The ClpXP Protease Is Responsible for the Degradation of the Epsilon Antidote to the Zeta Toxin of the Streptococcal pSM19035 Plasmid

Brzozowska, Iwona; Zielenkiewicz, Urszula

doi:10.1074/jbc.m113.519488

Cited by 12 publications

(16 citation statements)

References 59 publications

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“…The lack of conservation in sequences, even in patterns of hydrophobicity, is striking, particularly for families with many members, such as the RelB/DinJ antitoxins. The Epsilon antitoxin was degraded ~20 amino acids upstream of the C -terminus [ 102 ], which for most antitoxins would overlap with their toxin binding sites. It would seem that toxin binding would then hinder protease-mediated degradation, as was noted for the CcdA antitoxin [ 103 ].…”

Section: Proteases Shape the Ta Module Proteomementioning

confidence: 99%

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Toxin-Antitoxin Modules Are Pliable Switches Activated by Multiple Protease Pathways

Muthuramalingam

White

Bourne

2016

Toxins

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Toxin-antitoxin (TA) modules are bacterial regulatory switches that facilitate conflicting outcomes for cells by promoting a pro-survival phenotypic adaptation and/or by directly mediating cell death, all through the toxin activity upon degradation of antitoxin. Intensive study has revealed specific details of TA module functions, but significant gaps remain about the molecular details of activation via antitoxin degradation used by different bacteria and in different environments. This review summarizes the current state of knowledge about the interaction of antitoxins with cellular proteases Lon and ClpP to mediate TA module activation. An understanding of these processes can answer long-standing questions regarding stochastic versus specific activation of TA modules and provide insight into the potential for manipulation of TA modules to alter bacterial growth.

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Section: Proteases Shape the Ta Module Proteomementioning

confidence: 99%

“…N.D. , no data available. * Reported analysis of cleaved products of Epsilon antitoxin from in vitro assays revealed major cleavage sites ~20 amino acids from the C -terminal residue, and these sites were enriched for Leu, Asn, Glu and Val residues [ 102 ].…”

Section: Figurementioning

confidence: 99%

Toxin-Antitoxin Modules Are Pliable Switches Activated by Multiple Protease Pathways

Muthuramalingam

White

Bourne

2016

Toxins

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“…Bacillus subtilis DegS-DegU 2-component regulatory system is regulated by the ClpXP-Spx regulated proteolysis system (Shiwa et al 2015). ClpXP protease was responsible for Epsilon antitoxin degradation (Brzozowska and Zielenkiewicz 2014). The canonical E. coli protease La (Lon-Ec), a conserved and much studied protease important for homeostasis by targeting abnormal proteins and unstable regulatory proteins, was previously shown to play a role in host infection (Coleman et al 2009).…”

Section: Discussionmentioning

confidence: 98%

RAP-PCR fingerprinting reveals time-dependent expression of development-related genes following differentiation process of Bacillus thuringiensis

Huang

Gelbič

et al. 2015

Can. J. Microbiol.

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Gene expression profiles are important data to reveal the functions of genes putatively involved in crucial biological processes. RNA arbitrarily primed polymerase chain reaction (RAP-PCR) and specifically primed reverse transcription polymerase chain reaction (RT-PCR) were combined to screen differentially expressed genes following development of a commercial Bacillus thuringiensis subsp. kurstaki strain 8010 (serotype 3a3b). Six differentially expressed transcripts (RAP1 to RAP6) were obtained. RAP1 encoded a putative triple helix repeat-containing collagen or an exosporium protein H related to spore pathogenicity. RAP2 was homologous to a ClpX protease and an ATP-dependent protease La (LonB), which likely acted as virulence factors. RAP3 was homologous to a beta subunit of propionyl-CoA carboxylase required for the development of Myxococcus xanthus. RAP4 had homology to a quinone oxidoreductase involved in electron transport and ATP formation. RAP5 showed significant homology to a uridine kinase that mediates phosphorylation of uridine and azauridine. RAP6 shared high sequence identity with 3-methyl-2-oxobutanoate-hydroxymethyltransferase (also known as ketopantoate hydroxymethyltransferase or PanB) involved in the operation of the tricarboxylic acid cycle. The findings described here would help to elucidate the molecular mechanisms underlying the differentiation process of B. thuringiensis and unravel novel pathogenic genes.

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“…The N -terminally hexa-histidine-tagged His6-Epsilon protein was overproduced in E. coli BL21 (DE3) and purified under native conditions, as previously described [ 37 ]. Purified protein eluted from a Ni-TED column was dialyzed against the storage buffer (25 mM Tris pH 8, 100 mM KCl, 5 mM DTT, 1 mM MgCl 2 , 50% glycerol) and then stored at −20 °C.…”

Section: Methodsmentioning

confidence: 99%

“…The molecular basis for the function of the εζ system is the difference in the half-life times of these two proteins [ 36 ]. While the Zeta toxin is stable, the free Epsilon antitoxin is a labile protein that is vulnerable to degradation by cellular proteases [ 37 ]. Under normal cellular conditions, excess antitoxin stabilizes the TA complex, thereby neutralizing the Zeta toxin.…”

Section: Introductionmentioning

confidence: 99%

Mapping Protein–Protein Interactions of the Resistance-Related Bacterial Zeta Toxin–Epsilon Antitoxin Complex (ε2ζ2) with High Affinity Peptide Ligands Using Fluorescence Polarization

Fernández‐Bachiller

Brzozowska

Odolczyk

et al. 2016

Toxins

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Toxin–antitoxin systems constitute a native survival strategy of pathogenic bacteria and thus are potential targets of antibiotic drugs. Here, we target the Zeta–Epsilon toxin–antitoxin system, which is responsible for the stable maintenance of certain multiresistance plasmids in Gram-positive bacteria. Peptide ligands were designed on the basis of the ε2ζ2 complex. Three α helices of Zeta forming the protein–protein interaction (PPI) site were selected and peptides were designed conserving the residues interacting with Epsilon antitoxin while substituting residues binding intramolecularly to other parts of Zeta. Designed peptides were synthesized with an N-terminal fluoresceinyl-carboxy-residue for binding assays and provided active ligands, which were used to define the hot spots of the ε2ζ2 complex. Further shortening and modification of the binding peptides provided ligands with affinities <100 nM, allowing us to determine the most relevant PPIs and implement a robust competition binding assay.

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The ClpXP Protease Is Responsible for the Degradation of the Epsilon Antidote to the Zeta Toxin of the Streptococcal pSM19035 Plasmid

Cited by 12 publications

References 59 publications

Toxin-Antitoxin Modules Are Pliable Switches Activated by Multiple Protease Pathways

Toxin-Antitoxin Modules Are Pliable Switches Activated by Multiple Protease Pathways

RAP-PCR fingerprinting reveals time-dependent expression of development-related genes following differentiation process of Bacillus thuringiensis

Mapping Protein–Protein Interactions of the Resistance-Related Bacterial Zeta Toxin–Epsilon Antitoxin Complex (ε2ζ2) with High Affinity Peptide Ligands Using Fluorescence Polarization

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