and Plesiomonas were determined. The factors which may induce changes in bacterial populations during different stages of fife are the followings: quality of water in rearing ponds, different conditions for growth stages, suitable time for colonization of bacterial flora in rearing pond, water temperature increase in fingerlings size and feeding condition. (author)

  11. Effects of the Bacterial Extract OM-85 on Phagocyte Functions and the Stress Response

    Science.gov (United States)

    Baladi, S.; Kantengwa, S.; Donati, Y. R. A.; Polla, B. S.

    1994-01-01

    The effects of the bacterial extract OM-85 on the respiratory burst, intracellular calcium and the stress response have been investigated in human peripheral blood monocytes from normal donors. Activation of the respiratory burst during bacterial phagocytosis has been previously associated with heat shock/stress proteins synthesis. Whereas OM-85 stimulated superoxide production and increased Ca2+ mobilization, it fared to induce synthesis of classical HSPs. The lack of stress protein induction was observed even in the presence of iron which potentiates both oxidative injury and stress protein induction during bacterial phagocytosis. However OM-85 induced a 75–78 kDa protein, which is likely to be a glucose regulated protein (GRP78), and enhanced intracellular expression of interleukin-lβ precursor. PMID:18472933

  12. Bacterial translocation: impact of probiotics

    OpenAIRE

    Jeppsson, Bengt; Mangell, Peter; Adawi, Diya; Molin, Göran

    2004-01-01

    There is a considerable amount of data in humans showing that patients who cannot take in nutrients enterally have more organ failure in the intensive care unit, a less favourable prognosis, and a higher frequency of septicaemia, in particular involving bacterial species from the intestinal tract. However, there is little evidence that this is connected with translocation of bacterial species in humans. Animal data more uniformly imply the existence of such a connection. The main focus of thi...

  13. Bacterial cellulose/boehmite composites

    International Nuclear Information System (INIS)

    Salvi, Denise T.B. de; Barud, Hernane S.; Messaddeq, Younes; Ribeiro, Sidney J.L.; Caiut, Jose Mauricio A.

    2011-01-01

    Composites based on bacterial cellulose membranes and boehmite were obtained. SEM results indicate that the bacterial cellulose (BC) membranes are totally covered by boehmite and obtained XRD patterns suggest structural changes due to this boehmite addition. Thermal stability is accessed through TG curves and is dependent on boehmite content. Transparency is high comparing to pure BC as can be seen through UV-vis absorption spectroscopy. (author)

  14. Arsenic uptake in bacterial calcite

    Science.gov (United States)

    Catelani, Tiziano; Perito, Brunella; Bellucci, Francesco; Lee, Sang Soo; Fenter, Paul; Newville, Matthew; Rimondi, Valentina; Pratesi, Giovanni; Costagliola, Pilario

    2018-02-01

    Bio-mediated processes for arsenic (As) uptake in calcite were investigated by means of X-ray Diffraction (XRD) and X-ray Absorption Spectroscopy (XAS) coupled with X-ray Fluorescence measurements. The environmental bacterial strain Bacillus licheniformis BD5, sampled at the Bullicame Hot Springs (Viterbo, Central Italy), was used to synthesize calcite from As-enriched growth media. Both liquid and solid cultures were applied to simulate planktonic and biofilm community environments, respectively. Bacterial calcite samples cultured in liquid media had an As enrichment factor (Kd) 50 times higher than that from solid media. The XRD analysis revealed an elongation of the crystal lattice along the c axis (by 0.03 Å) for biogenic calcite, which likely resulted from the substitution of larger arsenate for carbonate in the crystal. The XAS data also showed a clear difference in the oxidation state of sorbed As between bacterial and abiotic calcite. Abiotic chemical processes yielded predominantly As(V) uptake whereas bacterial precipitation processes led to the uptake of both As(III) and As(V). The presence of As(III) in bacterial calcite is proposed to result from subsequent reduction of arsenate to arsenite by bacterial activities. To the best of our knowledge, this is the first experimental observation of the incorporation of As(III) in the calcite crystal lattice, revealing a critical role of biochemical processes for the As cycling in nature.

  15. Arsenic uptake in bacterial calcite

    Energy Technology Data Exchange (ETDEWEB)

    Catelani, Tiziano; Perito, Brunella; Bellucci, Francesco; Lee, Sang Soo; Fenter, Paul; Newville, Matthew G.; Rimondi, Valentina; Pratesi, Giovanni; Costagliola, Pilario

    2018-02-01

    Bio-mediated processes for arsenic (As) uptake in calcite were investigated by means of X-ray Diffraction (XRD) and Xray Absorption Spectroscopy (XAS) coupled with X-ray Fluorescence measurements. The environmental bacterial strain Bacillus licheniformis BD5, sampled at the Bullicame Hot Springs (Viterbo, Central Italy), was used to synthesize calcite from As-enriched growth media. Both liquid and solid cultures were applied to simulate planktonic and biofilm community environments, respectively. Bacterial calcite samples cultured in liquid media had an As enrichment factor (Kd) 50 times higher than that from solid media. The XRD analysis revealed an elongation of the crystal lattice along the c axis (by 0.03Å) for biogenic calcite, which likely resulted from the substitution of larger arsenate for carbonate in the crystal. The XAS data also showed a clear difference in the oxidation state of sorbed As between bacterial and abiotic calcite. Abiotic chemical processes yielded predominantly As(V) uptake whereas bacterial precipitation processes led to the uptake of both As(III) and As(V). The presence of As(III) in bacterial calcite is proposed to result from subsequent reduction of arsenate to arsenite by bacterial activities. To the best of our knowledge, this is the first experimental observation of the incorporation of As(III) in the calcite crystal lattice, revealing a critical role of biochemical processes for the As cycling in nature.

  16. Magnetotactic Bacterial Cages as Safe and Smart Gene Delivery Vehicles

    KAUST Repository

    Alsaiari, Shahad K.

    2016-07-27

    In spite of the huge advances in the area of synthetic carriers, their efficiency still poorly compares to natural vectors. Herein, we report the use of unmodified magnetotactic bacteria as a guidable delivery vehicle for DNA functionalized gold nanoparticles (AuNPs). High cargo loading is established under anaerobic conditions (bacteria is alive) through endocytosis where AuNPs are employed as transmembrane proteins mimics (facilitate endocytosis) as well as imaging agents to verify and quantify loading and release. The naturally bio-mineralized magnetosomes, within the bacteria, induce heat generation inside bacteria through magnetic hyperthermia. Most importantly after exposing the system to air (bacteria is dead) the cell wall stays intact providing an efficient bacterial vessel. Upon incubation with THP-1 cells, the magnetotactic bacterial cages (MBCs) adhere to the cell wall and are directly engulfed through the phagocytic activity of these cells. Applying magnetic hyperthermia leads to the dissociation of the bacterial microcarrier and eventual release of cargo.

  17. Magnetotactic Bacterial Cages as Safe and Smart Gene Delivery Vehicles

    KAUST Repository

    Alsaiari, Shahad K.; Ezzedine, Alaa H.; Abdallah, Abdallah; Sougrat, Rachid; Khashab, Niveen M.

    2016-01-01

    In spite of the huge advances in the area of synthetic carriers, their efficiency still poorly compares to natural vectors. Herein, we report the use of unmodified magnetotactic bacteria as a guidable delivery vehicle for DNA functionalized gold nanoparticles (AuNPs). High cargo loading is established under anaerobic conditions (bacteria is alive) through endocytosis where AuNPs are employed as transmembrane proteins mimics (facilitate endocytosis) as well as imaging agents to verify and quantify loading and release. The naturally bio-mineralized magnetosomes, within the bacteria, induce heat generation inside bacteria through magnetic hyperthermia. Most importantly after exposing the system to air (bacteria is dead) the cell wall stays intact providing an efficient bacterial vessel. Upon incubation with THP-1 cells, the magnetotactic bacterial cages (MBCs) adhere to the cell wall and are directly engulfed through the phagocytic activity of these cells. Applying magnetic hyperthermia leads to the dissociation of the bacterial microcarrier and eventual release of cargo.

  18. Growth mechanics of bacterial cell wall and morphology of bacteria

    Science.gov (United States)

    Jiang, Hongyuan; Sun, Sean

    2010-03-01

    The peptidoglycan cell wall of bacteria is responsible for maintaining the cell shape and integrity. During the bacterial life cycle, the growth of the cell wall is affected by mechanical stress and osmotic pressure internal to the cell. We develop a theory to describe cell shape changes under the influence of mechanical forces. We find that the theory predicts a steady state size and shape for bacterial cells ranging from cocci to spirillum. Moreover, the theory suggest a mechanism by which bacterial cytoskeletal proteins such as MreB and crescentin can maintain the shape of the cell. The theory can also explain the several recent experiments on growing bacteria in micro-environments.

  19. Energetics of bacterial photosynthesis.

    Science.gov (United States)

    Lebard, David N; Matyushov, Dmitry V

    2009-09-10

    We report the results of extensive numerical simulations and theoretical calculations of electronic transitions in the reaction center of Rhodobacter sphaeroides photosynthetic bacterium. The energetics and kinetics of five electronic transitions related to the kinetic scheme of primary charge separation have been analyzed and compared to experimental observations. Nonergodic formulation of the reaction kinetics is required for the calculation of the rates due to a severe breakdown of the system ergodicity on the time scale of primary charge separation, with the consequent inapplicability of the standard canonical prescription to calculate the activation barrier. Common to all reactions studied is a significant excess of the charge-transfer reorganization energy from the width of the energy gap fluctuations over that from the Stokes shift of the transition. This property of the hydrated proteins, breaking the linear response of the thermal bath, allows the reaction center to significantly reduce the reaction free energy of near-activationless electron hops and thus raise the overall energetic efficiency of the biological charge-transfer chain. The increase of the rate of primary charge separation with cooling is explained in terms of the temperature variation of induction solvation, which dominates the average donor-acceptor energy gap for all electronic transitions in the reaction center. It is also suggested that the experimentally observed break in the Arrhenius slope of the primary recombination rate, occurring near the temperature of the dynamical transition in proteins, can be traced back to a significant drop of the solvent reorganization energy close to that temperature.

  20. Isolation, characterization, and aggregation of a structured bacterial matrix precursor.

    Science.gov (United States)

    Chai, Liraz; Romero, Diego; Kayatekin, Can; Akabayov, Barak; Vlamakis, Hera; Losick, Richard; Kolter, Roberto

    2013-06-14

    Biofilms are surface-associated groups of microbial cells that are embedded in an extracellular matrix (ECM). The ECM is a network of biopolymers, mainly polysaccharides, proteins, and nucleic acids. ECM proteins serve a variety of structural roles and often form amyloid-like fibers. Despite the extensive study of the formation of amyloid fibers from their constituent subunits in humans, much less is known about the assembly of bacterial functional amyloid-like precursors into fibers. Using dynamic light scattering, atomic force microscopy, circular dichroism, and infrared spectroscopy, we show that our unique purification method of a Bacillus subtilis major matrix protein component results in stable oligomers that retain their native α-helical structure. The stability of these oligomers enabled us to control the external conditions that triggered their aggregation. In particular, we show that stretched fibers are formed on a hydrophobic surface, whereas plaque-like aggregates are formed in solution under acidic pH conditions. TasA is also shown to change conformation upon aggregation and gain some β-sheet structure. Our studies of the aggregation of a bacterial matrix protein from its subunits shed new light on assembly processes of the ECM within bacterial biofilms.

  1. Bacterial Prostatitis: Bacterial Virulence, Clinical Outcomes, and New Directions.

    Science.gov (United States)

    Krieger, John N; Thumbikat, Praveen

    2016-02-01

    Four prostatitis syndromes are recognized clinically: acute bacterial prostatitis, chronic bacterial prostatitis, chronic prostatitis/chronic pelvic pain syndrome, and asymptomatic prostatitis. Because Escherichia coli represents the most common cause of bacterial prostatitis, we investigated the importance of bacterial virulence factors and antimicrobial resistance in E. coli strains causing prostatitis and the potential association of these characteristics with clinical outcomes. A structured literature review revealed that we have limited understanding of the virulence-associated characteristics of E. coli causing acute prostatitis. Therefore, we completed a comprehensive microbiological and molecular investigation of a unique strain collection isolated from healthy young men. We also considered new data from an animal model system suggesting certain E. coli might prove important in the etiology of chronic prostatitis/chronic pelvic pain syndrome. Our human data suggest that E. coli needs multiple pathogenicity-associated traits to overcome anatomic and immune responses in healthy young men without urological risk factors. The phylogenetic background and accumulation of an exceptional repertoire of extraintestinal pathogenic virulence-associated genes indicate that these E. coli strains belong to a highly virulent subset of uropathogenic variants. In contrast, antibiotic resistance confers little added advantage to E. coli strains in these healthy outpatients. Our animal model data also suggest that certain pathogenic E. coli may be important in the etiology of chronic prostatitis/chronic pelvic pain syndrome through mechanisms that are dependent on the host genetic background and the virulence of the bacterial strain.

  2. Bacterial dynamics in a microphytobenthic biofilm: A tidal mesocosm approach

    Science.gov (United States)

    Agogué, Hélène; Mallet, Clarisse; Orvain, Francis; De Crignis, Margot; Mornet, Françoise; Dupuy, Christine

    2014-09-01

    In intertidal mudflats, during low tide exposure, microphytobenthos (MPB) migrate vertically through the surface sediment and form, with the heterotrophic bacteria, a transient biofilm. Inside this biofilm, multiple interactions exist between MPB and bacteria. These micro-organisms secrete a wide range of extracellular polymeric substances (EPS), which are major components of the biofilm matrix. In this study, we used a tidal mesocosm experiment in order to decipher the interactions of the MPB-EPS-bacteria complex within the biofilm. We tried to determine if the EPS could control bacterial activities and/or production and/or richness according to the age of the biofilm and to the immersion/emersion period. The dynamics of biomasses of MPB and prokaryotes, the bacterial production, the hydrolysis of predominating organic constituents in the dissolved organic carbon (DOC) pool (i.e., carbohydrates and polypeptides), and the bacterial structure were studied in relation to the different EPS fractions (carbohydrates and proteins: colloidal and bound) dynamics during 8 days. Our experiment had emphasized the influence of the environmental conditions (light, immersion/emersion) on the interactions within the biofilm and also on the effects on biofilm aging. Bacterial production was always inhibited by the bound EPS-carbohydrate, especially during low tide. Our results suggest that the concentration and composition of EPS had a major role in the bacterial/MPB interactions: these interactions can be either positive or negative in order to regulate the productive phases of MPB and bacteria.

  3. Composing a Tumor Specific Bacterial Promoter.

    Directory of Open Access Journals (Sweden)

    Igor V Deyneko

    Full Text Available Systemically applied Salmonella enterica spp. have been shown to invade and colonize neoplastic tissues where it retards the growth of many tumors. This offers the possibility to use the bacteria as a vehicle for the tumor specific delivery of therapeutic molecules. Specificity of such delivery is solely depending on promoter sequences that control the production of a target molecule. We have established the functional structure of bacterial promoters that are transcriptionally active exclusively in tumor tissues after systemic application. We observed that the specific transcriptional activation is accomplished by a combination of a weak basal promoter and a strong FNR binding site. This represents a minimal set of control elements required for such activation. In natural promoters, additional DNA remodeling elements are found that alter the level of transcription quantitatively. Inefficiency of the basal promoter ensures the absence of transcription outside tumors. As a proof of concept, we compiled an artificial promoter sequence from individual motifs representing FNR and basal promoter and showed specific activation in a tumor microenvironment. Our results open possibilities for the generation of promoters with an adjusted level of expression of target proteins in particular for applications in bacterial tumor therapy.

  4. Bacterial Enzymes and Antibiotic Resistance- Oral Presentation

    Energy Technology Data Exchange (ETDEWEB)

    Maltz, Lauren [SLAC National Accelerator Lab., Menlo Park, CA (United States)

    2015-08-25

    By using protein crystallography and X-ray diffraction, structures of bacterial enzymes were solved to gain a better understanding of how enzymatic modification acts as an antibacterial resistance mechanism. Aminoglycoside phosphotransferases (APHs) are one of three aminoglycoside modifying enzymes that confer resistance to the aminoglycoside antibiotics via enzymatic modification, rendering many drugs obsolete. Specifically, the APH(2”) family vary in their substrate specificities and also in their preference for the phosphate donor (ADP versus GDP). By solving the structures of members of the APH(2”) family of enzymes, we can see how domain movements are important to their substrate specificity. Our structure of the ternary complex of APH(2”)-IIIa with GDP and kanamycin, when compared to the known structures of APH(2”)-IVa, reveals that there are real physical differences between these two enzymes, a structural finding that explains why the two enzymes differ in their preferences for certain aminoglycosides. Another important group of bacterial resistance enzymes are the Class D β-lactamases. Oxacillinase carbapenemases (OXAs) are part of this enzyme class and have begun to confer resistance to ‘last resort’ drugs, most notably carbapenems. Our structure of OXA-143 shows that the conformational flexibility of a conserved hydrophobic residue in the active site (Val130) serves to control the entry of a transient water molecule responsible for a key step in the enzyme’s mechanism. Our results provide insight into the structural mechanisms of these two different enzymes.

  5. The neglected intrinsic resistome of bacterial pathogens.

    Directory of Open Access Journals (Sweden)

    Alicia Fajardo

    Full Text Available Bacteria with intrinsic resistance to antibiotics are a worrisome health problem. It is widely believed that intrinsic antibiotic resistance of bacterial pathogens is mainly the consequence of cellular impermeability and activity of efflux pumps. However, the analysis of transposon-tagged Pseudomonas aeruginosa mutants presented in this article shows that this phenotype emerges from the action of numerous proteins from all functional categories. Mutations in some genes make P. aeruginosa more susceptible to antibiotics and thereby represent new targets. Mutations in other genes make P. aeruginosa more resistant and therefore define novel mechanisms for mutation-driven acquisition of antibiotic resistance, opening a new research field based in the prediction of resistance before it emerges in clinical environments. Antibiotics are not just weapons against bacterial competitors, but also natural signalling molecules. Our results demonstrate that antibiotic resistance genes are not merely protective shields and offer a more comprehensive view of the role of antibiotic resistance genes in the clinic and in nature.

  6. Use of the [14C]Leucine Incorporation Technique To Measure Bacterial Production in River Sediments and the Epiphyton

    OpenAIRE

    Fischer, Helmut; Pusch, Martin

    1999-01-01

    Bacterial production is a key parameter for the understanding of carbon cycling in aquatic ecosystems, yet it remains difficult to measure in many aquatic habitats. We therefore tested the applicability of the [14C]leucine incorporation technique for the measurement of bulk bacterial production in various habitats of a lowland river ecosystem. To evaluate the method, we determined (i) extraction efficiencies of bacterial protein from the sediments, (ii) substrate saturation of leucine in sedi...

  7. Abdominal radiation causes bacterial translocation

    International Nuclear Information System (INIS)

    Guzman-Stein, G.; Bonsack, M.; Liberty, J.; Delaney, J.P.

    1989-01-01

    The purpose of this study was to determine if a single dose of radiation to the rat abdomen leads to bacterial translocation into the mesenteric lymph nodes (MLN). A second issue addressed was whether translocation correlates with anatomic damage to the mucosa. The radiated group (1100 cGy) which received anesthesia also was compared with a control group and a third group which received anesthesia alone but no abdominal radiation. Abdominal radiation lead to 100% positive cultures of MLN between 12 hr and 4 days postradiation. Bacterial translocation was almost nonexistent in the control and anesthesia group. Signs of inflammation and ulceration of the intestinal mucosa were not seen until Day 3 postradiation. Mucosal damage was maximal by Day 4. Bacterial translocation onto the MLN after a single dose of abdominal radiation was not apparently dependent on anatomical, histologic damage of the mucosa

  8. Bacterial Degradation of Aromatic Compounds

    Directory of Open Access Journals (Sweden)

    Qing X. Li

    2009-01-01

    Full Text Available Aromatic compounds are among the most prevalent and persistent pollutants in the environment. Petroleum-contaminated soil and sediment commonly contain a mixture of polycyclic aromatic hydrocarbons (PAHs and heterocyclic aromatics. Aromatics derived from industrial activities often have functional groups such as alkyls, halogens and nitro groups. Biodegradation is a major mechanism of removal of organic pollutants from a contaminated site. This review focuses on bacterial degradation pathways of selected aromatic compounds. Catabolic pathways of naphthalene, fluorene, phenanthrene, fluoranthene, pyrene, and benzo[a]pyrene are described in detail. Bacterial catabolism of the heterocycles dibenzofuran, carbazole, dibenzothiophene, and dibenzodioxin is discussed. Bacterial catabolism of alkylated PAHs is summarized, followed by a brief discussion of proteomics and metabolomics as powerful tools for elucidation of biodegradation mechanisms.

  9. Bacterial computing with engineered populations.

    Science.gov (United States)

    Amos, Martyn; Axmann, Ilka Maria; Blüthgen, Nils; de la Cruz, Fernando; Jaramillo, Alfonso; Rodriguez-Paton, Alfonso; Simmel, Friedrich

    2015-07-28

    We describe strategies for the construction of bacterial computing platforms by describing a number of results from the recently completed bacterial computing with engineered populations project. In general, the implementation of such systems requires a framework containing various components such as intracellular circuits, single cell input/output and cell-cell interfacing, as well as extensive analysis. In this overview paper, we describe our approach to each of these, and suggest possible areas for future research. © 2015 The Author(s) Published by the Royal Society. All rights reserved.

  10. Transport of Magnesium by a Bacterial Nramp-Related Gene

    Science.gov (United States)

    Rodionov, Dmitry A.; Freedman, Benjamin G.; Senger, Ryan S.; Winkler, Wade C.

    2014-01-01

    Magnesium is an essential divalent metal that serves many cellular functions. While most divalent cations are maintained at relatively low intracellular concentrations, magnesium is maintained at a higher level (∼0.5–2.0 mM). Three families of transport proteins were previously identified for magnesium import: CorA, MgtE, and MgtA/MgtB P-type ATPases. In the current study, we find that expression of a bacterial protein unrelated to these transporters can fully restore growth to a bacterial mutant that lacks known magnesium transporters, suggesting it is a new importer for magnesium. We demonstrate that this transport activity is likely to be specific rather than resulting from substrate promiscuity because the proteins are incapable of manganese import. This magnesium transport protein is distantly related to the Nramp family of proteins, which have been shown to transport divalent cations but have never been shown to recognize magnesium. We also find gene expression of the new magnesium transporter to be controlled by a magnesium-sensing riboswitch. Importantly, we find additional examples of riboswitch-regulated homologues, suggesting that they are a frequent occurrence in bacteria. Therefore, our aggregate data discover a new and perhaps broadly important path for magnesium import and highlight how identification of riboswitch RNAs can help shed light on new, and sometimes unexpected, functions of their downstream genes. PMID:24968120

  11. Bacterial translocation in an experimental intestinal obstruction model: C-reactive protein reliability? Translocação bacteriana no modelo experimental de obstrução intestinal: A proteína C-reativa é confiável?

    Directory of Open Access Journals (Sweden)

    Saleh Ibrahim El-Awady

    2009-04-01

    Full Text Available BACKGROUND: Bacterial translocation occurs in preseptic conditions such as intestinal obstruction through unclear mechanism. The C-reactive protein is an acute phase reactant and a marker of ischemia. METHODS: 45 albino male rats were divided into 3 groups each 15 rats. GI control, GII simple intestinal-obstruction and GIII strangulated obstruction. Outcome measures were: (1 Bacteriologic count and typing for intestinal contents, intestinal wall, liver, mesenteric lymph nodes and blood (cardiac and portal (2 Histopathologic: mucosal injury score, inflammatory cell infiltrate in the wall, MLN, liver, (3 Biochemical: serum CRP, IL-10, mucosal stress pattern (glutathione peroxidase-malonyldialdhyde tissue levels. RESULTS: (1 Intestinal obstruction associates with BT precursors (Bact-overgrowth, mucosal-acidosis, immuno-incomptence, (2 Bacterial translocation (frequency and density was found higher in strangulated I.O, that was mainly enteric (aerobic and anaerobic and mostly E.coli, (3 The pathogen commonality supports the gut origin hypothesis but the systemic inflammatory response goes with the cytokine generating one. (4 The CRP median values for GI, II, III were 0.5, 6.9, 8.5 mg/L, for BT +ve 8 mg/L and 0.75 mg/L for BT -ve rats. CONCLUSION: Bacterial translocation occurs bi-directional (systemic-portal in intestinal obstruction and the resultant inflammatory response pathogenesis is mostly 3 hit model. The CRP is a non selective marker of suspected I.O cases. However, it is a reliable marker of BT, BT density and vascular compromise during I.O.OBJETIVO: Translocação bacteriana ocorre em condições pré-sépticas como na obstrução intestinal por mecanismo não esclarecido. A proteína C-reativa é um marcador de ischemia em fase aguda. A proposição é investigar os possíveis efeitos da obstrução intestinal no equilíbrio ecológico microbiano. MÉTODOS: 45 ratos machos albinos foram distribuídos em três grupos de 15 ratos. GI

  12. Community-acquired bacterial meningitis

    NARCIS (Netherlands)

    van de Beek, Diederik; Brouwer, Matthijs; Hasbun, Rodrigo; Koedel, Uwe; Whitney, Cynthia G.; Wijdicks, Eelco

    2016-01-01

    Meningitis is an inflammation of the meninges and subarachnoid space that can also involve the brain cortex and parenchyma. It can be acquired spontaneously in the community - community-acquired bacterial meningitis - or in the hospital as a complication of invasive procedures or head trauma

  13. Food irradiation and bacterial toxins

    International Nuclear Information System (INIS)

    Tranter, H.S.; Modi, N.K.; Hambleton, P.; Melling, J.; Rose, S.; Stringer, M.F.

    1987-01-01

    The authors' findings indicate that irradiation confers no advantage over heat processing in respect of bacterial toxins (clostridium botulinum, neurotoxin A and staphylococcal enterotoxin A). It follows that irradiation at doses less than the ACINF recommended upper limit of 10 kGy could not be used to improve the ambient temperature shelf life on non-acid foods. (author)

  14. How carotenoids protect bacterial photosynthesis.

    OpenAIRE

    Cogdell, R J; Howard, T D; Bittl, R; Schlodder, E; Geisenheimer, I; Lubitz, W

    2000-01-01

    The essential function of carotenoids in photosynthesis is to act as photoprotective agents, preventing chlorophylls and bacteriochlorophylls from sensitizing harmful photodestructive reactions in the presence of oxygen. Based upon recent structural studies on reaction centres and antenna complexes from purple photosynthetic bacteria, the detailed organization of the carotenoids is described. Then with specific reference to bacterial antenna complexes the details of the photoprotective role, ...

  15. Biotechnological applications of bacterial cellulases

    Czech Academy of Sciences Publication Activity Database

    Menéndez, E.; García-Fraile, Paula; Rivas, R.

    2015-01-01

    Roč. 2, č. 3 (2015), s. 163-182 ISSN 2306-5354 R&D Projects: GA MŠk(CZ) EE2.3.30.0003 Institutional support: RVO:61388971 Keywords : Biotechnological applications * Bacterial cellulases * Cellulose degradation Subject RIV: EE - Microbiology, Virology

  16. bacterial flora and antibiotic sensitivity

    African Journals Online (AJOL)

    Purulent pelvic collections are common pathologies observed in contemporary gynaecological practice. They may originate from chronic pelvic inflammatory disease, from abortions or following normal deliveries. This study was designed to compare the bacterial flora in purulent pelvic collections obtained from HIV infected ...

  17. Genotoxicological Evaluation of NUTRALYS Pea Protein Isolate

    OpenAIRE

    Aouatif, Chentouf; Looten, Ph.; Parvathi, M. V. S.; Raja Ganesh, S.; Paranthaman, V.

    2013-01-01

    NUTRALYS Pea Protein Isolate, a protein supplement, is a high-quality source of protein which is primarily emulsifying functional protein. We evaluated the genotoxic potential of NUTRALYS isolated from dry yellow pea, using three established genotoxicity tests (AMES test in vitro chromosomal aberration test, and in vivo micronucleus test) employing OECD guidelines under GLP conditions. In the bacterial reverse mutation test, NUTRALYS did not show positive responses in strains detecting point ...

  18. Bacterial contribution to iodine volatilization in the environment

    Energy Technology Data Exchange (ETDEWEB)

    Amachi, S; Kasahara, M; Fujii, T [Chiba Univ., Dept. of Bioresources Chemistry, Matsudo, Chiba (Japan); Muramatsu, Y [National Inst. of Radiological Sciences, Chiba (Japan)

    2003-09-01

    The roles of microorganisms in iodine volatilization from the environment were studied. More than 100 bacterial strains were isolated from various environments such as soils, seawater and marine sediments, and were examined their capacities for volatilizing iodine. Approximately 40% of these bacteria showed significant capacities for volatilizing iodine. Gas chromatographic determinations revealed that the chemical species of gaseous iodine is methyl iodide (CH{sub 3}I). Phylogenetic analysis based on 16S ribosomal DNA showed that these 'iodine-volatilizing bacteria' are widely distributed through the bacterial domain. The iodide-methylating reaction was mediated by an enzyme protein with S-adenosyl-L-methionine (SAM) as the methyl donor. We then estimated bacterial contribution to iodine volatilization from soils. Iodine in soils was volatilized mainly as CH{sub 3}I. CH{sub 3}I emission was enhanced in the presence of glucose or yeast extract, but was inhibited by autoclaving of soils. Little CH{sub 3}I was produced under anaerobic conditions. Furthermore, the addition of streptomycin and tetracycline, antibiotics which inhibit bacterial growth, strongly inhibited CH{sub 3}I emission, while a fungal inhibitor cycloheximide caused little effect. These results suggest that iodine in soils is volatilized as CH{sub 3}I mainly by the action of aerobic soil bacteria. Similar experiment was carried out by using sea water samples. The emission of iodine from sea waters occurred biologically, and bacterial (and also other microbial) contribution was confirmed. Our results suggest that iodine is methylated and volatilized into the atmosphere as a result of bacterial activities. Since bacteria are so abundant and widespread in the environments, they may significantly contribute to global iodine volatilization. This indicates that if {sup 129}I would be released from nuclear facilities, weapons testing or ground storage of nuclear wastes, the pathway of volatilization by

  19. The actin homologue MreB organizes the bacterial cell membrane

    OpenAIRE

    Strahl, Henrik; Bürmann, Frank; Hamoen, Leendert W.

    2014-01-01

    The eukaryotic cortical actin cytoskeleton creates specific lipid domains, including lipid rafts, which determine the distribution of many membrane proteins. Here we show that the bacterial actin homologue MreB displays a comparable activity. MreB forms membrane-associated filaments that coordinate bacterial cell wall synthesis. We noticed that the MreB cytoskeleton influences fluorescent staining of the cytoplasmic membrane. Detailed analyses combining an array of mutants, using specific lip...

  20. RNA search engines empower the bacterial intranet.

    Science.gov (United States)

    Dendooven, Tom; Luisi, Ben F

    2017-08-15

    RNA acts not only as an information bearer in the biogenesis of proteins from genes, but also as a regulator that participates in the control of gene expression. In bacteria, small RNA molecules (sRNAs) play controlling roles in numerous processes and help to orchestrate complex regulatory networks. Such processes include cell growth and development, response to stress and metabolic change, transcription termination, cell-to-cell communication, and the launching of programmes for host invasion. All these processes require recognition of target messenger RNAs by the sRNAs. This review summarizes recent results that have provided insights into how bacterial sRNAs are recruited into effector ribonucleoprotein complexes that can seek out and act upon target transcripts. The results hint at how sRNAs and their protein partners act as pattern-matching search engines that efficaciously regulate gene expression, by performing with specificity and speed while avoiding off-target effects. The requirements for efficient searches of RNA patterns appear to be common to all domains of life. © 2017 The Author(s).

  1. Prostatitis-bacterial - self-care

    Science.gov (United States)

    ... this page: //medlineplus.gov/ency/patientinstructions/000395.htm Prostatitis - bacterial - self-care To use the sharing features ... enable JavaScript. You have been diagnosed with bacterial prostatitis . This is an infection of the prostate gland. ...

  2. Adjunctive Corticosteroids in Adults with Bacterial Meningitis

    NARCIS (Netherlands)

    van de Beek, Diederik; de Gans, Jan

    2005-01-01

    Bacterial meningitis is a complex disorder in which neurologic injury is caused, in part, by the causative organism and, in part, by the host's own inflammatory response. In studies of experimental bacterial meningitis, adjuvant treatment with corticosteroids, specifically dexamethasone, has

  3. Antimicrobial susceptibility in community-acquired bacterial ...

    African Journals Online (AJOL)

    Objectives: To determine the antimicrobial susceptibility patterns of Streptococcus pneumoniae and Haemophilus influenzae, two bacterial pathogens commonly associated with communityacquired pneumonia. Design: Cross-sectional study. Setting: Bacterial isolates were obtained from adults suspected to have ...

  4. Endocarditis in adults with bacterial meningitis

    NARCIS (Netherlands)

    Lucas, Marjolein J.; Brouwer, Matthijs C.; van der Ende, Arie; van de Beek, Diederik

    2013-01-01

    Endocarditis may precede or complicate bacterial meningitis, but the incidence and impact of endocarditis in bacterial meningitis are unknown. We assessed the incidence and clinical characteristics of patients with meningitis and endocarditis from a nationwide cohort study of adults with

  5. Membrane Protein Production in Lactococcus lactis for Functional Studies.

    Science.gov (United States)

    Seigneurin-Berny, Daphne; King, Martin S; Sautron, Emiline; Moyet, Lucas; Catty, Patrice; André, François; Rolland, Norbert; Kunji, Edmund R S; Frelet-Barrand, Annie

    2016-01-01

    Due to their unique properties, expression and study of membrane proteins in heterologous systems remains difficult. Among the bacterial systems available, the Gram-positive lactic bacterium, Lactococcus lactis, traditionally used in food fermentations, is nowadays widely used for large-scale production and functional characterization of bacterial and eukaryotic membrane proteins. The aim of this chapter is to describe the different possibilities for the functional characterization of peripheral or intrinsic membrane proteins expressed in Lactococcus lactis.

  6. Bacterial Urease and its Role in Long-Lasting Human Diseases

    Science.gov (United States)

    Konieczna, Iwona; Żarnowiec, Paulina; Kwinkowski, Marek; Kolesińska, Beata; Frączyk, Justyna; Kamiński, Zbigniew; Kaca, Wiesław

    2012-01-01

    Urease is a virulence factor found in various pathogenic bacteria. It is essential in colonization of a host organism and in maintenance of bacterial cells in tissues. Due to its enzymatic activity, urease has a toxic effect on human cells. The presence of ureolytic activity is an important marker of a number of bacterial infections. Urease is also an immunogenic protein and is recognized by antibodies present in human sera. The presence of such antibodies is connected with progress of several long-lasting diseases, like rheumatoid arthritis, atherosclerosis or urinary tract infections. In bacterial ureases, motives with a sequence and/or structure similar to human proteins may occur. This phenomenon, known as molecular mimicry, leads to the appearance of autoantibodies, which take part in host molecules destruction. Detection of antibodies-binding motives (epitopes) in bacterial proteins is a complex process. However, organic chemistry tools, such as synthetic peptide libraries, are helpful in both, epitope mapping as well as in serologic investigations. In this review, we present a synthetic report on a molecular organization of bacterial ureases - genetic as well as structural. We characterize methods used in detecting urease and ureolytic activity, including techniques applied in disease diagnostic processes and in chemical synthesis of urease epitopes. The review also provides a summary of knowledge about a toxic effect of bacterial ureases on human body and about occurrence of anti-urease antibodies in long-lasting diseases. PMID:23305365

  7. Bacterial cells with improved tolerance to polyamines

    DEFF Research Database (Denmark)

    2017-01-01

    Provided are bacterial cells genetically modified to improve their tolerance to certain commodity chemicals, such as polyamines, and methods of preparing and using such bacterial cells for production of polyamines and other compounds.......Provided are bacterial cells genetically modified to improve their tolerance to certain commodity chemicals, such as polyamines, and methods of preparing and using such bacterial cells for production of polyamines and other compounds....

  8. Bacterial cells with improved tolerance to polyols

    DEFF Research Database (Denmark)

    2017-01-01

    The present invention relates to bacterial cells genetically modified to improve their tolerance to certain commodity chemicals, such as diols and other polyols, and to methods of preparing and using such bacterial cells for production of polyols and other compounds.......The present invention relates to bacterial cells genetically modified to improve their tolerance to certain commodity chemicals, such as diols and other polyols, and to methods of preparing and using such bacterial cells for production of polyols and other compounds....

  9. [Validation of a clinical prediction rule to distinguish bacterial from aseptic meningitis].

    Science.gov (United States)

    Agüero, Gonzalo; Davenport, María C; Del Valle, María de la P; Gallegos, Paulina; Kannemann, Ana L; Bokser, Vivian; Ferrero, Fernando

    2010-02-01

    Despite most meningitis are not bacterial, antibiotics are usually administered on admission because bacterial meningitis is difficult to be rule-out. Distinguishing bacterial from aseptic meningitis on admission could avoid inappropriate antibiotic use and hospitalization. We aimed to validate a clinical prediction rule to distinguish bacterial from aseptic meningitis in children, on arriving to the emergency room. This prospective study included patients aged or = 1000 cells/mm(3), CSF protein > or = 80 mg/dl, peripheral blood absolute neutrophil count > or = 10.000/mm(3), seizure = 1 point each. Sensitivity (S), specificity (E), positive and negative predictive values (PPV and NPV), positive and negative likelihood ratios (PLR and NLR) of the BMS to predict bacterial meningitis were calculated. Seventy patients with meningitis were included (14 bacterial meningitis). When BMS was calculated, 25 patients showed a BMS= 0 points, 11 BMS= 1 point, and 34 BMS > or = 2 points. A BMS = 0 showed S: 100%, E: 44%, VPP: 31%, VPN: 100%, RVP: 1,81 RVN: 0. A BMS > or = 2 predicted bacterial meningitis with S: 100%, E: 64%, VPP: 41%, VPN: 100%, PLR: 2.8, NLR:0. Using BMS was simple, and allowed identifying children with very low risk of bacterial meningitis. It could be a useful tool to assist clinical decision making.

  10. The Bacterial Sequential Markov Coalescent.

    Science.gov (United States)

    De Maio, Nicola; Wilson, Daniel J

    2017-05-01

    Bacteria can exchange and acquire new genetic material from other organisms directly and via the environment. This process, known as bacterial recombination, has a strong impact on the evolution of bacteria, for example, leading to the spread of antibiotic resistance across clades and species, and to the avoidance of clonal interference. Recombination hinders phylogenetic and transmission inference because it creates patterns of substitutions (homoplasies) inconsistent with the hypothesis of a single evolutionary tree. Bacterial recombination is typically modeled as statistically akin to gene conversion in eukaryotes, i.e. , using the coalescent with gene conversion (CGC). However, this model can be very computationally demanding as it needs to account for the correlations of evolutionary histories of even distant loci. So, with the increasing popularity of whole genome sequencing, the need has emerged for a faster approach to model and simulate bacterial genome evolution. We present a new model that approximates the coalescent with gene conversion: the bacterial sequential Markov coalescent (BSMC). Our approach is based on a similar idea to the sequential Markov coalescent (SMC)-an approximation of the coalescent with crossover recombination. However, bacterial recombination poses hurdles to a sequential Markov approximation, as it leads to strong correlations and linkage disequilibrium across very distant sites in the genome. Our BSMC overcomes these difficulties, and shows a considerable reduction in computational demand compared to the exact CGC, and very similar patterns in simulated data. We implemented our BSMC model within new simulation software FastSimBac. In addition to the decreased computational demand compared to previous bacterial genome evolution simulators, FastSimBac provides more general options for evolutionary scenarios, allowing population structure with migration, speciation, population size changes, and recombination hotspots. FastSimBac is

  11. Bacterial reproductive pathogens of cats and dogs.

    Science.gov (United States)

    Graham, Elizabeth M; Taylor, David J

    2012-05-01

    With the notable exception of Brucella canis, exogenous bacterial pathogens are uncommon causes of reproductive disease in cats and dogs. Most bacterial reproductive infections are endogenous, and predisposing factors for infection are important. This article reviews the etiology, pathogenesis, clinical presentation, diagnosis, treatment, and public health significance of bacterial reproductive pathogens in cats and dogs.

  12. Focal Targeting of the Bacterial Envelope by Antimicrobial Peptides

    Directory of Open Access Journals (Sweden)

    Rafi eRashid

    2016-06-01

    Full Text Available Antimicrobial peptides (AMPs are utilized by both eukaryotic and prokaryotic organisms. AMPs such as the human beta defensins, human neutrophil peptides, human cathelicidin, and many bacterial bacteriocins are cationic and capable of binding to anionic regions of the bacterial surface. Cationic AMPs (CAMPs target anionic lipids (e.g. phosphatidylglycerol (PG and cardiolipins (CL in the cell membrane and anionic components (e.g. lipopolysaccharide (LPS and lipoteichoic acid (LTA of the cell envelope. Bacteria have evolved mechanisms to modify these same targets in order to resist CAMP killing, e.g. lysinylation of PG to yield cationic lysyl-PG and alanylation of LTA. Since CAMPs offer a promising therapeutic alternative to conventional antibiotics, which are becoming less effective due to rapidly emerging antibiotic resistance, there is a strong need to improve our understanding about the AMP mechanism of action. Recent literature suggests that AMPs often interact with the bacterial cell envelope at discrete foci. Here we review recent AMP literature, with an emphasis on focal interactions with bacteria, including (1 CAMP disruption mechanisms, (2 delocalization of membrane proteins and lipids by CAMPs, and (3 CAMP sensing systems and resistance mechanisms. We conclude with new approaches for studying the bacterial membrane, e.g., lipidomics, high resolution imaging and non-detergent-based membrane domain extraction.

  13. D-amino acids inhibit initial bacterial adhesion: thermodynamic evidence.

    Science.gov (United States)

    Xing, Su-Fang; Sun, Xue-Fei; Taylor, Alicia A; Walker, Sharon L; Wang, Yi-Fu; Wang, Shu-Guang

    2015-04-01

    Bacterial biofilms are structured communities of cells enclosed in a self-produced hydrated polymeric matrix that can adhere to inert or living surfaces. D-Amino acids were previously identified as self-produced compounds that mediate biofilm disassembly by causing the release of the protein component of the polymeric matrix. However, whether exogenous D-amino acids could inhibit initial bacterial adhesion is still unknown. Here, the effect of the exogenous amino acid D-tyrosine on initial bacterial adhesion was determined by combined use of chemical analysis, force spectroscopic measurement, and theoretical predictions. The surface thermodynamic theory demonstrated that the total interaction energy increased with more D-tyrosine, and the contribution of Lewis acid-base interactions relative to the change in the total interaction energy was much greater than the overall nonspecific interactions. Finally, atomic force microscopy analysis implied that the hydrogen bond numbers and adhesion forces decreased with the increase in D-tyrosine concentrations. D-Tyrosine contributed to the repulsive nature of the cell and ultimately led to the inhibition of bacterial adhesion. This study provides a new way to regulate biofilm formation by manipulating the contents of D-amino acids in natural or engineered systems. © 2014 Wiley Periodicals, Inc.

  14. Identification of Dominant Immunogenic Bacteria and Bacterial Proteins in Periodontitis

    DEFF Research Database (Denmark)

    Agerbæk, Mette Rylev; Haubek, Dorte; Birkelund, Svend

    Marginal periodontitis is considered an infectious disease that triggers host inflammatory responses resulting in destruction of the periodontium. A complex biofilm of bacteria is associated with periodontitis. Some species have been identified as putative pathogens such as Porphyromonas gingivalis...

  15. The Bacterial Actin MamK

    Science.gov (United States)

    Ozyamak, Ertan; Kollman, Justin; Agard, David A.; Komeili, Arash

    2013-01-01

    It is now recognized that actin-like proteins are widespread in bacteria and, in contrast to eukaryotic actins, are highly diverse in sequence and function. The bacterial actin, MamK, represents a clade, primarily found in magnetotactic bacteria, that is involved in the proper organization of subcellular organelles, termed magnetosomes. We have previously shown that MamK from Magnetospirillum magneticum AMB-1 (AMB-1) forms dynamic filaments in vivo. To gain further insights into the molecular mechanisms that underlie MamK dynamics and function, we have now studied the in vitro properties of MamK. We demonstrate that MamK is an ATPase that, in the presence of ATP, assembles rapidly into filaments that disassemble once ATP is depleted. The mutation of a conserved active site residue (E143A) abolishes ATPase activity of MamK but not its ability to form filaments. Filament disassembly depends on both ATPase activity and potassium levels, the latter of which results in the organization of MamK filaments into bundles. These data are consistent with observations indicating that accessory factors are required to promote filament disassembly and for spatial organization of filaments in vivo. We also used cryo-electron microscopy to obtain a high resolution structure of MamK filaments. MamK adopts a two-stranded helical filament architecture, but unlike eukaryotic actin and other actin-like filaments, subunits in MamK strands are unstaggered giving rise to a unique filament architecture. Beyond extending our knowledge of the properties and function of MamK in magnetotactic bacteria, this study emphasizes the functional and structural diversity of bacterial actins in general. PMID:23204522

  16. The actin homologue MreB organizes the bacterial cell membrane.

    Science.gov (United States)

    Strahl, Henrik; Bürmann, Frank; Hamoen, Leendert W

    2014-03-07

    The eukaryotic cortical actin cytoskeleton creates specific lipid domains, including lipid rafts, which determine the distribution of many membrane proteins. Here we show that the bacterial actin homologue MreB displays a comparable activity. MreB forms membrane-associated filaments that coordinate bacterial cell wall synthesis. We noticed that the MreB cytoskeleton influences fluorescent staining of the cytoplasmic membrane. Detailed analyses combining an array of mutants, using specific lipid staining techniques and spectroscopic methods, revealed that MreB filaments create specific membrane regions with increased fluidity (RIFs). Interference with these fluid lipid domains (RIFs) perturbs overall lipid homeostasis and affects membrane protein localization. The influence of MreB on membrane organization and fluidity may explain why the active movement of MreB stimulates membrane protein diffusion. These novel MreB activities add additional complexity to bacterial cell membrane organization and have implications for many membrane-associated processes.

  17. Extensive Identification of Bacterial Riboflavin Transporters and Their Distribution across Bacterial Species

    Science.gov (United States)

    Merino, Enrique; Bonomi, Hernán Ruy; Goldbaum, Fernando Alberto; García-Angulo, Víctor Antonio

    2015-01-01

    Riboflavin, the precursor for the cofactors flavin mononucleotide (FMN) and flavin adenine dinucleotide, is an essential metabolite in all organisms. While the functions for de novo riboflavin biosynthesis and riboflavin import may coexist in bacteria, the extent of this co-occurrence is undetermined. The RibM, RibN, RfuABCD and the energy-coupling factor-RibU bacterial riboflavin transporters have been experimentally characterized. In addition, ImpX, RfnT and RibXY are proposed as riboflavin transporters based on positional clustering with riboflavin biosynthetic pathway (RBP) genes or conservation of the FMN riboswitch regulatory element. Here, we searched for the FMN riboswitch in bacterial genomes to identify genes encoding riboflavin transporters and assessed their distribution among bacteria. Two new putative riboflavin transporters were identified: RibZ in Clostridium and RibV in Mesoplasma florum. Trans-complementation of an Escherichia coli riboflavin auxotroph strain confirmed the riboflavin transport activity of RibZ from Clostridium difficile, RibXY from Chloroflexus aurantiacus, ImpX from Fusobacterium nucleatum and RfnT from Ochrobactrum anthropi. The analysis of the genomic distribution of all known bacterial riboflavin transporters revealed that most occur in species possessing the RBP and that some bacteria may even encode functional riboflavin transporters from two different families. Our results indicate that some species possess ancestral riboflavin transporters, while others possess transporters that appear to have evolved recently. Moreover, our data suggest that unidentified riboflavin transporters also exist. The present study doubles the number of experimentally characterized riboflavin transporters and suggests a specific, non-accessory role for these proteins in riboflavin-prototrophic bacteria. PMID:25938806

  18. Bacterial cheating limits antibiotic resistance

    Science.gov (United States)

    Xiao Chao, Hui; Yurtsev, Eugene; Datta, Manoshi; Artemova, Tanya; Gore, Jeff

    2012-02-01

    The widespread use of antibiotics has led to the evolution of resistance in bacteria. Bacteria can gain resistance to the antibiotic ampicillin by acquiring a plasmid carrying the gene beta-lactamase, which inactivates the antibiotic. This inactivation may represent a cooperative behavior, as the entire bacterial population benefits from removing the antibiotic. The cooperative nature of this growth suggests that a cheater strain---which does not contribute to breaking down the antibiotic---may be able to take advantage of cells cooperatively inactivating the antibiotic. Here we find experimentally that a ``sensitive'' bacterial strain lacking the plasmid conferring resistance can invade a population of resistant bacteria, even in antibiotic concentrations that should kill the sensitive strain. We observe stable coexistence between the two strains and find that a simple model successfully explains the behavior as a function of antibiotic concentration and cell density. We anticipate that our results will provide insight into the evolutionary origin of phenotypic diversity and cooperative behaviors.

  19. Bacterial streamers in curved microchannels

    Science.gov (United States)

    Rusconi, Roberto; Lecuyer, Sigolene; Guglielmini, Laura; Stone, Howard

    2009-11-01

    Biofilms, generally identified as microbial communities embedded in a self-produced matrix of extracellular polymeric substances, are involved in a wide variety of health-related problems ranging from implant-associated infections to disease transmissions and dental plaque. The usual picture of these bacterial films is that they grow and develop on surfaces. However, suspended biofilm structures, or streamers, have been found in natural environments (e.g., rivers, acid mines, hydrothermal hot springs) and are always suggested to stem from a turbulent flow. We report the formation of bacterial streamers in curved microfluidic channels. By using confocal laser microscopy we are able to directly image and characterize the spatial and temporal evolution of these filamentous structures. Such streamers, which always connect the inner corners of opposite sides of the channel, are always located in the middle plane. Numerical simulations of the flow provide evidences for an underlying hydrodynamic mechanism behind the formation of the streamers.

  20. Collective Functionality through Bacterial Individuality

    Science.gov (United States)

    Ackermann, Martin

    According to the conventional view, the properties of an organism are a product of nature and nurture - of its genes and the environment it lives in. Recent experiments with unicellular organisms have challenged this view: several molecular mechanisms generate phenotypic variation independently of environmental signals, leading to variation in clonal groups. My presentation will focus on the causes and consequences of this microbial individuality. Using examples from bacterial genetic model systems, I will first discuss different molecular and cellular mechanisms that give rise to bacterial individuality. Then, I will discuss the consequences of individuality, and focus on how phenotypic variation in clonal populations of bacteria can promote interactions between individuals, lead to the division of labor, and allow clonal groups of bacteria to cope with environmental uncertainty. Variation between individuals thus provides clonal groups with collective functionality.