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...

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

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.

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...

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...

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.

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.

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

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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.

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

Rat fetuin: distribution of protein and mRNA in embryonic and neonatal rat tissues

DEFF Research Database (Denmark)

Terkelsen, O B; Jahnen-Dechent, W; Nielsen, Henrik

1998-01-01

Fetuin is a serum protein widely distributed in the animal kingdom and found in all mammalian species so far investigated. It is mainly a fetal protein, in the sense that the highest concentrations are found in serum and body fluids of embryos and fetuses. In order to elucidate possible biological......-nucleotides-long digoxigenin-labeled riboprobe. Fetuin was unevenly distributed in all organ systems during development, with the most pronounced expression at E 10Fetuin is a serum protein widely distributed in the animal kingdom and found in all mammalian species so far investigated. It is mainly a fetal...

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].

Distribution and diversity of bacterial secretion systems across metagenomic datasets.

Science.gov (United States)

Barret, Matthieu; Egan, Frank; O'Gara, Fergal

2013-02-01

Bacteria can manipulate their surrounding environment through the secretion of proteins into other living organisms and into the extracellular milieu. In Gram stain negative bacteria this process is mediated by different types of secretion systems from type I through type VI secretion system (T1SS-T6SS). In this study the prevalence of these secretion systems in 312 publicly available microbiomes derived from a wide range of ecosystems was investigated by a gene-centric approach. Our analysis demonstrates that some secretion systems are over-represented in some specific samples. In addition, some T3SS and T6SS phylogenetic clusters were specifically enriched in particular ecological niches, which could indicate specific bacterial adaptation to these environments. © 2012 Society for Applied Microbiology and Blackwell Publishing Ltd.

Escherichia coli type III secretion system 2 (ETT2) is widely distributed in avian pathogenic Escherichia coli isolates from Eastern China.

Science.gov (United States)

Wang, S; Liu, X; Xu, X; Zhao, Y; Yang, D; Han, X; Tian, M; Ding, C; Peng, D; Yu, S

2016-10-01

Pathogens utilize type III secretion systems to deliver effector proteins, which facilitate bacterial infections. The Escherichia coli type III secretion system 2 (ETT2) which plays a crucial role in bacterial virulence, is present in the majority of E. coli strains, although ETT2 has undergone widespread mutational attrition. We investigated the distribution and characteristics of ETT2 in avian pathogenic E. coli (APEC) isolates and identified five different ETT2 isoforms, including intact ETT2, in 57·6% (141/245) of the isolates. The ETT2 locus was present in the predominant APEC serotypes O78, O2 and O1. All of the ETT2 loci in the serotype O78 isolates were degenerate, whereas an intact ETT2 locus was mostly present in O1 and O2 serotype strains, which belong to phylogenetic groups B2 and D, respectively. Interestingly, a putative second type III secretion-associated locus (eip locus) was present only in the isolates with an intact ETT2. Moreover, ETT2 was more widely distributed in APEC isolates and exhibited more isoforms compared to ETT2 in human extraintestinal pathogenic E. coli, suggesting that APEC might be a potential risk to human health. However, there was no distinct correlation between ETT2 and other virulence factors in APEC.

Quantifying Multistate Cytoplasmic Molecular Diffusion in Bacterial Cells via Inverse Transform of Confined Displacement Distribution.

Science.gov (United States)

Chen, Tai-Yen; Jung, Won; Santiago, Ace George; Yang, Feng; Krzemiński, Łukasz; Chen, Peng

2015-11-12

Single-molecule tracking (SMT) of fluorescently tagged cytoplasmic proteins can provide valuable information on the underlying biological processes in living cells via subsequent analysis of the displacement distributions; however, the confinement effect originated from the small size of a bacterial cell skews the protein's displacement distribution and complicates the quantification of the intrinsic diffusive behaviors. Using the inverse transformation method, we convert the skewed displacement distribution (for both 2D and 3D imaging conditions) back to that in free space for systems containing one or multiple (non)interconverting Brownian diffusion states, from which we can reliably extract the number of diffusion states as well as their intrinsic diffusion coefficients and respective fractional populations. We further demonstrate a successful application to experimental SMT data of a transcription factor in living E. coli cells. This work allows a direct quantitative connection between cytoplasmic SMT data with diffusion theory for analyzing molecular diffusive behavior in live bacteria.

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

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

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

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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

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

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)

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.

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 ...

Genus-wide comparison of Pseudovibrio bacterial genomes reveal diverse adaptations to different marine invertebrate hosts.

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Alex, Anoop; Antunes, Agostinho

2018-01-01

Bacteria belonging to the genus Pseudovibrio have been frequently found in association with a wide variety of marine eukaryotic invertebrate hosts, indicative of their versatile and symbiotic lifestyle. A recent comparison of the sponge-associated Pseudovibrio genomes has shed light on the mechanisms influencing a successful symbiotic association with sponges. In contrast, the genomic architecture of Pseudovibrio bacteria associated with other marine hosts has received less attention. Here, we performed genus-wide comparative analyses of 18 Pseudovibrio isolated from sponges, coral, tunicates, flatworm, and seawater. The analyses revealed a certain degree of commonality among the majority of sponge- and coral-associated bacteria. Isolates from other marine invertebrate host, tunicates, exhibited a genetic repertoire for cold adaptation and specific metabolic abilities including mucin degradation in the Antarctic tunicate-associated bacterium Pseudovibrio sp. Tun.PHSC04_5.I4. Reductive genome evolution was simultaneously detected in the flatworm-associated bacteria and the sponge-associated bacterium P. axinellae AD2, through the loss of major secretion systems (type III/VI) and virulence/symbioses factors such as proteins involved in adhesion and attachment to the host. Our study also unraveled the presence of a CRISPR-Cas system in P. stylochi UST20140214-052 a flatworm-associated bacterium possibly suggesting the role of CRISPR-based adaptive immune system against the invading virus particles. Detection of mobile elements and genomic islands (GIs) in all bacterial members highlighted the role of horizontal gene transfer for the acquisition of novel genetic features, likely enhancing the bacterial ecological fitness. These findings are insightful to understand the role of genome diversity in Pseudovibrio as an evolutionary strategy to increase their colonizing success across a wide range of marine eukaryotic hosts.

Structural Aspects of Bacterial Outer Membrane Protein Assembly.

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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.

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...

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

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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

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.

Structural adaptations of proteins to different biological membranes

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Pogozheva, Irina D.; Tristram-Nagle, Stephanie; Mosberg, Henry I.; Lomize, Andrei L.

2013-01-01

To gain insight into adaptations of proteins to their membranes, intrinsic hydrophobic thicknesses, distributions of different chemical groups and profiles of hydrogen-bonding capacities (α and β) and the dipolarity/polarizability parameter (π*) were calculated for lipid-facing surfaces of 460 integral α-helical, β-barrel and peripheral proteins from eight types of biomembranes. For comparison, polarity profiles were also calculated for ten artificial lipid bilayers that have been previously studied by neutron and X-ray scattering. Estimated hydrophobic thicknesses are 30-31 Å for proteins from endoplasmic reticulum, thylakoid, and various bacterial plasma membranes, but differ for proteins from outer bacterial, inner mitochondrial and eukaryotic plasma membranes (23.9, 28.6 and 33.5 Å, respectively). Protein and lipid polarity parameters abruptly change in the lipid carbonyl zone that matches the calculated hydrophobic boundaries. Maxima of positively charged protein groups correspond to the location of lipid phosphates at 20-22 Å distances from the membrane center. Locations of Tyr atoms coincide with hydrophobic boundaries, while distributions maxima of Trp rings are shifted by 3-4 Å toward the membrane center. Distributions of Trp atoms indicate the presence of two 5-8 Å-wide midpolar regions with intermediate π* values within the hydrocarbon core, whose size and symmetry depend on the lipid composition of membrane leaflets. Midpolar regions are especially asymmetric in outer bacterial membranes and cell membranes of mesophilic but not hyperthermophilic archaebacteria, indicating the larger width of the central nonpolar region in the later case. In artificial lipid bilayers, midpolar regions are observed up to the level of acyl chain double bonds. PMID:23811361

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...

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.

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.

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

Changes in bacterial composition of biofilm in a metropolitan drinking water distribution system.

Science.gov (United States)

Revetta, R P; Gomez-Alvarez, V; Gerke, T L; Santo Domingo, J W; Ashbolt, N J

2016-07-01

This study examined the development of bacterial biofilms within a metropolitan distribution system. The distribution system is fed with different source water (i.e. groundwater, GW and surface water, SW) and undergoes different treatment processes in separate facilities. The biofilm community was characterized using 16S rRNA gene clone libraries and functional potential analysis, generated from total DNA extracted from coupons in biofilm annular reactors fed with onsite drinking water for up to 18 months. Differences in the bacterial community structure were observed between GW and SW. Representatives that explained the dissimilarity were associated with the classes Betaproteobacteria, Alphaproteobacteria, Actinobacteria, Gammaproteobacteria and Firmicutes. After 9 months the biofilm bacterial community from both GW and SW were dominated by Mycobacterium species. The distribution of the dominant operational taxonomic unit (OTU) (Mycobacterium) positively correlated with the drinking water distribution system (DWDS) temperature. In this study, the biofilm community structure observed between GW and SW were dissimilar, while communities from different locations receiving SW did not show significant differences. The results suggest that source water and/or the water quality shaped by their respective treatment processes may play an important role in shaping the bacterial communities in the distribution system. In addition, several bacterial groups were present in all samples, suggesting that they are an integral part of the core microbiota of this DWDS. These results provide an ecological insight into biofilm bacterial structure in chlorine-treated drinking water influenced by different water sources and their respective treatment processes. Published 2016. This article is a U.S. Government work and is in the public domain in the USA.

Biogeographical distribution and diversity of bacterial communities in surface sediments of the South China Sea.

Science.gov (United States)

Li, Tao; Wang, Peng

2013-05-01

This paper aims at an investigation of the features of bacterial communities in surface sediments of the South China Sea (SCS). In particular, biogeographical distribution patterns and the phylogenetic diversity of bacteria found in sediments collected from a coral reef platform, a continental slope, and a deep-sea basin were determined. Bacterial diversity was measured by an observation of 16S rRNA genes, and 18 phylogenetic groups were identified in the bacterial clone library. Planctomycetes, Deltaproteobacteria, candidate division OP11, and Alphaproteobacteria made up the majority of the bacteria in the samples, with their mean bacterial clones being 16%, 15%, 12%, and 9%, respectively. By comparison, the bacterial communities found in the SCS surface sediments were significantly different from other previously observed deep-sea bacterial communities. This research also emphasizes the fact that geographical factors have an impact on the biogeographical distribution patterns of bacterial communities. For instance, canonical correspondence analyses illustrated that the percentage of sand weight and water depth are important factors affecting the bacterial community composition. Therefore, this study highlights the importance of adequately determining the relationship between geographical factors and the distribution of bacteria in the world's seas and oceans.

Comparative study to develop a single method for retrieving wide class of recombinant proteins from classical inclusion bodies.

Science.gov (United States)

Padhiar, Arshad Ahmed; Chanda, Warren; Joseph, Thomson Patrick; Guo, Xuefang; Liu, Min; Sha, Li; Batool, Samana; Gao, Yifan; Zhang, Wei; Huang, Min; Zhong, Mintao

2018-03-01

The formation of inclusion bodies (IBs) is considered as an Achilles heel of heterologous protein expression in bacterial hosts. Wide array of techniques has been developed to recover biochemically challenging proteins from IBs. However, acquiring the active state even from the same protein family was found to be an independent of single established method. Here, we present a new strategy for the recovery of wide sub-classes of recombinant protein from harsh IBs. We found that numerous methods and their combinations for reducing IB formation and producing soluble proteins were not effective, if the inclusion bodies were harsh in nature. On the other hand, different practices with mild solubilization buffers were able to solubilize IBs completely, yet the recovery of active protein requires large screening of refolding buffers. With the integration of previously reported mild solubilization techniques, we proposed an improved method, which comprised low sarkosyl concentration, ranging from 0.05 to 0.1% coupled with slow freezing (- 1 °C/min) and fast thaw (room temperature), resulting in greater solubility and the integrity of solubilized protein. Dilution method was employed with single buffer to restore activity for every sub-class of recombinant protein. Results showed that the recovered protein's activity was significantly higher compared with traditional solubilization/refolding approach. Solubilization of IBs by the described method was proved milder in nature, which restored native-like conformation of proteins within IBs.

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.

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.

DistAMo: A web-based tool to characterize DNA-motif distribution on bacterial chromosomes

Directory of Open Access Journals (Sweden)

Patrick eSobetzko

2016-03-01

Full Text Available Short DNA motifs are involved in a multitude of functions such as for example chromosome segregation, DNA replication or mismatch repair. Distribution of such motifs is often not random and the specific chromosomal pattern relates to the respective motif function. Computational approaches which quantitatively assess such chromosomal motif patterns are necessary. Here we present a new computer tool DistAMo (Distribution Analysis of DNA Motifs. The algorithm uses codon redundancy to calculate the relative abundance of short DNA motifs from single genes to entire chromosomes. Comparative genomics analyses of the GATC-motif distribution in γ-proteobacterial genomes using DistAMo revealed that (i genes beside the replication origin are enriched in GATCs, (ii genome-wide GATC distribution follows a distinct pattern and (iii genes involved in DNA replication and repair are enriched in GATCs. These features are specific for bacterial chromosomes encoding a Dam methyltransferase. The new software is available as a stand-alone or as an easy-to-use web-based server version at http://www.computational.bio.uni-giessen.de/distamo.

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.

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.

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...

Isotopomer distributions in amino acids from a highly expressed protein as a proxy for those from total protein

Energy Technology Data Exchange (ETDEWEB)

Shaikh, Afshan; Shaikh, Afshan S.; Tang, Yinjie; Mukhopadhyay, Aindrila; Keasling, Jay D.

2008-06-27

{sup 13}C-based metabolic flux analysis provides valuable information about bacterial physiology. Though many biological processes rely on the synergistic functions of microbial communities, study of individual organisms in a mixed culture using existing flux analysis methods is difficult. Isotopomer-based flux analysis typically relies on hydrolyzed amino acids from a homogeneous biomass. Thus metabolic flux analysis of a given organism in a mixed culture requires its separation from the mixed culture. Swift and efficient cell separation is difficult and a major hurdle for isotopomer-based flux analysis of mixed cultures. Here we demonstrate the use of a single highly-expressed protein to analyze the isotopomer distribution of amino acids from one organism. Using the model organism E. coli expressing a plasmid-borne, his-tagged Green Fluorescent Protein (GFP), we show that induction of GFP does not affect E. coli growth kinetics or the isotopomer distribution in nine key metabolites. Further, the isotopomer labeling patterns of amino acids derived from purified GFP and total cell protein are indistinguishable, indicating that amino acids from a purified protein can be used to infer metabolic fluxes of targeted organisms in a mixed culture. This study provides the foundation to extend isotopomer-based flux analysis to study metabolism of individual strains in microbial communities.

Investigation of protein distribution in solid lipid particles and its impact on protein release using coherent anti-Stokes Raman scattering microscopy

DEFF Research Database (Denmark)

Christophersen, Philip C.; Birch, Ditlev; Saarinen, Jukka

2015-01-01

The aim of this study was to gain new insights into protein distribution in solid lipid microparticles (SLMs) and subsequent release mechanisms using a novel label-free chemical imaging method, coherent anti-Stokes Raman scattering (CARS) microscopy. Lysozyme-loaded SLMs were prepared using...... in the solid lipid matrix, which required full lipolysis of the entire matrix to release lysozyme completely. Therefore, SLMs with lysozyme incorporated in an aqueous solution released lysozyme much faster than with lysozyme incorporated as a solid. In conclusion, CARS microscopy was an efficient and non......-destructive method for elucidating the distribution of lysozyme in SLMs. The interpretation of protein distribution and release during lipolysis enabled elucidation of protein release mechanisms. In future, CARS microscopy analysis could facilitate development of a wide range of protein-lipid matrices with tailor...

An Architecture for a Wide Area Distributed System

NARCIS (Netherlands)

Homburg, P.; Steen, M.R. van; Tanenbaum, A.S.

1996-01-01

Distributed systems provide sharing of resources and information over a computer network. A key design issue that makes these systems attractive is that all aspects related to distribution are transparent to users. Unfortunately, general-purpose wide area distributed systems that allow users to

The utility of protein structure as a predictor of site-wise dN/dS varies widely among HIV-1 proteins.

Science.gov (United States)

Meyer, Austin G; Wilke, Claus O

2015-10-06

Protein structure acts as a general constraint on the evolution of viral proteins. One widely recognized structural constraint explaining evolutionary variation among sites is the relative solvent accessibility (RSA) of residues in the folded protein. In influenza virus, the distance from functional sites has been found to explain an additional portion of the evolutionary variation in the external antigenic proteins. However, to what extent RSA and distance from a reference site in the protein can be used more generally to explain protein adaptation in other viruses and in the different proteins of any given virus remains an open question. To address this question, we have carried out an analysis of the distribution and structural predictors of site-wise dN/dS in HIV-1. Our results indicate that the distribution of dN/dS in HIV follows a smooth gamma distribution, with no special enrichment or depletion of sites with dN/dS at or above one. The variation in dN/dS can be partially explained by RSA and distance from a reference site in the protein, but these structural constraints do not act uniformly among the different HIV-1 proteins. Structural constraints are highly predictive in just one of the three enzymes and one of three structural proteins in HIV-1. For these two proteins, the protease enzyme and the gp120 structural protein, structure explains between 30 and 40% of the variation in dN/dS. Finally, for the gp120 protein of the receptor-binding complex, we also find that glycosylation sites explain just 2% of the variation in dN/dS and do not explain gp120 evolution independently of either RSA or distance from the apical surface. © 2015 The Author(s).

Bacterial community dynamics and product distribution during pH-adjusted fermentation of vegetable wastes.

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Ye, N-F; Lü, F; Shao, L-M; Godon, J-J; He, P-J

2007-10-01

To estimate the effect of pH on the structures of bacterial community during fermentation of vegetable wastes and to investigate the relationship between bacterial community dynamics and product distribution. The bacterial communities in five batch tests controlled at different pH values [uncontrolled (about pH 4), 5, 6, 7 and 8] were monitored by denaturing gradient gel electrophoresis (DGGE) and single-strand conformation polymorphism (SSCP). The two fingerprinting methods provided consistent results and principal component analysis indicated a close similarity of bacterial community at pH 7 and 8 in addition to those at pH 4-6. This clustering also corresponded to dominant metabolic pathway. Thus, pH 7-8 shifted from alcohol-forming to acid-forming, especially butyric acid, whereas both alcohol-forming and acid-forming dominated at pH 5-6, and at pH 4, fermentation was inhibited. Shannon-weaver index was calculated to analyse the DGGE profiles, which revealed that the bacterial diversities at pH 7 and 8 were the highest while those at pH 5 and 4 (uncontrolled) were the lowest. According to sequencing results of the bands excised from DGGE gels, lactic acid bacteria and Clostridium sp. were predominant at all pH values, but varieties in species were observed as pH changed and time prolonged. The bacterial community during fermentation was materially influenced by pH and the diverse product distribution was related to the shift of different bacterial population. The study reveals that the impact of pH on fermentation product distribution is implemented primarily by changes of bacterial community. It also provides information about the comparison of two fingerprinting methods, DGGE and SSCP.

Distribution and life strategies of two bacterial populations in a eutrophic lake

Science.gov (United States)

Weinbauer; Hofle

1998-10-01

Monoclonal antibodies and epifluorescence microscopy were used to determine the depth distribution of two indigenous bacterial populations in the stratified Lake Plusssee and characterize their life strategies. Populations of Comamonas acidovorans PX54 showed a depth distribution with maximum abundances in the oxic epilimnion, whereas Aeromonas hydrophila PU7718 showed a depth distribution with maximum abundances in the anoxic thermocline layer (metalimnion), i. e., in the water layer with the highest microbial activity. Resistance of PX54 to protist grazing and high metabolic versatility and growth rate of PU7718 were the most important life strategy traits for explaining the depth distribution of the two bacterial populations. Maximum abundance of PX54 was 16,000 cells per ml, and maximum abundance of PU7718 was 20,000 cells per ml. Determination of bacterial productivity in dilution cultures with different-size fractions of dissolved organic matter (DOM) from lake water indicates that low-molecular-weight (LMW) DOM is less bioreactive than total DOM (TDOM). The abundance and growth rate of PU7718 were highest in the TDOM fractions, whereas those of PX54 were highest in the LMW DOM fraction, demonstrating that PX54 can grow well on the less bioreactive DOM fraction. We estimated that 13 to 24% of the entire bacterial community and 14% of PU7718 were removed by viral lysis, whereas no significant effect of viral lysis on PX54 could be detected. Growth rates of PX54 (0.11 to 0.13 h-1) were higher than those of the entire bacterial community (0.04 to 0.08 h-1) but lower than those of PU7718 (0.26 to 0.31 h-1). In undiluted cultures, the growth rates were significantly lower, pointing to density effects such as resource limitation or antibiosis, and the effects were stronger for PU7718 and the entire bacterial community than for PX54. Life strategy characterizations based on data from literature and this study revealed that the fast-growing and metabolically

Widely distributed SEP events and pseudostreamers

Science.gov (United States)

Panasenco, O.; Panasenco, A.; Velli, M.

2017-12-01

Our analysis of the pseudostreamer magnetic topology reveals new interesting implications for understanding SEP acceleration in CMEs. The possible reasons for the wide distribution of some SEP events can be the presence of pseudostreamers in the vicinity of the SEP source region which creates conditions for the existence of strong longitudinal spread of energetic particles as well as an anomalous longitudinal solar wind magnetic field component. We reconstructed the 3D magnetic configurations of pseudostreamers with a potential field source surface (PFSS) model, which uses as a lower boundary condition the magnetic field derived from an evolving surface-flux transport model. In order to estimate the possible magnetic connections between the spacecraft and the SEP source region, we used the Parker spiral, ENLIL and PFSS models. We found that in cases of the wide SEP distributions a specific configuration of magnetic field appears to exist at low solar latitudes all the way around the sun, we named this phenomenon a pseudostreamers belt. It appears that the presence of the well developed pseudostreamer or, rather multiple pseudostreamers, organized into the pseudostreamer belt can be considered as a very favorable condition for wide SEP events.

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

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

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...

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.

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

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.

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.

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)

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.

Wide-band segmented power distribution networks

NARCIS (Netherlands)

Tereshchenko, O.V.; Buesink, Frederik Johannes Karel; Leferink, Frank Bernardus Johannes

2013-01-01

This paper discusses a novel design of Power Distribution Network (PDN). By physical structuring of the power plane into repetitive symmetrical and asymmetrical segments of varying size, suppression of the propagation of unwanted noise throughout the PDN over a wide frequency range is achieved.

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.

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...

Exact protein distributions for stochastic models of gene expression using partitioning of Poisson processes.

Science.gov (United States)

Pendar, Hodjat; Platini, Thierry; Kulkarni, Rahul V

2013-04-01

Stochasticity in gene expression gives rise to fluctuations in protein levels across a population of genetically identical cells. Such fluctuations can lead to phenotypic variation in clonal populations; hence, there is considerable interest in quantifying noise in gene expression using stochastic models. However, obtaining exact analytical results for protein distributions has been an intractable task for all but the simplest models. Here, we invoke the partitioning property of Poisson processes to develop a mapping that significantly simplifies the analysis of stochastic models of gene expression. The mapping leads to exact protein distributions using results for mRNA distributions in models with promoter-based regulation. Using this approach, we derive exact analytical results for steady-state and time-dependent distributions for the basic two-stage model of gene expression. Furthermore, we show how the mapping leads to exact protein distributions for extensions of the basic model that include the effects of posttranscriptional and posttranslational regulation. The approach developed in this work is widely applicable and can contribute to a quantitative understanding of stochasticity in gene expression and its regulation.

Exact protein distributions for stochastic models of gene expression using partitioning of Poisson processes

Science.gov (United States)

Pendar, Hodjat; Platini, Thierry; Kulkarni, Rahul V.

2013-04-01

Stochasticity in gene expression gives rise to fluctuations in protein levels across a population of genetically identical cells. Such fluctuations can lead to phenotypic variation in clonal populations; hence, there is considerable interest in quantifying noise in gene expression using stochastic models. However, obtaining exact analytical results for protein distributions has been an intractable task for all but the simplest models. Here, we invoke the partitioning property of Poisson processes to develop a mapping that significantly simplifies the analysis of stochastic models of gene expression. The mapping leads to exact protein distributions using results for mRNA distributions in models with promoter-based regulation. Using this approach, we derive exact analytical results for steady-state and time-dependent distributions for the basic two-stage model of gene expression. Furthermore, we show how the mapping leads to exact protein distributions for extensions of the basic model that include the effects of posttranscriptional and posttranslational regulation. The approach developed in this work is widely applicable and can contribute to a quantitative understanding of stochasticity in gene expression and its regulation.

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.

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.

Long-Term Bacterial Dynamics in a Full-Scale Drinking Water Distribution System.

Science.gov (United States)

Prest, E I; Weissbrodt, D G; Hammes, F; van Loosdrecht, M C M; Vrouwenvelder, J S

2016-01-01

Large seasonal variations in microbial drinking water quality can occur in distribution networks, but are often not taken into account when evaluating results from short-term water sampling campaigns. Temporal dynamics in bacterial community characteristics were investigated during a two-year drinking water monitoring campaign in a full-scale distribution system operating without detectable disinfectant residual. A total of 368 water samples were collected on a biweekly basis at the water treatment plant (WTP) effluent and at one fixed location in the drinking water distribution network (NET). The samples were analysed for heterotrophic plate counts (HPC), Aeromonas plate counts, adenosine-tri-phosphate (ATP) concentrations, and flow cytometric (FCM) total and intact cell counts (TCC, ICC), water temperature, pH, conductivity, total organic carbon (TOC) and assimilable organic carbon (AOC). Multivariate analysis of the large dataset was performed to explore correlative trends between microbial and environmental parameters. The WTP effluent displayed considerable seasonal variations in TCC (from 90 × 103 cells mL-1 in winter time up to 455 × 103 cells mL-1 in summer time) and in bacterial ATP concentrations (water temperature variations. These fluctuations were not detected with HPC and Aeromonas counts. The water in the network was predominantly influenced by the characteristics of the WTP effluent. The increase in ICC between the WTP effluent and the network sampling location was small (34 × 103 cells mL-1 on average) compared to seasonal fluctuations in ICC in the WTP effluent. Interestingly, the extent of bacterial growth in the NET was inversely correlated to AOC concentrations in the WTP effluent (Pearson's correlation factor r = -0.35), and positively correlated with water temperature (r = 0.49). Collecting a large dataset at high frequency over a two year period enabled the characterization of previously undocumented seasonal dynamics in the distribution

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.

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...

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.

CHANGES IN BACTERIAL COMPOSITION OF BIOFILM IN A METROPOLITAN DRINKING WATER DISTRIBUTION SYSTEM

Science.gov (United States)

This study examined the development of bacterial biofilms within a metropolitan distribution system. The distribution system is fed with different source water (i.e., groundwater, GW and surface water, SW) and undergoes different treatment processes in separate facilities. The b...

Photosynthesis Is Widely Distributed among Proteobacteria as Demonstrated by the Phylogeny of PufLM Reaction Center Proteins

Directory of Open Access Journals (Sweden)

Johannes F. Imhoff

2018-01-01

Full Text Available Two different photosystems for performing bacteriochlorophyll-mediated photosynthetic energy conversion are employed in different bacterial phyla. Those bacteria employing a photosystem II type of photosynthetic apparatus include the phototrophic purple bacteria (Proteobacteria, Gemmatimonas and Chloroflexus with their photosynthetic relatives. The proteins of the photosynthetic reaction center PufL and PufM are essential components and are common to all bacteria with a type-II photosynthetic apparatus, including the anaerobic as well as the aerobic phototrophic Proteobacteria. Therefore, PufL and PufM proteins and their genes are perfect tools to evaluate the phylogeny of the photosynthetic apparatus and to study the diversity of the bacteria employing this photosystem in nature. Almost complete pufLM gene sequences and the derived protein sequences from 152 type strains and 45 additional strains of phototrophic Proteobacteria employing photosystem II were compared. The results give interesting and comprehensive insights into the phylogeny of the photosynthetic apparatus and clearly define Chromatiales, Rhodobacterales, Sphingomonadales as major groups distinct from other Alphaproteobacteria, from Betaproteobacteria and from Caulobacterales (Brevundimonas subvibrioides. A special relationship exists between the PufLM sequences of those bacteria employing bacteriochlorophyll b instead of bacteriochlorophyll a. A clear phylogenetic association of aerobic phototrophic purple bacteria to anaerobic purple bacteria according to their PufLM sequences is demonstrated indicating multiple evolutionary lines from anaerobic to aerobic phototrophic purple bacteria. The impact of pufLM gene sequences for studies on the environmental diversity of phototrophic bacteria is discussed and the possibility of their identification on the species level in environmental samples is pointed out.

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

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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.

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.

A systematic approach for the assessment of bacterial growth-controlling factors linked to biological stability of drinking water in distribution systems

KAUST Repository

Prest, E. I.

2016-01-06

A systematic approach is presented for the assessment of (i) bacterial growth-controlling factors in drinking water and (ii) the impact of distribution conditions on the extent of bacterial growth in full-scale distribution systems. The approach combines (i) quantification of changes in autochthonous bacterial cell concentrations in full-scale distribution systems with (ii) laboratoryscale batch bacterial growth potential tests of drinking water samples under defined conditions. The growth potential tests were done by direct incubation of water samples, without modification of the original bacterial flora, and with flow cytometric quantification of bacterial growth. This method was shown to be reproducible (ca. 4% relative standard deviation) and sensitive (detection of bacterial growth down to 5 μg L-1 of added assimilable organic carbon). The principle of step-wise assessment of bacterial growth-controlling factors was demonstrated on bottled water, shown to be primarily carbon limited at 133 (±18) × 103 cells mL-1 and secondarily limited by inorganic nutrients at 5,500 (±1,700) × 103 cells mL-1. Analysis of the effluent of a Dutch full-scale drinking water treatment plant showed (1) bacterial growth inhibition as a result of end-point chlorination, (2) organic carbon limitation at 192 (±72) × 103 cells mL-1 and (3) inorganic nutrient limitation at 375 (±31) × 103 cells mL-1. Significantly lower net bacterial growth was measured in the corresponding full-scale distribution system (176 (±25) × 103 cells mL-1) than in the laboratory-scale growth potential test of the same water (294 (±35) × 103 cells mL-1), highlighting the influence of distribution on bacterial growth. The systematic approach described herein provides quantitative information on the effect of drinking water properties and distribution system conditions on biological stability, which can assist water utilities in decision-making on treatment or distribution system improvements to

Long-Term Bacterial Dynamics in a Full-Scale Drinking Water Distribution System

KAUST Repository

Prest, E. I.; Weissbrodt, D. G.; Hammes, F.; Van Loosdrecht, M. C M; Vrouwenvelder, Johannes S.

2016-01-01

Large seasonal variations in microbial drinking water quality can occur in distribution networks, but are often not taken into account when evaluating results from short-term water sampling campaigns. Temporal dynamics in bacterial community characteristics were investigated during a two-year drinking water monitoring campaign in a full-scale distribution system operating without detectable disinfectant residual. A total of 368 water samples were collected on a biweekly basis at the water treatment plant (WTP) effluent and at one fixed location in the drinking water distribution network (NET). The samples were analysed for heterotrophic plate counts (HPC), Aeromonas plate counts, adenosine-tri-phosphate (ATP) concentrations, and flow cytometric (FCM) total and intact cell counts (TCC, ICC), water temperature, pH, conductivity, total organic carbon (TOC) and assimilable organic carbon (AOC). Multivariate analysis of the large dataset was performed to explore correlative trends between microbial and environmental parameters. The WTP effluent displayed considerable seasonal variations in TCC (from 90 × 103 cells mL-1 in winter time up to 455 × 103 cells mL-1 in summer time) and in bacterial ATP concentrations (<1–3.6 ng L-1), which were congruent with water temperature variations. These fluctuations were not detected with HPC and Aeromonas counts. The water in the network was predominantly influenced by the characteristics of the WTP effluent. The increase in ICC between the WTP effluent and the network sampling location was small (34 × 103 cells mL-1 on average) compared to seasonal fluctuations in ICC in the WTP effluent. Interestingly, the extent of bacterial growth in the NET was inversely correlated to AOC concentrations in the WTP effluent (Pearson’s correlation factor r = -0.35), and positively correlated with water temperature (r = 0.49). Collecting a large dataset at high frequency over a two year period enabled the characterization of previously

Long-Term Bacterial Dynamics in a Full-Scale Drinking Water Distribution System.

Directory of Open Access Journals (Sweden)

E I Prest

Full Text Available Large seasonal variations in microbial drinking water quality can occur in distribution networks, but are often not taken into account when evaluating results from short-term water sampling campaigns. Temporal dynamics in bacterial community characteristics were investigated during a two-year drinking water monitoring campaign in a full-scale distribution system operating without detectable disinfectant residual. A total of 368 water samples were collected on a biweekly basis at the water treatment plant (WTP effluent and at one fixed location in the drinking water distribution network (NET. The samples were analysed for heterotrophic plate counts (HPC, Aeromonas plate counts, adenosine-tri-phosphate (ATP concentrations, and flow cytometric (FCM total and intact cell counts (TCC, ICC, water temperature, pH, conductivity, total organic carbon (TOC and assimilable organic carbon (AOC. Multivariate analysis of the large dataset was performed to explore correlative trends between microbial and environmental parameters. The WTP effluent displayed considerable seasonal variations in TCC (from 90 × 103 cells mL-1 in winter time up to 455 × 103 cells mL-1 in summer time and in bacterial ATP concentrations (<1-3.6 ng L-1, which were congruent with water temperature variations. These fluctuations were not detected with HPC and Aeromonas counts. The water in the network was predominantly influenced by the characteristics of the WTP effluent. The increase in ICC between the WTP effluent and the network sampling location was small (34 × 103 cells mL-1 on average compared to seasonal fluctuations in ICC in the WTP effluent. Interestingly, the extent of bacterial growth in the NET was inversely correlated to AOC concentrations in the WTP effluent (Pearson's correlation factor r = -0.35, and positively correlated with water temperature (r = 0.49. Collecting a large dataset at high frequency over a two year period enabled the characterization of previously

Long-Term Bacterial Dynamics in a Full-Scale Drinking Water Distribution System

KAUST Repository

Prest, E. I.

2016-10-28

Large seasonal variations in microbial drinking water quality can occur in distribution networks, but are often not taken into account when evaluating results from short-term water sampling campaigns. Temporal dynamics in bacterial community characteristics were investigated during a two-year drinking water monitoring campaign in a full-scale distribution system operating without detectable disinfectant residual. A total of 368 water samples were collected on a biweekly basis at the water treatment plant (WTP) effluent and at one fixed location in the drinking water distribution network (NET). The samples were analysed for heterotrophic plate counts (HPC), Aeromonas plate counts, adenosine-tri-phosphate (ATP) concentrations, and flow cytometric (FCM) total and intact cell counts (TCC, ICC), water temperature, pH, conductivity, total organic carbon (TOC) and assimilable organic carbon (AOC). Multivariate analysis of the large dataset was performed to explore correlative trends between microbial and environmental parameters. The WTP effluent displayed considerable seasonal variations in TCC (from 90 × 103 cells mL-1 in winter time up to 455 × 103 cells mL-1 in summer time) and in bacterial ATP concentrations (<1–3.6 ng L-1), which were congruent with water temperature variations. These fluctuations were not detected with HPC and Aeromonas counts. The water in the network was predominantly influenced by the characteristics of the WTP effluent. The increase in ICC between the WTP effluent and the network sampling location was small (34 × 103 cells mL-1 on average) compared to seasonal fluctuations in ICC in the WTP effluent. Interestingly, the extent of bacterial growth in the NET was inversely correlated to AOC concentrations in the WTP effluent (Pearson’s correlation factor r = -0.35), and positively correlated with water temperature (r = 0.49). Collecting a large dataset at high frequency over a two year period enabled the characterization of previously

HIV Genome-Wide Protein Associations: a Review of 30 Years of Research

Science.gov (United States)

2016-01-01

SUMMARY The HIV genome encodes a small number of viral proteins (i.e., 16), invariably establishing cooperative associations among HIV proteins and between HIV and host proteins, to invade host cells and hijack their internal machineries. As a known example, the HIV envelope glycoprotein GP120 is closely associated with GP41 for viral entry. From a genome-wide perspective, a hypothesis can be worked out to determine whether 16 HIV proteins could develop 120 possible pairwise associations either by physical interactions or by functional associations mediated via HIV or host molecules. Here, we present the first systematic review of experimental evidence on HIV genome-wide protein associations using a large body of publications accumulated over the past 3 decades. Of 120 possible pairwise associations between 16 HIV proteins, at least 34 physical interactions and 17 functional associations have been identified. To achieve efficient viral replication and infection, HIV protein associations play essential roles (e.g., cleavage, inhibition, and activation) during the HIV life cycle. In either a dispensable or an indispensable manner, each HIV protein collaborates with another viral protein to accomplish specific activities that precisely take place at the proper stages of the HIV life cycle. In addition, HIV genome-wide protein associations have an impact on anti-HIV inhibitors due to the extensive cross talk between drug-inhibited proteins and other HIV proteins. Overall, this study presents for the first time a comprehensive overview of HIV genome-wide protein associations, highlighting meticulous collaborations between all viral proteins during the HIV life cycle. PMID:27357278

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.

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

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.

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.

Mass spectrometry analysis of proteome-wide proteolytic post-translational degradation of proteins

OpenAIRE

Shen, Yufeng; Hixson, Kim K.; Tolić, Nikola; Camp, David G.; Purvine, Samuel O.; Moore, Ronald J.; Smith, Richard D.

2008-01-01

Protein proteolytic degradation is an essential component to proper cell function and its life cycle. Here, we study the protein degradation in yeast Saccharomyces cerevisiae cells on a proteome-wide scale by detection of the intermediate peptides produced from the intracellular degradation of proteins using sequencing-based tandem mass spectrometry. By tracing the detected ~1,100 peptides and their ~200 protein substrate origins we obtain evidence for new insights into the proteome-wide prot...

Do bacterial cell numbers follow a theoretical Poisson distribution? Comparison of experimentally obtained numbers of single cells with random number generation via computer simulation.

Science.gov (United States)

Koyama, Kento; Hokunan, Hidekazu; Hasegawa, Mayumi; Kawamura, Shuso; Koseki, Shigenobu

2016-12-01

We investigated a bacterial sample preparation procedure for single-cell studies. In the present study, we examined whether single bacterial cells obtained via 10-fold dilution followed a theoretical Poisson distribution. Four serotypes of Salmonella enterica, three serotypes of enterohaemorrhagic Escherichia coli and one serotype of Listeria monocytogenes were used as sample bacteria. An inoculum of each serotype was prepared via a 10-fold dilution series to obtain bacterial cell counts with mean values of one or two. To determine whether the experimentally obtained bacterial cell counts follow a theoretical Poisson distribution, a likelihood ratio test between the experimentally obtained cell counts and Poisson distribution which parameter estimated by maximum likelihood estimation (MLE) was conducted. The bacterial cell counts of each serotype sufficiently followed a Poisson distribution. Furthermore, to examine the validity of the parameters of Poisson distribution from experimentally obtained bacterial cell counts, we compared these with the parameters of a Poisson distribution that were estimated using random number generation via computer simulation. The Poisson distribution parameters experimentally obtained from bacterial cell counts were within the range of the parameters estimated using a computer simulation. These results demonstrate that the bacterial cell counts of each serotype obtained via 10-fold dilution followed a Poisson distribution. The fact that the frequency of bacterial cell counts follows a Poisson distribution at low number would be applied to some single-cell studies with a few bacterial cells. In particular, the procedure presented in this study enables us to develop an inactivation model at the single-cell level that can estimate the variability of survival bacterial numbers during the bacterial death process. Copyright © 2016 Elsevier Ltd. All rights reserved.

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.

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

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.

Mathematical modeling and comparison of protein size distribution in different plant, animal, fungal and microbial species reveals a negative correlation between protein size and protein number, thus providing insight into the evolution of proteomes

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Tiessen Axel

2012-02-01

Full Text Available Abstract Background The sizes of proteins are relevant to their biochemical structure and for their biological function. The statistical distribution of protein lengths across a diverse set of taxa can provide hints about the evolution of proteomes. Results Using the full genomic sequences of over 1,302 prokaryotic and 140 eukaryotic species two datasets containing 1.2 and 6.1 million proteins were generated and analyzed statistically. The lengthwise distribution of proteins can be roughly described with a gamma type or log-normal model, depending on the species. However the shape parameter of the gamma model has not a fixed value of 2, as previously suggested, but varies between 1.5 and 3 in different species. A gamma model with unrestricted shape parameter described best the distributions in ~48% of the species, whereas the log-normal distribution described better the observed protein sizes in 42% of the species. The gamma restricted function and the sum of exponentials distribution had a better fitting in only ~5% of the species. Eukaryotic proteins have an average size of 472 aa, whereas bacterial (320 aa and archaeal (283 aa proteins are significantly smaller (33-40% on average. Average protein sizes in different phylogenetic groups were: Alveolata (628 aa, Amoebozoa (533 aa, Fornicata (543 aa, Placozoa (453 aa, Eumetazoa (486 aa, Fungi (487 aa, Stramenopila (486 aa, Viridiplantae (392 aa. Amino acid composition is biased according to protein size. Protein length correlated negatively with %C, %M, %K, %F, %R, %W, %Y and positively with %D, %E, %Q, %S and %T. Prokaryotic proteins had a different protein size bias for %E, %G, %K and %M as compared to eukaryotes. Conclusions Mathematical modeling of protein length empirical distributions can be used to asses the quality of small ORFs annotation in genomic releases (detection of too many false positive small ORFs. There is a negative correlation between average protein size and total number of

Long-Term Bacterial Dynamics in a Full-Scale Drinking Water Distribution System

NARCIS (Netherlands)

Prest, E.I.E.D.; Weissbrodt, D.G.; Hammes, F; van Loosdrecht, Mark C.M.; Vrouwenvelder, J.S.

2016-01-01

Large seasonal variations in microbial drinking water quality can occur in distribution networks, but are often not taken into account when evaluating results from short-term water sampling campaigns. Temporal dynamics in bacterial community characteristics were investigated during a two-year

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...

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.

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

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.

Plant 115-kDa actin-filament bundling protein, P-115-ABP, is a homologue of plant villin and is widely distributed in cells.

Science.gov (United States)

Yokota, Etsuo; Vidali, Luis; Tominaga, Motoki; Tahara, Hiroshi; Orii, Hidefumi; Morizane, Yosuke; Hepler, Peter K; Shimmen, Teruo

2003-10-01

In many cases, actin filaments are arranged into bundles and serve as tracks for cytoplasmic streaming in plant cells. We have isolated an actin-filament bundling protein, which is composed of 115-kDa polypeptide (P-115-ABP), from the germinating pollen of lily, Lilium longiflorum [Nakayasu et al. (1998) BIOCHEM: Biophys. Res. Commun. 249: 61]. P-115-ABP shared similar antigenicity with a plant 135-kDa actin-filament bundling protein (P-135-ABP), a plant homologue of villin. A full-length cDNA clone (ABP115; accession no. AB097407) was isolated from an expression cDNA library of lily pollen by immuno-screening using antisera against P-115-ABP and P-135-ABP. The amino acid sequence of P-115-ABP deduced from this clone showed high homology with those of P-135-ABP and four villin isoforms of Arabidopsis thaliana (AtVLN1, AtVLN2, AtVLN3 and AtVLN4), especially AtVLN4, indicating that P-115-ABP can also be classified as a plant villin. The P-115-ABP isolated biochemically from the germinating lily pollen was able to arrange F-actin filaments with uniform polarity into bundles and this bundling activity was suppressed by Ca2+-calmodulin (CaM), similar to the actin-filament bundling properties of P-135-ABP. The P-115-ABP type of plant villin was widely distributed in plant cells, from algae to land plants. In root hair cells of Hydrocharis dubia, this type of plant villin was co-localized with actin-filament bundles in the transvacuolar strands and the sub-cortical regions. Microinjection of the antiserum against P-115-ABP into living root hair cells caused the disappearance of transvaculor strands and alteration of the route of cytoplasmic streaming. In internodal cells of Chara corallina in which the P-135-ABP type of plant villin is lacking, the P-115-ABP type showed co-localization with actin-filament cables anchored on the intracellular surface of chloroplasts. These results indicated that plant villins are widely distributed and involved in the organization of actin

Dynamics of bacterial communities before and after distribution in a full-scale drinking water network.

Science.gov (United States)

El-Chakhtoura, Joline; Prest, Emmanuelle; Saikaly, Pascal; van Loosdrecht, Mark; Hammes, Frederik; Vrouwenvelder, Hans

2015-05-01

Understanding the biological stability of drinking water distribution systems is imperative in the framework of process control and risk management. The objective of this research was to examine the dynamics of the bacterial community during drinking water distribution at high temporal resolution. Water samples (156 in total) were collected over short time-scales (minutes/hours/days) from the outlet of a treatment plant and a location in its corresponding distribution network. The drinking water is treated by biofiltration and disinfectant residuals are absent during distribution. The community was analyzed by 16S rRNA gene pyrosequencing and flow cytometry as well as conventional, culture-based methods. Despite a random dramatic event (detected with pyrosequencing and flow cytometry but not with plate counts), the bacterial community profile at the two locations did not vary significantly over time. A diverse core microbiome was shared between the two locations (58-65% of the taxa and 86-91% of the sequences) and found to be dependent on the treatment strategy. The bacterial community structure changed during distribution, with greater richness detected in the network and phyla such as Acidobacteria and Gemmatimonadetes becoming abundant. The rare taxa displayed the highest dynamicity, causing the major change during water distribution. This change did not have hygienic implications and is contingent on the sensitivity of the applied methods. The concept of biological stability therefore needs to be revised. Biostability is generally desired in drinking water guidelines but may be difficult to achieve in large-scale complex distribution systems that are inherently dynamic. Copyright © 2015 Elsevier Ltd. All rights reserved.

Dynamics of bacterial communities before and after distribution in a full-scale drinking water network

KAUST Repository

El Chakhtoura, Joline

2015-05-01

Understanding the biological stability of drinking water distribution systems is imperative in the framework of process control and risk management. The objective of this research was to examine the dynamics of the bacterial community during drinking water distribution at high temporal resolution. Water samples (156 in total) were collected over short time-scales (minutes/hours/days) from the outlet of a treatment plant and a location in its corresponding distribution network. The drinking water is treated by biofiltration and disinfectant residuals are absent during distribution. The community was analyzed by 16S rRNA gene pyrosequencing and flow cytometry as well as conventional, culture-based methods. Despite a random dramatic event (detected with pyrosequencing and flow cytometry but not with plate counts), the bacterial community profile at the two locations did not vary significantly over time. A diverse core microbiome was shared between the two locations (58-65% of the taxa and 86-91% of the sequences) and found to be dependent on the treatment strategy. The bacterial community structure changed during distribution, with greater richness detected in the network and phyla such as Acidobacteria and Gemmatimonadetes becoming abundant. The rare taxa displayed the highest dynamicity, causing the major change during water distribution. This change did not have hygienic implications and is contingent on the sensitivity of the applied methods. The concept of biological stability therefore needs to be revised. Biostability is generally desired in drinking water guidelines but may be difficult to achieve in large-scale complex distribution systems that are inherently dynamic.

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.

The bacterial actin MreB rotates, and rotation depends on cell-wall assembly.

Science.gov (United States)

van Teeffelen, Sven; Wang, Siyuan; Furchtgott, Leon; Huang, Kerwyn Casey; Wingreen, Ned S; Shaevitz, Joshua W; Gitai, Zemer

2011-09-20

Bacterial cells possess multiple cytoskeletal proteins involved in a wide range of cellular processes. These cytoskeletal proteins are dynamic, but the driving forces and cellular functions of these dynamics remain poorly understood. Eukaryotic cytoskeletal dynamics are often driven by motor proteins, but in bacteria no motors that drive cytoskeletal motion have been identified to date. Here, we quantitatively study the dynamics of the Escherichia coli actin homolog MreB, which is essential for the maintenance of rod-like cell shape in bacteria. We find that MreB rotates around the long axis of the cell in a persistent manner. Whereas previous studies have suggested that MreB dynamics are driven by its own polymerization, we show that MreB rotation does not depend on its own polymerization but rather requires the assembly of the peptidoglycan cell wall. The cell-wall synthesis machinery thus either constitutes a novel type of extracellular motor that exerts force on cytoplasmic MreB, or is indirectly required for an as-yet-unidentified motor. Biophysical simulations suggest that one function of MreB rotation is to ensure a uniform distribution of new peptidoglycan insertion sites, a necessary condition to maintain rod shape during growth. These findings both broaden the view of cytoskeletal motors and deepen our understanding of the physical basis of bacterial morphogenesis.

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.

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.

Mass distribution and spatial organization of the linear bacterial motor of Spiroplasma citri R8A2.

Science.gov (United States)

Trachtenberg, Shlomo; Andrews, S Brian; Leapman, Richard D

2003-03-01

In the simple, helical, wall-less bacterial genus Spiroplasma, chemotaxis and motility are effected by a linear, contractile motor arranged as a flat cytoskeletal ribbon attached to the inner side of the membrane along the shortest helical line. With scanning transmission electron microscopy and diffraction analysis, we determined the hierarchical and spatial organization of the cytoskeleton of Spiroplasma citri R8A2. The structural unit appears to be a fibril, approximately 5 nm wide, composed of dimers of a 59-kDa protein; each ribbon is assembled from seven fibril pairs. The functional unit of the intact ribbon is a pair of aligned fibrils, along which pairs of dimers form tetrameric ring-like repeats. On average, isolated and purified ribbons contain 14 fibrils or seven well-aligned fibril pairs, which are the same structures observed in the intact cell. Scanning transmission electron microscopy mass analysis and sodium dodecyl sulfate-polyacrylamide gel electrophoresis of purified cytoskeletons indicate that the 59-kDa protein is the only constituent of the ribbons.

Bioinformatic evidence for a widely distributed, ribosomally produced electron carrier precursor, its maturation proteins, and its nicotinoprotein redox partners

Directory of Open Access Journals (Sweden)

Haft Daniel H

2011-01-01

Full Text Available Abstract Background Enzymes in the radical SAM (rSAM domain family serve in a wide variety of biological processes, including RNA modification, enzyme activation, bacteriocin core peptide maturation, and cofactor biosynthesis. Evolutionary pressures and relationships to other cellular constituents impose recognizable grammars on each class of rSAM-containing system, shaping patterns in results obtained through various comparative genomics analyses. Results An uncharacterized gene cluster found in many Actinobacteria and sporadically in Firmicutes, Chloroflexi, Deltaproteobacteria, and one Archaeal plasmid contains a PqqE-like rSAM protein family that includes Rv0693 from Mycobacterium tuberculosis. Members occur clustered with a strikingly well-conserved small polypeptide we designate "mycofactocin," similar in size to bacteriocins and PqqA, precursor of pyrroloquinoline quinone (PQQ. Partial Phylogenetic Profiling (PPP based on the distribution of these markers identifies the mycofactocin cluster, but also a second tier of high-scoring proteins. This tier, strikingly, is filled with up to thirty-one members per genome from three variant subfamilies that occur, one each, in three unrelated classes of nicotinoproteins. The pattern suggests these variant enzymes require not only NAD(P, but also the novel gene cluster. Further study was conducted using SIMBAL, a PPP-like tool, to search these nicotinoproteins for subsequences best correlated across multiple genomes to the presence of mycofactocin. For both the short chain dehydrogenase/reductase (SDR and iron-containing dehydrogenase families, aligning SIMBAL's top-scoring sequences to homologous solved crystal structures shows signals centered over NAD(P-binding sites rather than over substrate-binding or active site residues. Previous studies on some of these proteins have revealed a non-exchangeable NAD cofactor, such that enzymatic activity in vitro requires an artificial electron acceptor such

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.

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.

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.

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)

Spatial and vertical distribution of bacterial community in the northern South China Sea.

Science.gov (United States)

Sun, Fu-Lin; Wang, You-Shao; Wu, Mei-Lin; Sun, Cui-Ci; Cheng, Hao

2015-10-01

Microbial communities are highly diverse in coastal oceans and response rapidly with changing environments. Learning about this will help us understand the ecology of microbial populations in marine ecosystems. This study aimed to assess the spatial and vertical distributions of the bacterial community in the northern South China Sea. Multi-dimensional scaling analyses revealed structural differences of the bacterial community among sampling sites and vertical depth. Result also indicated that bacterial community in most sites had higher diversity in 0-75 m depths than those in 100-200 m depths. Bacterial community of samples was positively correlation with salinity and depth, whereas was negatively correlation with temperature. Proteobacteria and Cyanobacteria were the dominant groups, which accounted for the majority of sequences. The α-Proteobacteria was highly diverse, and sequences belonged to Rhodobacterales bacteria were dominant in all characterized sequences. The current data indicate that the Rhodobacterales bacteria, especially Roseobacter clade are the diverse group in the tropical waters.

Temporal distribution of bacterial community structure in the Changjiang Estuary hypoxia area and the adjacent East China Sea

International Nuclear Information System (INIS)

Liu Min; Huang Huiqin; Bao Shixiang; Xiao Tian; Zhang Wuchang; Wu Ying; Zhou Feng

2012-01-01

Bacterial community structure and the effects of environmental factors on the microbial community distribution were investigated in the Changjiang Estuary hypoxia area and its adjacent area in the East China Sea (ECS) in June, August and October, 2006. Profiles of bacterial communities were generated by denaturing gradient gel electrophoresis (DGGE) of 16S rRNA genes followed by DNA sequence analysis. The dominant bacterial groups were affiliated to Gammaproteobacteria, Cytophaga–Flavobacteria–Bacteroides (CFB), Deltaproteobacteria, Cyanobacteria and Firmicutes, which were mostly from the marine seawater ecosystem. Effects of environmental factors on the bacterial community distribution were analyzed by the ordination technique of canonical correspondence analysis (CCA). The environmental factors significantly influencing bacterial community structure were different in the three months; dissolved organic carbon (DOC) and temperature in June and nitrite in August. No environmental variables displayed significant influence on the bacterial community at the 5% level in October. The seasonal environmental heterogeneity in the Changjiang Estuary and the adjacent ECS, such as seasonal hydrodynamic conditions and riverine input of nutrients, might be the reason for the difference in the key environmental factors determining the bacterial community in the three months. (letter)

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.

The actin homologue MreB organizes the bacterial cell membrane

NARCIS (Netherlands)

Strahl, H.; Burmann, F.; Hamoen, L.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

A systematic approach for the assessment of bacterial growth-controlling factors linked to biological stability of drinking water in distribution systems

KAUST Repository

Prest, E. I.; Hammes, F.; Kotzsch, S.; van Loosdrecht, M. C. M.; Vrouwenvelder, Johannes S.

2016-01-01

A systematic approach is presented for the assessment of (i) bacterial growth-controlling factors in drinking water and (ii) the impact of distribution conditions on the extent of bacterial growth in full-scale distribution systems. The approach

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.

Allowance for effects of thermodynamic nonideality in sedimentation equilibrium distributions reflecting protein dimerization.

Science.gov (United States)

Wills, Peter R; Scott, David J; Winzor, Donald J

2012-03-01

This reexamination of a high-speed sedimentation equilibrium distribution for α-chymotrypsin under slightly acidic conditions (pH 4.1, I(M) 0.05) has provided experimental support for the adequacy of nearest-neighbor considerations in the allowance for effects of thermodynamic nonideality in the characterization of protein self-association over a moderate concentration range (up to 8 mg/mL). A widely held but previously untested notion about allowance for thermodynamic nonideality effects is thereby verified experimentally. However, it has also been shown that a greater obstacle to better characterization of protein self-association is likely to be the lack of a reliable estimate of monomer net charge, a parameter that has a far more profound effect on the magnitude of the measured equilibrium constant than any deficiency in current procedures for incorporating the effects of thermodynamic nonideality into the analysis of sedimentation equilibrium distributions reflecting reversible protein self-association. Copyright © 2011 Elsevier Inc. All rights reserved.

Protein kinase A regulatory subunit distribution in medulloblastoma

International Nuclear Information System (INIS)

Mucignat-Caretta, Carla; Denaro, Luca; Redaelli, Marco; D'Avella, Domenico; Caretta, Antonio

2010-01-01

Previous studies showed a differential distribution of the four regulatory subunits of cAMP-dependent protein kinases inside the brain, that changed in rodent gliomas: therefore, the distribution of these proteins inside the brain can give information on the functional state of the cells. Our goal was to examine human brain tumors to provide evidence for a differential distribution of protein kinase A in different tumors. The distribution of detergent insoluble regulatory (R1 and R2) and catalytic subunits of cAMP dependent kinases was examined in pediatric brain tumors by immunohistochemistry and fluorescent cAMP analogues binding. R2 is organized in large single dots in medulloblastomas, while it has a different appearance in other tumors. Fluorescent cAMP labelling was observed only in medulloblastoma. A different distribution of cAMP dependent protein kinases has been observed in medulloblastoma

Distributions of Bacterial Generalists among the Guts of Birds, Fish, and Mammals - abstract

Science.gov (United States)

Complex distributions of bacterial taxa within diverse animal microbiomes have inspired ecological and biogeographical approaches to revealing the functions of taxa that may be most important for host health. Of particular interest are bacteria that find many diverse habitats sui...

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...

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.

Recombinant Expression Screening of P. aeruginosa Bacterial Inner Membrane Proteins

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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

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

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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.

Bacterial community radial-spatial distribution in biofilms along pipe wall in chlorinated drinking water distribution system of East China.

Science.gov (United States)

Liu, Jingqing; Ren, Hongxing; Ye, Xianbei; Wang, Wei; Liu, Yan; Lou, Liping; Cheng, Dongqing; He, Xiaofang; Zhou, Xiaoyan; Qiu, Shangde; Fu, Liusong; Hu, Baolan

2017-01-01

Biofilms in the pipe wall may lead to water quality deterioration and biological instability in drinking water distribution systems (DWDSs). In this study, bacterial community radial-spatial distribution in biofilms along the pipe wall in a chlorinated DWDS of East China was investigated. Three pipes of large diameter (300, 600, and 600 mm) were sampled in this DWDS, including a ductile cast iron pipe (DCIP) with pipe age of 11 years and two gray cast iron pipes (GCIP) with pipe ages of 17 and 19 years, and biofilms in the upper, middle, and lower parts of each pipe wall were collected. Real-time quantitative polymerase chain reaction (qPCR) and culture-based method were used to quantify bacteria. 454 pyrosequencing was used for bacterial community analysis. The results showed that the biofilm density and total solid (TS) and volatile solid (VS) contents increased gradually from the top to the bottom along the pipe wall. Microorganisms were concentrated in the upper and lower parts of the pipe wall, together accounting for more than 80 % of the total biomass in the biofilms. The bacterial communities in biofilms were significantly different in different areas of the pipe wall and had no strong interaction. Compared with the upper and lower parts of the pipe wall, the bacterial community in the middle of the pipe wall was distributed evenly and had the highest diversity. The 16S rRNA genes of various possible pathogens, including Escherichia coli, Staphylococcus epidermidis, Pseudomonas aeruginosa, and Salmonella enterica, were detected in the biofilms, and the abundances of these possible pathogens were highest in the middle of the pipe wall among three areas. The detachment of the biofilms is the main reason for the deterioration of the water quality in DWDSs. The results of this study suggest that the biofilms in the middle of the pipe wall have highly potential risk for drinking water safety, which provides new ideas for the study of the microbial ecology in

Metagenomes Reveal Global Distribution of Bacterial Steroid Catabolism in Natural, Engineered, and Host Environments

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Johannes Holert

2018-01-01

Full Text Available Steroids are abundant growth substrates for bacteria in natural, engineered, and host-associated environments. This study analyzed the distribution of the aerobic 9,10-seco steroid degradation pathway in 346 publically available metagenomes from diverse environments. Our results show that steroid-degrading bacteria are globally distributed and prevalent in particular environments, such as wastewater treatment plants, soil, plant rhizospheres, and the marine environment, including marine sponges. Genomic signature-based sequence binning recovered 45 metagenome-assembled genomes containing a majority of 9,10-seco pathway genes. Only Actinobacteria and Proteobacteria were identified as steroid degraders, but we identified several alpha- and gammaproteobacterial lineages not previously known to degrade steroids. Actino- and proteobacterial steroid degraders coexisted in wastewater, while soil and rhizosphere samples contained mostly actinobacterial ones. Actinobacterial steroid degraders were found in deep ocean samples, while mostly alpha- and gammaproteobacterial ones were found in other marine samples, including sponges. Isolation of steroid-degrading bacteria from sponges confirmed their presence. Phylogenetic analysis of key steroid degradation proteins suggested their biochemical novelty in genomes from sponges and other environments. This study shows that the ecological significance as well as taxonomic and biochemical diversity of bacterial steroid degradation has so far been largely underestimated, especially in the marine environment.

Bacterial communities in the collection and chlorinated distribution sections of a drinking water system in Budapest, Hungary.

Science.gov (United States)

Homonnay, Zalán G; Török, György; Makk, Judit; Brumbauer, Anikó; Major, Eva; Márialigeti, Károly; Tóth, Erika

2014-07-01

Bacterial communities of a bank-filtered drinking water system were investigated by aerobic cultivation and clone library analysis. Moreover, bacterial communities were compared using sequence-aided terminal restriction fragment length polymorphism (T-RFLP) fingerprinting at ten characteristic points located at both the collecting and the distributing part of the water supply system. Chemical characteristics of the samples were similar, except for the presence of chlorine residuals in the distribution system and increased total iron concentration in two of the samples. Assimilable organic carbon (AOC) concentration increased within the collection system, it was reduced by chlorination and it increased again in the distribution system. Neither fecal indicators nor pathogens were detected by standard cultivation techniques. Chlorination reduced bacterial diversity and heterotrophic plate counts. Community structures were found to be significantly different before and after chlorination: the diverse communities in wells and the collection system were dominated by chemolithotrophic (e.g., Gallionella and Nitrospira) and oligocarbophilic-heterotrophic bacteria (e.g., Sphingomonas, Sphingopyxis, and Bradyrhizobium). After chlorination in the distribution system, the most characteristic bacterium was related to the facultative methylotrophic Methylocella spp. Communities changed within the distribution system too, Mycobacterium spp. or Sphingopyxis spp. became predominant in certain samples. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Distribution and Evolution of Yersinia Leucine-Rich Repeat Proteins

Science.gov (United States)

Hu, Yueming; Huang, He; Hui, Xinjie; Cheng, Xi; White, Aaron P.

2016-01-01

Leucine-rich repeat (LRR) proteins are widely distributed in bacteria, playing important roles in various protein-protein interaction processes. In Yersinia, the well-characterized type III secreted effector YopM also belongs to the LRR protein family and is encoded by virulence plasmids. However, little has been known about other LRR members encoded by Yersinia genomes or their evolution. In this study, the Yersinia LRR proteins were comprehensively screened, categorized, and compared. The LRR proteins encoded by chromosomes (LRR1 proteins) appeared to be more similar to each other and different from those encoded by plasmids (LRR2 proteins) with regard to repeat-unit length, amino acid composition profile, and gene expression regulation circuits. LRR1 proteins were also different from LRR2 proteins in that the LRR1 proteins contained an E3 ligase domain (NEL domain) in the C-terminal region or an NEL domain-encoding nucleotide relic in flanking genomic sequences. The LRR1 protein-encoding genes (LRR1 genes) varied dramatically and were categorized into 4 subgroups (a to d), with the LRR1a to -c genes evolving from the same ancestor and LRR1d genes evolving from another ancestor. The consensus and ancestor repeat-unit sequences were inferred for different LRR1 protein subgroups by use of a maximum parsimony modeling strategy. Structural modeling disclosed very similar repeat-unit structures between LRR1 and LRR2 proteins despite the different unit lengths and amino acid compositions. Structural constraints may serve as the driving force to explain the observed mutations in the LRR regions. This study suggests that there may be functional variation and lays the foundation for future experiments investigating the functions of the chromosomally encoded LRR proteins of Yersinia. PMID:27217422

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.

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.

An operon for production of bioactive gibberellin A4 phytohormone with wide distribution in the bacterial rice leaf streak pathogen Xanthomonas oryzae pv. oryzicola.

Science.gov (United States)

Nagel, Raimund; Turrini, Paula C G; Nett, Ryan S; Leach, Jan E; Verdier, Valérie; Van Sluys, Marie-Anne; Peters, Reuben J

2017-05-01

Phytopathogens have developed elaborate mechanisms to attenuate the defense response of their host plants, including convergent evolution of complex pathways for production of the GA phytohormones, which were actually first isolated from the rice fungal pathogen Gibberella fujikuroi. The rice bacterial pathogen Xanthomonas oryzae pv. oryzicola (Xoc) has been demonstrated to contain a biosynthetic operon with cyclases capable of producing the universal GA precursor ent-kaurene. Genetic (knock-out) studies indicate that the derived diterpenoid serves as a virulence factor for this rice leaf streak pathogen, serving to reduce the jasmonic acid-mediated defense response. Here the functions of the remaining genes in the Xoc operon are elucidated and the distribution of the operon in X. oryzae is investigated in over 100 isolates. The Xoc operon leads to production of the bioactive GA 4 , an additional step beyond production of the penultimate precursor GA 9 mediated by the homologous operons recently characterized from rhizobia. Moreover, this GA biosynthetic operon was found to be widespread in Xoc (> 90%), but absent in the other major X. oryzae pathovar. These results indicate selective pressure for production of GA 4 in the distinct lifestyle of Xoc, and the importance of GA to both fungal and bacterial pathogens of rice. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

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

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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.

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

Distribution Patterns of Polyphosphate Metabolism Pathway and Its Relationships With Bacterial Durability and Virulence

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Liang Wang

2018-04-01

Full Text Available Inorganic polyphosphate (polyP is a linear polymer of orthophosphate residues. It is reported to be present in all life forms. Experimental studies showed that polyP plays important roles in bacterial durability and virulence. Here we investigated the relationships of polyP with bacterial durability and virulence theoretically. Bacterial lifestyle, environmental persistence, virulence factors (VFs, and species evolution are all included in the analysis. The presence of seven genes involved in polyP metabolism (ppk1, ppk2, pap, surE, gppA, ppnK, and ppgK and 2595 core VFs were verified in 944 bacterial reference proteomes for distribution patterns via HMMER. Proteome size and VFs were compared in terms of gain and loss of polyP pathway. Literature mining and phylogenetic analysis were recruited to support the study. Our analyzes revealed that the presence of polyP metabolism is positively correlated with bacterial proteome size and the number of virulence genes. A potential relationship of polyP in bacterial lifestyle and environmental durability is suggested. Evolutionary analysis shows that polyP genes are randomly lost along the phylogenetic tree. In sum, based on our theoretical analysis, we confirmed that bacteria with polyP metabolism are associated with high environmental durability and more VFs.

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

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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.

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

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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.

Detecting Distributed Network Traffic Anomaly with Network-Wide Correlation Analysis

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Yang Dan

2008-12-01

Full Text Available Distributed network traffic anomaly refers to a traffic abnormal behavior involving many links of a network and caused by the same source (e.g., DDoS attack, worm propagation. The anomaly transiting in a single link might be unnoticeable and hard to detect, while the anomalous aggregation from many links can be prevailing, and does more harm to the networks. Aiming at the similar features of distributed traffic anomaly on many links, this paper proposes a network-wide detection method by performing anomalous correlation analysis of traffic signals' instantaneous parameters. In our method, traffic signals' instantaneous parameters are firstly computed, and their network-wide anomalous space is then extracted via traffic prediction. Finally, an anomaly is detected by a global correlation coefficient of anomalous space. Our evaluation using Abilene traffic traces demonstrates the excellent performance of this approach for distributed traffic anomaly detection.

Distribution of 10 periodontal bacterial species in children and adolescents over a 7-year period.

Science.gov (United States)

Nakano, K; Miyamoto, E; Tamura, K; Nemoto, H; Fujita, K; Nomura, R; Ooshima, T

2008-10-01

There is scant information available regarding the distribution of periodontal bacterial species in children and adolescents over an extended period. The purpose of this study was to compare bacterial profiles in the same individuals over a period of 7 years. Twenty-six children and adolescents from whom dental plaque and saliva specimens were obtained during both the first (1999-2000) and second (2006-2007) periods, were analyzed. Bacterial DNA was extracted from each specimen and the presence of 10 periodontal bacterial species was determined using a PCR method, with a focus on the red complex species of Porphyromonas gingivalis, Treponema denticola, and Tannerella forsythia. Subjects with red complex species in saliva specimens obtained during the second collection possessed a significantly higher number of total bacterial species than those without. The detection rate of the red complex species in the second collection period samples was significantly greater in subjects who had two or more species detected in samples taken during the first collection compared with the other subjects. Subjects possessing red complex species may be at possible risk for infection with a high number of periodontal bacterial species during adolescent and younger adult years.

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

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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

The Nucleoid Binding Protein H-NS Biases Genome-Wide Transposon Insertion Landscapes

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Satoshi Kimura

2016-08-01

Full Text Available Transposon insertion sequencing (TIS; also known as TnSeq is a potent approach commonly used to comprehensively define the genetic loci that contribute to bacterial fitness in diverse environments. A key presumption underlying analyses of TIS datasets is that loci with a low frequency of transposon insertions contribute to fitness. However, it is not known whether factors such as nucleoid binding proteins can alter the frequency of transposon insertion and thus whether TIS output may systematically reflect factors that are independent of the role of the loci in fitness. Here, we investigated whether the histone-like nucleoid structuring (H-NS protein, which preferentially associates with AT-rich sequences, modulates the frequency of Mariner transposon insertion in the Vibrio cholerae genome, using comparative analysis of TIS results from wild-type (wt and Δhns V. cholerae strains. These analyses were overlaid on gene classification based on GC content as well as on extant genome-wide identification of H-NS binding loci. Our analyses revealed a significant dearth of insertions within AT-rich loci in wt V. cholerae that was not apparent in the Δhns insertion library. Additionally, we observed a striking correlation between genetic loci that are overrepresented in the Δhns insertion library relative to their insertion frequency in wt V. cholerae and loci previously found to physically interact with H-NS. Collectively, our findings reveal that factors other than genetic fitness can systematically modulate the frequency of transposon insertions in TIS studies and add a cautionary note to interpretation of TIS data, particularly for AT-rich sequences.

Thermal proximity coaggregation for system-wide profiling of protein complex dynamics in cells.

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Tan, Chris Soon Heng; Go, Ka Diam; Bisteau, Xavier; Dai, Lingyun; Yong, Chern Han; Prabhu, Nayana; Ozturk, Mert Burak; Lim, Yan Ting; Sreekumar, Lekshmy; Lengqvist, Johan; Tergaonkar, Vinay; Kaldis, Philipp; Sobota, Radoslaw M; Nordlund, Pär

2018-03-09

Proteins differentially interact with each other across cellular states and conditions, but an efficient proteome-wide strategy to monitor them is lacking. We report the application of thermal proximity coaggregation (TPCA) for high-throughput intracellular monitoring of protein complex dynamics. Significant TPCA signatures observed among well-validated protein-protein interactions correlate positively with interaction stoichiometry and are statistically observable in more than 350 annotated human protein complexes. Using TPCA, we identified many complexes without detectable differential protein expression, including chromatin-associated complexes, modulated in S phase of the cell cycle. Comparison of six cell lines by TPCA revealed cell-specific interactions even in fundamental cellular processes. TPCA constitutes an approach for system-wide studies of protein complexes in nonengineered cells and tissues and might be used to identify protein complexes that are modulated in diseases. Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

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

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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.

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

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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.

[Distribution and spatial ordering of biopolymer molecules in resting bacterial spores].

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Duda, V I; Korolev, Iu N; El'-Registan, G I; Duzha, M V; Telegin, N L

1978-01-01

The presence, distribution and spatial arrangement of biopolymers in situ were studied in both a total intact spore and in a certain cellular layer using a spectroscopic technique of attenuated total refraction (ATR-IR) in the IR region. In contrast to vegetative cells, intact spores were characterized by isotropic distribution of protein components. This feature can be regarded as an index of the cryptobiotic state of spores. However, the distribution of protein components among individual layers of a spore was anisotropic. Bonds characterized by amide I and amide II bands were most often ordered in a layer which comprised cellular structures from the exosporium to the inner spore membrane.

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

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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.

Plant ice-binding (antifreeze) proteins

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Proteins that determine the temperature at which ice crystals will form in water-based solutions in cells and tissues, that bind to growing ice crystals, thus affecting their size, and that impact ice re-crystallization have been widely-documented and studied in many plant, bacterial, fungal, insect...

Biofilm bacterial communities in urban drinking water distribution systems transporting waters with different purification strategies.

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Wu, Huiting; Zhang, Jingxu; Mi, Zilong; Xie, Shuguang; Chen, Chao; Zhang, Xiaojian

2015-02-01

Biofilm formation in drinking water distribution systems (DWDS) has many adverse consequences. Knowledge of microbial community structure of DWDS biofilm can aid in the design of an effective control strategy. However, biofilm bacterial community in real DWDS and the impact of drinking water purification strategy remain unclear. The present study investigated the composition and diversity of biofilm bacterial community in real DWDSs transporting waters with different purification strategies (conventional treatment and integrated treatment). High-throughput Illumina MiSeq sequencing analysis illustrated a large shift in the diversity and structure of biofilm bacterial community in real DWDS. Proteobacteria, Firmicutes, Bacteroidetes, Actinobacteria, Nitrospirae, and Cyanobacteria were the major components of biofilm bacterial community. Proteobacteria (mainly Alphaproteobacteria, Betaproteobacteria, and Gammaproteobacteria) predominated in each DWDS biofilm, but the compositions of the dominant proteobacterial classes and genera and their proportions varied among biofilm samples. Drinking water purification strategy could shape DWDS biofilm bacterial community. Moreover, Pearson's correlation analysis indicated that Actinobacteria was positively correlated with the levels of total alkalinity and dissolved organic carbon in tap water, while Firmicutes had a significant positive correlation with nitrite nitrogen.

Dynamics of bacterial communities before and after distribution in a full-scale drinking water network

KAUST Repository

El Chakhtoura, Joline; Prest, Emmanuelle I E C; Saikaly, Pascal; van Loosdrecht, Mark C.M.; Hammes, Frederik A.; Vrouwenvelder, Johannes S.

2015-01-01

Understanding the biological stability of drinking water distribution systems is imperative in the framework of process control and risk management. The objective of this research was to examine the dynamics of the bacterial community during

Identification of Fic-1 as an enzyme that inhibits bacterial DNA replication by AMPylating GyrB, promoting filament formation.

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Lu, Canhua; Nakayasu, Ernesto S; Zhang, Li-Qun; Luo, Zhao-Qing

2016-01-26

The morphology of bacterial cells is important for virulence, evasion of the host immune system, and coping with environmental stresses. The widely distributed Fic proteins (filamentation induced by cAMP) are annotated as proteins involved in cell division because of the presence of the HPFx[D/E]GN[G/K]R motif. We showed that the presence of Fic-1 from Pseudomonas fluorescens significantly reduced the yield of plasmid DNA when expressed in Escherichia coli or P. fluorescens. Fic-1 interacted with GyrB, a subunit of DNA gyrase, which is essential for bacterial DNA replication. Fic-1 catalyzed the AMPylation of GyrB at Tyr(109), a residue critical for binding ATP, and exhibited auto-AMPylation activity. Mutation of the Fic-1 auto-AMPylated site greatly reduced AMPylation activity toward itself and toward GyrB. Fic-1-dependent AMPylation of GyrB triggered the SOS response, indicative of DNA replication stress or DNA damage. Fic-1 also promoted the formation of elongated cells when the SOS response was blocked. We identified an α-inhibitor protein that we named anti-Fic-1 (AntF), encoded by a gene immediately upstream of Fic-1. AntF interacted with Fic-1, inhibited the AMPylation activity of Fic-1 for GyrB in vitro, and blocked Fic-1-mediated inhibition of DNA replication in bacteria, suggesting that Fic-1 and AntF comprise a toxin-antitoxin module. Our work establishes Fic-1 as an AMPylating enzyme that targets GyrB to inhibit DNA replication and may target other proteins to regulate bacterial morphology. Copyright © 2016, American Association for the Advancement of Science.

Spatial Distribution of Bacterial Communities Driven by Multiple Environmental Factors in a Beach Wetland of the Largest Freshwater Lake in China

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Xia eDing

2015-02-01

Full Text Available The spatial distributions of bacterial communities may be driven by multiple environmental factors. Thus, understanding the relationships between bacterial distribution and environmental factors is critical for understanding wetland stability and the functioning of freshwater lakes. However, little research on the bacterial communities in deep sediment layers exists. In this study, thirty clone libraries of 16S rRNA were constructed from a beach wetland of the Poyang Lake along both horizontal (distance to the water-land junction and vertical (sediment depth gradients to assess the effects of sediment properties on bacterial community structure and diversity. Our results showed that bacterial diversity increased along the horizontal gradient and decreased along the vertical gradient. The heterogeneous sediment properties along gradients substantially affected the dominant bacterial groups at the phylum and species levels. For example, the NH4+ concentration decreased with increasing depth, which was positively correlated with the relative abundance of Alphaproteobacteria. The changes in bacterial diversity and dominant bacterial groups showed that the top layer had a different bacterial community structure than the deeper layers. Principal component analysis revealed that both gradients, not each gradient independently, contributed to the shift in the bacterial community structure. A multiple linear regression model explained the changes in bacterial diversity and richness along the depth and distance gradients. Overall, our results suggest that spatial gradients associated with sediment properties shaped the bacterial communities in the Poyang Lake beach wetland.

Modulating nanoparticle superlattice structure using proteins with tunable bond distributions

International Nuclear Information System (INIS)

McMillan, Janet R.; Brodin, Jeffrey D.; Millan, Jaime A.; Lee, Byeongdu; Olvera de la Cruz, Monica; Mirkin, Chad A.

2017-01-01

Here, we investigate the use of proteins with tunable DNA modification distributions to modulate nanoparticle superlattice structure. Using Beta-galactosidase (βgal) as a model system, we have employed the orthogonal chemical reactivities of surface amines and thiols to synthesize protein-DNA conjugates with 36 evenly distributed or 8 specifically positioned oligonucleotides. When assembled into crystalline superlattices with AuNPs, we find that the distribution of DNA modifications modulates the favored structure: βgal with uniformly distributed DNA bonding elements results in body-centered cubic crystals, whereas DNA functionalization of cysteines results in AB 2 packing. We probe the role of protein oligonucleotide number and conjugate size on this observation, which revealed the importance of oligonucleotide distribution and number in this observed assembly behavior. These results indicate that proteins with defined DNA-modification patterns are powerful tools to control the nanoparticle superlattices architecture, and establish the importance of oligonucleotide distribution in the assembly behavior of protein-DNA conjugates.

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

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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

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.

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

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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.

Genome-wide analysis of protein-protein interactions and involvement of viral proteins in SARS-CoV replication.

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Ji'an Pan

Full Text Available Analyses of viral protein-protein interactions are an important step to understand viral protein functions and their underlying molecular mechanisms. In this study, we adopted a mammalian two-hybrid system to screen the genome-wide intraviral protein-protein interactions of SARS coronavirus (SARS-CoV and therefrom revealed a number of novel interactions which could be partly confirmed by in vitro biochemical assays. Three pairs of the interactions identified were detected in both directions: non-structural protein (nsp 10 and nsp14, nsp10 and nsp16, and nsp7 and nsp8. The interactions between the multifunctional nsp10 and nsp14 or nsp16, which are the unique proteins found in the members of Nidovirales with large RNA genomes including coronaviruses and toroviruses, may have important implication for the mechanisms of replication/transcription complex assembly and functions of these viruses. Using a SARS-CoV replicon expressing a luciferase reporter under the control of a transcription regulating sequence, it has been shown that several viral proteins (N, X and SUD domains of nsp3, and nsp12 provided in trans stimulated the replicon reporter activity, indicating that these proteins may regulate coronavirus replication and transcription. Collectively, our findings provide a basis and platform for further characterization of the functions and mechanisms of coronavirus proteins.

Wide-area-distributed storage system for a multimedia database

Science.gov (United States)

Ueno, Masahiro; Kinoshita, Shigechika; Kuriki, Makato; Murata, Setsuko; Iwatsu, Shigetaro

1998-12-01

We have developed a wide-area-distribution storage system for multimedia databases, which minimizes the possibility of simultaneous failure of multiple disks in the event of a major disaster. It features a RAID system, whose member disks are spatially distributed over a wide area. Each node has a device, which includes the controller of the RAID and the controller of the member disks controlled by other nodes. The devices in the node are connected to a computer, using fiber optic cables and communicate using fiber-channel technology. Any computer at a node can utilize multiple devices connected by optical fibers as a single 'virtual disk.' The advantage of this system structure is that devices and fiber optic cables are shared by the computers. In this report, we first described our proposed system, and a prototype was used for testing. We then discussed its performance; i.e., how to read and write throughputs are affected by data-access delay, the RAID level, and queuing.

Elucidating the impact of micro-scale heterogeneous bacterial distribution on biodegradation

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Schmidt, Susanne I.; Kreft, Jan-Ulrich; Mackay, Rae; Picioreanu, Cristian; Thullner, Martin

2018-06-01

Groundwater microorganisms hardly ever cover the solid matrix uniformly-instead they form micro-scale colonies. To which extent such colony formation limits the bioavailability and biodegradation of a substrate is poorly understood. We used a high-resolution numerical model of a single pore channel inhabited by bacterial colonies to simulate the transport and biodegradation of organic substrates. These high-resolution 2D simulation results were compared to 1D simulations that were based on effective rate laws for bioavailability-limited biodegradation. We (i) quantified the observed bioavailability limitations and (ii) evaluated the applicability of previously established effective rate concepts if microorganisms are heterogeneously distributed. Effective bioavailability reductions of up to more than one order of magnitude were observed, showing that the micro-scale aggregation of bacterial cells into colonies can severely restrict the bioavailability of a substrate and reduce in situ degradation rates. Effective rate laws proved applicable for upscaling when using the introduced effective colony sizes.

Characterization of replication and conjugation of plasmid pWTY27 from a widely distributed Streptomyces species

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Wang Tao

2012-11-01

Full Text Available Abstract Background Streptomyces species are widely distributed in natural habitats, such as soils, lakes, plants and some extreme environments. Replication loci of several Streptomyces theta-type plasmids have been reported, but are not characterized in details. Conjugation loci of some Streptomyces rolling-circle-type plasmids are identified and mechanism of conjugal transferring are described. Results We report the detection of a widely distributed Streptomyces strain Y27 and its indigenous plasmid pWTY27 from fourteen plants and four soil samples cross China by both culturing and nonculturing methods. The complete nucleotide sequence of pWTY27 consisted of 14,288 bp. A basic locus for plasmid replication comprised repAB genes and an adjacent iteron sequence, to a long inverted-repeat (ca. 105 bp of which the RepA protein bound specifically in vitro, suggesting that RepA may recognize a second structure (e.g. a long stem-loop of the iteron DNA. A plasmid containing the locus propagated in linear mode when the telomeres of a linear plasmid were attached, indicating a bi-directional replication mode for pWTY27. As for rolling-circle plasmids, a single traA gene and a clt sequence (covering 16 bp within traA and its adjacent 159 bp on pWTY27 were required for plasmid transfer. TraA recognized and bound specifically to the two regions of the clt sequence, one containing all the four DC1 of 7 bp (TGACACC and one DC2 (CCCGCCC and most of IC1, and another covering two DC2 and part of IC1, suggesting formation of a high-ordered DNA-protein complex. Conclusions This work (i isolates a widespread Streptomyces strain Y27 and sequences its indigenous theta-type plasmid pWTY27; (ii identifies the replication and conjugation loci of pWTY27 and; (iii characterizes the binding sequences of the RepA and TraA proteins.

Characterization of replication and conjugation of plasmid pWTY27 from a widely distributed Streptomyces species

Science.gov (United States)

2012-01-01

Background Streptomyces species are widely distributed in natural habitats, such as soils, lakes, plants and some extreme environments. Replication loci of several Streptomyces theta-type plasmids have been reported, but are not characterized in details. Conjugation loci of some Streptomyces rolling-circle-type plasmids are identified and mechanism of conjugal transferring are described. Results We report the detection of a widely distributed Streptomyces strain Y27 and its indigenous plasmid pWTY27 from fourteen plants and four soil samples cross China by both culturing and nonculturing methods. The complete nucleotide sequence of pWTY27 consisted of 14,288 bp. A basic locus for plasmid replication comprised repAB genes and an adjacent iteron sequence, to a long inverted-repeat (ca. 105 bp) of which the RepA protein bound specifically in vitro, suggesting that RepA may recognize a second structure (e.g. a long stem-loop) of the iteron DNA. A plasmid containing the locus propagated in linear mode when the telomeres of a linear plasmid were attached, indicating a bi-directional replication mode for pWTY27. As for rolling-circle plasmids, a single traA gene and a clt sequence (covering 16 bp within traA and its adjacent 159 bp) on pWTY27 were required for plasmid transfer. TraA recognized and bound specifically to the two regions of the clt sequence, one containing all the four DC1 of 7 bp (TGACACC) and one DC2 (CCCGCCC) and most of IC1, and another covering two DC2 and part of IC1, suggesting formation of a high-ordered DNA-protein complex. Conclusions This work (i) isolates a widespread Streptomyces strain Y27 and sequences its indigenous theta-type plasmid pWTY27; (ii) identifies the replication and conjugation loci of pWTY27 and; (iii) characterizes the binding sequences of the RepA and TraA proteins. PMID:23134842

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.

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.

A species-specific nucleosomal signature defines a periodic distribution of amino acids in proteins.

Science.gov (United States)

Quintales, Luis; Soriano, Ignacio; Vázquez, Enrique; Segurado, Mónica; Antequera, Francisco

2015-04-01

Nucleosomes are the basic structural units of chromatin. Most of the yeast genome is organized in a pattern of positioned nucleosomes that is stably maintained under a wide range of physiological conditions. In this work, we have searched for sequence determinants associated with positioned nucleosomes in four species of fission and budding yeasts. We show that mononucleosomal DNA follows a highly structured base composition pattern, which differs among species despite the high degree of histone conservation. These nucleosomal signatures are present in transcribed and non-transcribed regions across the genome. In the case of open reading frames, they correctly predict the relative distribution of codons on mononucleosomal DNA, and they also determine a periodicity in the average distribution of amino acids along the proteins. These results establish a direct and species-specific connection between the position of each codon around the histone octamer and protein composition.

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.

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.

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...

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.

BPhyOG: An interactive server for genome-wide inference of bacterial phylogenies based on overlapping genes

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Lin Kui

2007-07-01

Full Text Available Abstract Background Overlapping genes (OGs in bacterial genomes are pairs of adjacent genes of which the coding sequences overlap partly or entirely. With the rapid accumulation of sequence data, many OGs in bacterial genomes have now been identified. Indeed, these might prove a consistent feature across all microbial genomes. Our previous work suggests that OGs can be considered as robust markers at the whole genome level for the construction of phylogenies. An online, interactive web server for inferring phylogenies is needed for biologists to analyze phylogenetic relationships among a set of bacterial genomes of interest. Description BPhyOG is an online interactive server for reconstructing the phylogenies of completely sequenced bacterial genomes on the basis of their shared overlapping genes. It provides two tree-reconstruction methods: Neighbor Joining (NJ and Unweighted Pair-Group Method using Arithmetic averages (UPGMA. Users can apply the desired method to generate phylogenetic trees, which are based on an evolutionary distance matrix for the selected genomes. The distance between two genomes is defined by the normalized number of their shared OG pairs. BPhyOG also allows users to browse the OGs that were used to infer the phylogenetic relationships. It provides detailed annotation for each OG pair and the features of the component genes through hyperlinks. Users can also retrieve each of the homologous OG pairs that have been determined among 177 genomes. It is a useful tool for analyzing the tree of life and overlapping genes from a genomic standpoint. Conclusion BPhyOG is a useful interactive web server for genome-wide inference of any potential evolutionary relationship among the genomes selected by users. It currently includes 177 completely sequenced bacterial genomes containing 79,855 OG pairs, the annotation and homologous OG pairs of which are integrated comprehensively. The reliability of phylogenies complemented by

In Silico Phylogenetic Analysis and Molecular Modelling Study of 2-Haloalkanoic Acid Dehalogenase Enzymes from Bacterial and Fungal Origin

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Raghunath Satpathy

2016-01-01

Full Text Available 2-Haloalkanoic acid dehalogenase enzymes have broad range of applications, starting from bioremediation to chemical synthesis of useful compounds that are widely distributed in fungi and bacteria. In the present study, a total of 81 full-length protein sequences of 2-haloalkanoic acid dehalogenase from bacteria and fungi were retrieved from NCBI database. Sequence analysis such as multiple sequence alignment (MSA, conserved motif identification, computation of amino acid composition, and phylogenetic tree construction were performed on these primary sequences. From MSA analysis, it was observed that the sequences share conserved lysine (K and aspartate (D residues in them. Also, phylogenetic tree indicated a subcluster comprised of both fungal and bacterial species. Due to nonavailability of experimental 3D structure for fungal 2-haloalkanoic acid dehalogenase in the PDB, molecular modelling study was performed for both fungal and bacterial sources of enzymes present in the subcluster. Further structural analysis revealed a common evolutionary topology shared between both fungal and bacterial enzymes. Studies on the buried amino acids showed highly conserved Leu and Ser in the core, despite variation in their amino acid percentage. Additionally, a surface exposed tryptophan was conserved in all of these selected models.

Identification of prophages in bacterial genomes by dinucleotide relative abundance difference.

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K V Srividhya

Full Text Available BACKGROUND: Prophages are integrated viral forms in bacterial genomes that have been found to contribute to interstrain genetic variability. Many virulence-associated genes are reported to be prophage encoded. Present computational methods to detect prophages are either by identifying possible essential proteins such as integrases or by an extension of this technique, which involves identifying a region containing proteins similar to those occurring in prophages. These methods suffer due to the problem of low sequence similarity at the protein level, which suggests that a nucleotide based approach could be useful. METHODOLOGY: Earlier dinucleotide relative abundance (DRA have been used to identify regions, which deviate from the neighborhood areas, in genomes. We have used the difference in the dinucleotide relative abundance (DRAD between the bacterial and prophage DNA to aid location of DNA stretches that could be of prophage origin in bacterial genomes. Prophage sequences which deviate from bacterial regions in their dinucleotide frequencies are detected by scanning bacterial genome sequences. The method was validated using a subset of genomes with prophage data from literature reports. A web interface for prophage scan based on this method is available at http://bicmku.in:8082/prophagedb/dra.html. Two hundred bacterial genomes which do not have annotated prophages have been scanned for prophage regions using this method. CONCLUSIONS: The relative dinucleotide distribution difference helps detect prophage regions in genome sequences. The usefulness of this method is seen in the identification of 461 highly probable loci pertaining to prophages which have not been annotated so earlier. This work emphasizes the need to extend the efforts to detect and annotate prophage elements in genome sequences.

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.

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

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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.

Distributing Congestion Management System Information Using the World Wide Web

Science.gov (United States)

1997-01-01

The Internet is a unique medium for the distribution of information, and it provides a tremendous opportunity to take advantage of peoples innate interest in transportation issues as they relate to their own lives. In particular, the World Wide Web (...

Genome-wide Selective Sweeps in Natural Bacterial Populations Revealed by Time-series Metagenomics

Energy Technology Data Exchange (ETDEWEB)

Chan, Leong-Keat; Bendall, Matthew L.; Malfatti, Stephanie; Schwientek, Patrick; Tremblay, Julien; Schackwitz, Wendy; Martin, Joel; Pati, Amrita; Bushnell, Brian; Foster, Brian; Kang, Dongwan; Tringe, Susannah G.; Bertilsson, Stefan; Moran, Mary Ann; Shade, Ashley; Newton, Ryan J.; Stevens, Sarah; McMcahon, Katherine D.; Mamlstrom, Rex R.

2014-05-12

Multiple evolutionary models have been proposed to explain the formation of genetically and ecologically distinct bacterial groups. Time-series metagenomics enables direct observation of evolutionary processes in natural populations, and if applied over a sufficiently long time frame, this approach could capture events such as gene-specific or genome-wide selective sweeps. Direct observations of either process could help resolve how distinct groups form in natural microbial assemblages. Here, from a three-year metagenomic study of a freshwater lake, we explore changes in single nucleotide polymorphism (SNP) frequencies and patterns of gene gain and loss in populations of Chlorobiaceae and Methylophilaceae. SNP analyses revealed substantial genetic heterogeneity within these populations, although the degree of heterogeneity varied considerably among closely related, co-occurring Methylophilaceae populations. SNP allele frequencies, as well as the relative abundance of certain genes, changed dramatically over time in each population. Interestingly, SNP diversity was purged at nearly every genome position in one of the Chlorobiaceae populations over the course of three years, while at the same time multiple genes either swept through or were swept from this population. These patterns were consistent with a genome-wide selective sweep, a process predicted by the ecotype model? of diversification, but not previously observed in natural populations.

Genome-wide Selective Sweeps in Natural Bacterial Populations Revealed by Time-series Metagenomics

Energy Technology Data Exchange (ETDEWEB)

Chan, Leong-Keat; Bendall, Matthew L.; Malfatti, Stephanie; Schwientek, Patrick; Tremblay, Julien; Schackwitz, Wendy; Martin, Joel; Pati, Amrita; Bushnell, Brian; Foster, Brian; Kang, Dongwan; Tringe, Susannah G.; Bertilsson, Stefan; Moran, Mary Ann; Shade, Ashley; Newton, Ryan J.; Stevens, Sarah; McMahon, Katherine D.; Malmstrom, Rex R.

2014-06-18

Multiple evolutionary models have been proposed to explain the formation of genetically and ecologically distinct bacterial groups. Time-series metagenomics enables direct observation of evolutionary processes in natural populations, and if applied over a sufficiently long time frame, this approach could capture events such as gene-specific or genome-wide selective sweeps. Direct observations of either process could help resolve how distinct groups form in natural microbial assemblages. Here, from a three-year metagenomic study of a freshwater lake, we explore changes in single nucleotide polymorphism (SNP) frequencies and patterns of gene gain and loss in populations of Chlorobiaceae and Methylophilaceae. SNP analyses revealed substantial genetic heterogeneity within these populations, although the degree of heterogeneity varied considerably among closely related, co-occurring Methylophilaceae populations. SNP allele frequencies, as well as the relative abundance of certain genes, changed dramatically over time in each population. Interestingly, SNP diversity was purged at nearly every genome position in one of the Chlorobiaceae populations over the course of three years, while at the same time multiple genes either swept through or were swept from this population. These patterns were consistent with a genome-wide selective sweep, a process predicted by the ‘ecotype model’ of diversification, but not previously observed in natural populations.

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.

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.

Proteogenomics of rare taxonomic phyla: A prospective treasure trove of protein coding genes.

Science.gov (United States)

Kumar, Dhirendra; Mondal, Anupam Kumar; Kutum, Rintu; Dash, Debasis

2016-01-01

Sustainable innovations in sequencing technologies have resulted in a torrent of microbial genome sequencing projects. However, the prokaryotic genomes sequenced so far are unequally distributed along their phylogenetic tree; few phyla contain the majority, the rest only a few representatives. Accurate genome annotation lags far behind genome sequencing. While automated computational prediction, aided by comparative genomics, remains a popular choice for genome annotation, substantial fraction of these annotations are erroneous. Proteogenomics utilizes protein level experimental observations to annotate protein coding genes on a genome wide scale. Benefits of proteogenomics include discovery and correction of gene annotations regardless of their phylogenetic conservation. This not only allows detection of common, conserved proteins but also the discovery of protein products of rare genes that may be horizontally transferred or taxonomy specific. Chances of encountering such genes are more in rare phyla that comprise a small number of complete genome sequences. We collated all bacterial and archaeal proteogenomic studies carried out to date and reviewed them in the context of genome sequencing projects. Here, we present a comprehensive list of microbial proteogenomic studies, their taxonomic distribution, and also urge for targeted proteogenomics of underexplored taxa to build an extensive reference of protein coding genes. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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.

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

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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.

Dispersal networks for enhancing bacterial degradation in heterogeneous environments

International Nuclear Information System (INIS)

Banitz, Thomas; Wick, Lukas Y.; Fetzer, Ingo; Frank, Karin; Harms, Hauke; Johst, Karin

2011-01-01

Successful biodegradation of organic soil pollutants depends on their bioavailability to catabolically active microorganisms. In particular, environmental heterogeneities often limit bacterial access to pollutants. Experimental and modelling studies revealed that fungal networks can facilitate bacterial dispersal and may thereby improve pollutant bioavailability. Here, we investigate the influence of such bacterial dispersal networks on biodegradation performance under spatially heterogeneous abiotic conditions using a process-based simulation model. To match typical situations in polluted soils, two types of abiotic conditions are studied: heterogeneous bacterial dispersal conditions and heterogeneous initial resource distributions. The model predicts that networks facilitating bacterial dispersal can enhance biodegradation performance for a wide range of these conditions. Additionally, the time horizon over which this performance is assessed and the network's spatial configuration are key factors determining the degree of biodegradation improvement. Our results support the idea of stimulating the establishment of fungal mycelia for enhanced bioremediation of polluted soils. - Highlights: → Bacterial dispersal networks can considerably improve biodegradation performance. → They facilitate bacterial access to dispersal-limited areas and remote resources. → Abiotic conditions, time horizon and network structure govern the improvements. → Stimulating the establishment of fungal mycelia promises enhanced soil remediation. - Simulation modelling demonstrates that fungus-mediated bacterial dispersal can considerably improve the bioavailability of organic pollutants under spatially heterogeneous abiotic conditions typical for water-unsaturated soils.

Dispersal networks for enhancing bacterial degradation in heterogeneous environments

Energy Technology Data Exchange (ETDEWEB)

Banitz, Thomas, E-mail: thomas.banitz@ufz.de [Department of Ecological Modelling, UFZ - Helmholtz Centre for Environmental Research, Permoserstr. 15, 04318 Leipzig (Germany); Wick, Lukas Y.; Fetzer, Ingo [Department of Environmental Microbiology, UFZ - Helmholtz Centre for Environmental Research, Permoserstr. 15, 04318 Leipzig (Germany); Frank, Karin [Department of Ecological Modelling, UFZ - Helmholtz Centre for Environmental Research, Permoserstr. 15, 04318 Leipzig (Germany); Harms, Hauke [Department of Environmental Microbiology, UFZ - Helmholtz Centre for Environmental Research, Permoserstr. 15, 04318 Leipzig (Germany); Johst, Karin [Department of Ecological Modelling, UFZ - Helmholtz Centre for Environmental Research, Permoserstr. 15, 04318 Leipzig (Germany)

2011-10-15

Successful biodegradation of organic soil pollutants depends on their bioavailability to catabolically active microorganisms. In particular, environmental heterogeneities often limit bacterial access to pollutants. Experimental and modelling studies revealed that fungal networks can facilitate bacterial dispersal and may thereby improve pollutant bioavailability. Here, we investigate the influence of such bacterial dispersal networks on biodegradation performance under spatially heterogeneous abiotic conditions using a process-based simulation model. To match typical situations in polluted soils, two types of abiotic conditions are studied: heterogeneous bacterial dispersal conditions and heterogeneous initial resource distributions. The model predicts that networks facilitating bacterial dispersal can enhance biodegradation performance for a wide range of these conditions. Additionally, the time horizon over which this performance is assessed and the network's spatial configuration are key factors determining the degree of biodegradation improvement. Our results support the idea of stimulating the establishment of fungal mycelia for enhanced bioremediation of polluted soils. - Highlights: > Bacterial dispersal networks can considerably improve biodegradation performance. > They facilitate bacterial access to dispersal-limited areas and remote resources. > Abiotic conditions, time horizon and network structure govern the improvements. > Stimulating the establishment of fungal mycelia promises enhanced soil remediation. - Simulation modelling demonstrates that fungus-mediated bacterial dispersal can considerably improve the bioavailability of organic pollutants under spatially heterogeneous abiotic conditions typical for water-unsaturated soils.

A novel firmicute protein family related to the actinobacterial resuscitation-promoting factors by non-orthologous domain displacement

Directory of Open Access Journals (Sweden)

Finan Christopher L

2005-03-01

Full Text Available Abstract Background In Micrococcus luteus growth and resuscitation from starvation-induced dormancy is controlled by the production of a secreted growth factor. This autocrine resuscitation-promoting factor (Rpf is the founder member of a family of proteins found throughout and confined to the actinobacteria (high G + C Gram-positive bacteria. The aim of this work was to search for and characterise a cognate gene family in the firmicutes (low G + C Gram-positive bacteria and obtain information about how they may control bacterial growth and resuscitation. Results In silico analysis of the accessory domains of the Rpf proteins permitted their classification into several subfamilies. The RpfB subfamily is related to a group of firmicute proteins of unknown function, represented by YabE of Bacillus subtilis. The actinobacterial RpfB and firmicute YabE proteins have very similar domain structures and genomic contexts, except that in YabE, the actinobacterial Rpf domain is replaced by another domain, which we have called Sps. Although totally unrelated in both sequence and secondary structure, the Rpf and Sps domains fulfil the same function. We propose that these proteins have undergone "non-orthologous domain displacement", a phenomenon akin to "non-orthologous gene displacement" that has been described previously. Proteins containing the Sps domain are widely distributed throughout the firmicutes and they too fall into a number of distinct subfamilies. Comparative analysis of the accessory domains in the Rpf and Sps proteins, together with their weak similarity to lytic transglycosylases, provide clear evidence that they are muralytic enzymes. Conclusions The results indicate that the firmicute Sps proteins and the actinobacterial Rpf proteins are cognate and that they control bacterial culturability via enzymatic modification of the bacterial cell envelope.

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

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...

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

Local and Widely Distributed EEG Activity in Schizophrenia With Prevalence of Negative Symptoms.

Science.gov (United States)

Grin-Yatsenko, Vera A; Ponomarev, Valery A; Pronina, Marina V; Poliakov, Yury I; Plotnikova, Irina V; Kropotov, Juri D

2017-09-01

We evaluated EEG frequency abnormalities in resting state (eyes closed and eyes open) EEG in a group of chronic schizophrenia patients as compared with healthy subjects. The study included 3 methods of analysis of deviation of EEG characteristics: genuine EEG, current source density (CSD), and group independent component (gIC). All 3 methods have shown that the EEG in schizophrenia patients is characterized by enhanced low-frequency (delta and theta) and high-frequency (beta) activity in comparison with the control group. However, the spatial pattern of differences was dependent on the type of method used. Comparative analysis has shown that increased EEG power in schizophrenia patients apparently concerns both widely spatially distributed components and local components of signal. Furthermore, the observed differences in the delta and theta range can be described mainly by the local components, and those in the beta range mostly by spatially widely distributed ones. The possible nature of the widely distributed activity is discussed.

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.

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

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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

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

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.

Characterization and detection of a widely distributed gene cluster that predicts anaerobic choline utilization by human gut bacteria.

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Martínez-del Campo, Ana; Bodea, Smaranda; Hamer, Hilary A; Marks, Jonathan A; Haiser, Henry J; Turnbaugh, Peter J; Balskus, Emily P

2015-04-14

Elucidation of the molecular mechanisms underlying the human gut microbiota's effects on health and disease has been complicated by difficulties in linking metabolic functions associated with the gut community as a whole to individual microorganisms and activities. Anaerobic microbial choline metabolism, a disease-associated metabolic pathway, exemplifies this challenge, as the specific human gut microorganisms responsible for this transformation have not yet been clearly identified. In this study, we established the link between a bacterial gene cluster, the choline utilization (cut) cluster, and anaerobic choline metabolism in human gut isolates by combining transcriptional, biochemical, bioinformatic, and cultivation-based approaches. Quantitative reverse transcription-PCR analysis and in vitro biochemical characterization of two cut gene products linked the entire cluster to growth on choline and supported a model for this pathway. Analyses of sequenced bacterial genomes revealed that the cut cluster is present in many human gut bacteria, is predictive of choline utilization in sequenced isolates, and is widely but discontinuously distributed across multiple bacterial phyla. Given that bacterial phylogeny is a poor marker for choline utilization, we were prompted to develop a degenerate PCR-based method for detecting the key functional gene choline TMA-lyase (cutC) in genomic and metagenomic DNA. Using this tool, we found that new choline-metabolizing gut isolates universally possessed cutC. We also demonstrated that this gene is widespread in stool metagenomic data sets. Overall, this work represents a crucial step toward understanding anaerobic choline metabolism in the human gut microbiota and underscores the importance of examining this microbial community from a function-oriented perspective. Anaerobic choline utilization is a bacterial metabolic activity that occurs in the human gut and is linked to multiple diseases. While bacterial genes responsible for

Use of the [(14)C]leucine incorporation technique to measure bacterial production in river sediments and the epiphyton.

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Fischer, H; Pusch, M

1999-10-01

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 [(14)C]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 sediments, the biofilms on aquatic plants (epiphyton), and the pelagic zone, (iii) bacterial activities at different leucine concentrations, (iv) specificity of leucine uptake by bacteria, and (v) the effect of the incubation technique (perfused-core incubation versus slurry incubation) on leucine incorporation into protein. Bacterial protein was best extracted from sediments and precipitated by hot trichloroacetic acid treatment following ultrasonication. For epiphyton, an alkaline-extraction procedure was most efficient. Leucine incorporation saturation occurred at 1 microM in epiphyton and 100 nM in the pelagic zone. Saturation curves in sediments were difficult to model but showed the first level of leucine saturation at 50 microM. Increased uptake at higher leucine concentrations could be partly attributed to eukaryotes. Addition of micromolar concentrations of leucine did not enhance bacterial electron transport activity or DNA replication activity. Similar rates of leucine incorporation into protein calculated for whole sediment cores were observed after slurry and perfused-core incubations, but the rates exhibited strong vertical gradients after the core incubation. We conclude that the leucine incorporation method can measure bacterial production in a wide range of aquatic habitats, including fluvial sediments, if substrate saturation and isotope dilution are determined.

Use of the [14C]Leucine Incorporation Technique To Measure Bacterial Production in River Sediments and the Epiphyton

Science.gov (United States)

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 sediments, the biofilms on aquatic plants (epiphyton), and the pelagic zone, (iii) bacterial activities at different leucine concentrations, (iv) specificity of leucine uptake by bacteria, and (v) the effect of the incubation technique (perfused-core incubation versus slurry incubation) on leucine incorporation into protein. Bacterial protein was best extracted from sediments and precipitated by hot trichloroacetic acid treatment following ultrasonication. For epiphyton, an alkaline-extraction procedure was most efficient. Leucine incorporation saturation occurred at 1 μM in epiphyton and 100 nM in the pelagic zone. Saturation curves in sediments were difficult to model but showed the first level of leucine saturation at 50 μM. Increased uptake at higher leucine concentrations could be partly attributed to eukaryotes. Addition of micromolar concentrations of leucine did not enhance bacterial electron transport activity or DNA replication activity. Similar rates of leucine incorporation into protein calculated for whole sediment cores were observed after slurry and perfused-core incubations, but the rates exhibited strong vertical gradients after the core incubation. We conclude that the leucine incorporation method can measure bacterial production in a wide range of aquatic habitats, including fluvial sediments, if substrate saturation and isotope dilution are determined. PMID:10508068

Bacterial Cell Mechanics.

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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.

Bacterial communities associated with an occurrence of colored water in an urban drinking water distribution system.

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Wu, Hui Ting; Mi, Zi Long; Zhang, Jing Xu; Chen, Chao; Xie, Shu Guang

2014-08-01

This study aimed to investigate bacterial community in an urban drinking water distribution system (DWDS) during an occurrence of colored water. Variation in the bacterial community diversity and structure was observed among the different waters, with the predominance of Proteobacteria. While Verrucomicrobia was also a major phylum group in colored water. Limnobacter was the major genus group in colored water, but Undibacterium predominated in normal tap water. The coexistence of Limnobacter as well as Sediminibacterium and Aquabacterium might contribute to the formation of colored water. Copyright © 2014 The Editorial Board of Biomedical and Environmental Sciences. Published by China CDC. All rights reserved.

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

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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

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

Genome-Wide Association Study Identifies NBS-LRR-Encoding Genes Related with Anthracnose and Common Bacterial Blight in the Common Bean.

Science.gov (United States)

Wu, Jing; Zhu, Jifeng; Wang, Lanfen; Wang, Shumin

2017-01-01

Nucleotide-binding site and leucine-rich repeat (NBS-LRR) genes represent the largest and most important disease resistance genes in plants. The genome sequence of the common bean ( Phaseolus vulgaris L.) provides valuable data for determining the genomic organization of NBS-LRR genes. However, data on the NBS-LRR genes in the common bean are limited. In total, 178 NBS-LRR-type genes and 145 partial genes (with or without a NBS) located on 11 common bean chromosomes were identified from genome sequences database. Furthermore, 30 NBS-LRR genes were classified into Toll/interleukin-1 receptor (TIR)-NBS-LRR (TNL) types, and 148 NBS-LRR genes were classified into coiled-coil (CC)-NBS-LRR (CNL) types. Moreover, the phylogenetic tree supported the division of these PvNBS genes into two obvious groups, TNL types and CNL types. We also built expression profiles of NBS genes in response to anthracnose and common bacterial blight using qRT-PCR. Finally, we detected nine disease resistance loci for anthracnose (ANT) and seven for common bacterial blight (CBB) using the developed NBS-SSR markers. Among these loci, NSSR24, NSSR73, and NSSR265 may be located at new regions for ANT resistance, while NSSR65 and NSSR260 may be located at new regions for CBB resistance. Furthermore, we validated NSSR24, NSSR65, NSSR73, NSSR260, and NSSR265 using a new natural population. Our results provide useful information regarding the function of the NBS-LRR proteins and will accelerate the functional genomics and evolutionary studies of NBS-LRR genes in food legumes. NBS-SSR markers represent a wide-reaching resource for molecular breeding in the common bean and other food legumes. Collectively, our results should be of broad interest to bean scientists and breeders.

Engineering a novel multifunctional green fluorescent protein tag for a wide variety of protein research.

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Takuya Kobayashi

Full Text Available BACKGROUND: Genetically encoded tag is a powerful tool for protein research. Various kinds of tags have been developed: fluorescent proteins for live-cell imaging, affinity tags for protein isolation, and epitope tags for immunological detections. One of the major problems concerning the protein tagging is that many constructs with different tags have to be made for different applications, which is time- and resource-consuming. METHODOLOGY/PRINCIPAL FINDINGS: Here we report a novel multifunctional green fluorescent protein (mfGFP tag which was engineered by inserting multiple peptide tags, i.e., octa-histidine (8xHis, streptavidin-binding peptide (SBP, and c-Myc tag, in tandem into a loop of GFP. When fused to various proteins, mfGFP monitored their localization in living cells. Streptavidin agarose column chromatography with the SBP tag successfully isolated the protein complexes in a native form with a high purity. Tandem affinity purification (TAP with 8xHis and SBP tags in mfGFP further purified the protein complexes. mfGFP was clearly detected by c-Myc-specific antibody both in immunofluorescence and immuno-electron microscopy (EM. These findings indicate that mfGFP works well as a multifunctional tag in mammalian cells. The tag insertion was also successful in other fluorescent protein, mCherry. CONCLUSIONS AND SIGNIFICANCE: The multifunctional fluorescent protein tag is a useful tool for a wide variety of protein research, and may have the advantage over other multiple tag systems in its higher expandability and compatibility with existing and future tag technologies.

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

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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.

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

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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

Bacterial Community Shift Drives Antibiotic Resistance Promotion during Drinking Water Chlorination.

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Jia, Shuyu; Shi, Peng; Hu, Qing; Li, Bing; Zhang, Tong; Zhang, Xu-Xiang

2015-10-20

For comprehensive insights into the effects of chlorination, a widely used disinfection technology, on bacterial community and antibiotic resistome in drinking water, this study applied high-throughput sequencing and metagenomic approaches to investigate the changing patterns of antibiotic resistance genes (ARGs) and bacterial community in a drinking water treatment and distribution system. At genus level, chlorination could effectively remove Methylophilus, Methylotenera, Limnobacter, and Polynucleobacter, while increase the relative abundance of Pseudomonas, Acidovorax, Sphingomonas, Pleomonas, and Undibacterium in the drinking water. A total of 151 ARGs within 15 types were detectable in the drinking water, and chlorination evidently increased their total relative abundance while reduced their diversity in the opportunistic bacteria (p < 0.05). Residual chlorine was identified as the key contributing factor driving the bacterial community shift and resistome alteration. As the dominant persistent ARGs in the treatment and distribution system, multidrug resistance genes (mainly encoding resistance-nodulation-cell division transportation system) and bacitracin resistance gene bacA were mainly carried by chlorine-resistant bacteria Pseudomonas and Acidovorax, which mainly contributed to the ARGs abundance increase. The strong correlation between bacterial community shift and antibiotic resistome alteration observed in this study may shed new light on the mechanism behind the chlorination effects on antibiotic resistance.

Isolation of cell-free bacterial inclusion bodies.

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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.

Spatial distribution of planktonic bacterial and archaeal communities in the upper section of the tidal reach in Yangtze River

Science.gov (United States)

Fan, Limin; Song, Chao; Meng, Shunlong; Qiu, Liping; Zheng, Yao; Wu, Wei; Qu, Jianhong; Li, Dandan; Zhang, Cong; Hu, Gengdong; Chen, Jiazhang

2016-01-01

Bacterioplankton and archaeaplankton communities play key roles in the biogeochemical processes of water, and they may be affected by many factors. In this study, we used high-throughput 16S rRNA gene sequencing to profile planktonic bacterial and archaeal community compositions in the upper section of the tidal reach in Yangtze River. We found that the predominant bacterial phyla in this river section were Proteobacteria, Firmicutes, and Actinobacteria, whereas the predominant archaeal classes were Halobacteria, Methanomicrobia, and unclassified Euryarchaeota. Additionally, the bacterial and archaeal community compositions, richnesses, functional profiles, and ordinations were affected by the spatial heterogeneity related to the concentration changes of sulphate or nitrate. Notably, the bacterial community was more sensitive than the archaeal community to changes in the spatial characteristics of this river section. These findings provide important insights into the distributions of bacterial and archaeal communities in natural water habitats. PMID:27966673

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% (...

Role of proteins in controlling selenium nanoparticle size

International Nuclear Information System (INIS)

Dobias, J; Suvorova, E I; Bernier-Latmani, R

2011-01-01

This work investigates the potential for harnessing the association of bacterial proteins to biogenic selenium nanoparticles (SeNPs) to control the size distribution and the morphology of the resultant SeNPs. We conducted a proteomic study and compared proteins associated with biogenic SeNPs produced by E. coli to chemically synthesized SeNPs as well as magnetite nanoparticles. We identified four proteins (AdhP, Idh, OmpC, AceA) that bound specifically to SeNPs and observed a narrower size distribution as well as more spherical morphology when the particles were synthesized chemically in the presence of proteins. A more detailed study of AdhP (alcohol dehydrogenase propanol-preferring) confirmed the strong affinity of this protein for the SeNP surface and revealed that this protein controlled the size distribution of the SeNPs and yielded a narrow size distribution with a three-fold decrease in the median size. These results support the assertion that protein may become an important tool in the industrial-scale synthesis of SeNPs of uniform size and properties.

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.

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

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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.

Role and regulation of the orphan AphA protein of quorum sensing in pathogenic Vibrios.

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Lu, Renfei; Osei-Adjei, George; Huang, Xinxiang; Zhang, Yiquan

2018-03-01

Quorum sensing (QS), a cell-to-cell communication process, is widely distributed in the bacterial kingdom. Bacteria use QS to control gene expression in response to cell density by detecting the signal molecules called autoinducers. AphA protein is the master QS regulator of vibrios operating at low cell density. It regulates the expression of a variety of genes, especially those encoding virulence factors, flagella/motility and biofilm formation. The role and regulation of AphA in vibrios, especially in human pathogenic vibrios, are summarized in this review. Clarification of the roles of AphA will help us to understand the pathogenesis of vibrios.

Distribution and biological role of the oligopeptide-binding protein (OppA) in Xanthomonas species.

Science.gov (United States)

Oshiro, Elisa E; Tavares, Milene B; Suzuki, Celso F; Pimenta, Daniel C; Angeli, Claudia B; de Oliveira, Julio C F; Ferro, Maria I T; Ferreira, Luis C S; Ferreira, Rita C C

2010-04-01

In this study we investigated the prevalence of the oppA gene, encoding the oligopeptide binding protein (OppA) of the major bacterial oligopeptide uptake system (Opp), in different species of the genus Xanthomonas. The oppA gene was detected in two Xanthomonas axonopodis strains among eight tested Xanthomonas species. The generation of an isogenic oppA-knockout derivative of the Xac 306 strain, showed that the OppA protein neither plays a relevant role in oligopeptide uptake nor contributes to the infectivity and multiplication of the bacterial strain in leaves of sweet orange (Citrus sinensis) and Rangpur lime (Citrus limonia). Taken together these results suggest that the oppA gene has a recent evolutionary history in the genus and does not contribute in the physiology or pathogenesis of X. axonopodis.

Distribution and biological role of the oligopeptide-binding protein (OppA in Xanthomonas species

Directory of Open Access Journals (Sweden)

Elisa E. Oshiro

2010-01-01

Full Text Available In this study we investigated the prevalence of the oppA gene, encoding the oligopeptide binding protein (OppA of the major bacterial oligopeptide uptake system (Opp, in different species of the genus Xanthomonas. The oppA gene was detected in two Xanthomonas axonopodis strains among eight tested Xanthomonas species. The generation of an isogenic oppA-knockout derivative of the Xac 306 strain, showed that the OppA protein neither plays a relevant role in oligopeptide uptake nor contributes to the infectivity and multiplication of the bacterial strain in leaves of sweet orange (Citrus sinensis and Rangpur lime (Citrus limonia. Taken together these results suggest that the oppA gene has a recent evolutionary history in the genus and does not contribute in the physiology or pathogenesis of X. axonopodis.

Calculation of X-ray scattering curves and electron distance distribution functions of biological macromolecules in solution using the PROTEIN DATA BANK

International Nuclear Information System (INIS)

Mueller, J.J.; Friedrichowicz, E.; Nothnagel, A.; Wunderlich, T.; Ziehlsdorf, E.; Damaschun, G.

1983-01-01

The wide angle X-ray scattering curve, the electron distance distribution function and the solvent excluded volume of a macromolecule in solution are calculated from the atomic coordinates contained in the PROTEIN DATA BANK. The structures and the projections of the excluded volumes are depicted using molecule graphic routines. The described computer programs are used to determine the three-dimensional structure of macromolecules in solution from wide angle X-ray scattering data. (author)

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...

Dietary Protein Intake and Distribution Patterns of Well-Trained Dutch Athletes.

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Gillen, Jenna B; Trommelen, Jorn; Wardenaar, Floris C; Brinkmans, Naomi Y J; Versteegen, Joline J; Jonvik, Kristin L; Kapp, Christoph; de Vries, Jeanne; van den Borne, Joost J G C; Gibala, Martin J; van Loon, Luc J C

2017-04-01

Dietary protein intake should be optimized in all athletes to ensure proper recovery and enhance the skeletal muscle adaptive response to exercise training. In addition to total protein intake, the use of specific proteincontaining food sources and the distribution of protein throughout the day are relevant for optimizing protein intake in athletes. In the present study, we examined the daily intake and distribution of various proteincontaining food sources in a large cohort of strength, endurance and team-sport athletes. Well-trained male (n=327) and female (n=226) athletes completed multiple web-based 24-hr dietary recalls over a 2-4 wk period. Total energy intake, the contribution of animal- and plant-based proteins to daily protein intake, and protein intake at six eating moments were determined. Daily protein intake averaged 108±33 and 90±24 g in men and women, respectively, which corresponded to relative intakes of 1.5±0.4 and 1.4±0.4 g/kg. Dietary protein intake was correlated with total energy intake in strength (r=0.71, p sport (r=0.77, p protein intake was 57% and 43%, respectively. The distribution of protein intake was 19% (19±8 g) at breakfast, 24% (25±13 g) at lunch and 38% (38±15 g) at dinner. Protein intake was below the recommended 20 g for 58% of athletes at breakfast, 36% at lunch and 8% at dinner. In summary, this survey of athletes revealed they habitually consume > 1.2 g protein/kg/d, but the distribution throughout the day may be suboptimal to maximize the skeletal muscle adaptive response to training.

Estimating bacterial diversity for ecological studies: methods, metrics, and assumptions.

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Julia Birtel

Full Text Available Methods to estimate microbial diversity have developed rapidly in an effort to understand the distribution and diversity of microorganisms in natural environments. For bacterial communities, the 16S rRNA gene is the phylogenetic marker gene of choice, but most studies select only a specific region of the 16S rRNA to estimate bacterial diversity. Whereas biases derived from from DNA extraction, primer choice and PCR amplification are well documented, we here address how the choice of variable region can influence a wide range of standard ecological metrics, such as species richness, phylogenetic diversity, β-diversity and rank-abundance distributions. We have used Illumina paired-end sequencing to estimate the bacterial diversity of 20 natural lakes across Switzerland derived from three trimmed variable 16S rRNA regions (V3, V4, V5. Species richness, phylogenetic diversity, community composition, β-diversity, and rank-abundance distributions differed significantly between 16S rRNA regions. Overall, patterns of diversity quantified by the V3 and V5 regions were more similar to one another than those assessed by the V4 region. Similar results were obtained when analyzing the datasets with different sequence similarity thresholds used during sequences clustering and when the same analysis was used on a reference dataset of sequences from the Greengenes database. In addition we also measured species richness from the same lake samples using ARISA Fingerprinting, but did not find a strong relationship between species richness estimated by Illumina and ARISA. We conclude that the selection of 16S rRNA region significantly influences the estimation of bacterial diversity and species distributions and that caution is warranted when comparing data from different variable regions as well as when using different sequencing techniques.

The neglected intrinsic resistome of bacterial pathogens.

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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.

Evidence of codon usage in the nearest neighbor spacing distribution of bases in bacterial genomes

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Higareda, M. F.; Geiger, O.; Mendoza, L.; Méndez-Sánchez, R. A.

2012-02-01

Statistical analysis of whole genomic sequences usually assumes a homogeneous nucleotide density throughout the genome, an assumption that has been proved incorrect for several organisms since the nucleotide density is only locally homogeneous. To avoid giving a single numerical value to this variable property, we propose the use of spectral statistics, which characterizes the density of nucleotides as a function of its position in the genome. We show that the cumulative density of bases in bacterial genomes can be separated into an average (or secular) plus a fluctuating part. Bacterial genomes can be divided into two groups according to the qualitative description of their secular part: linear and piecewise linear. These two groups of genomes show different properties when their nucleotide spacing distribution is studied. In order to analyze genomes having a variable nucleotide density, statistically, the use of unfolding is necessary, i.e., to get a separation between the secular part and the fluctuations. The unfolding allows an adequate comparison with the statistical properties of other genomes. With this methodology, four genomes were analyzed Burkholderia, Bacillus, Clostridium and Corynebacterium. Interestingly, the nearest neighbor spacing distributions or detrended distance distributions are very similar for species within the same genus but they are very different for species from different genera. This difference can be attributed to the difference in the codon usage.

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.

Genome-wide polysomal analysis of a yeast strain with mutated ribosomal protein S9

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Arava Yoav

2007-08-01

Full Text Available Abstract Background The yeast ribosomal protein S9 (S9 is located at the entrance tunnel of the mRNA into the ribosome. It is known to play a role in accurate decoding and its bacterial homolog (S4 has recently been shown to be involved in opening RNA duplexes. Here we examined the effects of changing the C terminus of S9, which is rich in acidic amino acids and extends out of the ribosome surface. Results We performed a genome-wide analysis to reveal effects at the transcription and translation levels of all yeast genes. While negligible relative changes were observed in steady-state mRNA levels, a significant number of mRNAs appeared to have altered ribosomal density. Notably, 40% of the genes having reliable signals changed their ribosomal association by more than one ribosome. Yet, no general correlations with physical or functional features of the mRNA were observed. Ribosome Density Mapping (RDM along four of the mRNAs with increased association revealed an increase in ribosomal density towards the end of the coding region for at least two of them. Read-through analysis did not reveal any increase in read-through of a premature stop codon by the mutant strain. Conclusion The ribosomal protein rpS9 appears to be involved in the translation of many mRNAs, since altering its C terminus led to a significant change in ribosomal association of many mRNAs. We did not find strong correlations between these changes and several physical features of the mRNA, yet future studies with advanced tools may allow such correlations to be determined. Importantly, our results indicate an accumulation of ribosomes towards the end of the coding regions of some mRNAs. This suggests an involvement of S9 in ribosomal dissociation during translation termination.

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...

Genome-Wide Association Study of the Genetic Determinants of Emphysema Distribution.

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Boueiz, Adel; Lutz, Sharon M; Cho, Michael H; Hersh, Craig P; Bowler, Russell P; Washko, George R; Halper-Stromberg, Eitan; Bakke, Per; Gulsvik, Amund; Laird, Nan M; Beaty, Terri H; Coxson, Harvey O; Crapo, James D; Silverman, Edwin K; Castaldi, Peter J; DeMeo, Dawn L

2017-03-15

Emphysema has considerable variability in the severity and distribution of parenchymal destruction throughout the lungs. Upper lobe-predominant emphysema has emerged as an important predictor of response to lung volume reduction surgery. Yet, aside from alpha-1 antitrypsin deficiency, the genetic determinants of emphysema distribution remain largely unknown. To identify the genetic influences of emphysema distribution in non-alpha-1 antitrypsin-deficient smokers. A total of 11,532 subjects with complete genotype and computed tomography densitometry data in the COPDGene (Genetic Epidemiology of Chronic Obstructive Pulmonary Disease [COPD]; non-Hispanic white and African American), ECLIPSE (Evaluation of COPD Longitudinally to Identify Predictive Surrogate Endpoints), and GenKOLS (Genetics of Chronic Obstructive Lung Disease) studies were analyzed. Two computed tomography scan emphysema distribution measures (difference between upper-third and lower-third emphysema; ratio of upper-third to lower-third emphysema) were tested for genetic associations in all study subjects. Separate analyses in each study population were followed by a fixed effect metaanalysis. Single-nucleotide polymorphism-, gene-, and pathway-based approaches were used. In silico functional evaluation was also performed. We identified five loci associated with emphysema distribution at genome-wide significance. These loci included two previously reported associations with COPD susceptibility (4q31 near HHIP and 15q25 near CHRNA5) and three new associations near SOWAHB, TRAPPC9, and KIAA1462. Gene set analysis and in silico functional evaluation revealed pathways and cell types that may potentially contribute to the pathogenesis of emphysema distribution. This multicohort genome-wide association study identified new genomic loci associated with differential emphysematous destruction throughout the lungs. These findings may point to new biologic pathways on which to expand diagnostic and therapeutic

Distribution of N-glycosylation sequons in proteins: how apart are they?

DEFF Research Database (Denmark)

Rao, Shyama Prasad; Buus, Ole Thomsen; Wollenweber, Bernd

2011-01-01

of experimentally confirmed eukaryotic N-glycoproteins we analyzed the relative position and distribution of sequons. N-Glycosylation probability was found to be lower in the termini of protein sequences compared to the mid region. N-glycosylated sequons were found much farther from C terminus compared to the N......N-glycosylation is a common protein modification process, which affects a number of properties of proteins. Little is known about the distribution of N-glycosylation sequons, for example, the distance between glycosylated sites and their position in the protein primary sequence. Using a large set......-terminus of the protein sequence and this effect was more pronounced for NXS sequons. The distribution of sequons, modeled based on balls-in-boxes classical occupancy, showed a near-maximum probability. Considerable proportion of sequons was found within a distance of ten amino acids, indicating that the steric hindrance...

The SPOR Domain, a Widely Conserved Peptidoglycan Binding Domain That Targets Proteins to the Site of Cell Division.

Science.gov (United States)

Yahashiri, Atsushi; Jorgenson, Matthew A; Weiss, David S

2017-07-15

Sporulation-related repeat (SPOR) domains are small peptidoglycan (PG) binding domains found in thousands of bacterial proteins. The name "SPOR domain" stems from the fact that several early examples came from proteins involved in sporulation, but SPOR domain proteins are quite diverse and contribute to a variety of processes that involve remodeling of the PG sacculus, especially with respect to cell division. SPOR domains target proteins to the division site by binding to regions of PG devoid of stem peptides ("denuded" glycans), which in turn are enriched in septal PG by the intense, localized activity of cell wall amidases involved in daughter cell separation. This targeting mechanism sets SPOR domain proteins apart from most other septal ring proteins, which localize via protein-protein interactions. In addition to SPOR domains, bacteria contain several other PG-binding domains that can exploit features of the cell wall to target proteins to specific subcellular sites. Copyright © 2017 American Society for Microbiology.

Connecting protein and mRNA burst distributions for stochastic models of gene expression

International Nuclear Information System (INIS)

Elgart, Vlad; Jia, Tao; Fenley, Andrew T; Kulkarni, Rahul

2011-01-01

The intrinsic stochasticity of gene expression can lead to large variability in protein levels for genetically identical cells. Such variability in protein levels can arise from infrequent synthesis of mRNAs which in turn give rise to bursts of protein expression. Protein expression occurring in bursts has indeed been observed experimentally and recent studies have also found evidence for transcriptional bursting, i.e. production of mRNAs in bursts. Given that there are distinct experimental techniques for quantifying the noise at different stages of gene expression, it is of interest to derive analytical results connecting experimental observations at different levels. In this work, we consider stochastic models of gene expression for which mRNA and protein production occurs in independent bursts. For such models, we derive analytical expressions connecting protein and mRNA burst distributions which show how the functional form of the mRNA burst distribution can be inferred from the protein burst distribution. Additionally, if gene expression is repressed such that observed protein bursts arise only from single mRNAs, we show how observations of protein burst distributions (repressed and unrepressed) can be used to completely determine the mRNA burst distribution. Assuming independent contributions from individual bursts, we derive analytical expressions connecting means and variances for burst and steady-state protein distributions. Finally, we validate our general analytical results by considering a specific reaction scheme involving regulation of protein bursts by small RNAs. For a range of parameters, we derive analytical expressions for regulated protein distributions that are validated using stochastic simulations. The analytical results obtained in this work can thus serve as useful inputs for a broad range of studies focusing on stochasticity in gene expression

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.

Classification of bacterial contamination using image processing and distributed computing.

Science.gov (United States)

Ahmed, W M; Bayraktar, B; Bhunia, A; Hirleman, E D; Robinson, J P; Rajwa, B

2013-01-01

Disease outbreaks due to contaminated food are a major concern not only for the food-processing industry but also for the public at large. Techniques for automated detection and classification of microorganisms can be a great help in preventing outbreaks and maintaining the safety of the nations food supply. Identification and classification of foodborne pathogens using colony scatter patterns is a promising new label-free technique that utilizes image-analysis and machine-learning tools. However, the feature-extraction tools employed for this approach are computationally complex, and choosing the right combination of scatter-related features requires extensive testing with different feature combinations. In the presented work we used computer clusters to speed up the feature-extraction process, which enables us to analyze the contribution of different scatter-based features to the overall classification accuracy. A set of 1000 scatter patterns representing ten different bacterial strains was used. Zernike and Chebyshev moments as well as Haralick texture features were computed from the available light-scatter patterns. The most promising features were first selected using Fishers discriminant analysis, and subsequently a support-vector-machine (SVM) classifier with a linear kernel was used. With extensive testing we were able to identify a small subset of features that produced the desired results in terms of classification accuracy and execution speed. The use of distributed computing for scatter-pattern analysis, feature extraction, and selection provides a feasible mechanism for large-scale deployment of a light scatter-based approach to bacterial classification.

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.

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

Biomarker-based classification of bacterial and fungal whole-blood infections in a genome-wide expression study

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Andreas eDix

2015-03-01

Full Text Available Sepsis is a clinical syndrome that can be caused by bacteria or fungi. Early knowledge on the nature of the causative agent is a prerequisite for targeted anti-microbial therapy. Besides currently used detection methods like blood culture and PCR-based assays, the analysis of the transcriptional response of the host to infecting organisms holds great promise. In this study, we aim to examine the transcriptional footprint of infections caused by the bacterial pathogens Staphylococcus aureus and Escherichia coli and the fungal pathogens Candida albicans and Aspergillus fumigatus in a human whole-blood model. Moreover, we use the expression information to build a random forest classifier to classify if a sample contains a bacterial, fungal, or mock-infection. After normalizing the transcription intensities using stably expressed reference genes, we filtered the gene set for biomarkers of bacterial or fungal blood infections. This selection is based on differential expression and an additional gene relevance measure. In this way, we identified 38 biomarker genes, including IL6, SOCS3, and IRG1 which were already associated to sepsis by other studies. Using these genes, we trained the classifier and assessed its performance. It yielded a 96% accuracy (sensitivities >93%, specificities >97% for a 10-fold stratified cross-validation and a 92% accuracy (sensitivities and specificities >83% for an additional test dataset comprising Cryptococcus neoformans infections. Furthermore, the classifier is robust to Gaussian noise, indicating correct class predictions on datasets of new species. In conclusion, this genome-wide approach demonstrates an effective feature selection process in combination with the construction of a well-performing classification model. Further analyses of genes with pathogen-dependent expression patterns can provide insights into the systemic host responses, which may lead to new anti-microbial therapeutic advances.

The Mineralosphere Concept: Mineralogical Control of the Distribution and Function of Mineral-associated Bacterial Communities.

Science.gov (United States)

Uroz, Stephane; Kelly, Laura Catherine; Turpault, Marie-Pierre; Lepleux, Cendrella; Frey-Klett, Pascale

2015-12-01

Soil is composed of a mosaic of different rocks and minerals, usually considered as an inert substrata for microbial colonization. However, recent findings suggest that minerals, in soils and elsewhere, favour the development of specific microbial communities according to their mineralogy, nutritive content, and weatherability. Based upon recent studies, we highlight how bacterial communities are distributed on the surface of, and in close proximity to, minerals. We also consider the potential role of the mineral-associated bacterial communities in mineral weathering and nutrient cycling in soils, with a specific focus on nutrient-poor and acidic forest ecosystems. We propose to define this microbial habitat as the mineralosphere, where key drivers of the microbial communities are the physicochemical properties of the minerals. Copyright © 2015 Elsevier Ltd. All rights reserved.

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.

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

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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.

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

World-wide distribution automation systems

International Nuclear Information System (INIS)

Devaney, T.M.

1994-01-01

A worldwide power distribution automation system is outlined. Distribution automation is defined and the status of utility automation is discussed. Other topics discussed include a distribution management system, substation feeder, and customer functions, potential benefits, automation costs, planning and engineering considerations, automation trends, databases, system operation, computer modeling of system, and distribution management systems

Use of Random and Site-Directed Mutagenesis to Probe Protein Structure-Function Relationships: Applied Techniques in the Study of Helicobacter pylori.

Science.gov (United States)

Whitmire, Jeannette M; Merrell, D Scott

2017-01-01

Mutagenesis is a valuable tool to examine the structure-function relationships of bacterial proteins. As such, a wide variety of mutagenesis techniques and strategies have been developed. This chapter details a selection of random mutagenesis methods and site-directed mutagenesis procedures that can be applied to an array of bacterial species. Additionally, the direct application of the techniques to study the Helicobacter pylori Ferric Uptake Regulator (Fur) protein is described. The varied approaches illustrated herein allow the robust investigation of the structural-functional relationships within a protein of interest.

Dietary Protein in Older Adults: Adequate Daily Intake but Potential for Improved Distribution

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Danielle K. Cardon-Thomas

2017-02-01

Full Text Available Daily distribution of dietary protein may be important in protecting against sarcopenia, specifically in terms of per meal amounts relative to a proposed threshold for maximal response. The aims of this study were to determine total and per meal protein intake in older adults, as well as identifying associations with physical activity and sedentary behavior. Three-day food diaries recorded protein intake in 38 participants. Protein distribution, coefficient of variation (CV, and per meal amounts were calculated. Accelerometry was used to collect physical activity data as well as volume and patterns of sedentary time. Average intake was 1.14 g·kg−1·day−1. Distribution was uneven (CV = 0.67, and 79% of participants reported <0.4 g·kg−1 protein content in at least 2/3 daily meals. Protein intake was significantly correlated with step count (r = 0.439, p = 0.007 and negatively correlated with sedentary time (r = −0.456, p = 0.005 and Gini index G, which describes the pattern of accumulation of sedentary time (r = −0.421, p = 0.011. Total daily protein intake was sufficient; however, distribution did not align with the current literature; increasing protein intake may help to facilitate optimization of distribution. Associations between protein and other risk factors for sarcopenia may also inform protective strategies.

Genome-wide identification and characterization of stress-associated protein (SAP gene family encoding A20/AN1 zinc-finger proteins in Medicago truncatula

Directory of Open Access Journals (Sweden)

Zhou Yong

2018-01-01

Full Text Available Stress associated proteins (SAPs play important roles in developmental processes, responses to various stresses and hormone stimulation in plants. However, little is known about the SAP gene family in Medicago truncatula. In this study, a total of 17 MtSAP genes encoding A20/AN1 zinc-finger proteins were characterized. Out of these 17 genes, 15 were distributed over all 8 chromosomes at different densities, and two segmental duplication events were detected. The phylogenetic analysis of these proteins and their orthologs from Arabidopsis and rice suggested that they could be classified into five out of the seven groups of SAP family genes, with genes in the same group showing similar structures and conserved domains. The cis-elements of the MtSAP promoters were studied, and many cis-elements related to stress and plant hormone responses were identified. We also investigated the stress-responsive expression patterns of the MtSAP genes under various stresses, including drought, exposure to NaCl and cold. The qRT-PCR results showed that numerous MtSAP genes exhibited transcriptional responses to multiple abiotic stresses. These results lay the foundation for further functional characterization of SAP genes. To the best of our knowledge, this is the first report of a genome-wide analysis of the SAP gene family in M. truncatula.

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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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.

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

Directory of Open Access Journals (Sweden)

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

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

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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.

Applications of whole-cell bacterial sensors in biotechnology and environmental science

Energy Technology Data Exchange (ETDEWEB)

Yagi, Kiyohito [Osaka Univ., Suita (Japan). Graduate School of Pharmaceutical Sciences

2007-01-15

Biosensors have major advantages over chemical or physical analyses with regard to specificity, sensitivity, and portability. Recently, many types of whole-cell bacterial biosensors have been developed using recombinant DNA technology. The bacteria are genetically engineered to respond to the presence of chemicals or physiological stresses by synthesizing a reporter protein, such as luciferase, {beta}-galactosidase, or green fluorescent protein. In addition to an overview of conventional biosensors, this minireview discusses a novel type of biosensor using a photosynthetic bacterium as the sensor strain and the crtA gene, which is responsible for carotenoid synthesis, as the reporter. Since bacteria possess a wide variety of stress-response mechanisms, including antioxidation, heat-shock responses, nutrient-starvation, and membrane-damage responses, DNA response elements for several stress-response proteins can be fused with various reporter genes to construct a versatile set of bacterial biosensors for a variety of analytes. Portable biosensors for on-site monitoring have been developed using a freeze-dried biosensing strain, and cell array biosensors have been designed for high-throughput analysis. Moreover, in the future, the use of single-cell biosensors will permit detailed analyses of samples. Signals from such sensors could be detected with digital imaging, epifluorescence microscopy, and/or flow cytometry. (orig.)

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

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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.

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

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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.

Printing Multistrain Bacterial Patterns with a Piezoelectric Inkjet Printer

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Merrin, Jack; Leibler, Stanislas; Chuang, John S.

2007-01-01

Many studies involving interacting microorganisms would benefit from simple devices able to deposit cells in precisely defined patterns. We describe an inexpensive bacterial piezoelectric inkjet printer (adapted from the design of the POSaM oligonucleotide microarrayer) that can be used to “print out” different strains of bacteria or chemicals in small droplets onto a flat surface at high resolution. The capabilities of this device are demonstrated by printing ordered arrays comprising two bacterial strains labeled with different fluorescent proteins. We also characterized several properties of this piezoelectric printer, such as the droplet volume (of the order of tens of pl), the distribution of number of cells in each droplet, and the dependence of droplet volume on printing frequency. We established the limits of the printing resolution, and determined that the printed viability of Escherichia coli exceeded 98.5%. PMID:17653283

A genome-wide association study identifies protein quantitative trait loci (pQTLs.

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David Melzer

2008-05-01

Full Text Available There is considerable evidence that human genetic variation influences gene expression. Genome-wide studies have revealed that mRNA levels are associated with genetic variation in or close to the gene coding for those mRNA transcripts - cis effects, and elsewhere in the genome - trans effects. The role of genetic variation in determining protein levels has not been systematically assessed. Using a genome-wide association approach we show that common genetic variation influences levels of clinically relevant proteins in human serum and plasma. We evaluated the role of 496,032 polymorphisms on levels of 42 proteins measured in 1200 fasting individuals from the population based InCHIANTI study. Proteins included insulin, several interleukins, adipokines, chemokines, and liver function markers that are implicated in many common diseases including metabolic, inflammatory, and infectious conditions. We identified eight Cis effects, including variants in or near the IL6R (p = 1.8x10(-57, CCL4L1 (p = 3.9x10(-21, IL18 (p = 6.8x10(-13, LPA (p = 4.4x10(-10, GGT1 (p = 1.5x10(-7, SHBG (p = 3.1x10(-7, CRP (p = 6.4x10(-6 and IL1RN (p = 7.3x10(-6 genes, all associated with their respective protein products with effect sizes ranging from 0.19 to 0.69 standard deviations per allele. Mechanisms implicated include altered rates of cleavage of bound to unbound soluble receptor (IL6R, altered secretion rates of different sized proteins (LPA, variation in gene copy number (CCL4L1 and altered transcription (GGT1. We identified one novel trans effect that was an association between ABO blood group and tumour necrosis factor alpha (TNF-alpha levels (p = 6.8x10(-40, but this finding was not present when TNF-alpha was measured using a different assay , or in a second study, suggesting an assay-specific association. Our results show that protein levels share some of the features of the genetics of gene expression. These include the presence of strong genetic effects in cis

Biodegradation in a Partially Saturated Sand Matrix: Compounding Effects of Water Content, Bacterial Spatial Distribution, and Motility

DEFF Research Database (Denmark)

Dechesne, Arnaud; Owsianiak, Mikolaj; Bazire, Alexis

2010-01-01

colonizing these zones or on pollutant mass transfer to neighboring zones containing degraders. In a model system, we quantified the role exerted by water on mineralization rate in the context of a heterogeneously distributed degradation potential. Alginate beads colonized by Pseudomonas putida KT2440 were......Bacterial pesticide degraders are generally heterogeneously distributed in soils, leaving soil volumes devoid of degradation potential. This is expected to have an impact on degradation rates because the degradation of pollutant molecules in such zones will be contingent either on degraders...... inserted at prescribed locations in sand microcosms so that the initial spatial distribution of the mineralization potential was controlled. The mineralization rate was strongly affected by the matric potential (decreasing rate with decreasing matric potential) and by the initial distribution...

Population-wide distributions of neural activity during perceptual decision-making

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Machens, Christian

2018-01-01

Cortical activity involves large populations of neurons, even when it is limited to functionally coherent areas. Electrophysiological recordings, on the other hand, involve comparatively small neural ensembles, even when modern-day techniques are used. Here we review results which have started to fill the gap between these two scales of inquiry, by shedding light on the statistical distributions of activity in large populations of cells. We put our main focus on data recorded in awake animals that perform simple decision-making tasks and consider statistical distributions of activity throughout cortex, across sensory, associative, and motor areas. We transversally review the complexity of these distributions, from distributions of firing rates and metrics of spike-train structure, through distributions of tuning to stimuli or actions and of choice signals, and finally the dynamical evolution of neural population activity and the distributions of (pairwise) neural interactions. This approach reveals shared patterns of statistical organization across cortex, including: (i) long-tailed distributions of activity, where quasi-silence seems to be the rule for a majority of neurons; that are barely distinguishable between spontaneous and active states; (ii) distributions of tuning parameters for sensory (and motor) variables, which show an extensive extrapolation and fragmentation of their representations in the periphery; and (iii) population-wide dynamics that reveal rotations of internal representations over time, whose traces can be found both in stimulus-driven and internally generated activity. We discuss how these insights are leading us away from the notion of discrete classes of cells, and are acting as powerful constraints on theories and models of cortical organization and population coding. PMID:23123501

Impulse radio ultra wide-band over multi-mode fiber for in-home signal distribution

DEFF Research Database (Denmark)

Caballero Jambrina, Antonio; Rodes, Roberto; Jensen, Jesper Bevensee

2009-01-01

We propose and experimentally demonstrate a high speed impulse radio ultra wide-band (IR-UWB) wireless link for in-home network signal distribution. The IR-UWB pulse is distributed over a multimode fiber to the transmitter antenna. Wireless transmitted bit-rates of 1 Gbps at 2 m and 2 Gbps at 1.5 m...

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)

Constraints on lateral gene transfer in promoting fimbrial usher protein diversity and function.

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Stubenrauch, Christopher J; Dougan, Gordon; Lithgow, Trevor; Heinz, Eva

2017-11-01

Fimbriae are long, adhesive structures widespread throughout members of the family Enterobacteriaceae. They are multimeric extrusions, which are moved out of the bacterial cell through an integral outer membrane protein called usher. The complex folding mechanics of the usher protein were recently revealed to be catalysed by the membrane-embedded translocation and assembly module (TAM). Here, we examine the diversity of usher proteins across a wide range of extraintestinal (ExPEC) and enteropathogenic (EPEC) Escherichia coli , and further focus on a so far undescribed chaperone-usher system, with this usher referred to as UshC. The fimbrial system containing UshC is distributed across a discrete set of EPEC types, including model strains like E2348/67, as well as ExPEC ST131, currently the most prominent multi-drug-resistant uropathogenic E. coli strain worldwide. Deletion of the TAM from a naive strain of E. coli results in a drastic time delay in folding of UshC, which can be observed for a protein from EPEC as well as for two introduced proteins from related organisms, Yersinia and Enterobacter We suggest that this models why the TAM machinery is essential for efficient folding of proteins acquired via lateral gene transfer. © 2017 The Authors.

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

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

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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.

Anthropogenic impact on biogenic substance distribution and bacterial community in sediment along the Yarlung Tsangpo River on Tibet Plateau, China

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Wang, C.; Peifang, W.; Wang, X.; Hou, J.; Miao, L.

2017-12-01

Lotic river system plays an important part in water-vapor transfer and biogenic substances migration and transformation. Anthropogenic activities, including wastewater discharging and river damming, have altered river ecosystem and continuum. However, as the longest alpine river in China and suffered from increasing anthropogenic activities, the Yarlung Tsangpo River has been rarely studied. Recently, more attention has also been paid to the bacteria in river sediment as they make vital contributions to the biogeochemical nutrient cycling. Here, the distribution of biogenic substances, including nitrogen, phosphorus, silicon and carbon, was explored in both water and sediment of the Yarlung Tsangpo River. By using the next generation 16S rRNA sequencing, the bacterial diversity and structure in river sediment were presented. The results indicated that the nutrient concentrations increased in densely populated sites, revealing that biogenic substance distribution corresponded with the intensity of anthropogenic activity along the river. Nitrogen, phosphorus, silicon and carbon in water and sediment were all retained by the Zangmu Dam which is the only dam in the mainstream of the river. Moreover, the river damming decreased the biomass and diversity of bacteria in sediment, but no significant alteration of community structure was observed upstream and downstream of the dam. The most dominant bacteria all along the river was Proteobacteria. Meanwhile, Verrucomicrobia and Firmicutes also dominated the community composition in upstream and downstream of the river, respectively. In addition, total organic carbon (TOC) was proved to be the most important environmental factor shaping the bacterial community in river sediment. Our study offered the preliminary insights into the biogenic substance distribution and bacterial community in sediment along an alpine river which was affected by anthropogenic activities. In the future, more studies are needed to reveal the

An Even Distribution of Protein Intake Daily Promotes Protein Adequacy but Does Not Influence Nutritional Status in Institutionalized Elderly.

Science.gov (United States)

Tieland, Michael; Beelen, Janne; Laan, Anna C M; Poon, Shirley; de Groot, Lisette C P G M; Seeman, Ego; Wang, Xiaofang; Iuliano, Sandra

2018-01-01

Although it has been established that sufficient protein is required to maintain good nutritional status and support healthy aging, it is not clear if the pattern of protein consumption may also influence nutritional status, especially in institutionalized elderly who are at risk of malnutrition. Therefore, we aim to determine the association between protein intake distribution and nutritional status in institutionalized elderly people. Cross-sectional study among 481 institutionalized older adults. Dietary data from 481 ambulant elderly people (68.8% female, mean age 87.5 ± 6.3 years) residing in 52 aged-care facilities in Victoria, Australia, were assessed over 2 days using plate waste analysis. Nutritional status was determined using the Mini-Nutritional Assessment tool and serum (n = 208) analyzed for albumin, hemoglobin, and IGF-1. Protein intake distribution was classified as: spread (even distribution across 3 meals, n = 65), pulse (most protein consumed in one meal, n = 72) or intermediate (n = 344). Regression analysis was used to investigate associations. Mean protein intakes were higher in the spread (60.5 ± 2.0 g/d) than intermediate group (56.0 ± 0.8 g/d, P = .037), and tended to be higher than those in the pulse group (55.9 ± 1.9 g/d, P = .097). Residents with an even distribution of protein intake achieved a higher level of the recommended daily intake for protein (96.2 ± 30.0%) than the intermediate (86.3 ± 26.2%, P = .008) and pulse (87.4 ± 30.5%, P = .06) groups, and also achieved a greater level of their estimated energy requirements (intermediate; P = .039, pulse; P = .001). Nutritional status (Mini-Nutritional Assessment score) did not differ between groups (pulse; 20.5 ± 4.5, intermediate; 21.0 ± 2.5, spread; 20.5 ± 3.5), nor did any other indices of nutritional status. Meeting protein requirements is required before protein distribution may influence nutritional status in institutionalized

Protein-only, antimicrobial peptide-containing recombinant nanoparticles with inherent built-in antibacterial activity.

Science.gov (United States)

Serna, Naroa; Sánchez-García, Laura; Sánchez-Chardi, Alejandro; Unzueta, Ugutz; Roldán, Mónica; Mangues, Ramón; Vázquez, Esther; Villaverde, Antonio

2017-09-15

The emergence of bacterial antibiotic resistances is a serious concern in human and animal health. In this context, naturally occurring cationic antimicrobial peptides (AMPs) might play a main role in a next generation of drugs against bacterial infections. Taking an innovative approach to design self-organizing functional proteins, we have generated here protein-only nanoparticles with intrinsic AMP microbicide activity. Using a recombinant version of the GWH1 antimicrobial peptide as building block, these materials show a wide antibacterial activity spectrum in absence of detectable toxicity on mammalian cells. The GWH1-based nanoparticles combine clinically appealing properties of nanoscale materials with full biocompatibility, structural and functional plasticity and biological efficacy exhibited by proteins. Because of the largely implemented biological fabrication of recombinant protein drugs, the protein-based platform presented here represents a novel and scalable strategy in antimicrobial drug design, that by solving some of the limitations of AMPs offers a promising alternative to conventional antibiotics. The low molecular weight antimicrobial peptide GWH1 has been engineered to oligomerize as self-assembling protein-only nanoparticles of around 50nm. In this form, the peptide exhibits potent and broad antibacterial activities against both Gram-positive and Gram-negative bacteria, without any harmful effect over mammalian cells. As a solid proof-of-concept, this finding strongly supports the design and biofabrication of nanoscale antimicrobial materials with in-built functionalities. The protein-based homogeneous composition offer advantages over alternative materials explored as antimicrobial agents, regarding biocompatibility, biodegradability and environmental suitability. Beyond the described prototype, this transversal engineering concept has wide applicability in the design of novel nanomedicines for advanced treatments of bacterial infections

Biomimicry of quorum sensing using bacterial lifecycle model.

Science.gov (United States)

Niu, Ben; Wang, Hong; Duan, Qiqi; Li, Li

2013-01-01

Recent microbiologic studies have shown that quorum sensing mechanisms, which serve as one of the fundamental requirements for bacterial survival, exist widely in bacterial intra- and inter-species cell-cell communication. Many simulation models, inspired by the social behavior of natural organisms, are presented to provide new approaches for solving realistic optimization problems. Most of these simulation models follow population-based modelling approaches, where all the individuals are updated according to the same rules. Therefore, it is difficult to maintain the diversity of the population. In this paper, we present a computational model termed LCM-QS, which simulates the bacterial quorum-sensing (QS) mechanism using an individual-based modelling approach under the framework of Agent-Environment-Rule (AER) scheme, i.e. bacterial lifecycle model (LCM). LCM-QS model can be classified into three main sub-models: chemotaxis with QS sub-model, reproduction and elimination sub-model and migration sub-model. The proposed model is used to not only imitate the bacterial evolution process at the single-cell level, but also concentrate on the study of bacterial macroscopic behaviour. Comparative experiments under four different scenarios have been conducted in an artificial 3-D environment with nutrients and noxious distribution. Detailed study on bacterial chemotatic processes with quorum sensing and without quorum sensing are compared. By using quorum sensing mechanisms, artificial bacteria working together can find the nutrient concentration (or global optimum) quickly in the artificial environment. Biomimicry of quorum sensing mechanisms using the lifecycle model allows the artificial bacteria endowed with the communication abilities, which are essential to obtain more valuable information to guide their search cooperatively towards the preferred nutrient concentrations. It can also provide an inspiration for designing new swarm intelligence optimization algorithms

Dietary protein intake and distribution patterns of well-trained Dutch athletes

NARCIS (Netherlands)

Gillen, Jenna B.; Trommelen, Jorn; Wardenaar, Floris C.; Brinkmans, Naomi Y.J.; Versteegen, Joline J.; Jonvik, Kristin L.; Kapp, Christoph; Vries, de Jeanne; Borne, van den Joost J.G.C.; Gibala, Martin J.; Loon, van Luc J.C.

2017-01-01

Dietary protein intake should be optimized in all athletes to ensure proper recovery and enhance the skeletal muscle adaptive response to exercise training. In addition to total protein intake, the use of specific proteincontaining food sources and the distribution of protein throughout the day

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.

Mechanical Homogenization Increases Bacterial Homogeneity in Sputum

Science.gov (United States)

Stokell, Joshua R.; Khan, Ammad

2014-01-01

Sputum obtained from patients with cystic fibrosis (CF) is highly viscous and often heterogeneous in bacterial distribution. Adding dithiothreitol (DTT) is the standard method for liquefaction prior to processing sputum for molecular detection assays. To determine if DTT treatment homogenizes the bacterial distribution within sputum, we measured the difference in mean total bacterial abundance and abundance of Burkholderia multivorans between aliquots of DTT-treated sputum samples with and without a mechanical homogenization (MH) step using a high-speed dispersing element. Additionally, we measured the effect of MH on bacterial abundance. We found a significant difference between the mean bacterial abundances in aliquots that were subjected to only DTT treatment and those of the aliquots which included an MH step (all bacteria, P = 0.04; B. multivorans, P = 0.05). There was no significant effect of MH on bacterial abundance in sputum. Although our results are from a single CF patient, they indicate that mechanical homogenization increases the homogeneity of bacteria in sputum. PMID:24759710

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.

Spatial distribution of bacterial communities on volumetric and planar anodes in single-chamber air-cathode microbial fuel cells

KAUST Repository

Vargas, Ignacio T.

2013-05-29

Pyrosequencing was used to characterize bacterial communities in air-cathode microbial fuel cells across a volumetric (graphite fiber brush) and a planar (carbon cloth) anode, where different physical and chemical gradients would be expected associated with the distance between anode location and the air cathode. As expected, the stable operational voltage and the coulombic efficiency (CE) were higher for the volumetric anode than the planar anode (0.57V and CE=22% vs. 0.51V and CE=12%). The genus Geobacter was the only known exoelectrogen among the observed dominant groups, comprising 57±4% of recovered sequences for the brush and 27±5% for the carbon-cloth anode. While the bacterial communities differed between the two anode materials, results showed that Geobacter spp. and other dominant bacterial groups were homogenously distributed across both planar and volumetric anodes. This lends support to previous community analysis interpretations based on a single biofilm sampling location in these systems. © 2013 Wiley Periodicals, Inc.

Phylogenetic distribution and membrane topology of the LytR-CpsA-Psr protein family

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Berger-Bächi Brigitte

2008-12-01

Full Text Available Abstract Background The bacterial cell wall is the target of many antibiotics and cell envelope constituents are critical to host-pathogen interactions. To combat resistance development and virulence, a detailed knowledge of the individual factors involved is essential. Members of the LytR-CpsA-Psr family of cell envelope-associated attenuators are relevant for β-lactam resistance, biofilm formation, and stress tolerance, and they are suggested to play a role in cell wall maintenance. However, their precise function is still unknown. This study addresses the occurrence as well as sequence-based characteristics of the LytR-CpsA-Psr proteins. Results A comprehensive list of LytR-CpsA-Psr proteins was established, and their phylogenetic distribution and clustering into subgroups was determined. LytR-CpsA-Psr proteins were present in all Gram-positive organisms, except for the cell wall-deficient Mollicutes and one strain of the Clostridiales. In contrast, the majority of Gram-negatives did not contain LytR-CpsA-Psr family members. Despite high sequence divergence, the LytR-CpsA-Psr domains of different subclusters shared a highly similar, predicted mixed a/β-structure, and conserved charged residues. PhoA fusion experiments, using MsrR of Staphylococcus aureus, confirmed membrane topology predictions and extracellular location of its LytR-CpsA-Psr domain. Conclusion The LytR-CpsA-Psr domain is unique to bacteria. The presence of diverse subgroups within the LytR-CpsA-Psr family might indicate functional differences, and could explain variations in phenotypes of respective mutants reported. The identified conserved structural elements and amino acids are likely to be important for the function of the domain and will help to guide future studies of the LytR-CpsA-Psr proteins.

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.

Frequency of inhibitors of daphnid trypsin in the widely distributed cyanobacterial genus Planktothrix

DEFF Research Database (Denmark)

Rohrlack, T.; Christoffersen, K.; Friberg-Jensen, U.

2005-01-01

on the frequency of such compounds in the widely distributed cyanobacterial genus Planktothrix. Of the 89 Planktothrix strains analysed, about 70% produced inhibitors of daphnid trypsin. The strains tested positive represented three common Planktothrix species and were isolated from diverse localities...

The dynamic nature and territory of transcriptional machinery in the bacterial chromosome

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Ding Jun Jin

2015-05-01

Full Text Available Our knowledge of the regulation of genes involved in bacterial growth and stress responses is extensive; however, we have only recently begun to understand how environmental cues influence the dynamic, three-dimensional distribution of RNA polymerase (RNAP in Escherichia coli on the level of single cell, using wide-field fluorescence microscopy and state-of-the-art imaging techniques. Live-cell imaging using either an agarose-embedding procedure or a microfluidic system further underscores the dynamic nature of the distribution of RNAP in response to changes in the environment. A general agreement between live-cell and fixed-cell images has validated the formaldehyde-fixing procedure, which is a technical breakthrough in the study of the cell biology of RNAP. In this review we use a systems biology perspective to summarize the advances in the cell biology of RNAP in E. coli, including the discoveries of the bacterial nucleolus, the spatial compartmentalization of the transcription machinery at the periphery of the nucleoid, and the segregation of the chromosome territories for the two major cellular functions of transcription and replication in fast-growing cells. Our understanding of the coupling of transcription and bacterial chromosome (or nucleoid structure is also summarized. Using E. coli as a simple model system, co-imaging of RNAP with DNA and other factors during growth and stress responses will continue to be a useful tool for studying bacterial growth and adaptation in changing environment.

Eukaryote-wide sequence analysis of mitochondrial β-barrel outer membrane proteins

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Fujita Naoya

2011-01-01

Full Text Available Abstract Background The outer membranes of mitochondria are thought to be homologous to the outer membranes of Gram negative bacteria, which contain 100's of distinct families of β-barrel membrane proteins (BOMPs often forming channels for transport of nutrients or drugs. However, only four families of mitochondrial BOMPs (MBOMPs have been confirmed to date. Although estimates as high as 100 have been made in the past, the number of yet undiscovered MBOMPs is an open question. Fortunately, the recent discovery of a membrane integration signal (the β-signal for MBOMPs gave us an opportunity to look for undiscovered MBOMPs. Results We present the results of a comprehensive survey of eukaryotic protein sequences intended to identify new MBOMPs. Our search employs recent results on β-signals as well as structural information and a novel BOMP predictor trained on both bacterial and mitochondrial BOMPs. Our principal finding is circumstantial evidence suggesting that few MBOMPs remain to be discovered, if one assumes that, like known MBOMPs, novel MBOMPs will be monomeric and β-signal dependent. In addition to this, our analysis of MBOMP homologs reveals some exceptions to the current model of the β-signal, but confirms its consistent presence in the C-terminal region of MBOMP proteins. We also report a β-signal independent search for MBOMPs against the yeast and Arabidopsis proteomes. We find no good candidates MBOMPs in yeast but the Arabidopsis results are less conclusive. Conclusions Our results suggest there are no remaining MBOMPs left to discover in yeast; and if one assumes all MBOMPs are β-signal dependent, few MBOMP families remain undiscovered in any sequenced organism.

The Measurement and Interpretation of Dietary Protein Distribution During a Rugby Preseason.

Science.gov (United States)

MacKenzie, Kristen; Slater, Gary; King, Neil; Byrne, Nuala

2015-08-01

Evidence suggests that increasing protein distribution may be desirable to promote muscle protein synthesis (MPS) in combination with resistance exercise. However, there is a threshold above which additional protein consumption has limited benefit for MPS and may promote protein loss due to increased oxidation. This study aimed to measure daily protein intake and protein distribution in a cohort of rugby players. Twenty-five developing elite rugby union athletes (20.5 ± 2.3 years, 100.2 ± 13.3 kg, 184.4 ± 7.4 cm) were assessed at the start and end of a rugby preseason. Using a 7-day food diary the reported daily protein intake was 2.2 ± 0.7 g · kg · day(-1) which exceeds daily recommendations. The reported carbohydrate intake was 3.6 ± 1.3 g · kg · day(-1) which may reflect a suboptimal intake or dietary underreporting. In general, the rugby athletes were regularly consuming more than 20 g of protein; 3.8 ± 0.9 times per day (68 ± 18% of eating occasions). In addition to documenting current dietary intakes, an excess protein estimation score was calculated to determine how frequently the rugby athletes consumed protein above a known effective dose with a margin of error. 2.0 ± 0.9 eating occasions contained protein in excess of doses (20 g) known to promote MPS. Therefore, it is currently unclear whether the consumption of regular large doses of protein will benefit rugby athletes via increasing protein distribution, or whether high protein intakes may have unintended effects including a reduction in carbohydrate and/or energy intake.

A genome-wide association study of seed protein and oil content in soybean.

Science.gov (United States)

Hwang, Eun-Young; Song, Qijian; Jia, Gaofeng; Specht, James E; Hyten, David L; Costa, Jose; Cregan, Perry B

2014-01-02

Association analysis is an alternative to conventional family-based methods to detect the location of gene(s) or quantitative trait loci (QTL) and provides relatively high resolution in terms of defining the genome position of a gene or QTL. Seed protein and oil concentration are quantitative traits which are determined by the interaction among many genes with small to moderate genetic effects and their interaction with the environment. In this study, a genome-wide association study (GWAS) was performed to identify quantitative trait loci (QTL) controlling seed protein and oil concentration in 298 soybean germplasm accessions exhibiting a wide range of seed protein and oil content. A total of 55,159 single nucleotide polymorphisms (SNPs) were genotyped using various methods including Illumina Infinium and GoldenGate assays and 31,954 markers with minor allele frequency >0.10 were used to estimate linkage disequilibrium (LD) in heterochromatic and euchromatic regions. In euchromatic regions, the mean LD (r2) rapidly declined to 0.2 within 360 Kbp, whereas the mean LD declined to 0.2 at 9,600 Kbp in heterochromatic regions. The GWAS results identified 40 SNPs in 17 different genomic regions significantly associated with seed protein. Of these, the five SNPs with the highest associations and seven adjacent SNPs were located in the 27.6-30.0 Mbp region of Gm20. A major seed protein QTL has been previously mapped to the same location and potential candidate genes have recently been identified in this region. The GWAS results also detected 25 SNPs in 13 different genomic regions associated with seed oil. Of these markers, seven SNPs had a significant association with both protein and oil. This research indicated that GWAS not only identified most of the previously reported QTL controlling seed protein and oil, but also resulted in narrower genomic regions than the regions reported as containing these QTL. The narrower GWAS-defined genome regions will allow more precise

Effects of Acids, Bases, and Heteroatoms on Proximal Radial Distribution Functions for Proteins.

Science.gov (United States)

Nguyen, Bao Linh; Pettitt, B Montgomery

2015-04-14

The proximal distribution of water around proteins is a convenient method of quantifying solvation. We consider the effect of charged and sulfur-containing amino acid side-chain atoms on the proximal radial distribution function (pRDF) of water molecules around proteins using side-chain analogs. The pRDF represents the relative probability of finding any solvent molecule at a distance from the closest or surface perpendicular protein atom. We consider the near-neighbor distribution. Previously, pRDFs were shown to be universal descriptors of the water molecules around C, N, and O atom types across hundreds of globular proteins. Using averaged pRDFs, a solvent density around any globular protein can be reconstructed with controllable relative error. Solvent reconstruction using the additional information from charged amino acid side-chain atom types from both small models and protein averages reveals the effects of surface charge distribution on solvent density and improves the reconstruction errors relative to simulation. Solvent density reconstructions from the small-molecule models are as effective and less computationally demanding than reconstructions from full macromolecular models in reproducing preferred hydration sites and solvent density fluctuations.

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.

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

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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.

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.

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

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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.

Distribution, transition and thermodynamic stability of protein conformations in the denaturant-induced unfolding of proteins.

Science.gov (United States)

Bian, Liujiao; Ji, Xu

2014-01-01

Extensive and intensive studies on the unfolding of proteins require appropriate theoretical model and parameter to clearly illustrate the feature and characteristic of the unfolding system. Over the past several decades, four approaches have been proposed to describe the interaction between proteins and denaturants, but some ambiguity and deviations usually occur in the explanation of the experimental data. In this work, a theoretical model was presented to show the dependency of the residual activity ratio of the proteins on the molar denaturant concentration. Through the characteristic unfolding parameters ki and Δmi in this model, the distribution, transition and thermodynamic stability of protein conformations during the unfolding process can be quantitatively described. This model was tested with the two-state unfolding of bovine heart cytochrome c and the three-state unfolding of hen egg white lysozyme induced by both guanidine hydrochloride and urea, the four-state unfolding of bovine carbonic anhydrase b induced by guanidine hydrochloride and the unfolding of some other proteins induced by denaturants. The results illustrated that this model could be used accurately to reveal the distribution and transition of protein conformations in the presence of different concentrations of denaturants and to evaluate the unfolding tendency and thermodynamic stability of different conformations. In most denaturant-induced unfolding of proteins, the unfolding became increasingly hard in next transition step and the proteins became more unstable as they attained next successive stable conformation. This work presents a useful method for people to study the unfolding of proteins and may be used to describe the unfolding and refolding of other biopolymers induced by denaturants, inducers, etc.

Correlation of gene expression and protein production rate - a system wide study

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Arvas Mikko

2011-12-01

Full Text Available Abstract Background Growth rate is a major determinant of intracellular function. However its effects can only be properly dissected with technically demanding chemostat cultivations in which it can be controlled. Recent work on Saccharomyces cerevisiae chemostat cultivations provided the first analysis on genome wide effects of growth rate. In this work we study the filamentous fungus Trichoderma reesei (Hypocrea jecorina that is an industrial protein production host known for its exceptional protein secretion capability. Interestingly, it exhibits a low growth rate protein production phenotype. Results We have used transcriptomics and proteomics to study the effect of growth rate and cell density on protein production in chemostat cultivations of T. reesei. Use of chemostat allowed control of growth rate and exact estimation of the extracellular specific protein production rate (SPPR. We find that major biosynthetic activities are all negatively correlated with SPPR. We also find that expression of many genes of secreted proteins and secondary metabolism, as well as various lineage specific, mostly unknown genes are positively correlated with SPPR. Finally, we enumerate possible regulators and regulatory mechanisms, arising from the data, for this response. Conclusions Based on these results it appears that in low growth rate protein production energy is very efficiently used primarly for protein production. Also, we propose that flux through early glycolysis or the TCA cycle is a more fundamental determining factor than growth rate for low growth rate protein production and we propose a novel eukaryotic response to this i.e. the lineage specific response (LSR.

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.

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

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

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

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

Distribution of protein and RNA in the 30S ribosomal subunit

International Nuclear Information System (INIS)

Ramakrishnan, V.

1986-01-01

In Escherichia coli, the small ribosomal subunit has a sedimentation coefficient of 30S, and consists of a 16S RNA molecule of 1541 nucleotides complexed with 21 proteins. Over the last few years, a controversy has emerged regarding the spatial distribution of RNA and protein in the 30S subunit. Contrast variation with neutron scattering was used to suggest that the RNA was located in a central core of the subunit and the proteins mainly in the periphery, with virtually no separation between the centers of mass of protein and RNA. However, these findings are incompatible with the results of efforts to locate individual ribosomal proteins by immune electron microscopy and triangulation with interprotein distance measurements. The conflict between these two views is resolved in this report of small-angle neutron scattering measurements on 30S subunits with and without protein S1, and on subunits reconstituted from deuterated 16S RNA and unlabeled proteins. The results show that (i) the proteins and RNA are intermingled, with neither component dominating at the core or the periphery, and (ii) the spatial distribution of protein and RNA is asymmetrical, with a separation between their centers of mass of about 25 angstroms

Genome-Wide Identification and Analysis of Genes Encoding PHD-Finger Protein in Tomato

International Nuclear Information System (INIS)

Hayat, S.; Cheng, Z.; Chen, X.

2016-01-01

The PHD-finger proteins are conserved in eukaryotic organisms and are involved in a variety of important functions in different biological processes in plants. However, the function of PHD fingers are poorly known in tomato (Solanum lycopersicum L.). In current study, we identified 45 putative genes coding Phd finger protein in tomato distributed on 11 chromosomes except for chromosome 8. Some of the genes encode other conserved key domains besides Phd-finger. Phylogenetic analysis of these 45 proteins resulted in seven clusters. Most Phd finger proteins were predicted to PML body location. These PHD-finger genes displayed differential expression either in various organs, at different development stages and under stresses in tomato. Our study provides the first systematic analysis of PHD-finger genes and proteins in tomato. This preliminary study provides a very useful reference information for Phd-finger proteins in tomato. They will be helpful for cloning and functional study of tomato PHD-finger genes. (author)

Towards Spectral Library-free MALDI-TOF MS Bacterial Identification.

Science.gov (United States)

Cheng, Ding; Qiao, Liang; Horvatovich, Péter

2018-05-11

Bacterial identification is of great importance in clinical diagnosis, environmental monitoring and food safety control. Among various strategies, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has drawn significant interests, and has been clinically used. Nevertheless, current bioinformatics solutions use spectral libraries for the identification of bacterial strains. Spectral library generation requires acquisition of MALDI-TOF spectra from monoculture bacterial colonies, which is time-consuming and not possible for many species and strains. We propose a strategy for bacterial typing by MALDI-TOF using protein sequences from public database, i.e. UniProt. Ten genes were identified to encode proteins most often observed by MALD-TOF from bacteria through 500 times repeated a 10-fold double cross-validation procedure, using 403 MALDI-TOF spectra corresponding to 14 genera, 81 species and 403 strains, and the protein sequences of 1276 species in UniProt. The 10 genes were then used to annotate peaks on MALDI-TOF spectra of bacteria for bacterial identification. With the approach, bacteria can be identified at the genus level by searching against a database containing the protein sequences of 42 genera of bacteria from UniProt. Our approach identified 84.1% of the 403 spectra correctly at the genus level. Source code of the algorithm is available at https://github.com/dipcarbon/BacteriaMSLF.

Relative Roles of Deterministic and Stochastic Processes in Driving the Vertical Distribution of Bacterial Communities in a Permafrost Core from the Qinghai-Tibet Plateau, China.

Science.gov (United States)

Hu, Weigang; Zhang, Qi; Tian, Tian; Li, Dingyao; Cheng, Gang; Mu, Jing; Wu, Qingbai; Niu, Fujun; Stegen, James C; An, Lizhe; Feng, Huyuan

2015-01-01

Understanding the processes that influence the structure of biotic communities is one of the major ecological topics, and both stochastic and deterministic processes are expected to be at work simultaneously in most communities. Here, we investigated the vertical distribution patterns of bacterial communities in a 10-m-long soil core taken within permafrost of the Qinghai-Tibet Plateau. To get a better understanding of the forces that govern these patterns, we examined the diversity and structure of bacterial communities, and the change in community composition along the vertical distance (spatial turnover) from both taxonomic and phylogenetic perspectives. Measures of taxonomic and phylogenetic beta diversity revealed that bacterial community composition changed continuously along the soil core, and showed a vertical distance-decay relationship. Multiple stepwise regression analysis suggested that bacterial alpha diversity and phylogenetic structure were strongly correlated with soil conductivity and pH but weakly correlated with depth. There was evidence that deterministic and stochastic processes collectively drived bacterial vertically-structured pattern. Bacterial communities in five soil horizons (two originated from the active layer and three from permafrost) of the permafrost core were phylogenetically random, indicator of stochastic processes. However, we found a stronger effect of deterministic processes related to soil pH, conductivity, and organic carbon content that were structuring the bacterial communities. We therefore conclude that the vertical distribution of bacterial communities was governed primarily by deterministic ecological selection, although stochastic processes were also at work. Furthermore, the strong impact of environmental conditions (for example, soil physicochemical parameters and seasonal freeze-thaw cycles) on these communities underlines the sensitivity of permafrost microorganisms to climate change and potentially subsequent

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.

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.

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

Extensive genomic plasticity in Pseudomonas aeruginosa revealed by identification and distribution studies of novel genes among clinical isolates.

Science.gov (United States)

Shen, Kai; Sayeed, Sameera; Antalis, Patricia; Gladitz, John; Ahmed, Azad; Dice, Bethany; Janto, Benjamin; Dopico, Richard; Keefe, Randy; Hayes, Jay; Johnson, Sandra; Yu, Sujun; Ehrlich, Nathan; Jocz, Jennifer; Kropp, Laura; Wong, Ray; Wadowsky, Robert M; Slifkin, Malcolm; Preston, Robert A; Erdos, Geza; Post, J Christopher; Ehrlich, Garth D; Hu, Fen Z

2006-09-01

The distributed genome hypothesis (DGH) states that each strain within a bacterial species receives a unique distribution of genes from a population-based supragenome that is many times larger than the genome of any given strain. The observations that natural infecting populations are often polyclonal and that most chronic bacterial pathogens have highly developed mechanisms for horizontal gene transfer suggested the DGH and provided the means and the mechanisms to explain how chronic infections persist in the face of a mammalian host's adaptive defense mechanisms. Having previously established the validity of the DGH for obligate pathogens, we wished to evaluate its applicability to an opportunistic bacterial pathogen. This was accomplished by construction and analysis of a highly redundant pooled genomic library containing approximately 216,000 functional clones that was constructed from 12 low-passage clinical isolates of Pseudomonas aeruginosa, 6 otorrheic isolates and 6 from other body sites. Sequence analysis of 3,214 randomly picked clones (mean insert size, approximately 1.4 kb) from this library demonstrated that 348 (10.8%) of the clones were unique with respect to all genomic sequences of the P. aeruginosa prototype strain, PAO1. Hypothetical translations of the open reading frames within these unique sequences demonstrated protein homologies to a number of bacterial virulence factors and other proteins not previously identified in P. aeruginosa. PCR and reverse transcription-PCR-based assays were performed to analyze the distribution and expression patterns of a 70-open reading frame subset of these sequences among 11 of the clinical strains. These sequences were unevenly distributed among the clinical isolates, with nearly half (34/70) of the novel sequences being present in only one or two of the individual strains. Expression profiling revealed that a vast majority of these sequences are expressed, strongly suggesting they encode functional proteins.

Bacterial composition in a metropolitan drinking water distribution system utilizing different source waters.

Science.gov (United States)

Gomez-Alvarez, Vicente; Humrighouse, Ben W; Revetta, Randy P; Santo Domingo, Jorge W

2015-03-01

We investigated the bacterial composition of water samples from two service areas within a drinking water distribution system (DWDS), each associated with a different primary source of water (groundwater, GW; surface water, SW) and different treatment process. Community analysis based on 16S rRNA gene clone libraries indicated that Actinobacteria (Mycobacterium spp.) and α-Proteobacteria represented nearly 43 and 38% of the total sequences, respectively. Sequences closely related to Legionella, Pseudomonas, and Vibrio spp. were also identified. In spite of the high number of sequences (71%) shared in both areas, multivariable analysis revealed significant differences between the GW and SW areas. While the dominant phylotypes where not significantly contributing in the ordination of samples, the populations associated with the core of phylotypes (1-10% in each sample) significantly contributed to the differences between both service areas. Diversity indices indicate that the microbial community inhabiting the SW area is more diverse and contains more distantly related species coexisting with local assemblages as compared with the GW area. The bacterial community structure of SW and GW service areas were dissimilar, suggesting that their respective source water and/or water quality parameters shaped by the treatment processes may contribute to the differences in community structure observed.

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.)

The mechanical behavior of metal alloys with grain size distribution in a wide range of strain rates

Science.gov (United States)

Skripnyak, V. A.; Skripnyak, V. V.; Skripnyak, E. G.

2017-12-01

The paper discusses a multiscale simulation approach for the construction of grain structure of metals and alloys, providing high tensile strength with ductility. This work compares the mechanical behavior of light alloys and the influence of the grain size distribution in a wide range of strain rates. The influence of the grain size distribution on the inelastic deformation and fracture of aluminium and magnesium alloys is investigated by computer simulations in a wide range of strain rates. It is shown that the yield stress depends on the logarithm of the normalized strain rate for light alloys with a bimodal grain distribution and coarse-grained structure.

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

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

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.

An automated wide-field time-gated optically sectioning fluorescence lifetime imaging multiwell plate reader for high-content analysis of protein-protein interactions

Science.gov (United States)

Alibhai, Dominic; Kumar, Sunil; Kelly, Douglas; Warren, Sean; Alexandrov, Yuriy; Munro, Ian; McGinty, James; Talbot, Clifford; Murray, Edward J.; Stuhmeier, Frank; Neil, Mark A. A.; Dunsby, Chris; French, Paul M. W.

2011-03-01

We describe an optically-sectioned FLIM multiwell plate reader that combines Nipkow microscopy with wide-field time-gated FLIM, and its application to high content analysis of FRET. The system acquires sectioned FLIM images in fluorescent protein. It has been applied to study the formation of immature HIV virus like particles (VLPs) in live cells by monitoring Gag-Gag protein interactions using FLIM FRET of HIV-1 Gag transfected with CFP or YFP. VLP formation results in FRET between closely packed Gag proteins, as confirmed by our FLIM analysis that includes automatic image segmentation.

Genome-Wide Association Studies of the Human Gut Microbiota.

Directory of Open Access Journals (Sweden)

Emily R Davenport

Full Text Available The bacterial composition of the human fecal microbiome is influenced by many lifestyle factors, notably diet. It is less clear, however, what role host genetics plays in dictating the composition of bacteria living in the gut. In this study, we examined the association of ~200K host genotypes with the relative abundance of fecal bacterial taxa in a founder population, the Hutterites, during two seasons (n = 91 summer, n = 93 winter, n = 57 individuals collected in both. These individuals live and eat communally, minimizing variation due to environmental exposures, including diet, which could potentially mask small genetic effects. Using a GWAS approach that takes into account the relatedness between subjects, we identified at least 8 bacterial taxa whose abundances were associated with single nucleotide polymorphisms in the host genome in each season (at genome-wide FDR of 20%. For example, we identified an association between a taxon known to affect obesity (genus Akkermansia and a variant near PLD1, a gene previously associated with body mass index. Moreover, we replicate a previously reported association from a quantitative trait locus (QTL mapping study of fecal microbiome abundance in mice (genus Lactococcus, rs3747113, P = 3.13 x 10-7. Finally, based on the significance distribution of the associated microbiome QTLs in our study with respect to chromatin accessibility profiles, we identified tissues in which host genetic variation may be acting to influence bacterial abundance in the gut.

New Paenibacillus larvae bacterial isolates from honey bee colonies infected with American foulbrood disease in Egypt.

Science.gov (United States)

Masry, Saad Hamdy Daif; Kabeil, Sanaa Soliman; Hafez, Elsayed Elsayed

2014-03-04

The American foulbrood disease is widely distributed all over the world and causes a serious problem for the honeybee industry. Different infected larvae were collected from different apiaries, ground in phosphate saline buffer (PSB) and bacterial isolation was carried out on nutrient agar medium. Different colonies were observed and were characterized biologically. Two bacterial isolates (SH11 and SH33) were subjected to molecular identification using 16S rRNA gene and the sequence analysis revealed that the two isolates are Paenibacillus larvae with identity not exceeding 83%. The DNA sequence alignment between the other P. larvae bacterial strains and the two identified bacterial isolates showed that all the examined bacterial strains have the same ancestor, i.e. they have the same origin. The SH33 isolate was closely related to the P. larvae isolated from Germany, whereas the isolate SH11 was close to the P. larvae isolated from India. The phylogenetic tree constructed for 20 different Bacillus sp. and the two isolates SH11 and SH33 demonstrated that the two isolates are Bacillus sp. and they are new isolates. The bacterial isolates will be subjected to more tests for more confirmations.

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.

Genome-wide profiling of DNA-binding proteins using barcode-based multiplex Solexa sequencing.

Science.gov (United States)

Raghav, Sunil Kumar; Deplancke, Bart

2012-01-01

Chromatin immunoprecipitation (ChIP) is a commonly used technique to detect the in vivo binding of proteins to DNA. ChIP is now routinely paired to microarray analysis (ChIP-chip) or next-generation sequencing (ChIP-Seq) to profile the DNA occupancy of proteins of interest on a genome-wide level. Because ChIP-chip introduces several biases, most notably due to the use of a fixed number of probes, ChIP-Seq has quickly become the method of choice as, depending on the sequencing depth, it is more sensitive, quantitative, and provides a greater binding site location resolution. With the ever increasing number of reads that can be generated per sequencing run, it has now become possible to analyze several samples simultaneously while maintaining sufficient sequence coverage, thus significantly reducing the cost per ChIP-Seq experiment. In this chapter, we provide a step-by-step guide on how to perform multiplexed ChIP-Seq analyses. As a proof-of-concept, we focus on the genome-wide profiling of RNA Polymerase II as measuring its DNA occupancy at different stages of any biological process can provide insights into the gene regulatory mechanisms involved. However, the protocol can also be used to perform multiplexed ChIP-Seq analyses of other DNA-binding proteins such as chromatin modifiers and transcription factors.

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

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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.

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

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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

Spatial distribution of archaeal and bacterial ammonia oxidizers in the littoral buffer zone of a nitrogen-rich lake.

Science.gov (United States)

Wang, Yu; Zhu, Guibing; Ye, Lei; Feng, Xiaojuan; Op den Camp, Huub J M; Yin, Chengqing

2012-01-01

The spatial distribution and diversity of archaeal and bacterial ammonia oxidizers (AOA and AOB) were evaluated targeting amoA genes in the gradient of a littoral buffer zone which has been identified as a hot spot for N cycling. Here we found high spatial heterogeneity in the nitrification rate and abundance of ammonia oxidizers in the five sampling sites. The bacterial amoA gene was numerically dominant in most of the surface soil but decreased dramatically in deep layers. Higher nitrification potentials were detected in two sites near the land/water interface at 4.4-6.1 microg NO(2-)-N/(g dry weight soil x hr), while only 1.0-1.7 microg NO(2-)-N/(g dry weight soil x hr) was measured at other sites. The potential nitrification rates were proportional to the amoA gene abundance for AOB, but with no significant correlation with AOA. The NH4+ concentration was the most determinative parameter for the abundance of AOB and potential nitrification rates in this study. Higher richness in the surface layer was found in the analysis of biodiversity. Phylogenetic analysis revealed that most of the bacterial amoA sequences in surface soil were affiliated with the genus of Nitrosopira while the archaeal sequences were almost equally affiliated with Candidatus 'Nitrososphaera gargensis' and Candidatus 'Nitrosocaldus yellowstonii'. The spatial distribution of AOA and AOB indicated that bacteria may play a more important role in nitrification in the littoral buffer zone of a N-rich lake.

Distribution of protein components of wheat from different regions

African Journals Online (AJOL)

kesiena

2012-06-07

Jun 7, 2012 ... The distribution of wheat protein components in different regions was researched to ..... properties of wheat gliadins II. effects on dynamic rheoligical ... fractions properties of wheat dough depending on molecular size and.

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.

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...

Genome-wide identification of Streptococcus pneumoniae genes essential for bacterial replication during experimental meningitis

DEFF Research Database (Denmark)

Molzen, T E; Burghout, P; Bootsma, H J

2010-01-01

Meningitis is the most serious of invasive infections caused by the Gram-positive bacterium Streptococcus pneumoniae. Vaccines protect only against a limited number of serotypes, and evolving bacterial resistance to antimicrobials impedes treatment. Further insight into the molecular pathogenesis...... as targets for future therapy and prevention of pneumococcal meningitis, since their mutants were attenuated in both models of infection as well as in competitive growth in human cerebrospinal fluid in vitro.......Meningitis is the most serious of invasive infections caused by the Gram-positive bacterium Streptococcus pneumoniae. Vaccines protect only against a limited number of serotypes, and evolving bacterial resistance to antimicrobials impedes treatment. Further insight into the molecular pathogenesis...... genes mutants of which had become attenuated or enriched, respectively, during infection. The results point to essential roles for capsular polysaccharides, nutrient uptake, and amino acid biosynthesis in bacterial replication during experimental meningitis. The GAF phenotype of a subset of identified...

Genome-wide RNAi screen identifies novel host proteins required for alphavirus entry.

Directory of Open Access Journals (Sweden)

Yaw Shin Ooi

Full Text Available The enveloped alphaviruses include important and emerging human pathogens such as Chikungunya virus and Eastern equine encephalitis virus. Alphaviruses enter cells by clathrin-mediated endocytosis, and exit by budding from the plasma membrane. While there has been considerable progress in defining the structure and function of the viral proteins, relatively little is known about the host factors involved in alphavirus infection. We used a genome-wide siRNA screen to identify host factors that promote or inhibit alphavirus infection in human cells. Fuzzy homologue (FUZ, a protein with reported roles in planar cell polarity and cilia biogenesis, was required for the clathrin-dependent internalization of both alphaviruses and the classical endocytic ligand transferrin. The tetraspanin membrane protein TSPAN9 was critical for the efficient fusion of low pH-triggered virus with the endosome membrane. FUZ and TSPAN9 were broadly required for infection by the alphaviruses Sindbis virus, Semliki Forest virus, and Chikungunya virus, but were not required by the structurally-related flavivirus Dengue virus. Our results highlight the unanticipated functions of FUZ and TSPAN9 in distinct steps of alphavirus entry and suggest novel host proteins that may serve as targets for antiviral therapy.

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.

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

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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

Genome-Wide Characterization and Expression Analysis of Major Intrinsic Proteins during Abiotic and Biotic Stresses in Sweet Orange (Citrus sinensis L. Osb.).

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Martins, Cristina de Paula Santos; Pedrosa, Andresa Muniz; Du, Dongliang; Gonçalves, Luana Pereira; Yu, Qibin; Gmitter, Frederick G; Costa, Marcio Gilberto Cardoso

2015-01-01

The family of aquaporins (AQPs), or major intrinsic proteins (MIPs), includes integral membrane proteins that function as transmembrane channels for water and other small molecules of physiological significance. MIPs are classified into five subfamilies in higher plants, including plasma membrane (PIPs), tonoplast (TIPs), NOD26-like (NIPs), small basic (SIPs) and unclassified X (XIPs) intrinsic proteins. This study reports a genome-wide survey of MIP encoding genes in sweet orange (Citrus sinensis L. Osb.), the most widely cultivated Citrus spp. A total of 34 different genes encoding C. sinensis MIPs (CsMIPs) were identified and assigned into five subfamilies (CsPIPs, CsTIPs, CsNIPs, CsSIPs and CsXIPs) based on sequence analysis and also on their phylogenetic relationships with clearly classified MIPs of Arabidopsis thaliana. Analysis of key amino acid residues allowed the assessment of the substrate specificity of each CsMIP. Gene structure analysis revealed that the CsMIPs possess an exon-intron organization that is highly conserved within each subfamily. CsMIP loci were precisely mapped on every sweet orange chromosome, indicating a wide distribution of the gene family in the sweet orange genome. Investigation of their expression patterns in different tissues and upon drought and salt stress treatments, as well as with 'Candidatus Liberibacter asiaticus' infection, revealed a tissue-specific and coordinated regulation of the different CsMIP isoforms, consistent with the organization of the stress-responsive cis-acting regulatory elements observed in their promoter regions. A special role in regulating the flow of water and nutrients is proposed for CsTIPs and CsXIPs during drought stress, and for most CsMIPs during salt stress and the development of HLB disease. These results provide a valuable reference for further exploration of the CsMIPs functions and applications to the genetic improvement of both abiotic and biotic stress tolerance in citrus.

Genome-Wide Characterization and Expression Analysis of Major Intrinsic Proteins during Abiotic and Biotic Stresses in Sweet Orange (Citrus sinensis L. Osb..

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Cristina de Paula Santos Martins

Full Text Available The family of aquaporins (AQPs, or major intrinsic proteins (MIPs, includes integral membrane proteins that function as transmembrane channels for water and other small molecules of physiological significance. MIPs are classified into five subfamilies in higher plants, including plasma membrane (PIPs, tonoplast (TIPs, NOD26-like (NIPs, small basic (SIPs and unclassified X (XIPs intrinsic proteins. This study reports a genome-wide survey of MIP encoding genes in sweet orange (Citrus sinensis L. Osb., the most widely cultivated Citrus spp. A total of 34 different genes encoding C. sinensis MIPs (CsMIPs were identified and assigned into five subfamilies (CsPIPs, CsTIPs, CsNIPs, CsSIPs and CsXIPs based on sequence analysis and also on their phylogenetic relationships with clearly classified MIPs of Arabidopsis thaliana. Analysis of key amino acid residues allowed the assessment of the substrate specificity of each CsMIP. Gene structure analysis revealed that the CsMIPs possess an exon-intron organization that is highly conserved within each subfamily. CsMIP loci were precisely mapped on every sweet orange chromosome, indicating a wide distribution of the gene family in the sweet orange genome. Investigation of their expression patterns in different tissues and upon drought and salt stress treatments, as well as with 'Candidatus Liberibacter asiaticus' infection, revealed a tissue-specific and coordinated regulation of the different CsMIP isoforms, consistent with the organization of the stress-responsive cis-acting regulatory elements observed in their promoter regions. A special role in regulating the flow of water and nutrients is proposed for CsTIPs and CsXIPs during drought stress, and for most CsMIPs during salt stress and the development of HLB disease. These results provide a valuable reference for further exploration of the CsMIPs functions and applications to the genetic improvement of both abiotic and biotic stress tolerance in citrus.

Exploring bacterial outer membrane barrier to combat bad bugs

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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

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

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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

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

Influence of hydraulic regimes on bacterial community structure and composition in an experimental drinking water distribution system.

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Douterelo, I; Sharpe, R L; Boxall, J B

2013-02-01

Microbial biofilms formed on the inner-pipe surfaces of drinking water distribution systems (DWDS) can alter drinking water quality, particularly if they are mechanically detached from the pipe wall to the bulk water, such as due to changes in hydraulic conditions. Results are presented here from applying 454 pyrosequencing of the 16S ribosomal RNA (rRNA) gene to investigate the influence of different hydrological regimes on bacterial community structure and to study the potential mobilisation of material from the pipe walls to the network using a full scale, temperature-controlled experimental pipeline facility accurately representative of live DWDS. Analysis of pyrosequencing and water physico-chemical data showed that habitat type (water vs. biofilm) and hydraulic conditions influenced bacterial community structure and composition in our experimental DWDS. Bacterial community composition clearly differed between biofilms and bulk water samples. Gammaproteobacteria and Betaproteobacteria were the most abundant phyla in biofilms while Alphaproteobacteria was predominant in bulk water samples. This suggests that bacteria inhabiting biofilms, predominantly species belonging to genera Pseudomonas, Zooglea and Janthinobacterium, have an enhanced ability to express extracellular polymeric substances to adhere to surfaces and to favour co-aggregation between cells than those found in the bulk water. Highest species richness and diversity were detected in 28 days old biofilms with this being accentuated at highly varied flow conditions. Flushing altered the pipe-wall bacterial community structure but did not completely remove bacteria from the pipe walls, particularly under highly varied flow conditions, suggesting that under these conditions more compact biofilms were generated. This research brings new knowledge regarding the influence of different hydraulic regimes on the composition and structure of bacterial communities within DWDS and the implication that this

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.

Evaluation of protein adsorption onto a polyurethane nanofiber surface having different segment distributions

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Morita, Yuko; Koizumi, Gaku [Frontier Fiber Technology and Science, Graduate School of Engineering, University of Fukui (Japan); Sakamoto, Hiroaki, E-mail: hi-saka@u-fukui.ac.jp [Tenure-Track Program for Innovative Research, University of Fukui (Japan); Suye, Shin-ichiro [Frontier Fiber Technology and Science, Graduate School of Engineering, University of Fukui (Japan)

2017-02-01

Electrospinning is well known to be an effective method for fabricating polymeric nanofibers with a diameter of several hundred nanometers. Recently, the molecular-level orientation within nanofibers has attracted particular attention. Previously, we used atomic force microscopy to visualize the phase separation between soft and hard segments of a polyurethane (PU) nanofiber surface prepared by electrospinning. The unstretched PU nanofibers exhibited irregularly distributed hard segments, whereas hard segments of stretched nanofibers prepared with a high-speed collector exhibited periodic structures along the long-axis direction. PU was originally used to inhibit protein adsorption, but because the surface segment distribution was changed in the stretched nanofiber, here, we hypothesized that the protein adsorption property on the stretched nanofiber might be affected. We investigated protein adsorption onto PU nanofibers to elucidate the effects of segment distribution on the surface properties of PU nanofibers. The amount of adsorbed protein on stretched PU nanofibers was increased compared with that of unstretched nanofibers. These results indicate that the hard segment alignment on stretched PU nanofibers mediated protein adsorption. It is therefore expected that the amount of protein adsorption can be controlled by rotation of the collector. - Highlights: • The hard segments of stretched PU nanofibers exhibit periodic structures. • The adsorbed protein on stretched PU nanofibers was increased compared with PU film. • The hard segment alignment on stretched PU nanofibers mediated protein adsorption.

Distribution and uses of legume DNA clone resources

International Nuclear Information System (INIS)

Young, N.D.

2001-01-01

Since 1990, my lab has developed and distributed various DNA clone resources for the legumes. In the first several years, the focus was on members of the tropical genus, Vigna, including the widely cultivated species, mungbean (V. radiata) and cowpea (V. unguiculata). Both of these grain legumes play key roles in agriculture in developing countries of Asia (mungbean) and Africa (cowpea). Moreover, because there is substantial genome conservation among legumes, these genetic resources have also been utilized by a wide range of researchers in other crop species. In 1997, my lab began to focus on the development and distribution of a new generation of DNA clone resources; Bacterial Artificial Chromosomes (BAC). A library of these clones was constructed in soybean (Glycine max) the most important legume species worldwide in terms of economic value. Again, the library has become a valuable resource for the legume research community and has been widely used in studies of legume genomics. (author)

Spatial Patterning of Newly-Inserted Material during Bacterial Cell Growth

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Ursell, Tristan

2012-02-01

In the life cycle of a bacterium, rudimentary microscopy demonstrates that cell growth and elongation are essential characteristics of cellular reproduction. The peptidoglycan cell wall is the main load-bearing structure that determines both cell shape and overall size. However, simple imaging of cellular growth gives no indication of the spatial patterning nor mechanism by which material is being incorporated into the pre-existing cell wall. We employ a combination of high-resolution pulse-chase fluorescence microscopy, 3D computational microscopy, and detailed mechanistic simulations to explore how spatial patterning results in uniform growth and maintenance of cell shape. We show that growth is happening in discrete bursts randomly distributed over the cell surface, with a well-defined mean size and average rate. We further use these techniques to explore the effects of division and cell wall disrupting antibiotics, like cephalexin and A22, respectively, on the patterning of cell wall growth in E. coli. Finally, we explore the spatial correlation between presence of the bacterial actin-like cytoskeletal protein, MreB, and local cell wall growth. Together these techniques form a powerful method for exploring the detailed dynamics and involvement of antibiotics and cell wall-associated proteins in bacterial cell growth.[4pt] In collaboration with Kerwyn Huang, Stanford University.

Visualization and characterization of individual type III protein secretion machines in live bacteria.

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Zhang, Yongdeng; Lara-Tejero, María; Bewersdorf, Jörg; Galán, Jorge E

2017-06-06

Type III protein secretion machines have evolved to deliver bacterially encoded effector proteins into eukaryotic cells. Although electron microscopy has provided a detailed view of these machines in isolation or fixed samples, little is known about their organization in live bacteria. Here we report the visualization and characterization of the Salmonella type III secretion machine in live bacteria by 2D and 3D single-molecule switching superresolution microscopy. This approach provided access to transient components of this machine, which previously could not be analyzed. We determined the subcellular distribution of individual machines, the stoichiometry of the different components of this machine in situ, and the spatial distribution of the substrates of this machine before secretion. Furthermore, by visualizing this machine in Salmonella mutants we obtained major insights into the machine's assembly. This study bridges a major resolution gap in the visualization of this nanomachine and may serve as a paradigm for the examination of other bacterially encoded molecular machines.

A novel host-proteome signature for distinguishing between acute bacterial and viral infections.

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Kfir Oved

Full Text Available Bacterial and viral infections are often clinically indistinguishable, leading to inappropriate patient management and antibiotic misuse. Bacterial-induced host proteins such as procalcitonin, C-reactive protein (CRP, and Interleukin-6, are routinely used to support diagnosis of infection. However, their performance is negatively affected by inter-patient variability, including time from symptom onset, clinical syndrome, and pathogens. Our aim was to identify novel viral-induced host proteins that can complement bacterial-induced proteins to increase diagnostic accuracy. Initially, we conducted a bioinformatic screen to identify putative circulating host immune response proteins. The resulting 600 candidates were then quantitatively screened for diagnostic potential using blood samples from 1002 prospectively recruited patients with suspected acute infectious disease and controls with no apparent infection. For each patient, three independent physicians assigned a diagnosis based on comprehensive clinical and laboratory investigation including PCR for 21 pathogens yielding 319 bacterial, 334 viral, 112 control and 98 indeterminate diagnoses; 139 patients were excluded based on predetermined criteria. The best performing host-protein was TNF-related apoptosis-inducing ligand (TRAIL (area under the curve [AUC] of 0.89; 95% confidence interval [CI], 0.86 to 0.91, which was consistently up-regulated in viral infected patients. We further developed a multi-protein signature using logistic-regression on half of the patients and validated it on the remaining half. The signature with the highest precision included both viral- and bacterial-induced proteins: TRAIL, Interferon gamma-induced protein-10, and CRP (AUC of 0.94; 95% CI, 0.92 to 0.96. The signature was superior to any of the individual proteins (P<0.001, as well as routinely used clinical parameters and their combinations (P<0.001. It remained robust across different physiological systems

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.

Model to predict inhomogeneous protein-sugar distribution in powders prepared by spray drying

NARCIS (Netherlands)

Grasmeijer, Niels; Frijlink, Henderik W.; Hinrichs, Wouter L. J.

2016-01-01

A protein can be stabilized by spray drying an aqueous solution of the protein and a sugar, thereby incorporating the protein into a glassy sugar matrix. For optimal stability, the protein should be homogeneously distributed inside the sugar matrix. The aim of this study was to develop a model that

Genome-Wide Comparison of Magnaporthe Species Reveals a Host-Specific Pattern of Secretory Proteins and Transposable Elements.

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Meghana Deepak Shirke

Full Text Available Blast disease caused by the Magnaporthe species is a major factor affecting the productivity of rice, wheat and millets. This study was aimed at generating genomic information for rice and non-rice Magnaporthe isolates to understand the extent of genetic variation. We have sequenced the whole genome of the Magnaporthe isolates, infecting rice (leaf and neck, finger millet (leaf and neck, foxtail millet (leaf and buffel grass (leaf. Rice and finger millet isolates infecting both leaf and neck tissues were sequenced, since the damage and yield loss caused due to neck blast is much higher as compared to leaf blast. The genome-wide comparison was carried out to study the variability in gene content, candidate effectors, repeat element distribution, genes involved in carbohydrate metabolism and SNPs. The analysis of repeat element footprints revealed some genes such as naringenin, 2-oxoglutarate 3-dioxygenase being targeted by Pot2 and Occan, in isolates from different host species. Some repeat insertions were host-specific while other insertions were randomly shared between isolates. The distributions of repeat elements, secretory proteins, CAZymes and SNPs showed significant variation across host-specific lineages of Magnaporthe indicating an independent genome evolution orchestrated by multiple genomic factors.

Differences in bacterial composition between men's and women's restrooms and other common areas within a public building.

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Dobbler, Priscila Caroline Thiago; Laureano, Álvaro Macedo; Sarzi, Deise Schroder; Cañón, Ehidy Rocio Peña; Metz, Geferson Fernando; de Freitas, Anderson Santos; Takagaki, Beatriz Midori; D Oliveira, Cristiane Barbosa; Pylro, Victor Satler; Copetti, André Carlos; Victoria, Filipe; Redmile-Gordon, Marc; Morais, Daniel Kumazawa; Roesch, Luiz Fernando Wurdig

2018-04-01

Humans distribute a wide range of microorganisms around building interiors, and some of these are potentially pathogenic. Recent research established that humans are the main drivers of the indoor microbiome and up to now significant literature has been produced about this topic. Here we analyzed differences in bacterial composition between men's and women's restrooms and other common areas within the same public building. Bacterial DNA samples were collected from restrooms and halls of a three-floor building from the Federal University of Pampa, RS, Brazil. The bacterial community was characterized by amplification of the V4 region of the 16S rRNA gene and sequencing. Throughout all samples, the most abundant phylum was Proteobacteria, followed by Actinobacteria, Bacteroidetes and Firmicutes. Beta diversity metrics showed that the structure of the bacterial communities were different among the areas and floors tested, however, only 6-9% of the variation in bacterial communities was explained by the area and floors sampled. A few microorganisms showed significantly differential abundance between men's and women's restrooms, but in general, the bacterial communities from both places were very similar. Finally, significant differences among the microbial community profile from different floors were reported, suggesting that the type of use and occupant demographic within the building may directly influence bacterial dispersion and establishment.

Tissue distribution and developmental expression of type XVI collagen in the mouse.

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Lai, C H; Chu, M L

1996-04-01

The expression of a recently identified collagen, alpha 1 (XVI), in adult mouse tissue and developing mouse embryo was examined by immunohistochemistry and in situ hybridization. A polyclonal antiserum was raised against a recombinant fusion protein, which contained a segment of 161 amino acids in the N-terminal noncollagenous domain of the human alpha 1 (XVI) collagen. Immunoprecipitation of metabolically labelled human or mouse fibroblast cell lysates with this antibody revealed a major, bacterial collagenase sensitive polypeptide of approximately 210 kDa. The size agrees with the prediction from the full-length cDNA. Immunofluorescence examination of adult mouse tissues using the affinity purified antibody revealed a rather broad distribution of the protein. The heart, kidney, intestine, ovary, testis, eye, arterial walls and smooth muscles all exhibited significant levels of expression, while the skeletal muscle, lung and brain showed very restricted and low signals. During development, no significant expression of the mRNA or protein was observed in embryo of day 8 of gestation, but strong signals was detected in placental trophoblasts. Expression in embryos was detectable first after day 11 of gestation with weak positive signals appearing in the heart. In later stages of development, stronger RNA hybridizations were observed in a variety of tissues, particularly in atrial and ventricular walls of the developing heart, spinal root neural fibers and skin. These data demonstrate that type XVI collagen represents another collagenous component widely distributed in the extracellular matrix and may contribute to the structural integrity of various tissues.

CRISPR-mediated control of the bacterial initiation of replication.

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Wiktor, Jakub; Lesterlin, Christian; Sherratt, David J; Dekker, Cees

2016-05-05

Programmable control of the cell cycle has been shown to be a powerful tool in cell-biology studies. Here, we develop a novel system for controlling the bacterial cell cycle, based on binding of CRISPR/dCas9 to the origin-of-replication locus. Initiation of replication of bacterial chromosomes is accurately regulated by the DnaA protein, which promotes the unwinding of DNA at oriC We demonstrate that the binding of CRISPR/dCas9 to any position within origin or replication blocks the initiation of replication. Serial-dilution plating, single-cell fluorescence microscopy, and flow-cytometry experiments show that ongoing rounds of chromosome replication are finished upon CRISPR/dCas9 binding, but no new rounds are initiated. Upon arrest, cells stay metabolically active and accumulate cell mass. We find that elevating the temperature from 37 to 42°C releases the CRISR/dCas9 replication inhibition, and we use this feature to recover cells from the arrest. Our simple and robust method of controlling the bacterial cell cycle is a useful asset for synthetic biology and DNA-replication studies in particular. The inactivation of CRISPR/dCas9 binding at elevated temperatures may furthermore be of wide interest for CRISPR/Cas9 applications in genomic engineering. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

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

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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.

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

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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.

[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.

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.

Genome-wide association scan meta-analysis identifies three loci influencing adiposity and fat distribution

NARCIS (Netherlands)

C.M. Lindgren (Cecilia); I.M. Heid (Iris); J.C. Randall (Joshua); C. Lamina (Claudia); V. Steinthorsdottir (Valgerdur); L. Qi (Lu); E.K. Speliotes (Elizabeth); G. Thorleifsson (Gudmar); C.J. Willer (Cristen); B.M. Herrera (Blanca); A.U. Jackson (Anne); N. Lim (Noha); P. Scheet (Paul); N. Soranzo (Nicole); N. Amin (Najaf); Y.S. Aulchenko (Yurii); J.C. Chambers (John); A. Drong (Alexander); J. Luan; H.N. Lyon (Helen); F. Rivadeneira Ramirez (Fernando); S. Sanna (Serena); N.J. Timpson (Nicholas); M.C. Zillikens (Carola); H.Z. Jing; P. Almgren (Peter); S. Bandinelli (Stefania); A.J. Bennett (Amanda); R.N. Bergman (Richard); L.L. Bonnycastle (Lori); S. Bumpstead (Suzannah); S.J. Chanock (Stephen); L. Cherkas (Lynn); P.S. Chines (Peter); L. Coin (Lachlan); C. Cooper (Charles); G. Crawford (Gabe); A. Doering (Angela); A. Dominiczak (Anna); A.S.F. Doney (Alex); S. Ebrahim (Shanil); P. Elliott (Paul); M.R. Erdos (Michael); K. Estrada Gil (Karol); L. Ferrucci (Luigi); G. Fischer (Guido); N.G. Forouhi (Nita); C. Gieger (Christian); H. Grallert (Harald); C.J. Groves (Christopher); S.M. Grundy (Scott); C. Guiducci (Candace); D. Hadley (David); A. Hamsten (Anders); A.S. Havulinna (Aki); A. Hofman (Albert); R. Holle (Rolf); J.W. Holloway (John); T. Illig (Thomas); B. Isomaa (Bo); L.C. Jacobs (Leonie); K. Jameson (Karen); P. Jousilahti (Pekka); F. Karpe (Fredrik); J. Kuusisto (Johanna); J. Laitinen (Jaana); G.M. Lathrop (Mark); D.A. Lawlor (Debbie); M. Mangino (Massimo); W.L. McArdle (Wendy); T. Meitinger (Thomas); M.A. Morken (Mario); A.P. Morris (Andrew); P. Munroe (Patricia); N. Narisu (Narisu); A. Nordström (Anna); B.A. Oostra (Ben); C.N.A. Palmer (Colin); F. Payne (Felicity); J. Peden (John); I. Prokopenko (Inga); F. Renström (Frida); A. Ruokonen (Aimo); V. Salomaa (Veikko); M.S. Sandhu (Manjinder); L.J. Scott (Laura); A. Scuteri (Angelo); K. Silander (Kaisa); K. Song (Kijoung); X. Yuan (Xin); H.M. Stringham (Heather); A.J. Swift (Amy); T. Tuomi (Tiinamaija); M. Uda (Manuela); P. Vollenweider (Peter); G. Waeber (Gérard); C. Wallace (Chris); G.B. Walters (Bragi); M.N. Weedon (Michael); J.C.M. Witteman (Jacqueline); C. Zhang (Cuilin); M. Caulfield (Mark); F.S. Collins (Francis); G.D. Smith; I.N.M. Day (Ian); P.W. Franks (Paul); A.T. Hattersley (Andrew); F.B. Hu (Frank); M.-R. Jarvelin (Marjo-Riitta); A. Kong (Augustine); J.S. Kooner (Jaspal); M. Laakso (Markku); E. Lakatta (Edward); V. Mooser (Vincent); L. Peltonen (Leena Johanna); N.J. Samani (Nilesh); T.D. Spector (Timothy); D.P. Strachan (David); T. Tanaka (Toshiko); J. Tuomilehto (Jaakko); A.G. Uitterlinden (André); P. Tikka-Kleemola (Päivi); N.J. Wareham (Nick); H. Watkins (Hugh); D. Waterworth (Dawn); M. Boehnke (Michael); P. Deloukas (Panagiotis); L. Groop (Leif); D.J. Hunter (David); U. Thorsteinsdottir (Unnur); D. Schlessinger (David); H.E. Wichmann (Erich); T.M. Frayling (Timothy); G.R. Abecasis (Gonçalo); J.N. Hirschhorn (Joel); R.J.F. Loos (Ruth); J-A. Zwart (John-Anker); K.L. Mohlke (Karen); I.E. Barroso (Inês); M.I. McCarthy (Mark)

2009-01-01

textabstractTo identify genetic loci influencing central obesity and fat distribution, we performed a meta-analysis of 16 genome-wide association studies (GWAS, N = 38,580) informative for adult waist circumference (WC) and waist-hip ratio (WHR). We selected 26 SNPs for follow-up, for which the

A walk into the LuxR regulators of Actinobacteria: phylogenomic distribution and functional diversity.

Directory of Open Access Journals (Sweden)

Catarina Lopes Santos

Full Text Available LuxR regulators are a widely studied group of bacterial helix-turn-helix (HTH transcription factors involved in the regulation of many genes coding for important traits at an ecological and medical level. This regulatory family is particularly known by their involvement in quorum-sensing (QS mechanisms, i.e., in the bacterial ability to communicate through the synthesis and binding of molecular signals. However, these studies have been mainly focused on gram-negative organisms, and the presence of LuxR regulators in the gram-positive Actinobacteria phylum is still poorly explored. In this manuscript, the presence of LuxR regulators among Actinobacteria was assayed using a domain-based strategy. A total of 991 proteins having one LuxR domain were identified in 53 genome-sequenced actinobacterial species, of which 59% had an additional domain. In most cases (53% this domain was REC (receiver domain, suggesting that LuxR regulators in Actinobacteria may either function as single transcription factors or as part of two-component systems. The frequency, distribution and evolutionary stability of each of these sub-families of regulators was analyzed and contextualized regarding the ecological niche occupied by each organism. The results show that the presence of extra-domains in the LuxR-regulators was likely driven by a general need to physically uncouple the signal sensing from the signal transduction. Moreover, the total frequency of LuxR regulators was shown to be dependent on genetic, metabolic and ecological variables. Finally, the functional annotation of the LuxR regulators revealed that the bacterial ecological niche has biased the specialization of these proteins. In the case of pathogens, our results suggest that LuxR regulators can be involved in virulence and are therefore promising targets for future studies in the health-related biotechnology field.

A Walk into the LuxR Regulators of Actinobacteria: Phylogenomic Distribution and Functional Diversity

Science.gov (United States)

Santos, Catarina Lopes; Correia-Neves, Margarida; Moradas-Ferreira, Pedro; Mendes, Marta Vaz

2012-01-01

LuxR regulators are a widely studied group of bacterial helix-turn-helix (HTH) transcription factors involved in the regulation of many genes coding for important traits at an ecological and medical level. This regulatory family is particularly known by their involvement in quorum-sensing (QS) mechanisms, i.e., in the bacterial ability to communicate through the synthesis and binding of molecular signals. However, these studies have been mainly focused on Gram-negative organisms, and the presence of LuxR regulators in the Gram-positive Actinobacteria phylum is still poorly explored. In this manuscript, the presence of LuxR regulators among Actinobacteria was assayed using a domain-based strategy. A total of 991 proteins having one LuxR domain were identified in 53 genome-sequenced actinobacterial species, of which 59% had an additional domain. In most cases (53%) this domain was REC (receiver domain), suggesting that LuxR regulators in Actinobacteria may either function as single transcription factors or as part of two-component systems. The frequency, distribution and evolutionary stability of each of these sub-families of regulators was analyzed and contextualized regarding the ecological niche occupied by each organism. The results show that the presence of extra-domains in the LuxR-regulators was likely driven by a general need to physically uncouple the signal sensing from the signal transduction. Moreover, the total frequency of LuxR regulators was shown to be dependent on genetic, metabolic and ecological variables. Finally, the functional annotation of the LuxR regulators revealed that the bacterial ecological niche has biased the specialization of these proteins. In the case of pathogens, our results suggest that LuxR regulators can be involved in virulence and are therefore promising targets for future studies in the health-related biotechnology field. PMID:23056438

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.

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 ...

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.

UNUSUALLY WIDE BINARIES: ARE THEY WIDE OR UNUSUAL?

International Nuclear Information System (INIS)

Kraus, Adam L.; Hillenbrand, Lynne A.

2009-01-01

We describe an astrometric and spectroscopic campaign to confirm the youth and association of a complete sample of candidate wide companions in Taurus and Upper Sco. Our survey found 15 new binary systems (three in Taurus and 12 in Upper Sco) with separations of 3''-30'' (500-5000 AU) among all of the known members with masses of 2.5-0.012 M sun . The total sample of 49 wide systems in these two regions conforms to only some expectations from field multiplicity surveys. Higher mass stars have a higher frequency of wide binary companions, and there is a marked paucity of wide binary systems near the substellar regime. However, the separation distribution appears to be log-flat, rather than declining as in the field, and the mass ratio distribution is more biased toward similar-mass companions than the initial mass function or the field G-dwarf distribution. The maximum separation also shows no evidence of a limit at ∼ sun . We attribute this result to the post-natal dynamical sculpting that occurs for most field systems; our binary systems will escape to the field intact, but most field stars are formed in denser clusters and undergo significant dynamical evolution. In summary, only wide binary systems with total masses ∼ sun appear to be 'unusually wide'.

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

Protein phosphorylation systems in postmortem human brain

International Nuclear Information System (INIS)

Walaas, S.I.; Perdahl-Wallace, E.; Winblad, B.; Greengard, P.

1989-01-01

Protein phosphorylation systems regulated by cyclic adenosine 3',5'-monophosphate (cyclic AMP), or calcium in conjunction with calmodulin or phospholipid/diacylglycerol, have been studied by phosphorylation in vitro of particulate and soluble fractions from human postmortem brain samples. One-dimensional or two-dimensional gel electrophoretic protein separations were used for analysis. Protein phosphorylation catalyzed by cyclic AMP-dependent protein kinase was found to be highly active in both particulate and soluble preparations throughout the human CNS, with groups of both widely distributed and region-specific substrates being observed in different brain nuclei. Dopamine-innervated parts of the basal ganglia and cerebral cortex contained the phosphoproteins previously observed in rodent basal ganglia. In contrast, calcium/phospholipid-dependent and calcium/calmodulin-dependent protein phosphorylation systems were less prominent in human postmortem brain than in rodent brain, and only a few widely distributed substrates for these protein kinases were found. Protein staining indicated that postmortem proteolysis, particularly of high-molecular-mass proteins, was prominent in deeply located, subcortical regions in the human brain. Our results indicate that it is feasible to use human postmortem brain samples, when obtained under carefully controlled conditions, for qualitative studies on brain protein phosphorylation. Such studies should be of value in studies on human neurological and/or psychiatric disorders

CNF1-like deamidase domains: common Lego bricks among cancer-promoting immunomodulatory bacterial virulence factors.

Science.gov (United States)

Ho, Mengfei; Mettouchi, Amel; Wilson, Brenda A; Lemichez, Emmanuel

2018-05-03

Alterations of the cellular proteome over time due to spontaneous or toxin-mediated enzymatic deamidation of glutamine (Gln) and asparagine (Asn) residues contribute to bacterial infection and might represent a source of aging-related diseases. Here, we put into perspective what is known about the mode of action of the CNF1 toxin from pathogenic E. coli, a paradigm of bacterial deamidases that activate Rho GTPases, to illustrate the importance of determining whether exposure to these factors are risk factors in the etiology age-related diseases, such as cancer. In particular, through in silico analysis of the distribution of the CNF1-like deamidase active site Gly-Cys-(Xaa)n-His sequence motif in bacterial genomes, we unveil the wide distribution of the super-family of CNF-like toxins and CNF-like deamidase domains among members of the enterobacteriacae and in association with a large variety of toxin delivery systems. We extent our discussion with recent findings concerning cellular systems that control activated Rac1 GTPase stability and provide protection against cancer. These findings point to the urgency for developing holistic approaches toward personalized medicine that include monitoring for asymptomatic carriage of pathogenic toxin-producing bacteria and that ultimately might lead to improved public health and increased lifespans.

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

Distribution and interaction patterns of bacterial communities in an ornithogenic soil of Seymour Island, Antarctica.

Science.gov (United States)

Rampelotto, Pabulo Henrique; Barboza, Anthony Diego Muller; Pereira, Antônio Batista; Triplett, Eric W; Schaefer, Carlos Ernesto G R; de Oliveira Camargo, Flávio Anastácio; Roesch, Luiz Fernando Wurdig

2015-04-01

Next-generation, culture-independent sequencing offers an excellent opportunity to examine network interactions among different microbial species. In this study, soil bacterial communities from a penguin rookery site at Seymour Island were analyzed for abundance, structure, diversity, and interaction networks to identify interaction patterns among the various taxa at three soil depths. The analysis revealed the presence of eight phyla distributed in different proportions among the surface layer (0-8 cm), middle layer (20-25 cm), and bottom (35-40 cm). The bottom layer presented the highest values of bacterial richness, diversity, and evenness when compared to surface and middle layers. The network analysis revealed the existence of a unique pattern of interactions in which the soil microbial network formed a clustered topology, rather than a modular structure as is usually found in biological communities. In addition, specific taxa were identified as important players in microbial community structure. Furthermore, simulation analyses indicated that the loss of potential keystone groups of microorganisms might alter the patterns of interactions within the microbial community. These findings provide new insights for assessing the consequences of environmental disturbances at the whole-community level in Antarctica.

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.

Structural insights and ab initio sequencing within the DING proteins family

International Nuclear Information System (INIS)

Elias, Mikael; Liebschner, Dorothee; Gotthard, Guillaume; Chabriere, Eric

2011-01-01

DING proteins constitute a recently discovered protein family that is ubiquitous in eukaryotes. The structural insights and the physiological involvements of these intriguing proteins are hereby deciphered. DING proteins constitute an intriguing family of phosphate-binding proteins that was identified in a wide range of organisms, from prokaryotes and archae to eukaryotes. Despite their seemingly ubiquitous occurrence in eukaryotes, their encoding genes are missing from sequenced genomes. Such a lack has considerably hampered functional studies. In humans, these proteins have been related to several diseases, like atherosclerosis, kidney stones, inflammation processes and HIV inhibition. The human phosphate binding protein is a human representative of the DING family that was serendipitously discovered from human plasma. An original approach was developed to determine ab initio the complete and exact sequence of this 38 kDa protein by utilizing mass spectrometry and X-ray data in tandem. Taking advantage of this first complete eukaryotic DING sequence, a immunohistochemistry study was undertaken to check the presence of DING proteins in various mice tissues, revealing that these proteins are widely expressed. Finally, the structure of a bacterial representative from Pseudomonas fluorescens was solved at sub-angstrom resolution, allowing the molecular mechanism of the phosphate binding in these high-affinity proteins to be elucidated

Structural insights and ab initio sequencing within the DING proteins family

Energy Technology Data Exchange (ETDEWEB)

Elias, Mikael, E-mail: mikael.elias@weizmann.ac.il [Weizmann Institute of Science, Rehovot (Israel); Liebschner, Dorothee [CRM2, Nancy Université (France); Gotthard, Guillaume; Chabriere, Eric [AFMB, Université Aix-Marseille II (France)

2011-01-01

DING proteins constitute a recently discovered protein family that is ubiquitous in eukaryotes. The structural insights and the physiological involvements of these intriguing proteins are hereby deciphered. DING proteins constitute an intriguing family of phosphate-binding proteins that was identified in a wide range of organisms, from prokaryotes and archae to eukaryotes. Despite their seemingly ubiquitous occurrence in eukaryotes, their encoding genes are missing from sequenced genomes. Such a lack has considerably hampered functional studies. In humans, these proteins have been related to several diseases, like atherosclerosis, kidney stones, inflammation processes and HIV inhibition. The human phosphate binding protein is a human representative of the DING family that was serendipitously discovered from human plasma. An original approach was developed to determine ab initio the complete and exact sequence of this 38 kDa protein by utilizing mass spectrometry and X-ray data in tandem. Taking advantage of this first complete eukaryotic DING sequence, a immunohistochemistry study was undertaken to check the presence of DING proteins in various mice tissues, revealing that these proteins are widely expressed. Finally, the structure of a bacterial representative from Pseudomonas fluorescens was solved at sub-angstrom resolution, allowing the molecular mechanism of the phosphate binding in these high-affinity proteins to be elucidated.

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.

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.

Role of environmental factors for the vertical distribution (0–1000 m of marine bacterial communities in the NW Mediterranean Sea

Directory of Open Access Journals (Sweden)

J. F. Ghiglione

2008-12-01

Full Text Available Bacterioplankton plays a central role in energy and matter fluxes in the sea, yet the factors that constrain its variation in marine systems are still poorly understood. Here we use the explanatory power of direct multivariate gradient analysis to evaluate the driving forces exerted by environmental parameters on bacterial community distribution in the water column. We gathered and analysed data from a one month sampling period from the surface to 1000 m depth at the JGOFS-DYFAMED station (NW Mediterranean Sea. This station is characterized by very poor horizontal advection currents which makes it an ideal model to test hypotheses on the causes of vertical stratification of bacterial communities. Capillary electrophoresis single strand conformation polymorphism (CE-SSCP fingerprinting profiles analyzed using multivariate statistical methods demonstrated a vertical zonation of bacterial assemblages in three layers, above, in or just below the chlorophyll maximum and deeper, that remained stable during the entire sampling period. Through the use of direct gradient multivariate ordination analyses we demonstrate that a complex array of biogeochemical parameters is the driving force behind bacterial community structure shifts in the water column. Physico-chemical parameters such as phosphate, nitrate, salinity and to a lesser extent temperature, oxygen, dissolved organic carbon and photosynthetically active radiation acted in synergy to explain bacterial assemblages changes with depth. Analysis of lipid biomarkers of organic matter sources and fates suggested that bacterial community structure in the surface layers was in part explained by lipids of chloroplast origin. Further detailed analysis of pigment-based phytoplankton diversity gave evidence of a compartmentalized influence of several phytoplankton groups on bacterial community structure in the first 150 m depth.

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

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.

A size-structured model of bacterial growth and reproduction.

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Ellermeyer, S F; Pilyugin, S S

2012-01-01

We consider a size-structured bacterial population model in which the rate of cell growth is both size- and time-dependent and the average per capita reproduction rate is specified as a model parameter. It is shown that the model admits classical solutions. The population-level and distribution-level behaviours of these solutions are then determined in terms of the model parameters. The distribution-level behaviour is found to be different from that found in similar models of bacterial population dynamics. Rather than convergence to a stable size distribution, we find that size distributions repeat in cycles. This phenomenon is observed in similar models only under special assumptions on the functional form of the size-dependent growth rate factor. Our main results are illustrated with examples, and we also provide an introductory study of the bacterial growth in a chemostat within the framework of our model.

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.

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

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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.

Discovery of novel bacterial toxins by genomics and computational biology.

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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.

Bacterial Urease and its Role in Long-Lasting Human Diseases

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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

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...

Evaluation of bacterial run and tumble motility parameters through trajectory analysis

Science.gov (United States)

Liang, Xiaomeng; Lu, Nanxi; Chang, Lin-Ching; Nguyen, Thanh H.; Massoudieh, Arash

2018-04-01

In this paper, a method for extraction of the behavior parameters of bacterial migration based on the run and tumble conceptual model is described. The methodology is applied to the microscopic images representing the motile movement of flagellated Azotobacter vinelandii. The bacterial cells are considered to change direction during both runs and tumbles as is evident from the movement trajectories. An unsupervised cluster analysis was performed to fractionate each bacterial trajectory into run and tumble segments, and then the distribution of parameters for each mode were extracted by fitting mathematical distributions best representing the data. A Gaussian copula was used to model the autocorrelation in swimming velocity. For both run and tumble modes, Gamma distribution was found to fit the marginal velocity best, and Logistic distribution was found to represent better the deviation angle than other distributions considered. For the transition rate distribution, log-logistic distribution and log-normal distribution, respectively, was found to do a better job than the traditionally agreed exponential distribution. A model was then developed to mimic the motility behavior of bacteria at the presence of flow. The model was applied to evaluate its ability to describe observed patterns of bacterial deposition on surfaces in a micro-model experiment with an approach velocity of 200 μm/s. It was found that the model can qualitatively reproduce the attachment results of the micro-model setting.

Distribution in rat tissues of modulator-binding protein of particulate nature

International Nuclear Information System (INIS)

Sobue, K.; Muramoto, Y.; Kakiuchi, S.; Yamazaki, R.

1979-01-01

Studies on Ca 2+ -activatable cyclic nucleotide phosphodiesterase led to the discovery of a protein modulator that is required for the activation of this enzyme by Ca 2+ . Later, this protein has been shown to cause the Ca 2+ -dependent activation of several enzymes that include phosphodiesterase, adenylate cyclase, a protein kinase from muscles, phosphorylase b kinase, actomyosin ATPase, membranous ATPase from erythrocytes and nerve synapses. Thus, modulator protein appears to be an intracellular mediator of actions of Ca 2+ . The present work shows the distribution of this particulate modulator-binding component in rat tissues. This paper also describes the labeling of modulator protein with tritium without deteriorating its biological activities and application of this 3 H-modulator protein to the determination of the Ca ++ dependent binding of modulator protein with membranous protein. This technique proves to be useful in studying enzymes or proteins whose functions are regulated by Ca ++ /modulator protein system. (Auth.)

Distribution of PASTA domains in penicillin-binding proteins and serine/threonine kinases of Actinobacteria.

Science.gov (United States)

Ogawara, Hiroshi

2016-09-01

PASTA domains (penicillin-binding protein and serine/threonine kinase-associated domains) have been identified in penicillin-binding proteins and serine/threonine kinases of Gram-positive Firmicutes and Actinobacteria. They are believed to bind β-lactam antibiotics, and be involved in peptidoglycan metabolism, although their biological function is not definitively clarified. Actinobacteria, especially Streptomyces species, are distinct in that they undergo complex cellular differentiation and produce various antibiotics including β-lactams. This review focuses on the distribution of PASTA domains in penicillin-binding proteins and serine/threonine kinases in Actinobacteria. In Actinobacteria, PASTA domains are detectable exclusively in class A but not in class B penicillin-binding proteins, in sharp contrast to the cases in other bacteria. In penicillin-binding proteins, PASTA domains distribute independently from taxonomy with some distribution bias. Particularly interesting thing is that no Streptomyces species have penicillin-binding protein with PASTA domains. Protein kinases in Actinobacteria possess 0 to 5 PASTA domains in their molecules. Protein kinases in Streptomyces can be classified into three groups: no PASTA domain, 1 PASTA domain and 4 PASTA domain-containing groups. The 4 PASTA domain-containing groups can be further divided into two subgroups. The serine/threonine kinases in different groups may perform different functions. The pocket region in one of these subgroup is more dense and extended, thus it may be involved in binding of ligands like β-lactams more efficiently.

Protein Intake and Distribution in Relation to Physical Functioning and Quality of Life in Community-Dwelling Elderly People: Acknowledging the Role of Physical Activity

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Dominique S. M. ten Haaf

2018-04-01

Full Text Available Increasing total protein intake and a spread protein intake distribution are potential strategies to attenuate sarcopenia related loss of physical function and quality of life. The aim of this cross-sectional study was to investigate whether protein intake and protein intake distribution are associated with muscle strength, physical function and quality of life in community-dwelling elderly people with a wide range of physical activity. Dietary and physical activity data were obtained from two studies (N = 140, age 81 ± 6, 64% male, with the following outcome measures: physical functioning (Short Physical Performance Battery (SPPB, comprising balance, gait speed and chair rise tests, handgrip strength and quality of life (EQ-5D-5L. Protein intake distribution was calculated for each participant as a coefficient of variance (CV = SD of grams of protein intake per main meal divided by the average total amount of proteins (grams of the main meals. Based on the CV, participants were divided into tertiles and classified as spread, intermediate or pulse. The average total protein intake was 1.08 ± 0.29 g/kg/day. Total protein intake was not associated with outcome measures using multivariate regression analyses. Individuals with a spread protein diet during the main meals (CV < 0.43 had higher gait speed compared to those with an intermediate diet (CV 0.43–0.62 (β = −0.42, p = 0.035, whereas a spread and pulse protein diet were not associated with SPPB total score, chair rise, grip strength and Quality-Adjusted Life Year (QALY. The interaction of higher physical activity and higher total protein intake was significantly associated with higher quality of life (β = 0.71, p = 0.049. While this interaction was not associated with SPPB or grip strength, the association with quality of life emphasizes the need for a higher total protein intake together with an active lifestyle in the elderly.

Genome wide binding (ChIP-Seq of murine Bapx1 and Sox9 proteins in vivo and in vitro

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Sumantra Chatterjee

2016-12-01

Full Text Available This work pertains to GEO submission GSE36672, in vivo and in vitro genome wide binding (ChIP-Seq of Bapx1/Nkx3.2 and Sox9 proteins. We have previously shown that data from a genome wide binding assay combined with transcriptional profiling is an insightful means to divulge the mechanisms directing cell type specification and the generation of tissues and subsequent organs [1]. Our earlier work identified the role of the DNA-binding homeodomain containing protein Bapx1/Nkx3.2 in midgestation murine embryos. Microarray analysis of EGFP-tagged cells (both wildtype and null was integrated using ChIP-Seq analysis of Bapx1/Nkx3.2 and Sox9 DNA-binding proteins in living tissue.

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).

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.

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...

Bacterial Community Associated with Healthy and Diseased Pacific White Shrimp (Litopenaeus vannamei) Larvae and Rearing Water across Different Growth Stages.

Science.gov (United States)

Zheng, Yanfen; Yu, Min; Liu, Jiwen; Qiao, Yanlu; Wang, Long; Li, Zhitao; Zhang, Xiao-Hua; Yu, Mingchao

2017-01-01

Bacterial communities are called another "organ" for aquatic animals and their important influence on the health of host has drawn increasing attention. Thus, it is important to study the relationships between aquatic animals and bacterial communities. Here, bacterial communities associated with Litopenaeus vannamei larvae at different healthy statuses (diseased and healthy) and growth stages (i.e., zoea, mysis, and early postlarvae periods) were examined using 454-pyrosequencing of the 16S rRNA gene. Bacterial communities with significant difference were observed between healthy and diseased rearing water, and several bacterial groups, such as genera Nautella and Kordiimonas could also distinguish healthy and diseased shrimp. Rhodobacteraceae was widely distributed in rearing water at all growth stages but there were several stage-specific groups, indicating that bacterial members in rearing water assembled into distinct communities throughout the larval development. However, Gammaproteobacteria , mainly family Enterobacteriaceae , was the most abundant group (accounting for more than 85%) in shrimp larvae at all growth stages. This study compared bacterial communities associated with healthy and diseased L . vannamei larvae and rearing water, and identified several health- and growth stage-specific bacterial groups, which might be provided as indicators for monitoring the healthy status of shrimp larvae in hatchery.

Bacterial Community Associated with Healthy and Diseased Pacific White Shrimp (Litopenaeus vannamei Larvae and Rearing Water across Different Growth Stages

Directory of Open Access Journals (Sweden)

Yanfen Zheng

2017-07-01

Full Text Available Bacterial communities are called another “organ” for aquatic animals and their important influence on the health of host has drawn increasing attention. Thus, it is important to study the relationships between aquatic animals and bacterial communities. Here, bacterial communities associated with Litopenaeus vannamei larvae at different healthy statuses (diseased and healthy and growth stages (i.e., zoea, mysis, and early postlarvae periods were examined using 454-pyrosequencing of the 16S rRNA gene. Bacterial communities with significant difference were observed between healthy and diseased rearing water, and several bacterial groups, such as genera Nautella and Kordiimonas could also distinguish healthy and diseased shrimp. Rhodobacteraceae was widely distributed in rearing water at all growth stages but there were several stage-specific groups, indicating that bacterial members in rearing water assembled into distinct communities throughout the larval development. However, Gammaproteobacteria, mainly family Enterobacteriaceae, was the most abundant group (accounting for more than 85% in shrimp larvae at all growth stages. This study compared bacterial communities associated with healthy and diseased L. vannamei larvae and rearing water, and identified several health- and growth stage-specific bacterial groups, which might be provided as indicators for monitoring the healthy status of shrimp larvae in hatchery.

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

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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.

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.

Sulfonamide and tetracycline resistance genes in total- and culturable-bacterial assemblages in South African aquatic environments

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Satoru eSuzuki

2015-08-01

Full Text Available Antibiotic resistant bacteria (ARB are ubiquitous in the natural environment. The introduction of effluent derived antibiotic resistance genes (ARGs into aquatic environments is of concern in the spreading of genetic risk. This study showed the prevalence of sulfonamide and tetracycline resistance genes, sul1, sul2, sul3 and tet(M, in the total bacterial assemblage and colony forming bacterial assemblage in river and estuarine water and sewage treatment plants (STP in South Africa. There was no correlation between antibiotic concentrations and ARGs, suggesting the targeted ARGs are spread in a wide area without connection to selection pressure. Among sul genes, sul1 and sul2 were major genes in the total (over 10-2 copies/16S and colony forming bacteria assemblages (approx 10-1 copies/16S. In urban waters, the sul3 gene was mostly not detectable in total and culturable assemblages, suggesting sul3 is not abundant. tet(M was found in natural assemblages with 10-3 copies/16S level in STP, but was not detected in colony forming bacteria, suggesting the non-culturable (yet-to-be cultured bacterial community in urban surface waters and STP effluent possess the tet(M gene. Sulfamethoxazole resistant (SMXr and oxytetracycline resistant (OTCr bacterial communities in urban waters possessed not only sul1 and sul2 but also sul3 and tet(M genes. These genes are widely distributed in SMXr and OTCr bacteria. In conclusion, urban river and estuarine water and STP effluent in the Durban area were highly contaminated with ARGs, and the yet-to-be cultured bacterial community may act as a non-visible ARG reservoir in certain situations.

Impact of transgenic potatoes expressing anti-bacterial agents on bacterial endophytes is comparable with the effects of plant genotype, soil type and pathogen infection

NARCIS (Netherlands)

Rasche, F; Velvis, H; Zachow, C; Berg, G; Van Elsas, JD; Sessitsch, A

1. Blackleg and soft rot disease of potatoes Solanum tuberosum L., mainly caused by the bacterial pathogen Erwinia carotovora ssp. atrospetica (Eca), lead to enormous yield losses world-wide. Genetically modified (GM) potatoes producing anti-bacterial agents, such as cecropin/attacin and T4

Impact of transgenic potatoes expressing anti-bacterial agents on bacterial endophytes is comparable with the effects of plant genotype, soil type and pathogen infection

NARCIS (Netherlands)

Rasche, F.; Velvis, H.; Zachow, C.; Berg, G.; Elsas, van J.D.; Sessitsch, A.

2006-01-01

1. Blackleg and soft rot disease of potatoes Solanum tuberosum L., mainly caused by the bacterial pathogen Erwinia carotovora ssp. atrospetica (Eca), lead to enormous yield losses world-wide. Genetically modified (GM) potatoes producing anti-bacterial agents, such as cecropin/attacin and T4

Two Paralogous Families of a Two-Gene Subtilisin Operon Are Widely Distributed in Oral Treponemes

Science.gov (United States)

Correia, Frederick F.; Plummer, Alvin R.; Ellen, Richard P.; Wyss, Chris; Boches, Susan K.; Galvin, Jamie L.; Paster, Bruce J.; Dewhirst, Floyd E.

2003-01-01

Certain oral treponemes express a highly proteolytic phenotype and have been associated with periodontal diseases. The periodontal pathogen Treponema denticola produces dentilisin, a serine protease of the subtilisin family. The two-gene operon prcA-prtP is required for expression of active dentilisin (PrtP), a putative lipoprotein attached to the treponeme's outer membrane or sheath. The purpose of this study was to examine the diversity and structure of treponemal subtilisin-like proteases in order to better understand their distribution and function. The complete sequences of five prcA-prtP operons were determined for Treponema lecithinolyticum, “Treponema vincentii,” and two canine species. Partial operon sequences were obtained for T. socranskii subsp. 04 as well as 450- to 1,000-base fragments of prtP genes from four additional treponeme strains. Phylogenetic analysis demonstrated that the sequences fall into two paralogous families. The first family includes the sequence from T. denticola. Treponemes possessing this operon family express chymotrypsin-like protease activity and can cleave the substrate N-succinyl-alanyl-alanyl-prolyl-phenylalanine-p-nitroanilide (SAAPFNA). Treponemes possessing the second paralog family do not possess chymotrypsin-like activity or cleave SAAPFNA. Despite examination of a range of protein and peptide substrates, the specificity of the second protease family remains unknown. Each of the fully sequenced prcA and prtP genes contains a 5′ hydrophobic leader sequence with a treponeme lipobox. The two paralogous families of treponeme subtilisins represent a new subgroup within the subtilisin family of proteases and are the only subtilisin lipoprotein family. The present study demonstrated that the subtilisin paralogs comprising a two-gene operon are widely distributed among treponemes. PMID:14617650

The relationship of protein conservation and sequence length

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Panchenko Anna R

2002-11-01

Full Text Available Abstract Background In general, the length of a protein sequence is determined by its function and the wide variance in the lengths of an organism's proteins reflects the diversity of specific functional roles for these proteins. However, additional evolutionary forces that affect the length of a protein may be revealed by studying the length distributions of proteins evolving under weaker functional constraints. Results We performed sequence comparisons to distinguish highly conserved and poorly conserved proteins from the bacterium Escherichia coli, the archaeon Archaeoglobus fulgidus, and the eukaryotes Saccharomyces cerevisiae, Drosophila melanogaster, and Homo sapiens. For all organisms studied, the conserved and nonconserved proteins have strikingly different length distributions. The conserved proteins are, on average, longer than the poorly conserved ones, and the length distributions for the poorly conserved proteins have a relatively narrow peak, in contrast to the conserved proteins whose lengths spread over a wider range of values. For the two prokaryotes studied, the poorly conserved proteins approximate the minimal length distribution expected for a diverse range of structural folds. Conclusions There is a relationship between protein conservation and sequence length. For all the organisms studied, there seems to be a significant evolutionary trend favoring shorter proteins in the absence of other, more specific functional constraints.

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.

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.

Bacterial community dynamics during the early stages of biofilm formation in a chlorinated experimental drinking water distribution system: implications for drinking water discolouration.

Science.gov (United States)

Douterelo, I; Sharpe, R; Boxall, J

2014-07-01

To characterize bacterial communities during the early stages of biofilm formation and their role in water discolouration in a fully representative, chlorinated, experimental drinking water distribution systems (DWDS). Biofilm development was monitored in an experimental DWDS over 28 days; subsequently the system was disturbed by raising hydraulic conditions to simulate pipe burst, cleaning or other system conditions. Biofilm cell cover was monitored by fluorescent microscopy and a fingerprinting technique used to assess changes in bacterial community. Selected samples were analysed by cloning and sequencing of the 16S rRNA gene. Fingerprinting analysis revealed significant changes in the bacterial community structure over time (P < 0·05). Cell coverage increased over time accompanied by an increase in bacterial richness and diversity. Shifts in the bacterial community structure were observed along with an increase in cell coverage, bacterial richness and diversity. Species related to Pseudomonas spp. and Janthinobacterium spp. dominated the process of initial attachment. Based on fingerprinting results, the hydraulic regimes did not affect the bacteriological composition of biofilms, but they did influence their mechanical stability. This study gives a better insight into the early stages of biofilm formation in DWDS and will contribute to the improvement of management strategies to control the formation of biofilms and the risk of discolouration. © 2014 The Authors. published by John Wiley & Sons Ltd on behalf of Society for Applied Microbiology.

Bacterial community dynamics during the early stages of biofilm formation in a chlorinated experimental drinking water distribution system: implications for drinking water discolouration

Science.gov (United States)

Douterelo, I; Sharpe, R; Boxall, J

2014-01-01

Aims To characterize bacterial communities during the early stages of biofilm formation and their role in water discolouration in a fully representative, chlorinated, experimental drinking water distribution systems (DWDS). Methods and Results Biofilm development was monitored in an experimental DWDS over 28 days; subsequently the system was disturbed by raising hydraulic conditions to simulate pipe burst, cleaning or other system conditions. Biofilm cell cover was monitored by fluorescent microscopy and a fingerprinting technique used to assess changes in bacterial community. Selected samples were analysed by cloning and sequencing of the 16S rRNA gene. Fingerprinting analysis revealed significant changes in the bacterial community structure over time (P < 0·05). Cell coverage increased over time accompanied by an increase in bacterial richness and diversity. Conclusions Shifts in the bacterial community structure were observed along with an increase in cell coverage, bacterial richness and diversity. Species related to Pseudomonas spp. and Janthinobacterium spp. dominated the process of initial attachment. Based on fingerprinting results, the hydraulic regimes did not affect the bacteriological composition of biofilms, but they did influence their mechanical stability. Significance and Importance of the Study This study gives a better insight into the early stages of biofilm formation in DWDS and will contribute to the improvement of management strategies to control the formation of biofilms and the risk of discolouration. PMID:24712449

Protein interactions in genome maintenance as novel antibacterial targets.

Directory of Open Access Journals (Sweden)

Aimee H Marceau

Full Text Available Antibacterial compounds typically act by directly inhibiting essential bacterial enzyme activities. Although this general mechanism of action has fueled traditional antibiotic discovery efforts for decades, new antibiotic development has not kept pace with the emergence of drug resistant bacterial strains. These limitations have severely restricted the therapeutic tools available for treating bacterial infections. Here we test an alternative antibacterial lead-compound identification strategy in which essential protein-protein interactions are targeted rather than enzymatic activities. Bacterial single-stranded DNA-binding proteins (SSBs form conserved protein interaction "hubs" that are essential for recruiting many DNA replication, recombination, and repair proteins to SSB/DNA nucleoprotein substrates. Three small molecules that block SSB/protein interactions are shown to have antibacterial activity against diverse bacterial species. Consistent with a model in which the compounds target multiple SSB/protein interactions, treatment of Bacillus subtilis cultures with the compounds leads to rapid inhibition of DNA replication and recombination, and ultimately to cell death. The compounds also have unanticipated effects on protein synthesis that could be due to a previously unknown role for SSB/protein interactions in translation or to off-target effects. Our results highlight the potential of targeting protein-protein interactions, particularly those that mediate genome maintenance, as a powerful approach for identifying new antibacterial compounds.

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

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.

Genome-wide identification and comparative analysis of squamosa-promoter binding proteins (sbp) transcription factor family in gossypium raimondii and arabidopsis thaliana

International Nuclear Information System (INIS)

Ali, M.A.; Alia, K.B.; Atif, R.M.; Rasulj, I.; Nadeem, H.U.; Shahid, A.; Azeem, F

2017-01-01

SQUAMOSA-Promoter Binding Proteins (SBP) are class of transcription factors that play vital role in regulation of plant tissue growth and development. The genes encoding these proteins have not yet been identified in diploid cotton. Thus here, a comprehensive genome wide analysis of SBP genes/proteins was carried out to identify the genes encoding SBP proteins in Gossypium raimondii and Arabidopsis thaliana. We identified 17 SBP genes from Arabidopsis thaliana genome and 30 SBP genes from Gossypium raimondii. Chromosome localization studies revealed the uneven distribution of SBP encoding genes both in the genomes of A. thaliana and G. raimondii. In cotton, five SBP genes were located on chromosome no. 2, while no gene was found on chromosome 9. In A. thaliana, maximum seven SBP genes were identified on chromosome 9, while chromosome 4 did not have any SBP gene. Thus, the SBP gene family might have expanded as a result of segmental as well as tandem duplications in these species. The comparative phylogenetic analysis of Arabidopsis and cotton SBPs revealed the presence of eight groups. The gene structure analysis of SBP encoding genes revealed the presence of one to eleven inrons in both Arabidopsis and G. raimondii. The proteins sharing the same phyletic group mostly demonstrated the similar intron-exon occurrence pattern; and share the common conserved domains. The SBP DNA-binding domain shared 24 absolutely conserved residues in Arabidopsis. The present study can serve as a base for the functional characterization of SBP gene family in Gossypium raimondii. (author)

Ribosome-dependent ATPase interacts with conserved membrane protein in Escherichia coli to modulate protein synthesis and oxidative phosphorylation.

Directory of Open Access Journals (Sweden)

Mohan Babu

Full Text Available Elongation factor RbbA is required for ATP-dependent deacyl-tRNA release presumably after each peptide bond formation; however, there is no information about the cellular role. Proteomic analysis in Escherichia coli revealed that RbbA reciprocally co-purified with a conserved inner membrane protein of unknown function, YhjD. Both proteins are also physically associated with the 30S ribosome and with members of the lipopolysaccharide transport machinery. Genome-wide genetic screens of rbbA and yhjD deletion mutants revealed aggravating genetic interactions with mutants deficient in the electron transport chain. Cells lacking both rbbA and yhjD exhibited reduced cell division, respiration and global protein synthesis as well as increased sensitivity to antibiotics targeting the ETC and the accuracy of protein synthesis. Our results suggest that RbbA appears to function together with YhjD as part of a regulatory network that impacts bacterial oxidative phosphorylation and translation efficiency.

Assessing protein conformational sampling methods based on bivariate lag-distributions of backbone angles

KAUST Repository

Maadooliat, Mehdi; Gao, Xin; Huang, Jianhua Z.

2012-01-01

Despite considerable progress in the past decades, protein structure prediction remains one of the major unsolved problems in computational biology. Angular-sampling-based methods have been extensively studied recently due to their ability to capture the continuous conformational space of protein structures. The literature has focused on using a variety of parametric models of the sequential dependencies between angle pairs along the protein chains. In this article, we present a thorough review of angular-sampling-based methods by assessing three main questions: What is the best distribution type to model the protein angles? What is a reasonable number of components in a mixture model that should be considered to accurately parameterize the joint distribution of the angles? and What is the order of the local sequence-structure dependency that should be considered by a prediction method? We assess the model fits for different methods using bivariate lag-distributions of the dihedral/planar angles. Moreover, the main information across the lags can be extracted using a technique called Lag singular value decomposition (LagSVD), which considers the joint distribution of the dihedral/planar angles over different lags using a nonparametric approach and monitors the behavior of the lag-distribution of the angles using singular value decomposition. As a result, we developed graphical tools and numerical measurements to compare and evaluate the performance of different model fits. Furthermore, we developed a web-tool (http://www.stat.tamu. edu/~madoliat/LagSVD) that can be used to produce informative animations. © The Author 2012. Published by Oxford University Press.

Assessing protein conformational sampling methods based on bivariate lag-distributions of backbone angles

KAUST Repository

Maadooliat, Mehdi

2012-08-27

Despite considerable progress in the past decades, protein structure prediction remains one of the major unsolved problems in computational biology. Angular-sampling-based methods have been extensively studied recently due to their ability to capture the continuous conformational space of protein structures. The literature has focused on using a variety of parametric models of the sequential dependencies between angle pairs along the protein chains. In this article, we present a thorough review of angular-sampling-based methods by assessing three main questions: What is the best distribution type to model the protein angles? What is a reasonable number of components in a mixture model that should be considered to accurately parameterize the joint distribution of the angles? and What is the order of the local sequence-structure dependency that should be considered by a prediction method? We assess the model fits for different methods using bivariate lag-distributions of the dihedral/planar angles. Moreover, the main information across the lags can be extracted using a technique called Lag singular value decomposition (LagSVD), which considers the joint distribution of the dihedral/planar angles over different lags using a nonparametric approach and monitors the behavior of the lag-distribution of the angles using singular value decomposition. As a result, we developed graphical tools and numerical measurements to compare and evaluate the performance of different model fits. Furthermore, we developed a web-tool (http://www.stat.tamu. edu/~madoliat/LagSVD) that can be used to produce informative animations. © The Author 2012. Published by Oxford University Press.

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...

Protein Structure Classification and Loop Modeling Using Multiple Ramachandran Distributions

KAUST Repository

Najibi, Seyed Morteza; Maadooliat, Mehdi; Zhou, Lan; Huang, Jianhua Z.; Gao, Xin

2017-01-01

Recently, the study of protein structures using angular representations has attracted much attention among structural biologists. The main challenge is how to efficiently model the continuous conformational space of the protein structures based on the differences and similarities between different Ramachandran plots. Despite the presence of statistical methods for modeling angular data of proteins, there is still a substantial need for more sophisticated and faster statistical tools to model the large-scale circular datasets. To address this need, we have developed a nonparametric method for collective estimation of multiple bivariate density functions for a collection of populations of protein backbone angles. The proposed method takes into account the circular nature of the angular data using trigonometric spline which is more efficient compared to existing methods. This collective density estimation approach is widely applicable when there is a need to estimate multiple density functions from different populations with common features. Moreover, the coefficients of adaptive basis expansion for the fitted densities provide a low-dimensional representation that is useful for visualization, clustering, and classification of the densities. The proposed method provides a novel and unique perspective to two important and challenging problems in protein structure research: structure-based protein classification and angular-sampling-based protein loop structure prediction.