Analysis of the catalytic site of the actin ADP‐ribosylating <i>Clostridium perfringens</i> iota toxin

Damme, Josef Van; Jung, Martin; Hofmann, Fred; Just, Ingo; Vandekerckhove, Joël; Aktories, Klaus

doi:10.1016/0014-5793(96)00052-x

Cited by 32 publications

(21 citation statements)

References 49 publications

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“…Damme et al (13) reported the Glu-378 in i a was photolabeled by NAD ϩ but that Glu-380 was not. Therefore, the Glu-378 and -380 residues seem to play different roles in the ARTase activity of i a .…”

Section: Discussionmentioning

confidence: 99%

Characterization of the Enzymatic Component of Clostridium perfringens Iota-Toxin

et al. 2000

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The iota a component (i a ) of Clostridium perfringens ADP ribosylates nonmuscle ␤/␥ actin and skeletal muscle ␣-actin. Replacement of Arg-295 in i a with alanine led to a complete loss of NAD ؉ -glycohydrolase (NADase) and ADP-ribosyltransferase (ARTase); that of the residue with lysine caused a drastic reduction in NADase and ARTase activities (<0.1% of the wild-type activities) but did not completely diminish them. Substitution of alanine for Glu-378 and Glu-380 caused a complete loss of NADase and ARTase. However, exchange of Glu-378 to aspartic acid or glutamine resulted in little effect on NADase activity but a drastic reduction in ARTase activity (<0.1% of the wild-type activity). Exchange of Glu-380 to aspartic acid caused a drastic reduction in NADase and ARTase activities (<0.1% of the wild-type activities) but did not completely diminish them; that of the residue to glutamine caused a complete loss of ARTase activity. Replacement of Ser-338 with alanine resulted in 0.7 to 2.3% wild-type activities, and that of Ser-340 and Thr-339 caused a reduction in these activities of 5 to 30% wild-type activities. The kinetic analysis showed that Arg-295 and Ser-338 also play an important role in the binding of NAD ؉ to i a , that Arg-295, Glu-380, and Ser-338 play a crucial role in the catalytic rate of NADase activity, and that these three amino acid residues and Glu-378 are essential for ARTase activity. The effect of amino acid replacement in i a on ARTase activity was similar to that on lethal and cytotoxic activities, suggesting that lethal and cytotoxic activities in i a are dependent on ARTase activity.

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Section: Discussionmentioning

confidence: 99%

Characterization of the Enzymatic Component of Clostridium perfringens Iota-Toxin

et al. 2000

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“…X-ray crystallography of other bacterial ADP-ribosyltransferases such as B. pertussis pertussis toxin (415), C. diphtheriae diphtheria toxin (82), E. coli heat-labile enterotoxin (404), P. aeruginosa exotoxin A (235), and VIP2 (171) ing an 387 EXE 389 motif found in prokaryotic as well as eukaryotic ADP-ribosyltransferases (444). However, the sequence homology of these toxins within the C terminus is low.…”

Section: Botulinum C2ii and C2imentioning

confidence: 99%

“…Like C2I (36), the initial glutamic acid within the 378 EXE 380 motif of Ia is important for ADPribosyltransferase, but not NAD-glycohydrolase, activity (287). Further analysis of Ia by site-directed mutagenesis or mass spectrometry of cyanogen bromide/trypsin-generated peptides reveals that C-terminal residues R 295 and E 380 , which are conserved among various ADP-ribosyltransferases (76,196,368,432), are also important for Ia catalysis (287,323,444) (Fig. 3A).…”

Section: Perfringens Ib and Iamentioning

confidence: 99%

Binary Bacterial Toxins: Biochemistry, Biology, and Applications of CommonClostridiumandBacillusProteins

Barth

Aktories

Popoff

et al. 2004

Microbiol Mol Biol Rev

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Certain pathogenic species of Bacillus and Clostridium have developed unique methods for intoxicating cells that employ the classic enzymatic “A-B” paradigm for protein toxins. The binary toxins produced by B. anthracis, B. cereus, C. botulinum, C. difficile, C. perfringens, and C. spiroforme consist of components not physically associated in solution that are linked to various diseases in humans, animals, or insects. The “B” components are synthesized as precursors that are subsequently activated by serine-type proteases on the targeted cell surface and/or in solution. Following release of a 20-kDa N-terminal peptide, the activated “B” components form homoheptameric rings that subsequently dock with an “A” component(s) on the cell surface. By following an acidified endosomal route and translocation into the cytosol, “A” molecules disable a cell (and host organism) via disruption of the actin cytoskeleton, increasing intracellular levels of cyclic AMP, or inactivation of signaling pathways linked to mitogen-activated protein kinase kinases. Recently, B. anthracis has gleaned much notoriety as a biowarfare/bioterrorism agent, and of primary interest has been the edema and lethal toxins, their role in anthrax, as well as the development of efficacious vaccines and therapeutics targeting these virulence factors and ultimately B. anthracis. This review comprehensively surveys the literature and discusses the similarities, as well as distinct differences, between each Clostridium and Bacillus binary toxin in terms of their biochemistry, biology, genetics, structure, and applications in science and medicine. The information may foster future studies that aid novel vaccine and drug development, as well as a better understanding of a conserved intoxication process utilized by various gram-positive, spore-forming bacteria

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“…The 47-kDa enzymatic component, designated iota a (Ia), docks with and is translocated by the 80-kDa iota b (Ib) into a target cell. Iota-toxin belongs to an A-B toxin family possessing nonlinked proteins like Clostridium botulinum C2 toxin (12), Clostridium spiroforme iota-like toxin (23, 29), anthrax toxin from Bacillus anthracis (14), and the vegetative insecticidal protein (VIP) produced by Bacillus cereus (11).The current understanding of iota-toxin is mainly centered around the Ia molecule (16,18,34,35), while the structurefunction relationship of Ib in iota toxicity is poorly understood. Like many clostridial toxins, Ia and Ib are activated by proteases (10).…”

mentioning

confidence: 99%

“…The current understanding of iota-toxin is mainly centered around the Ia molecule (16,18,34,35), while the structurefunction relationship of Ib in iota toxicity is poorly understood. Like many clostridial toxins, Ia and Ib are activated by proteases (10).…”

mentioning

confidence: 99%

Clostridium perfringens Iota-Toxin: Mapping of Receptor Binding and Ia Docking Domains on Ib

et al. 2001

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Clostridium perfringens iota-toxin is a binary toxin consisting of iota a (Ia), an ADP-ribosyltransferase that modifies actin, and iota b (Ib), which binds to a cell surface protein and translocates Ia into a target cell. Fusion proteins of recombinant Ib and truncated variants were tested for binding to Vero cells and docking with Ia via fluorescence-activated cytometry and cytotoxicity experiments. C-terminal residues (656 to 665) of Ib were critical for cell surface binding, and truncated Ib variants containing >200 amino acids of the C terminus were effective Ib competitors and prevented iota cytotoxicity. The N-terminal domain (residues 1 to 106) of Ib was important for Ia docking, yet this region was not an effective competitor of iota cytotoxicity. Further studies showed that Ib lacking just the N-terminal 27 residues did not facilitate Ia entry into a target cell and subsequent cytotoxicity. Five monoclonal antibodies against Ib were also tested with each truncated Ib variant for epitope and structural mapping by surface plasmon resonance and an enzyme-linked immunosorbent assay. Each antibody bound to a linear epitope within the N terminus (residues 28 to 66) or the C terminus (residues 632 to 655). Antibodies that target the C terminus neutralized in vitro cytotoxicity and delayed the lethal effects of iota-toxin in mice.Clostridium perfringens type E produces iota-toxin, a binary toxin that consists of two nonlinked proteins implicated in sporadic diarrheic outbreaks among animals (6, 28, 32). The 47-kDa enzymatic component, designated iota a (Ia), docks with and is translocated by the 80-kDa iota b (Ib) into a target cell. Iota-toxin belongs to an A-B toxin family possessing nonlinked proteins like Clostridium botulinum C2 toxin (12), Clostridium spiroforme iota-like toxin (23, 29), anthrax toxin from Bacillus anthracis (14), and the vegetative insecticidal protein (VIP) produced by Bacillus cereus (11).The current understanding of iota-toxin is mainly centered around the Ia molecule (16,18,34,35), while the structurefunction relationship of Ib in iota toxicity is poorly understood. Like many clostridial toxins, Ia and Ib are activated by proteases (10). The protoxin form of Ib contains a 20-kDa aminoterminal peptide released after proteolysis. Upon binding to a protein receptor (30), activated Ib docks with Ia and facilitates entry of Ia into a cell via receptor-mediated endocytosis (23). Once inside a cell, an Ia molecule ADP-ribosylates monomeric actin and consequently disrupts formation of actin filaments necessary for the cytoskeleton, with resultant cell rounding and death (18,35). Either Ia or Ib functionally complements the serologically related iota-like toxin proteins (Sa and Sb) produced by C. spiroforme (29). There is no biological synergy of either iota or iota-like components with other A-B proteins from anthrax (20) or C2 (22, 24, 25) toxins. However, there is a 34 and 41% sequence homology between Ib and the cellbinding proteins of anthrax toxin (protective antigen; PA) and C2 toxin (...

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Analysis of the catalytic site of the actin ADP‐ribosylating Clostridium perfringens iota toxin

Cited by 32 publications

References 49 publications

Characterization of the Enzymatic Component of Clostridium perfringens Iota-Toxin

Characterization of the Enzymatic Component of Clostridium perfringens Iota-Toxin

Binary Bacterial Toxins: Biochemistry, Biology, and Applications of CommonClostridiumandBacillusProteins

Clostridium perfringens Iota-Toxin: Mapping of Receptor Binding and Ia Docking Domains on Ib

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