scholarly journals Phylogeny Reveals Novel HipA-Homologous Kinase Families and Toxin – Antitoxin Gene Organizations

2021 ◽  
Author(s):  
Kenn Gerdes ◽  
Rene Bærentsen ◽  
Ditlev E. Brodersen
Keyword(s):  
Phylogenetic Analysis ◽  
Stringent Response ◽  
Protein Translation ◽  
Kinase Domain ◽  
Trna Synthetase ◽  
Antibiotic Tolerance ◽  
Bacterial Cells ◽  
E Coli ◽  
Cellular Translation ◽  
K 12

AbstractToxin – Antitoxin modules function in the genetic stability of mobile genetic elements, bacteriophage defense, and antibiotic tolerance. A gain-of-function mutation of the Escherichia coli K-12 hipBA module can induce antibiotic tolerance in a subpopulation of bacterial cells, a phenomenon known as persistence. HipA is a Ser/Thr kinase that phosphorylates and inactivates glutamyl tRNA synthetase, inhibiting cellular translation and inducing the stringent response. Additional characterized HipA homologues include HipT from pathogenic E. coli O127 and YjjJ of E. coli K-12, which are encoded by tri-cistronic hipBST and monocistronic operons, respectively. The apparent diversity of HipA homologues in bacterial genomes inspired us to investigate overall phylogeny. Here we present a comprehensive phylogenetic analysis of the Hip kinases in bacteria and archaea that expands on this diversity by revealing seven novel kinase families. Kinases of one family, encoded by monocistronic operons, consist of an N-terminal core kinase domain, a HipS-like domain and a HIRAN (HIP116 Rad5p N-terminal) domain. HIRAN domains bind single or double-stranded DNA ends. Moreover, five types of bicistronic kinase operons encode putative antitoxins with HipS-HIRAN, HipS, γδ-resolvase or Stl repressor-like domains. Finally, our analysis indicates that reversion of hipBA gene-order happened independently several times during evolution.ImportanceBacterial multidrug tolerance and persistence are problems of increasing scientific and medical significance. The first gene discovered to confer persistence was hipA, encoding the kinase toxin of the hipBA toxin-antitoxin (TA) module of E. coli. HipA-homologous kinases phosphorylate and thereby inactivate specific tRNA synthetases, thus inhibiting protein translation and cell proliferation. Here, we present a comprehensive phylogenetic analysis of bacterial Hip kinases and discover seven new families with novel operon structures and domains. Overall, Hip kinases are encoded by TA modules with at least 10 different genetic organizations, seven of which have not been described before. These results open up exciting avenues for the experimental analysis of the superfamily of Hip kinases.

scholarly journals The Real-Time-Based Assessment of the Microbial Killing by the Antimicrobial Compounds of Neutrophils

2011 ◽  
Vol 11 ◽  
pp. 2382-2390 ◽  
Author(s):  
J. T. Atosuo ◽  
E.-M. Lilius
Keyword(s):  
Real Time ◽  
Antimicrobial Agents ◽  
Hypochlorous Acid ◽  
Bacterial Cells ◽  
Photorhabdus Luminescens ◽  
Bacterial Luciferase ◽  
E Coli ◽  
High Concentrations ◽  
Time Basis ◽  
K 12

A recombinantEscherichia coliK-12 strain, transformed with a modified bacterial luciferase gene (luxABCDE) fromPhotorhabdus luminescens, was constructed in order to monitor the activity of various antimicrobial agents on a real-time basis. ThisE. coli-lux emitted, without any addition of substrate, constitutive bioluminescence (BL), which correlated to the number of viable bacterial cells. The decrease in BL signal correlated to the number of killed bacterial cells. Antimicrobial activity of hydrogen peroxide (H2O2) and myeloperoxidase (MPO) was assessed. In high concentrations, H2O2alone had a bacteriocidic function and MPO enhanced this killing by forming hypochlorous acid (HOCl). Taurine, the known HOCl scavenger, blocked the killing by MPO. WhenE. coli-lux was incubated with neutrophils, similar killing kinetics was recorded as in H2O2/MPO experiments. The opsonization of bacteria enhanced the killing, and the maximum rate of the MPO release from lysosomes coincided with the onset of the killing.

A new method for the isolation of methionyl transfer RNA synthetase mutants from Escherichia coli

1975 ◽  
Vol 21 (6) ◽  
pp. 754-758 ◽  
Author(s):  
John B. Armstrong ◽  
John A. Fairfield
Keyword(s):  
Escherichia Coli ◽  
Trna Synthetase ◽  
Transfer Rna ◽  
New Method ◽  
Wild Type ◽  
E Coli ◽  
Adenosyl Methionine ◽  
K 12 ◽  
Methionyl Trna Synthetase

Six methionine auxotrophs were isolated from an E. coli K-12 strain which required up to 100 times as much methionine for growth as a conventional auxotroph. In these mutants, the methionyl-tRNA synthetase had an increased Km for methionine. The Km value for the mutants ranged from 0.48 to 1.63 mM, compared to 0.078 mM for the wild type. The Km (methionine) for S-adenosyl methionine synthetase was not altered.

scholarly journals Phage Mediate Bacterial Self Recognition

2018 ◽  
Author(s):  
Sooyeon Song ◽  
Yunxue Guo ◽  
Jun-Seob Kim ◽  
Xiaoxue Wang ◽  
Thomas K. Wood
Keyword(s):  
Kin Recognition ◽  
Group Behavior ◽  
Lytic Phage ◽  
Bacterial Cells ◽  
Demarcation Line ◽  
E Coli ◽  
Self Recognition ◽  
The Family ◽  
K 12

AbstractCells are social, and self-recognition is an important and conserved aspect of group behavior where cells assist kin and antagonize non-kin to conduct group behavior such as foraging for food and biofilm formation. However, the role of the common bacterial cohabitant, phage, in kin recognition, has not been explored. Here we find that a boundary (demarcation line) is formed between different swimmingEscherichia colistrains but not between identical clones; hence, motile bacterial cells discriminate between self and non-self. The basis for this self-recognition is a novel, 49 kb, T1-type, lytic phage of the family siphoviridae (named here SW1) that controls formation of the demarcation line by utilizing one of the host’s cryptic prophage proteins, YfdM, to propagate. Critically, SW1 increases the fitness ofE. coliK-12 compared to the identical strain that lacks the phage. Therefore, bacteria use phage to recognize kin.

scholarly journals Rhs Elements Comprise Three Subfamilies Which Diverged Prior to Acquisition by Escherichia coli

1998 ◽  
Vol 180 (16) ◽  
pp. 4102-4110 ◽  
Author(s):  
Yong-Dong Wang ◽  
Sheng Zhao ◽  
Charles W. Hill
Keyword(s):  
Escherichia Coli ◽  
Phylogenetic Analysis ◽  
Open Reading Frame ◽  
Large Protein ◽  
Rich Region ◽  
Reading Frame ◽  
E Coli ◽  
The Core ◽  
Reference Collection ◽  
K 12

ABSTRACT The Rhs elements are complex genetic composites widely spread among Escherichia coli isolates. One of their components, a 3.7-kb, GC-rich core, maintains a single open reading frame that extends the full length of the core and then 400 to 600 bp beyond into an AT-rich region. Whereas Rhs cores are homologous, core extensions from different elements are dissimilar. Two new Rhs elements from strains of the ECOR reference collection have been characterized. RhsG (from strain ECOR-11) maps to min 5.3, and RhsH (from strain ECOR-45) maps to min 32.8, where it lies in tandem with RhsE. Comparison of strain K-12 to ECOR-11 indicates that RhsGwas once present in but has been largely deleted from an ancestor of K-12. Phylogenetic analysis shows that the cores from eight known elements fall into three subfamilies, RhsA-B-C-F,RhsD-E, and RhsG-H. Cores from different subfamilies diverge 22 to 29%. Analysis of substitutions that distinguish between subfamilies shows that the origin of the ancestral core as well as the process of subfamily separation occurred in a GC-rich background. Furthermore, each subfamily independently passed from the GC-rich background to a less GC-rich background such asE. coli. A new example of core-extension shuffling provides the first example of exchange between cores of different subfamilies. A novel component of RhsE and RhsG,vgr, encodes a large protein distinguished by 18 to 19 repetitions of a Val-Gly dipeptide occurring with a eight-residue periodicity.

scholarly journals Aqueous Mechano-Bactericidal Action of Acicular Aragonite Crystals

2021 ◽  
Author(s):  
Nobuaki Negishi ◽  
Tomohiro Inaba ◽  
Yukari Miyazaki ◽  
Genki Ishii ◽  
Yang Yingnan ◽  
...  
Keyword(s):  
Laser Scanning ◽  
Hard Water ◽  
Petri Dish ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Black Silicon ◽  
Bacterial Cells ◽  
Bactericidal Action ◽  
E Coli ◽  
K 12

Abstract Nanoneedle structures on dragonfly and cicada wing surfaces or black silicon nanoneedles demonstrate antibacterial phenomena, namely mechano-bactericidal action. These air-exposed, mechano-bactericidal surfaces serve to destroy adherent bacteria, but their bactericidal action in the water is no precedent to report. Calcium carbonate easily accumulates on surfaces during long-term exposure to hard water. We expect that aragonite nanoneedles, in particular, which grow on TiO2 during the photocatalytic treatment of calcium-rich groundwater, exhibit mechano-bactericidal action against bacteria in water. Here, we show that aragonite nanoneedles are grown on TiO2 ceramics from the calcium bicarbonate in mineral water exhibit mechano-bactericidal action against E. coli K-12 in water. Unmodified, calcite-modified as references and aragonite-modified TiO2 ceramics were exposed to water containing E. coli K-12 (in a petri dish), and their bactericidal action over time was investigated under static and agitated conditions. The surfaces of the materials were observed by scanning electron microscopy, and the live/dead bacterial cells were observed by confocal laser scanning microscopy. Further, the synergistic bactericidal performance achieved by mechano-bactericidal action and photocatalysis was demonstrated. Aragonite itself has a high biological affinity for the human body different from the other whisker-sharpen nano-materials, therefore, the mechano-bactericidal action of acicular aragonite in water is expected to inform the development of safe water purification systems for use in developing countries.

scholarly journals Characterization of the Avian PathogenicEscherichia coli Hemagglutinin Tsh, a Member of the Immunoglobulin A Protease-Type Family of Autotransporters

1999 ◽  
Vol 67 (2) ◽  
pp. 772-781 ◽  
Author(s):  
Christos Stathopoulos ◽  
David L. Provence ◽  
Roy Curtiss
Keyword(s):  
Amino Acid ◽  
Sequence Data ◽  
Single Gene ◽  
Shigella Flexneri ◽  
Immunoglobulin A ◽  
Site Directed Mutagenesis ◽  
Bacterial Cells ◽  
E Coli ◽  
K 12

ABSTRACT We reported earlier that a single gene, tsh, isolated from a strain of avian pathogenic Escherichia coli (APEC) was sufficient to confer on E. coli K-12 a hemagglutinin-positive phenotype and that the deduced sequence of the Tsh protein shared homology to the serine-type immunoglobulin A (IgA) proteases of Neisseria gonorrhoeae and Haemophilus influenzae. In this report we show that E. coli K-12 containing the recombinant tsh gene produced two proteins, a 106-kDa extracellular protein and a 33-kDa outer membrane protein, and was also able to agglutinate chicken erythrocytes. N-terminal sequence data indicated that the 106-kDa protein, designated Tshs, was derived from the N-terminal end of Tsh after the removal of a 52-amino-acid N-terminal signal peptide, while the 33-kDa protein, designated Tshβ, was derived from the C-terminal end of Tsh starting at residue N1101. The Tshsdomain contains the 7-amino-acid serine protease motif that includes the active-site serine (S259), found also in the secreted domains of the IgA proteases. However, site-directed mutagenesis of S259 did not abolish the hemagglutinin activity or the extracellular secretion of Tshs indicating that host-directed proteolysis was mediating the release of Tshs. Studies with an E. coli K-12ompT mutant strain showed that the surface protease OmpT was not needed for the secretion of Tshs. Tsh belongs to a subclass of the IgA protease family, which also includes EspC of enteropathogenic E. coli, EspP of enterohemorragic E. coli, and SepA and VirG of Shigella flexneri, which seem to involve a host endopeptidase to achieve extracellular release of their N-terminal domains. In proteolytic studies conducted in vitro, Tshs did not cleave the substrate of the IgA proteases, human IgA1 or chicken IgA, and did not show proteolytic activity in a casein-based assay. Correlation of Tsh expression and hemagglutination activity appears to be a very complex phenomenon, influenced by strain and environmental conditions. Nevertheless, for both APEC and recombinant E. coli K-12 strains containing thetsh gene, it was only the whole bacterial cells and not the cell-free supernatants that could confer hemagglutinin activity. Our results provide insights into the expression, secretion, and proteolytic features of the Tsh protein, which belongs to the growing family of gram-negative bacterial extracellular virulence factors, named autotransporters, which utilize a self-mediated mechanism to achieve export across the bacterial cell envelope.

scholarly journals The Global Regulatory hns Gene Negatively Affects Adhesion to Solid Surfaces by Anaerobically Grown Escherichia coli by Modulating Expression of Flagellar Genes and Lipopolysaccharide Production

2002 ◽  
Vol 184 (6) ◽  
pp. 1522-1529 ◽  
Author(s):  
Paolo Landini ◽  
Alexander J. B. Zehnder
Keyword(s):  
Escherichia Coli ◽  
Sharp Decrease ◽  
Luria Broth ◽  
Bacterial Cells ◽  
Gene Encoding ◽  
Aerobic Conditions ◽  
E Coli ◽  
Hydrophilic Surfaces ◽  
K 12 ◽  
Initial Binding

ABSTRACT The initial binding of bacterial cells to a solid surface is a critical and essential step in biofilm formation. In this report we show that stationary-phase cultures of Escherichia coli W3100 (a K-12 strain) can efficiently attach to sand columns when they are grown in Luria broth medium at 28°C in fully aerobic conditions. In contrast, growth in oxygen-limited conditions results in a sharp decrease in adhesion to hydrophilic substrates. We show that the production of lipopolysaccharide (LPS) and of flagella, as well as the transcription of the fliC gene, encoding the major flagellar subunit, increases under oxygen-limited conditions. Inactivation of the global regulatory hns gene counteracts increased production of LPS and flagella in response to anoxia and allows E. coli W3100 to attach to sand columns even when it is grown under oxygen-limited conditions. We propose that increased production of the FliC protein and of LPS in response to oxygen limitation results in the loss of the ability of E. coli W3100 to adhere to hydrophilic surfaces. Indeed, overexpression of the fliC gene results in a decreased adhesion to sand even when W3100 is grown in fully aerobic conditions. Our observations strongly suggest that anoxia is a negative environmental signal for adhesion in E. coli.

scholarly journals tRNA Synthetase Mutants of Escherichia coli K-12 Are Resistant to the Gyrase Inhibitor Novobiocin

1999 ◽  
Vol 181 (9) ◽  
pp. 2979-2983 ◽  
Author(s):  
Jovanovic Milija ◽  
Mirjana Lilic ◽  
Radmila Janjusevic ◽  
Goran Jovanovic ◽  
Dragutin J. Savic
Keyword(s):  
Escherichia Coli ◽  
Rna Polymerase ◽  
Stringent Response ◽  
Trna Synthetase ◽  
Protein Inactivation ◽  
Constitutive Production ◽  
K 12 ◽  
Novobiocin Resistance

ABSTRACT In previous studies we demonstrated that mutations in the genescysB, cysE, and cls(nov) affect resistance of Escherichia coli to novobiocin (J. Rakonjac, M. Milic, and D. J. Savic, Mol. Gen. Genet. 228:307–311, 1991; R. Ivanisevic, M. Milic, D. Ajdic, J. Rakonjac, and D. J. Savic, J. Bacteriol. 177:1766–1771, 1995). In this work we expand this list with mutations in rpoN (the gene for RNA polymerase subunit ς54) and the tRNA synthetase genes alaS, argS, ileS, and leuS. Similarly to resistance to the penicillin antibiotic mecillinam, resistance to novobiocin of tRNA synthetase mutants appears to depend upon the RelA-mediated stringent response. However, at this point the overlapping pathways of mecillinam and novobiocin resistance diverge. Under conditions of stringent response induction, either by the presence of tRNA synthetase mutations or by constitutive production of RelA protein, inactivation of thecls gene diminishes resistance to novobiocin but not to mecillinam.

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