Ola Johnsborg

Norwegian University of Life Sciences

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Leiv Sigve Håvarstein

Norwegian University of Life Sciences

Ingolf F. Nes

Norwegian University of Life Sciences

Vegard Eldholm

Norwegian Institute of Public Health

Dzung B. Diep

Norwegian University of Life Sciences

Jean‐Pierre Claverys

Université Toulouse III - Paul Sabatier

Daniela Vollmer

Newcastle University

Waldemar Vollmer

Newcastle University

Robert M. Cleverley

Newcastle University

Hilde Solheim Ohnstad

Norwegian University of Life Sciences

P. A. Risøen

University of Oslo

Cooperative Institutions

Norwegian University of Life Sciences

Newcastle University

The Research Council of Norway

Centre National de la Recherche Scientifique

Université Toulouse III - Paul Sabatier

Laboratoire de Microbiologie et Génétique Moléculaires

University of Oslo

Université de Toulouse

University of Kaiserslautern

University of Newcastle Australia

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Exploration of antimicrobial potential in LAB by genomics

Current Opinion in Biotechnology (2004)

Ingolf F. Nes Ola Johnsborg

Food Preservatives

Identification

10.1016/j.copbio.2004.02.001

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Citations (115)

New insights into the pneumococcal fratricide: relationship to clumping and identification of a novel immunity factor

Molecular Microbiology (2005)

Leiv Sigve Håvarstein Bernard Martin Ola Johnsborg Chantal Granadel Jean‐Pierre Claverys

Summary In 1971, Tomasz and Zanati discovered that competent pneumococci have a tendency to form aggregates when pelleted by centrifugation and resuspended in 0.01 N HCl by brief vortexing. Interestingly, no clumping was observed with parallel cultures of non‐competent cells treated in the same way. We set out to elucidate the mechanism behind this striking phenomenon, and were able to show that it depends on extracellular DNA that is presumably released by so‐called competence‐induced cell lysis. Competence‐induced cell lysis, which was first described a few years ago, seems to rely on the concerted action of several murein hydrolases. Our results confirmed and extended previous findings by showing that competence‐induced aggregation is abolished in a lytA–lytC double mutant, and absolutely requires CbpD and its N‐terminal CHAP amidase domain. Furthermore, we discovered a novel competence stimulating peptide (CSP)‐induced immunity protein, encoded by the early competence gene comM ( spr1762 ), which protects competent pneumococci against their own lysins. Together, the murein hydrolases and the immunity protein constitutes a CSP‐controlled mechanism that allows competent pneumococci to commit fratricide by killing non‐competent pneumococci sharing the same ecological niche. Through such predatory behaviour, pneumococci can get access to transforming DNA and nutrients, promote the release of virulence factors, and at the same time get rid of competitors.

Lysin

Amidase

10.1111/j.1365-2958.2005.05021.x

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Citations (144)

Pneumococcal CbpD is a murein hydrolase that requires a dual cell envelope binding specificity to kill target cells during fratricide

Molecular Microbiology (2010)

Vegard Eldholm Ola Johnsborg Daniel Straume Hilde Solheim Ohnstad Kari Helene Berg

Summary Pneumococci that are competent for natural genetic transformation express a number of proteins involved in binding, uptake, translocation and recombination of DNA. In addition, they attack and lyse non‐competent sister cells present in the same environment. This phenomenon has been termed fratricide. The key effector of pneumococcal fratricide is CbpD, a secreted protein encompassing an N‐terminal CHAP domain, two SH3b domains and a C‐terminal choline‐binding domain (CBD). CbpD is believed to degrade the cell wall of target cells, but experimental evidence supporting this hypothesis has been lacking. Here, we show that CbpD indeed has muralytic activity, and that this activity requires functional CBD and SH3b domains. To better understand the critical role played by the non‐catalytic C‐terminal region of CbpD, various translational fusions were constructed between the CBD and SH3b domains and green fluorescent protein (GFP). The results showed that the SH3b domains specifically recognize and bind peptidoglycan, while the CBD domain functions as a localization signal that directs CbpD to the septal region of the pneumococcal cell. Intriguingly, transmission electron microscopy analysis revealed that target cells attacked by CbpD ruptures at the septal region, in accordance with the binding specificity displayed by the CBD domain.

Cell envelope

10.1111/j.1365-2958.2010.07143.x

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Attachment of Capsular Polysaccharide to the Cell Wall inStreptococcus pneumoniae

Microbial Drug Resistance (2012)

Alice Eberhardt C.N. Hoyland Daniela Vollmer Stephanie Bisle Robert M. Cleverley

Streptococcus pneumoniae protects itself from components of the human immune defense system by a thick polysaccharide capsule, which in most serotypes is covalently attached to the cell wall peptidoglycan. Members of the LytR-Cps2A-Psr (LCP) protein family have recently been implicated in the attachment of anionic polymers to peptidoglycan in Gram-positive bacteria, based on genetic evidence from Bacillus subtilis mutant strains and on the crystal structure of S. pneumoniae Cps2A containing a tightly bound polyprenol (pyro)phosphate lipid. Here, we provide evidence that Cps2A and its two pneumococcal homologs, LytR and Psr, contribute to the maintenance of normal capsule levels and to the retention of the capsular polysaccharide at the cell wall in the capsular type 2 S. pneumoniae strain D39. GFP fusions of all three LCP proteins showed enhanced localization at mid-cell, indicating a role in cell wall growth. Single cps2A or psr mutants produced a reduced amount of capsule. A cps2A lytR double mutant showed greatly impaired growth and cell morphology and lost approximately half of the total capsule material into the culture supernatant. We also present the crystal structure of the B. subtilis LCP protein YwtF and provide crystallographic evidence for the phosphotransferase activity of Cps2A, supporting an enzymatic function in the attachment of capsular polysaccharides to cell wall peptidoglycan.

Bacterial capsule

Autolysin

Capsule

10.1089/mdr.2011.0232

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BOX Elements Modulate Gene Expression in Streptococcus pneumoniae : Impact on the Fine-Tuning of Competence Development

Journal of Bacteriology (2006)

Eivind Knutsen Ola Johnsborg Yves Quentin Jean‐Pierre Claverys Leiv Sigve Håvarstein

More than 100 BOX elements are randomly distributed in intergenic regions of the pneumococcal genome. Here we demonstrate that these elements can affect expression of neighboring genes and present evidence that they are mobile. Together, our findings show that BOX elements enhance genetic diversity and genomic plasticity in Streptococcus pneumoniae.

10.1128/jb.00850-06

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Citations (44)

Molecular and Genetic Characterization of Propionicin F, a Bacteriocin fromPropionibacterium freudenreichii

Applied and Environmental Microbiology (2004)

Dag Anders Brede Therese Faye Ola Johnsborg Inger Ødegård Ingolf F. Nes

ABSTRACT This work describes the purification and characterization of propionicin F, the first bacteriocin isolated from Propionibacterium freudenreichii . The bacteriocin has a bactericidal activity and is only active against strains of P. freudenreichii . Propionicin F appears to be formed through a processing pathway new to bacteriocins. The mass of the purified bacteriocin was determined by mass spectrometry, and the N-terminal amino acid sequence was determined by Edman degradation. Sequencing of pcfA , the bacteriocin structural gene, revealed that propionicin F corresponds to a 43-amino-acid peptide in the central part of a 255-amino-acid open reading frame, suggesting that mature propionicin F is excised from the probacteriocin by N- and C-terminal proteolytic modifications. DNA sequencing and Northern blot hybridizations revealed that pcfA is cotranscribed with genes encoding a putative proline peptidase and a protein from the radical S -adenosylmethionine family. A gene encoding an ABC transporter was also identified in close proximity to the bacteriocin structural gene. The potential role of these genes in propionicin F maturation and secretion is discussed.

Propionibacterium freudenreichii

10.1128/aem.70.12.7303-7310.2004

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Citations (54)

A Hydrophobic Patch in the Competence-Stimulating Peptide, a Pneumococcal Competence Pheromone, Is Essential for Specificity and Biological Activity

Journal of Bacteriology (2006)

Ola Johnsborg Per Eugen Kristiansen Trinelise Blomqvist Leiv Sigve Håvarstein

Induction of competence for natural genetic transformation in Streptococcus pneumoniae depends on pheromone-mediated cell-cell communication and a signaling pathway consisting of the competence-stimulating peptide (CSP), its membrane-embedded histidine kinase receptor ComD, and the cognate response regulator ComE. Extensive screening of pneumococcal isolates has revealed that two major CSP variants, CSP1 and CSP2, are found in members of this species. Even though the primary structures of CSP1 and CSP2 are about 50% identical, they are highly specific for their respective receptors, ComD1 and ComD2. In the present work, we have investigated the structural basis of this specificity by determining the three-dimensional structure of CSP1 from nuclear magnetic resonance data and comparing the agonist activity of a number of CSP1/CSP2 hybrid peptides toward the ComD1 and ComD2 receptors. Our results show that upon exposure to membrane-mimicking environments, the 17-amino-acid CSP1 pheromone adopts an amphiphilic alpha-helical configuration stretching from residue 6 to residue 12. Furthermore, the pattern of agonist activity displayed by the various hybrid peptides revealed that hydrophobic amino acids, some of which are situated on the nonpolar side of the alpha-helix, strongly contribute to CSP specificity. Together, these data indicate that the identified alpha-helix is an important structural feature of CSP1 which is essential for effective receptor recognition under natural conditions.

10.1128/jb.188.5.1744-1749.2006

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Citations (56)

Natural genetic transformation: prevalence, mechanisms and function

Research in Microbiology (2007)

Ola Johnsborg Vegard Eldholm Leiv Sigve Håvarstein

10.1016/j.resmic.2007.09.004

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Citations (324)

Properties and Biological Role of Streptococcal Fratricins

Applied and Environmental Microbiology (2012)

Kari Helene Berg Truls Johan Biørnstad Ola Johnsborg Leiv Sigve Håvarstein

Competence for natural genetic transformation is widespread in the genus Streptococcus. The current view is that all streptococcal species possess this property. In addition to the proteins required for DNA uptake and recombination, competent streptococci secrete muralytic enzymes termed fratricins. Since the synthesis and secretion of these cell wall-degrading enzymes are always coupled to competence development in streptococci, fratricins are believed to carry out an important function associated with natural transformation. This minireview summarizes what is known about the properties of fratricins and discusses their possible biological roles in streptococcal transformation.

10.1128/aem.00098-12

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Citations (33)

Fratricide in Streptococcus pneumoniae: contributions and role of the cell wall hydrolases CbpD, LytA and LytC

Microbiology (2009)

Vegard Eldholm Ola Johnsborg Kristine Haugen Hilde Solheim Ohnstad Leiv Sigve Håvarstein

Pneumococci that have developed the competent state kill and lyse non-competent sister cells and members of closely related species during co-cultivation in vitro. The key component in this process, called fratricide, is the product of the late competence gene cbpD. In addition, the peptidoglycan hydrolases LytA and LytC are required for efficient lysis of target cells. Here, we have investigated the relative contribution and possible role of each of the proteins mentioned above. Previous studies have shown that CbpD is produced exclusively by competent cells, whereas LytA and LytC can be provided by the competent attackers as well as the non-competent target cells. By using an improved assay to compare the effect of cis- versus trans-acting LytA and LytC, we were able to show that target cells are lysed much more efficiently when LytA and LytC are provided in cis, i.e. by the target cells themselves. Western analysis demonstrated that considerable amounts of LytC are present in the growth medium. In contrast, we were not able to detect any extracellular LytA. This finding indicates that LytA- and LytC-mediated fratricide represent different processes. In the absence of LytA and LytC, only a tiny fraction of the target cells were lysed, demonstrating that CbpD does not function efficiently on its own. However, in the presence of 1 mM EDTA, the fraction of target cells lysed directly by CbpD increased dramatically, indicating that divalent cations are involved in the regulation of fratricide under natural conditions.

10.1099/mic.0.026328-0

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Citations (128)