umu.sePublikationer
Ändra sökning
Avgränsa sökresultatet
12 51 - 82 av 82
RefereraExporteraLänk till träfflistan
Permanent länk
Referera
Referensformat
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Annat format
Fler format
Språk
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Annat språk
Fler språk
Utmatningsformat
  • html
  • text
  • asciidoc
  • rtf
Träffar per sida
  • 5
  • 10
  • 20
  • 50
  • 100
  • 250
Sortering
  • Standard (Relevans)
  • Författare A-Ö
  • Författare Ö-A
  • Titel A-Ö
  • Titel Ö-A
  • Publikationstyp A-Ö
  • Publikationstyp Ö-A
  • Äldst först
  • Nyast först
  • Skapad (Äldst först)
  • Skapad (Nyast först)
  • Senast uppdaterad (Äldst först)
  • Senast uppdaterad (Nyast först)
  • Disputationsdatum (tidigaste först)
  • Disputationsdatum (senaste först)
  • Standard (Relevans)
  • Författare A-Ö
  • Författare Ö-A
  • Titel A-Ö
  • Titel Ö-A
  • Publikationstyp A-Ö
  • Publikationstyp Ö-A
  • Äldst först
  • Nyast först
  • Skapad (Äldst först)
  • Skapad (Nyast först)
  • Senast uppdaterad (Äldst först)
  • Senast uppdaterad (Nyast först)
  • Disputationsdatum (tidigaste först)
  • Disputationsdatum (senaste först)
Markera
Maxantalet träffar du kan exportera från sökgränssnittet är 250. Vid större uttag använd dig av utsökningar.
  • 51. Hsu, Yen-Pang
    et al.
    Hall, Edward
    Booher, Garrett
    Murphy, Brennan
    Radkov, Atanas D.
    Yablonowski, Jacob
    Mulcahey, Caitlyn
    Alvarez, Laura
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Brun, Yves, V
    Kuru, Erkin
    VanNieuwenhze, Michael S.
    Fluorogenic D-amino acids enable real-time monitoring of peptidoglycan biosynthesis and high-throughput transpeptidation assays2019Ingår i: Nature Chemistry, ISSN 1755-4330, E-ISSN 1755-4349, Vol. 11, nr 4, s. 335-341Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Peptidoglycan is an essential cell wall component that maintains the morphology and viability of nearly all bacteria. Its biosynthesis requires periplasmic transpeptidation reactions, which construct peptide crosslinkages between polysaccharide chains to endow mechanical strength. However, tracking the transpeptidation reaction in vivo and in vitro is challenging, mainly due to the lack of efficient, biocompatible probes. Here, we report the design, synthesis and application of rotor-fluorogenic D-amino acids (RfDAAs), enabling real-time, continuous tracking of transpeptidation reactions. These probes allow peptidoglycan biosynthesis to be monitored in real time by visualizing transpeptidase reactions in live cells, as well as real-time activity assays of D,D- and L,D-transpeptidases and sortases in vitro. The unique ability of RfDAAs to become fluorescent when incorporated into peptidoglycan provides a powerful new tool to study peptidoglycan biosynthesis with high temporal resolution and prospectively enable high-throughput screening for inhibitors of peptidoglycan biosynthesis.

  • 52.
    Irazoki, Oihane
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Hernandez, Sara B.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Peptidoglycan Muropeptides: Release, Perception, and Functions as Signaling Molecules2019Ingår i: Frontiers in Microbiology, ISSN 1664-302X, E-ISSN 1664-302X, Vol. 10, artikel-id 500Artikel, forskningsöversikt (Refereegranskat)
    Abstract [en]

    Peptidoglycan (PG) is an essential molecule for the survival of bacteria, and thus, its biosynthesis and remodeling have always been in the spotlight when it comes to the development of antibiotics. The peptidoglycan polymer provides a protective function in bacteria, but at the same time is continuously subjected to editing activities that in some cases lead to the release of peptidoglycan fragments (i.e., muropeptides) to the environment. Several soluble muropeptides have been reported to work as signaling molecules. In this review, we summarize the mechanisms involved in muropeptide release (PG breakdown and PG recycling) and describe the known PG-receptor proteins responsible for PG sensing. Furthermore, we overview the role of muropeptides as signaling molecules, focusing on the microbial responses and their functions in the host beyond their immunostimulatory activity.

  • 53. Jacquier, Nicolas
    et al.
    Yadav, Akhilesh K.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Pillonel, Trestan
    Viollier, Patrick H.
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Greub, Gilbert
    A SpoIID Homolog Cleaves Glycan Strands at the Chlamydial Division Septum2019Ingår i: mBio, ISSN 2161-2129, E-ISSN 2150-7511, Vol. 10, nr 4, artikel-id e01128-19Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Chlamydiales species are obligate intracellular bacteria lacking a classical peptidoglycan sacculus but relying on peptidoglycan synthesis for cytokinesis. While septal peptidoglycan biosynthesis seems to be regulated by MreB actin and its membrane anchor RodZ rather than FtsZ tubulin in Chlamydiales, the mechanism of peptidoglycan remodeling is poorly understood. An amidase conserved in Chlamydiales is able to cleave peptide stems in peptidoglycan, but it is not clear how peptidoglycan glycan strands are cleaved since no classical lytic transglycosylase is encoded in chlamydial genomes. However, a protein containing a SpoIID domain, known to possess transglycosylase activity in Bacillus subtilis, is conserved in Chiamydiales. We show here that the SpoIID homologue of the Chlamydia-related pathogen Waddlia chondrophila is a septal peptidoglycan-binding protein. Moreover, we demonstrate that SpoIID acts as a lytic transglycosylase on peptidoglycan and as a muramidase on denuded glycan strands in vitro. As SpoIID-like proteins are widespread in nonsporulating bacteria, SpoIID might commonly be a septal peptidoglycan remodeling protein in bacteria, including obligate intracellular pathogens, and thus might represent a promising drug target. IMPORTANCE Chlamydiales species are obligate intracellular bacteria and important human pathogens that have a minimal division machinery lacking the proteins that are essential for bacterial division in other species, such as FtsZ. Chlamydial division requires synthesis of peptidoglycan, which forms a ring at the division septum and is rapidly turned over. However, little is known of peptidoglycan degradation, because many peptidoglycan-degrading enzymes are not encoded by chlamydial genomes. Here we show that an homologue of SpoIID, a peptidoglycan-degrading enzyme involved in sporulation of bacteria such as Bacillus subtilis, is expressed in Chlamydiales, localizes at the division septum, and degrades peptidoglycan in vitro, indicating that SpoIID is not only involved in sporulation but also likely implicated in division of some bacteria.

  • 54. Kudrin, Pavel
    et al.
    Varik, Vallo
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). University of Tartu, Institute of Technology, Tartu, Estonia.
    Oliveira, Sofia Raquel Alves
    Beljantseva, Jelena
    Santos, Teresa Del Peso
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Dzhygyr, Ievgen
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Rejman, Dominik
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Tenson, Tanel
    Hauryliuk, Vasili
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). University of Tartu, Institute of Technology, Tartu, Estonia.
    Subinhibitory Concentrations of Bacteriostatic Antibiotics Induce relA-Dependent and relA-Independent Tolerance to beta-Lactams2017Ingår i: Antimicrobial Agents and Chemotherapy, ISSN 0066-4804, E-ISSN 1098-6596, Vol. 61, nr 4, artikel-id e02173-16Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The nucleotide (p) ppGpp is a key regulator of bacterial metabolism, growth, stress tolerance, and virulence. During amino acid starvation, the Escherichia coli (p) ppGpp synthetase RelA is activated by deacylated tRNA in the ribosomal A-site. An increase in (p) ppGpp is believed to drive the formation of antibiotic-tolerant persister cells, prompting the development of strategies to inhibit (p) ppGpp synthesis. We show that in a biochemical system from purified E. coli components, the antibiotic thiostrepton efficiently inhibits RelA activation by the A-site tRNA. In bacterial cultures, the ribosomal inhibitors thiostrepton, chloramphenicol, and tetracycline all efficiently abolish accumulation of (p) ppGpp induced by the Ile-tRNA synthetase inhibitor mupirocin. This abolishment, however, does not reduce the persister level. In contrast, the combination of dihydrofolate reductase inhibitor trimethoprim with mupirocin, tetracycline, or chloramphenicol leads to ampicillin tolerance. The effect is independent of RelA functionality, specific to beta-lactams, and not observed with the fluoroquinolone norfloxacin. These results refine our understanding of (p) ppGpp's role in antibiotic tolerance and persistence and demonstrate unexpected drug interactions that lead to tolerance to bactericidal antibiotics.

  • 55.
    Kumar, Keshav
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Chromatographic analysis of peptidoglycan samples with the aid of a chemometric technique: introducing a novel analytical procedure to classify bacterial cell wall collection2019Ingår i: Analytical Methods, ISSN 1759-9660, E-ISSN 1759-9679, Vol. 11, nr 12, s. 1671-1679Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The technical development of liquid chromatography has provided the necessary sensitivity to characterise peptidoglycan samples. However, the analysis of large numbers of complex chromatographic data sets without the aid of a proper chemometric technique is a laborious task, carrying a high risk of losing important biochemical information. The present work describes the development of a simple analytical procedure using self-organising map (SOM) analysis to analyse the large number of complex chromatographic data sets from bacterial peptidoglycan samples. SOM analysis essentially maps the samples to a hexagonal sheet based on their compositional similarity, and thus provides an approach to classify the bacterial cell wall collection in an unsupervised manner. The utility of the proposed approach was successfully validated by analysing peptidoglycan samples belonging to the Alphaproteobacterium class. The classification results achieved with SOM analysis were found to correlate well with their relative similarity in peptidoglycan compositions. In summary, the SOM analysis-based analytical procedure is shown to be useful towards automatising the analyses of chromatographic data sets of peptidoglycan samples from bacterial collections.

  • 56.
    Kumar, Keshav
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi.
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi.
    Constraint randomised non-negative factor analysis (CRNNFA): an alternate chemometrics approach for analysing the biochemical data sets2017Ingår i: The Analyst, ISSN 0003-2654, E-ISSN 1364-5528, Vol. 142, nr 11, s. 1916-1928Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The present work introduces an alternate chemometrics approach constraint randomised non-negative factor analysis (CRNNFA) for analysing the bioanalytical data sets. The CRNNFA algorithm provides the outputs that are easy to interpret and correlate with the real chromatograms. The CRNNFA algorithm achieves termination when the iteration limit is reached circumventing the premature convergence. Theoretical and computational aspects of the proposed method are also described. The analytical and computational potential of CRNNFA are successfully tested by analysing the complex chromatograms of the peptidoglycan samples belonging to the Alphaproteobacterium members. The obtained results clearly show that CRNNFA can easily trace the compositional variability of the peptidoglycan samples. In summary, the proposed method in general can be a potential alternate approach for analysing the data sets obtained from different analytical and clinical fields.

  • 57.
    Kumar, Keshav
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Integrating network analysis with chromatography: introducing a novel chemometry-chromatography based analytical procedure to classify the bacterial cell wall collection2018Ingår i: Analytical Methods, ISSN 1759-9660, E-ISSN 1759-9679, Vol. 10, nr 10, s. 1172-1180Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The present work integrates network analysis with chromatography and proposes a novel analytical procedure to classify the bacterial cell wall collection. The network analysis model can capture the heterogeneity present in the datasets and hence can provide unsupervised classification. The proposed approach is successfully applied for classifying the peptidoglycan samples of certain bacterial collections belonging to the class of Alphaproteobacteria. The obtained classification results are found to correlate well with their relative similarity in the peptidoglycan compositions. In summary, the proposed network analysis approach can be helpful in automatizing the bacterial cell wall analysis. The proposed approach can be useful to accelerate the research related to understanding the morphology of bacterial cell walls, host-pathogen interaction and development of effective antibiotics.

  • 58.
    Kumar, Keshav
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Principal coordinate analysis assisted chromatographic analysis of bacterial cell wall collection: a robust classification approach2018Ingår i: Analytical Biochemistry, ISSN 0003-2697, E-ISSN 1096-0309, Vol. 550, s. 8-14Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    In the present work, Principal coordinate analysis (PCoA) is introduced to develop a robust model to classify the chromatographic data sets of peptidoglycan sample. PcoA captures the heterogeneity present in the data sets by using the dissimilarity matrix as input. Thus, in principle, it can even capture the subtle differences in the bacterial peptidoglycan composition and can provide a more robust and fast approach for classifying the bacterial collection and identifying the novel cell wall targets for further biological and clinical studies. The utility of the proposed approach is successfully demonstrated by analysing the two different kind of bacterial collections. The first set comprised of peptidoglycan sample belonging to different subclasses of Alphaproteobacteria. Whereas, the second set that is relatively more intricate for the chemometric analysis consist of different wild type Vibrio Cholerae and its mutants having subtle differences in their peptidoglycan composition. The present work clearly proposes a useful approach that can classify the chromatographic data sets of chromatographic peptidoglycan samples having subtle differences. Furthermore, present work clearly suggest that PCoA can be a method of choice in any data analysis workflow.

  • 59.
    Kumar, Keshav
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå centrum för molekylär medicin (UCMM). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Espaillat, Akbar
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå centrum för molekylär medicin (UCMM). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå centrum för molekylär medicin (UCMM). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    PG-metrics: a chemometric-based approach for classifying bacterial peptidoglycan data sets and uncovering their subjacent chemical variability2017Ingår i: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 12, nr 10, artikel-id e0186197Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Bacteria cells are protected from osmotic and environmental stresses by an exoskeleton-like polymeric structure called peptidoglycan ( PG) or murein sacculus. This structure is fundamental for bacteria's viability and thus, the mechanisms underlying cell wall assembly and how it is modulated serve as targets for many of our most successful antibiotics. Therefore, it is now more important than ever to understand the genetics and structural chemistry of the bacterial cell walls in order to find new and effective methods of blocking it for the treatment of disease. In the last decades, liquid chromatography and mass spectrometry have been demonstrated to provide the required resolution and sensitivity to characterize the fine chemical structure of PG. However, the large volume of data sets that can be produced by these instruments today are difficult to handle without a proper data analysis work-flow. Here, we present PG-metrics, a chemometric based pipeline that allows fast and easy classification of bacteria according to their muropeptide chromatographic profiles and identification of the subjacent PG chemical variability between e.g. bacterial species, growth conditions and, mutant libraries. The pipeline is successfully validated here using PG samples from different bacterial species and mutants in cell wall proteins. The obtained results clearly demonstrated that PG-metrics pipeline is a valuable bioanalytical tool that can lead us to cell wall classification and biomarker discovery.

  • 60.
    Kuru, Erkin
    et al.
    Indiana University, Bloomington, USA.
    Hughes, H Velocity
    Indiana University, Bloomington, USA.
    Brown, Pamela J
    Indiana University, Bloomington, USA.
    Hall, Edward
    Indiana University, Bloomington, USA.
    Tekkam, Srinivas
    Indiana University, Bloomington, USA.
    Cava, Felipe
    Universidad Autonoma de Madrid, Campus de Cantoblanco, Madrid, Spain.
    de Pedro, Miguel A
    Universidad Autonoma de Madrid, Campus de Cantoblanco, Madrid, Spain.
    Brun, Yves V
    Indiana University, Bloomington, USA.
    VanNieuwenhze, Michael S
    Indiana University, Bloomington, USA.
    In Situ probing of newly synthesized peptidoglycan in live bacteria with fluorescent D-amino acids2012Ingår i: Angewandte Chemie International Edition, ISSN 1433-7851, E-ISSN 1521-3773, Vol. 51, nr 50, s. 12519-12523Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Tracking a bug's life: Peptidoglycan (PG) of diverse bacteria is labeled by exploiting the tolerance of cells for incorporating different non-natural D-amino acids. These nontoxic D-amino acids preferably label the sites of active PG synthesis, thereby enabling fine spatiotemporal tracking of cell-wall dynamics in phylogenetically and morphologically diverse bacteria. HCC = 7-hydroxycoumarin, NBD = 7-nitrobenzofurazan, TAMRA = carboxytetramethylrhodamine.

  • 61.
    Lam, Hubert
    et al.
    Channing Laboratory, Brigham and Women’s Hospital, Harvard Medical School, and Howard Hughes Medical Institute, Boston, MA, USA.
    Oh, Dong-Chan
    Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA, USA.
    Cava, Felipe
    Channing Laboratory, Brigham and Women’s Hospital, Harvard Medical School, and Howard Hughes Medical Institute, Boston, MA, USA.
    Takacs, Constantin N
    Channing Laboratory, Brigham and Women’s Hospital, Harvard Medical School, and Howard Hughes Medical Institute, Boston, MA, USA.
    Clardy, Jon
    Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA, USA.
    de Pedro, Miguel A
    CBM ‘Severo Ochoa’ CSIC-UAM, Madrid, Spain.
    Waldor, Matthew K
    Channing Laboratory, Brigham and Women’s Hospital, Harvard Medical School, and Howard Hughes Medical Institute, Boston, MA, USA.
    D-amino acids govern stationary phase cell wall remodeling in bacteria2009Ingår i: Science, ISSN 0036-8075, E-ISSN 1095-9203, Vol. 325, nr 5947, s. 1552-1555Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    In all known organisms, amino acids are predominantly thought to be synthesized and used as their L-enantiomers. Here, we found that bacteria produce diverse D-amino acids as well, which accumulate at millimolar concentrations in supernatants of stationary phase cultures. In Vibrio cholerae, a dedicated racemase produced D-Met and D-Leu, whereas Bacillus subtilis generated D-Tyr and D-Phe. These unusual D-amino acids appear to modulate synthesis of peptidoglycan, a strong and elastic polymer that serves as the stress-bearing component of the bacterial cell wall. D-Amino acids influenced peptidoglycan composition, amount, and strength, both by means of their incorporation into the polymer and by regulating enzymes that synthesize and modify it. Thus, synthesis of D-amino acids may be a common strategy for bacteria to adapt to changing environmental conditions.

  • 62.
    Laptenko, Oleg
    et al.
    Department of Cell Biology, School of Osteopathic Medicine at Stratford, University of Medicine and Dentistry of New Jersey, Stratford, NJ, USA.
    Kim, Seung-Sup
    Department of Biochemistry, New York University School of Medicine, New York, NY, USA.
    Lee, Jookyung
    Department of Cell Biology, School of Osteopathic Medicine at Stratford, University of Medicine and Dentistry of New Jersey, Stratford, NJ, USA.
    Starodubtseva, Marina
    Department of Cell Biology, School of Osteopathic Medicine at Stratford, University of Medicine and Dentistry of New Jersey, Stratford, NJ, USA.
    Cava, Felipe
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    Berenguer, Jose
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    Kong, Xiang-Peng
    Department of Biochemistry, New York University School of Medicine, New York, NY, USA.
    Borukhov, Sergei
    Department of Cell Biology, School of Osteopathic Medicine at Stratford, University of Medicine and Dentistry of New Jersey, Stratford, NJ, USA.
    pH-dependent conformational switch activates the inhibitor of transcription elongation2006Ingår i: EMBO Journal, ISSN 0261-4189, E-ISSN 1460-2075, Vol. 25, nr 10, s. 2131-2141Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Gfh1, a transcription factor from Thermus thermophilus, inhibits all catalytic activities of RNA polymerase (RNAP). We characterized the Gfh1 structure, function and possible mechanism of action and regulation. Gfh1 inhibits RNAP by competing with NTPs for coordinating the active site Mg2+ ion. This coordination requires at least two aspartates at the tip of the Gfh1 N-terminal coiled-coil domain (NTD). The overall structure of Gfh1 is similar to that of the Escherichia coli transcript cleavage factor GreA, except for the flipped orientation of the C-terminal domain (CTD). We show that depending on pH, Gfh1-CTD exists in two alternative orientations. At pH above 7, it assumes an inactive 'flipped' orientation seen in the structure, which prevents Gfh1 from binding to RNAP. At lower pH, Gfh1-CTD switches to an active 'Gre-like' orientation, which enables Gfh1 to bind to and inhibit RNAP.

  • 63. LeRoux, Frederique
    et al.
    Wegner, K. Mathias
    Baker-Austin, Craig
    Vezzulli, Luigi
    Osorio, Carlos R.
    Amaro, Carmen
    Ritchie, Jennifer M.
    Defoirdt, Tom
    Destoumieux-Garzon, Delphine
    Blokesch, Melanie
    Mazel, Didier
    Jacq, Annick
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Gram, Lone
    Wendling, Carolin C.
    Strauch, Eckhard
    Kirschner, Alexander
    Huehn, Stephan
    The emergence of Vibrio pathogens in Europe: ecology, evolution, and pathogenesis (Paris, 11-12th March 2015)2015Ingår i: Frontiers in Microbiology, ISSN 1664-302X, E-ISSN 1664-302X, Vol. 6, artikel-id 830Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Global change has caused a worldwide increase in reports of Vibrio-associated diseases with ecosystem-wide impacts on humans and marine animals. In Europe, higher prevalence of human infections followed regional climatic trends with outbreaks occurring during episodes of unusually warm weather. Similar patterns were also observed in Vibrio-associated diseases affecting marine organisms such as fish, bivalves and corals. Basic knowledge is still lacking on the ecology and evolutionary biology of these bacteria as well as on their virulence mechanisms. Current limitations in experimental systems to study infection and the lack of diagnostic tools still prevent a better understanding of Vibrio emergence. A major challenge is to foster cooperation between fundamental and applied research in order to investigate the consequences of pathogen emergence in natural Vibrio populations and answer federative questions that meet societal needs. Here we report the proceedings of the first European workshop dedicated to these specific goals of the Vibrio research community by connecting current knowledge to societal issues related to ocean health and food security.

  • 64. Maciejewska, B
    et al.
    Roszniowski, B
    Espaillat, Akbar
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Kęsik-Szeloch, A
    Majkowska-Skrobek, G
    Kropinski, AM
    Briers, Y
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Lavigne, R
    Drulis-Kawa, Z
    Klebsiella phages representing a novel clade of viruses with an unknown DNA modification and biotechnologically interesting enzymes2017Ingår i: Applied Microbiology and Biotechnology, ISSN 0175-7598, E-ISSN 1432-0614, Vol. 101, nr 2, s. 673-684Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Lytic bacteriophages and phage-encoded endolysins (peptidoglycan hydrolases) provide a source for the development of novel antimicrobial strategies. In the present study, we focus on the closely related (96 % DNA sequence identity) environmental myoviruses vB_KpnM_KP15 (KP15) and vB_KpnM_KP27 (KP27) infecting multidrug-resistant Klebsiella pneumoniae and Klebsiella oxytoca strains. Their genome organisation and evolutionary relationship are compared to Enterobacter phage phiEap-3 and Klebsiella phages Matisse and Miro. Due to the shared and distinct evolutionary history of these phages, we propose to create a new phage genus BKp15virus^ within the Tevenvirinae subfamily. In silico genome analysis reveals two unique putative homing endonucleases of KP27 phage, probably involved in unrevealed mechanism of DNA modification and resistance to restriction digestion, resulting in a broader host spectrum. Additionally, we identified in KP15 and KP27 a complete set of lysis genes, containing holin, antiholin, spanin and endolysin. By turbidimetric assays on permeabilized Gram-negative strains, we verified the ability of the KP27 endolysin to destroy the bacterial peptidoglycan. We confirmed high stability, absence of toxicity on a human epithelial cell line and the enzymatic specificity of endolysin, which was found to possess endopeptidase activity, cleaving the peptide stem between L-alanine and D-glutamic acid.

  • 65. Maciejewska, Barbara
    et al.
    Zrubek, Karol
    Espaillat, Akbar
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Wisniewska, Magdalena
    Rembacz, Krzysztof P.
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Dubin, Grzegorz
    Drulis-Kawa, Zuzanna
    Modular endolysin of Burkholderia AP3 phage has the largest lysozyme-like catalytic subunit discovered to date and no catalytic aspartate residue2017Ingår i: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 7, artikel-id 14501Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Endolysins are peptidoglycan-degrading enzymes utilized by bacteriophages to release the progeny from bacterial cells. The lytic properties of phage endolysins make them potential antibacterial agents for medical and industrial applications. Here, we present a comprehensive characterization of phage AP3 modular endolysin (AP3gp15) containing cell wall binding domain and an enzymatic domain (DUF3380 by BLASTP), both widespread and conservative. Our structural analysis demonstrates the low similarity of an enzymatic domain to known lysozymes and an unusual catalytic centre characterized by only a single glutamic acid residue and no aspartic acid. Thus, our findings suggest distinguishing a novel class of muralytic enzymes having the activity and catalytic centre organization of DUF3380. The lack of amino acid sequence homology between AP3gp15 and other known muralytic enzymes may reflect the evolutionary convergence of analogous glycosidases. Moreover, the broad antibacterial spectrum, lack of cytotoxic effect on human cells and the stability characteristics of AP3 endolysin advocate for its future application development.

  • 66.
    Moell, Andrea
    et al.
    Boston, Massachusetts, USA .
    Doerr, Tobias
    Boston, Massachusetts, USA .
    Alvarez, Laura
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Chao, Michael C.
    Boston, Massachusetts, USA .
    Davis, Brigid M.
    Boston, Massachusetts, USA .
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Waldor, Matthew K.
    Boston, Massachusetts, USA .
    Cell Separation in Vibrio cholerae Is Mediated by a Single Amidase Whose Action Is Modulated by Two Nonredundant Activators2014Ingår i: Journal of Bacteriology, ISSN 0021-9193, E-ISSN 1098-5530, Vol. 196, nr 22, s. 3937-3948Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Synthesis and hydrolysis of septal peptidoglycan (PG) are critical processes at the conclusion of cell division that enable separation of daughter cells. Cleavage of septal PG is mediated by PG amidases, hydrolytic enzymes that release peptide side chains from the glycan strand. Most gammaproteobacteria, including Escherichia coli, encode several functionally redundant periplasmic amidases. However, members of the Vibrio genus, including the enteric pathogen Vibrio cholerae, encode only a single PG amidase, AmiB. Here, we show that V. cholerae AmiB is crucial for cell division and growth. Genetic and biochemical analyses indicated that AmiB is regulated by two activators, EnvC and NlpD, at least one of which is required for AmiB's localization to the cell division site. Localization of the activators (and thus of AmiB) is dependent upon the cell division protein FtsN. These factors mediate septal PG cleavage in E. coli as well; however, their precise roles vary between the two organisms in a number of ways. Notably, even though V. cholerae EnvC and NlpD appear to be functionally redundant under most growth conditions tested, NlpD is specifically required for intestinal colonization in the infant mouse model of cholera and for V. cholerae resistance against bile salts, perhaps due to environmental regulation of AmiB or its activators. Collectively, our findings reveal that although the cellular components that enable cleavage of septal PG appear to be generally conserved between E. coli and V. cholerae, they can be combined into diverse functional regulatory networks.

  • 67.
    Moreno, Renata
    et al.
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    Hidalgo, Aurelio
    Departamento de Biocatálisis, Instituto de Catálisis y Petroleoquímica-CSIC, Campus de Cantoblanco, Madrid, Spain.
    Cava, Felipe
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    Fernández-Lafuente, Roberto
    Departamento de Biocatálisis, Instituto de Catálisis y Petroleoquímica-CSIC, Campus de Cantoblanco, Madrid, Spain.
    Guisán, José Manuel
    Departamento de Biocatálisis, Instituto de Catálisis y Petroleoquímica-CSIC, Campus de Cantoblanco, Madrid, Spain.
    Berenguer, José
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    Use of an antisense RNA strategy to investigate the functional significance of Mn-catalase in the extreme thermophile Thermus thermophilus2004Ingår i: Journal of Bacteriology, ISSN 0021-9193, E-ISSN 1098-5530, Vol. 186, nr 22, s. 7804-7806Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The expression of an antisense RNA revealed that an Mn-catalase was required in Thermus thermophilus for aerobic but not for anaerobic growth. The antisense system is based on the constitutive expression of a "bicistronic" transcript consisting of the kanamycin resistance gene mRNA followed by the antisense RNA against the selected target.

  • 68.
    Moreno, Renata
    et al.
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    Zafra, Olga
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    Cava, Felipe
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    Berenguer, José
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    Development of a gene expression vector for Thermus thermophilus based on the promoter of the respiratory nitrate reductase2003Ingår i: Plasmid, ISSN 0147-619X, E-ISSN 1095-9890, Vol. 49, nr 1, s. 2-8Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    A specific expression system for Thermus spp. is described. Plasmid pMKE1 contains replicative origins for Escherichia coli and Thermus spp., a selection gene encoding a thermostable resistance to kanamycin, and a 720 bp DNA region containing the promoter (Pnar), and the regulatory sequences of the respiratory nitrate reductase operon of Thermus thermophilus HB8. Two genes, encoding a thermophilic beta-galactosidase and an alkaline phosphatase were cloned in pMKE1 as cytoplasmic and periplasmic reporters, respectively. The expression of the reporters was specifically induced by the combined action of nitrate and anoxia in facultative anaerobic derivatives of T. thermophilus HB27 to which the gene cluster for nitrate respiration was transferred by conjugation. Overexpressions in the range of approximately 200-fold were obtained for the cytoplasmic reporter, whereas that of the periplasmic reporter was limited to approximately 20-fold, with respect to their intrinsic respective activities.

  • 69. Moreno-Guzman, Maria
    et al.
    Garcia-Carmona, Laura
    Molinero-Fernandez, Agueda
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Lopez Gil, Miguel Angel
    Escarpa, Alberto
    Bi-enzymatic biosensor for on-site, fast and reliable electrochemical detection of relevant D-amino acids in bacterial samples2017Ingår i: Sensors and actuators. B, Chemical, ISSN 0925-4005, E-ISSN 1873-3077, Vol. 242, s. 95-101Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    In this work, a bi-enzymatic biosensor allowed the total content of D-amino acids (DAAs) determination in highly relevant matrices involving bacteria. The strategy is based on the unique coimmobilization of D-amino acid oxidase (DAAO) and horseradish peroxidase (HRP) enzymes onto a multi-walled carbon nanotubes (MWCNTs) and gold nanoparticles (AuNPs) modified screen-printed electrode (SPCE). The greater amount of AuNPs deposited and hence the greater loading of both enzymes was observed when they were deposited after the activation of the carboxylated MWCNTs with EDC/Sulfo-NHS chemistry. These platforms provided a fast (300s) and selective quantification of DAAs with excellent precision (RSD < 5%) and accuracy (Recoveries 100-104%) in bacterial samples. Collectively, the electrochemical bi-enzymatic biosensor become an universal, fast, sensitive and easy-to-use approach to determine total content of DAAs in complex matrices.

  • 70. Murphy, Shannon G.
    et al.
    Alvarez, Laura
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Adams, Myfanwy C.
    Liu, Shuning
    Chappie, Joshua S.
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Dorr, Tobias
    Endopeptidase Regulation as a Novel Function of the Zur-Dependent Zinc Starvation Response2019Ingår i: mBio, ISSN 2161-2129, E-ISSN 2150-7511, Vol. 10, nr 1, artikel-id e02620-18Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The cell wall is a strong, yet flexible, meshwork of peptidoglycan (PG) that gives a bacterium structural integrity. To accommodate a growing cell, the wall is remodeled by both PG synthesis and degradation. Vibrio cholerae encodes a group of three nearly identical zinc-dependent endopeptidases (EPs) that are predicted to hydrolyze PG to facilitate cell growth. Two of these (ShyA and ShyC) are conditionally essential housekeeping EPs, while the third (ShyB) is not expressed under standard laboratory conditions. To investigate the role of ShyB, we conducted a transposon screen to identify mutations that activate shyB transcription. We found that shyB is induced as part of the Zur-mediated zinc starvation response, a mode of regulation not previously reported for cell wall lytic enzymes. In vivo, ShyB alone was sufficient to sustain cell growth in low-zinc environments. In vitro, ShyB retained its D, D-endopeptidase activity against purified sacculi in the presence of the metal chelator EDTA at concentrations that inhibit ShyA and ShyC. This insensitivity to metal chelation is likely what enables ShyB to substitute for other EPs during zinc starvation. Our survey of transcriptomic data from diverse bacteria identified other candidate Zur-regulated EPs, suggesting that this adaptation to zinc starvation is employed by other Gram-negative bacteria. IMPORTANCE Bacteria encode a variety of adaptations that enable them to survive during zinc starvation, a condition which is encountered both in natural environments and inside the human host. In Vibrio cholerae, the causative agent of the diarrheal disease cholera, we have identified a novel member of this zinc starvation response, a cell wall hydrolase that retains function and is conditionally essential for cell growth in low-zinc environments. Other Gram-negative bacteria contain homologs that appear to be under similar regulatory control. These findings are significant because they represent, to our knowledge, the first evidence that zinc homeostasis influences cell wall turnover. Anti-infective therapies commonly target the bacterial cell wall; therefore, an improved understanding of how the cell wall adapts to host-induced zinc starvation could lead to new antibiotic development. Such therapeutic interventions are required to combat the rising threat of drug-resistant infections.

  • 71. Möll, Andrea
    et al.
    Dörr, Tobias
    Alvarez, Laura
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Davis, Brigid M
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Waldor, Matthew K
    A D, D-carboxypeptidase is required for Vibrio cholerae halotolerance2015Ingår i: Environmental Microbiology, ISSN 1462-2912, E-ISSN 1462-2920, Vol. 17, nr 2, s. 527-540Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The biological roles of low molecular weight penicillin-binding proteins (LMW PBP) have been difficult to discern in Gram-negative organisms. In Escherichia coli, mutants lacking these proteins often have no phenotype, and cells lacking all seven LMW PBPs remain viable. In contrast, we report here that Vibrio cholerae lacking DacA-1, a PBP5 homologue, displays slow growth, aberrant morphology and altered peptidoglycan (PG) homeostasis in Luria-Bertani (LB) medium, as well as a profound plating defect. DacA-1 alone among V.cholerae'sLMW PBPs is critical for bacterial growth; mutants lacking the related protein DacA-2 and/or homologues of PBP4 or PBP7 displayed normal growth and morphology. Remarkably, the growth and morphology of the dacA-1 mutant were unimpaired in LB media containing reduced concentrations of NaCl (100mM or less), and also within suckling mice, a model host for the study of cholera pathogenesis. Peptidoglycan from the dacA-1 mutant contained elevated pentapeptide levels in standard and low salt media, and comparative analyses suggest that DacA-1 is V.cholerae's principal DD-carboxypeptidase. The basis for the dacA-1 mutant's halosensitivity is unknown; nonetheless, the mutant's survival in biochemically uncharacterized environments (such as the suckling mouse intestine) can be used as a reporter of low Na+ content.

  • 72. Orrego, Alejandro H.
    et al.
    Lopez-Gallego, Fernando
    Espaillat, Akbar
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Guisan, Jose M.
    Rocha-Martin, Javier
    One‐step Synthesis of α‐Keto Acids from Racemic Amino Acids by A Versatile Immobilized Multienzyme Cell‐free System2018Ingår i: ChemCatChem, ISSN 1867-3880, E-ISSN 1867-3899, Vol. 10, nr 14, s. 3002-3011Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The elevated value of α‐keto acids has pushed scientists to explore more efficient and less expensive alternatives for their synthesis. In this work, an immobilized tri‐enzyme system that produced α‐keto acids in “one‐pot” from l‐ or racemic mixtures of diverse amino acids was presented. The system combined a broad‐spectrum amino acid racemase (BsrV), a d‐amino acid oxidase (DAAO) and catalase (CAT). BsrV racemized l‐amino acids into their d‐enantiomers, DAAO catalyzed the stereospecific oxidative deamination of the d‐amino acids into their corresponding α‐keto acids, ammonium ion, and H2O2. Finally, CAT converted the inactivating H2O2 into H2O and O2, which can be reused by the oxidase reaction. BsrV thermal stability was improved 3,300‐fold by immobilizing the enzyme on glyoxyl‐activated agarose beads. DAAO and CAT were co‐immobilized on agarose beads functionalized with glutaraldehyde groups for enhancing their stabilities and eliminating H2O2 in a much more effective way. To show the versatility of this system, racemic mixtures of amino acids were converted in their corresponding α‐keto acids. The coupling of the three immobilized enzymes permitted conversions of approximately 99 % through a dynamic kinetic resolution process. This system conserved 100 % of its initial effectiveness after 8 reaction cycles. Collectively, our innovative tri‐enzyme system for the synthesis of α‐keto acids opens the door for a cheapening in the production of many pharmaceutical and cosmetics.

  • 73.
    Perales, Celia
    et al.
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    Cava, Felipe
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    Meijer, Wilfried J J
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    Berenguer, José
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    Enhancement of DNA, cDNA synthesis and fidelity at high temperatures by a dimeric single-stranded DNA-binding protein2003Ingår i: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962, Vol. 31, nr 22, s. 6473-6480Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Bacterial single-stranded DNA-binding proteins (SSBs) are required for DNA replication and repair. We have over-expressed and purified the native form and two His-tagged fusions of the SSB from Thermus thermophilus (TthSSB). The three proteins were found as dimers in solution. They bound in vitro to single-stranded DNA specifically over a temperature range of 4-80 degrees C, and the wild-type protein could withstand incubation at 94 degrees C for 2 min. Addition of TthSSB to PCR halved the elongation time required for the DNA polymerases of T.thermophilus (Tth) and Pyrococcus furiosus (Pfu) to synthesise DNA fragments in PCRs. The presence of TthSSB increased the fidelity of the proof- reading-free DNA polymerase of T.thermophilus. TthSSB was also able to bind single-stranded RNA, allowing a dramatic enhancement of the reverse transcription activity of its cognate Tth DNA polymerase during cDNA synthesis.

  • 74. Siegrist, M. Sloan
    et al.
    Aditham, Arjun K.
    Espaillat, Akbar
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Cameron, Todd A.
    Whiteside, Sarah A.
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Portnoy, Daniel A.
    Bertozzi, Carolyn R.
    Host Actin Polymerization Tunes the Cell Division Cycle of an Intracellular Pathogen2015Ingår i: Cell reports, ISSN 2211-1247, E-ISSN 2211-1247, Vol. 11, nr 4, s. 499-507Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Growth and division are two of the most fundamental capabilities of a bacterial cell. While they are well described for model organisms growing in broth culture, very little is known about the cell division cycle of bacteria replicating in more complex environments. Using a D-alanine reporter strategy, we found that intracellular Listeria monocytogenes (Lm) spend a smaller proportion of their cell cycle dividing compared to Lm growing in broth culture. This alteration to the cell division cycle is independent of bacterial doubling time. Instead, polymerization of host-derived actin at the bacterial cell surface extends the non-dividing elongation period and compresses the division period. By decreasing the relative proportion of dividing Lm, actin polymerization biases the population toward cells with the highest propensity to form actin tails. Thus, there is a positive-feedback loop between the Lm cell division cycle and a physical interaction with the host cytoskeleton.

  • 75. Siegrist, M Sloan
    et al.
    Whiteside, Sarah
    Jewett, John C
    Aditham, Arjun
    Cava, Felipe
    Centro de Biologia Molecular Severo Ochoa, Universidad Autonoma de Madrid, Madrid, Spain.
    Bertozzi, Carolyn R
    (D)-amino acid chemical reporters reveal peptidoglycan dynamics of an intracellular pathogen2013Ingår i: ACS Chemical Biology, ISSN 1554-8929, E-ISSN 1554-8937, Vol. 8, nr 3, s. 500-505Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Peptidoglycan (PG) is an essential component of the bacterial cell wall. Although experiments with organisms in vitro have yielded a wealth of information on PG synthesis and maturation, it is unclear how these studies translate to bacteria replicating within host cells. We report a chemical approach for probing PG in vivo via metabolic labeling and bioorthogonal chemistry. A wide variety of bacterial species incorporated azide and alkyne-functionalized d-alanine into their cell walls, which we visualized by covalent reaction with click chemistry probes. The d-alanine analogues were specifically incorporated into nascent PG of the intracellular pathogen Listeria monocytogenes both in vitro and during macrophage infection. Metabolic incorporation of d-alanine derivatives and click chemistry detection constitute a facile, modular platform that facilitates unprecedented spatial and temporal resolution of PG dynamics in vivo.

  • 76. Terceti, Mateus S.
    et al.
    Rivas, Amable J.
    Alvarez, Laura
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Noia, Manuel
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Osorio, Carlos R.
    rstB Regulates Expression of the Photobacterium damselae subsp damselae Major Virulence Factors Damselysin, Phobalysin P and Phobalysin C2017Ingår i: Frontiers in Microbiology, ISSN 1664-302X, E-ISSN 1664-302X, Vol. 8, artikel-id 582Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The marine pathogenic bacterium Photobacterium damselae subsp. damselae causes septicemia in marine animals and in humans. The pPHDD1 plasmid-encoded hemolysins damselysin (Dly) and phobalysin P (PhlyP), and the chromosome-encoded hemolysin phobalysin C (PhlyC) constitute its main virulence factors. However, the mechanisms by which expression of these three hemolysins is regulated remain unknown. Here we report the isolation of a mini-Tn10 transposon mutant which showed a strong impairment in its hemolytic activity. The transposon disrupted a putative sensor histidine kinase gene vda_000600 (rstB), which together with vda_000601 (rstA) is predicted to encode a putative two-component regulatory system. This system showed to be homologous to the Vibrio cholerae CarSR/VprAB and Escherichia coli RstAB systems. Reconstruction of the mutant by allelic exchange of rstB showed equal impairment in hemolysis, and complementation with a plasmid expressing rstAB restored hemolysis to wild-type levels. Remarkably, we demonstrated by promoter expression analyses that the reduced hemolysis in the rstB mutant was accompanied by a strong decrease in transcription activities of the three hemolysin genes dly (damselysin), hlyA(pl) (phobalysin P) and hlyA(ch) (phobalysin C). Thus, RstB, encoded in the small chromosome, regulates plasmid and chromosomal virulence genes. We also found that reduced expression of the three virulence genes correlated with a strong decrease in virulence in a sea bass model, demonstrating that RstB constitutes a master regulator of the three P. damselae subsp. damselae hemolysins and plays critical roles in the pathogenicity of this bacterium. This study represents the first evidence of a direct role of a RstAB-like system in the regulation of bacterial toxins.

  • 77.
    van Teeseling, Muriel C. F.
    et al.
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    de Pedro, Miguel A.
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Determinants of Bacterial Morphology: From Fundamentals to Possibilities for Antimicrobial Targeting2017Ingår i: Frontiers in Microbiology, ISSN 1664-302X, E-ISSN 1664-302X, Vol. 8, artikel-id 1264Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Bacterial morphology is extremely diverse. Specific shapes are the consequence of adaptive pressures optimizing bacterial fitness. Shape affects critical biological functions, including nutrient acquisition, motility, dispersion, stress resistance and interactions with other organisms. Although the characteristic shape of a bacterial species remains unchanged for vast numbers of generations, periodical variations occur throughout the cell (division) and life cycles, and these variations can be influenced by environmental conditions. Bacterial morphology is ultimately dictated by the net-like peptidoglycan (PG) sacculus. The species-specific shape of the PG sacculus at any time in the cell cycle is the product of multiple determinants. Some morphological determinants act as a cytoskeleton to guide biosynthetic complexes spatiotemporally, whereas others modify the PG sacculus after biosynthesis. Accumulating evidence supports critical roles of morphogenetic processes in bacteria-host interactions, including pathogenesis. Here, we review the molecular determinants underlying morphology, discuss the evidence linking bacterial morphology to niche adaptation and pathogenesis, and examine the potential of morphological determinants as antimicrobial targets.

  • 78. van Teeseling, Muriel C. F.
    et al.
    Mesman, Rob J.
    Kuru, Erkin
    Espaillat, Akbar
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS).
    Brun, Yves V.
    VanNieuwenhze, Michael S.
    Kartal, Boran
    van Niftrik, Laura
    Anammox Planctomycetes have a peptidoglycan cell wall2015Ingår i: Nature Communications, ISSN 2041-1723, E-ISSN 2041-1723, Vol. 6, artikel-id 6878Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Planctomycetes are intriguing microorganisms that apparently lack peptidoglycan, a structure that controls the shape and integrity of almost all bacterial cells. Therefore, the planctomycetal cell envelope is considered exceptional and their cell plan uniquely compartmentalized. Anaerobic ammonium-oxidizing (anammox) Planctomycetes play a key role in the global nitrogen cycle by releasing fixed nitrogen back to the atmosphere as N-2. Here using a complementary array of state-of-the-art techniques including continuous culturing, cryo-transmission electron microscopy, peptidoglycan-specific probes and muropeptide analysis, we show that the anammox bacterium Kuenenia stuttgartiensis contains peptidoglycan. On the basis of the thickness, composition and location of peptidoglycan in K. stuttgartiensis, we propose to redefine Planctomycetes as Gram-negative bacteria. Our results demonstrate that Planctomycetes are not an exception to the universal presence of peptidoglycan in bacteria.

  • 79. Vanderlinde, Elizabeth M.
    et al.
    Strozen, Timothy G.
    Hernandez, Sara B.
    Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Howard, S. Peter
    Alterations in Peptidoglycan Cross-Linking Suppress the Secretin Assembly Defect Caused by Mutation of GspA in the Type II Secretion System2017Ingår i: Journal of Bacteriology, ISSN 0021-9193, E-ISSN 1098-5530, Vol. 199, nr 8, artikel-id UNSP e00617-16Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    In Gram-negative bacteria, the peptidoglycan (PG) cell wall is a significant structural barrier for outer membrane protein assembly. In Aeromonas hydrophila, outer membrane multimerization of the type II secretion system (T2SS) secretin ExeD requires the function of the inner membrane assembly factor complex ExeAB. The putative mechanism of the complex involves the reorganization of PG and localization of ExeD, whereby ExeA functions by interacting with PG to form a site for secretin assembly and ExeB forms an interaction with ExeD. This mechanism led us to hypothesize that increasing the pore size of PG would circumvent the requirement for ExeA in the assembly of the ExeD secretin. Growth of A. hydrophila in 270 mM Gly reduced PG cross-links by approximately 30% and led to the suppression of secretin assembly defects in exeA strains and in those expressing ExeA mutants by enabling localization of the secretin in the outer membrane. We also established a heterologous ExeD assembly system in Escherichia coli and showed that ExeAB and ExeC are the only A. hydrophila proteins required for the assembly of the ExeD secretin in E. coli and that ExeAB-independent assembly of ExeD can occur upon overexpression of the D, D-carboxypeptidase PBP 5. These results support an assembly model in which, upon binding to PG, ExeA induces multimerization and pore formation in the sacculus, which enables ExeD monomers to interact with ExeB and assemble into a secretin that both is inserted in the outer membrane and crosses the PG layer to interact with the inner membrane platform of the T2SS. IMPORTANCE The PG layer imposes a strict structural impediment for the assembly of macromolecular structures that span the cell envelope and serve as virulence factors in Gram-negative species. This work revealed that by decreasing PG crosslinking by growth in Gly, the absolute requirement for the PG-binding activity of ExeA in the assembly of the ExeD secretin was alleviated in A. hydrophila. In a heterologous assembly model in E. coli, expression of the carboxypeptidase PBP 5 could relieve the requirement for ExeAB in the assembly of the ExeD secretin. These results provide some mechanistic details of the ExeAB assembly complex function, in which the PG-binding and oligomerization functions of ExeAB are used to create a pore in the PG that is required for secretin assembly.

  • 80.
    Yadav, Akhilesh K.
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Espaillat, Akbar
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Cava, Felipe
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Bacterial Strategies to Preserve Cell Wall Integrity Against Environmental Threats2018Ingår i: Frontiers in Microbiology, ISSN 1664-302X, E-ISSN 1664-302X, Vol. 9, artikel-id 2064Artikel, forskningsöversikt (Refereegranskat)
    Abstract [en]

    Bacterial cells are surrounded by an exoskeleton-like structure, the cell wall, composed primarily of the peptidoglycan (PG) sacculus. This structure is made up of glycan strands cross-linked by short peptides generating a covalent mesh that shapes bacteria and prevents their lysis due to their high internal osmotic pressure. Even though the PG is virtually universal in bacteria, there is a notable degree of diversity in its chemical structure. Modifications in both the sugars and peptides are known to be instrumental for bacteria to cope with diverse environmental challenges. In this review, we summarize and discuss the cell wall strategies to withstand biotic and abiotic environmental insults such as the effect of antibiotics targeting cell wall enzymes, predatory PG hydrolytic proteins, and PG signaling systems. Finally we will discuss the opportunities that species-specific PG variability might open to develop antimicrobial therapies.

  • 81.
    Zafra, Olga
    et al.
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    Cava, Felipe
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    Blasco, Francis
    Laboratoire de Chimie Bactérienne, Institut de Biologie Structurale et Microbiologie, CNRS 31, Marseille, France.
    Magalon, Axel
    Laboratoire de Chimie Bactérienne, Institut de Biologie Structurale et Microbiologie, CNRS 31, Marseille, France.
    Berenguer, Jose
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    Membrane-associated maturation of the heterotetrameric nitrate reductase of Thermus thermophilus2005Ingår i: Journal of Bacteriology, ISSN 0021-9193, E-ISSN 1098-5530, Vol. 187, nr 12, s. 3990-3996Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The nar operon, coding for the respiratory nitrate reductase of Thermus thermophilus (NRT), encodes a di-heme b-type (NarJ) and a di-heme c-type (NarC) cytochrome. The role of both cytochromes and that of a putative chaperone (NarJ) in the synthesis and maturation of NRT was studied. Mutants of T. thermophilus lacking either NarI or NarC synthesized a soluble form of NarG, suggesting that a putative NarCI complex constitutes the attachment site for the enzyme. Interestingly, the NarG protein synthesized by both mutants was inactive in nitrate reduction and misfolded, showing that membrane attachment was required for enzyme maturation. Consistent with its putative role as a specific chaperone, inactive and misfolded NarG was synthesized by narJ mutants, but in contrast to its Escherichia coli homologue, NarJ was also required for the attachment of the thermophilic enzyme to the membrane. A bacterial two-hybrid system was used to demonstrate the putative interactions between the NRT proteins suggested by the analysis of the mutants. Strong interactions were detected between NarC and NarI and between NarG and NarJ. Weaker interaction signals were detected between NarI, but not NarC, and both NarG and NarH. These results lead us to conclude that the NRT is a heterotetrameric (NarC/NarI/NarG/NarH) enzyme, and we propose a model for its synthesis and maturation that is distinct from that of E. coli. In the synthesis of NRT, a NarCI membrane complex and a soluble NarGJH complex are synthesized in a first step. In a second step, both complexes interact at the cytoplasmic face of the membrane, where the enzyme is subsequently activated with the concomitant conformational change and release of the NarJ chaperone from the mature enzyme.

  • 82.
    Zafra, Olga
    et al.
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    Ramírez, Sandra
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    Castán, Pablo
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    Moreno, Renata
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    Cava, Felipe
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    Vallés, Cristina
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    Caro, Eddy
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    Berenguer, José
    Centro de Biología Molecular 'Severo Ochoa' CSIC-UAM, Campus de Cantoblanco, Madrid, Spain.
    A cytochrome c encoded by the nar operon is required for the synthesis of active respiratory nitrate reductase in Thermus thermophilus2002Ingår i: FEBS Letters, ISSN 0014-5793, E-ISSN 1873-3468, Vol. 523, nr 1-3, s. 99-102Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    A cytochrome c (NarC) is encoded as the first gene of the operon for nitrate respiration in Thermus thermophilus. NarC is required for anaerobic growth and for the synthesis of active nitrate reductase (NR). The alpha and delta subunits (NarG, NarJ) of the NR were constitutively expressed in narC::kat mutants, but NarG appeared in the soluble fraction instead of associated with the membranes. Our data demonstrate for NarC an essential role in the synthesis of active enzyme and for the attachment to the membrane of the respiratory NR from T. thermophilus.

12 51 - 82 av 82
RefereraExporteraLänk till träfflistan
Permanent länk
Referera
Referensformat
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Annat format
Fler format
Språk
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Annat språk
Fler språk
Utmatningsformat
  • html
  • text
  • asciidoc
  • rtf