MICROBIOLOGY: Enhanced: Fighting Anthrax with a Mutant Toxin

Science ◽  
2001 ◽  
Vol 292 (5517) ◽  
pp. 647-648
Author(s):  
S. Olsnes
Keyword(s):  
Mutant Toxin

Pore Formation of Thermostable Direct Hemolysin Secreted from Vibrio parahaemolyticus in Lipid Bilayers

2006 ◽  
Vol 25 (5) ◽  
pp. 409-418 ◽  
Author(s):  
Akira Takahashi ◽  
Chiyo Yamamoto ◽  
Toshio Kodama ◽  
Kanami Yamashita ◽  
Nagakatsu Harada ◽  
...  
Keyword(s):  
Lipid Bilayer ◽  
Lipid Bilayers ◽  
Lipid Membranes ◽  
Vibrio Parahaemolyticus ◽  
Cultured Cells ◽  
Ion Selectivity ◽  
Ion Permeability ◽  
Lower Extent ◽  
Thermostable Direct Hemolysin ◽  
Mutant Toxin

Vibrio parahaemolyticus secretes thermostable direct hemolysin (TDH), a major virulence factor. Earlier studies report that TDH is a pore-forming toxin. However, the characteristics of pores formed by TDH in the lipid bilayer, which is permeable to small ions, remain to be elucidated. Ion channel-like activities were observed in lipid bilayers containing TDH. Three types of conductance were identified. All the channels displayed relatively low ion selectivity, and similar ion permeability. The Cl− channel inhibitors, DIDS, glybenclamide, and NPPB, did not affect the channel activity of pores formed by TDH. R7, a mutant toxin of TDH, also forms pores with channel-like activity in lipid bilayers. The ion permeability of these channels is similar to that of TDH. R7 binds cultured cells and liposomes to a lower extent, compared to TDH. R7 does not display significant hemolytic activity and cell cytotoxicity, possibly owing to the difficulty of insertion into lipid membranes. Once R7 is assembled within lipid membranes, it may assume the same structure as TDH. The authors propose that the single glycine at position 62, substituted with serine in the R7 mutant toxin, plays an important role in TDH insertion into the lipid bilayer.

scholarly journals Native and Mutant Forms of Cholera Toxin and Heat-Labile Enterotoxin Effectively Enhance Protective Efficacy of Live Attenuated and Heat-Killed Shigella Vaccines

1999 ◽  
Vol 67 (11) ◽  
pp. 5841-5847 ◽  
Author(s):  
Antoinette B. Hartman ◽  
Lillian L. Van De Verg ◽  
Malabi M. Venkatesan
Keyword(s):  
Guinea Pig ◽  
Cholera Toxin ◽  
Protective Efficacy ◽  
Shigella Flexneri ◽  
Serum Antibody ◽  
Adjuvant Effect ◽  
O Antigen ◽  
Heat Labile ◽  
Mutant Forms ◽  
Mutant Toxin

Both native and mutant forms of cholera toxin (CT) and heat-labile enterotoxin (LT) are effective adjuvants for antigens and killed whole-cell preparations. To determine whether these toxin molecules could also boost the immunogenicity and efficacy of live attenuated vaccines directed against shigellosis, the guinea pig keratoconjunctivitis model was used to evaluate the adjuvant effect of these toxin molecules on EcSf2a-3, a ΔvirG ΔaroD Escherichia coli-Shigella flexneri 2a hybrid vaccine strain that was previously found to be less protective than its parent strain in the guinea pig model. Experiments using native and mutant toxin molecules showed that both CT and LT and mutant derivatives were effective as an adjuvant for EcSf2a-3 and that the mutant toxin molecules, which were developed to retain adjuvanticity without the toxicity associated with the native molecules, were as effective as the native toxin molecules as adjuvants. Protective efficacy was enhanced for both the oral and intranasal routes of immunization. Serum antibody response to theS. flexneri 2a O antigen, the primary antigen for protective immunity, was not dependent on the addition of an adjuvant. However, enumeration of the O-antigen-specific immunoglobulin G (IgG) and IgA antibody-secreting cells in the spleen and draining lymph nodes following intranasal immunization suggested that enhancement of the local immune response by the toxin molecules may contribute to the observed increase in protective efficacy. The efficacy of heat-killedS. flexneri 2a was enhanced only by mutant LT molecules. These results suggest that the best candidates for enhancing the efficacy of both live attenuated and heat-killed Shigellavaccines with minimal reactogenicity are the mutant toxin molecules.

scholarly journals Aggregation of Bacillus thuringiensisCry1A Toxins upon Binding to Target Insect Larval Midgut Vesicles

1999 ◽  
Vol 65 (6) ◽  
pp. 2503-2507 ◽  
Author(s):  
Arthur I. Aronson ◽  
Chaoxian Geng ◽  
Lan Wu
Keyword(s):  
Heliothis Virescens ◽  
Tight Binding ◽  
Amino Terminus ◽  
Irreversible Binding ◽  
Insect Larvae ◽  
Mild Conditions ◽  
Amphipathic Helices ◽  
Cry1ac Toxin ◽  
Toxin Receptor ◽  
Mutant Toxin

ABSTRACT During sporulation, Bacillus thuringiensis produces crystalline inclusions comprised of a mixture of δ-endotoxins. Following ingestion by insect larvae, these inclusion proteins are solubilized, and the protoxins are converted to toxins. These bind specifically to receptors on the surfaces of midgut apical cells and are then incorporated into the membrane to form ion channels. The steps required for toxin insertion into the membrane and possible oligomerization to form a channel have been examined. When bound to vesicles from the midguts of Manduca sexta larvae, the Cry1Ac toxin was largely resistant to digestion with protease K. Only about 60 amino acids were removed from the Cry1Ac amino terminus, which included primarily helix α1. Following incubation of the Cry1Ab or Cry1Ac toxins with vesicles, the preparations were solubilized by relatively mild conditions, and the toxin antigens were analyzed by immunoblotting. In both cases, most of the toxin formed a large, antigenic aggregate of ca. 200 kDa. These toxin aggregates did not include the toxin receptor aminopeptidase N, but interactions with other vesicle components were not excluded. No oligomerization occurred when inactive toxins with mutations in amphipathic helices (α5) and known to insert into the membrane were tested. Active toxins with other mutations in this helix did form oligomers. There was one exception; a very active helix α5 mutant toxin bound very well to membranes, but no oligomers were detected. Toxins with mutations in the loop connecting helices α2 and α3, which affected the irreversible binding to vesicles, also did not oligomerize. There was a greater extent of oligomerization of the Cry1Ac toxin with vesicles from theHeliothis virescens midgut than with those from theM. sexta midgut, which correlated with observed differences in toxicity. Tight binding of virtually the entire toxin molecule to the membrane and the subsequent oligomerization are both important steps in toxicity.

Saccharomyces cerevisiae spheroplasts are sensitive to the action of diphtheria toxin

1982 ◽  
Vol 2 (5) ◽  
pp. 588-592
Author(s):  
S Murakami ◽  
J W Bodley ◽  
D M Livingston
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Protein Synthesis ◽  
Diphtheria Toxin ◽  
Mammalian Cells ◽  
Cell Interaction ◽  
Yeast Cells ◽  
Intact Cells ◽  
Rna And Dna Synthesis ◽  
Mutant Toxin ◽  
Rna And Dna

Diphtheria toxin kills spheroplasts of Saccharomyces cerevisiae but not the intact yeast cells. After 2 h of exposure to ca. 10(-7) M toxin, less than 1% of spheroplasts were able to regenerate into intact cells. The same high levels of toxin inhibited the rate of protein synthesis by more than 90% within 1 h, whereas RNA and DNA synthesis were not inhibited until 4 h or exposure. Both killing and protein synthesis inhibition were dependent on toxin concentration. The nature of the toxin-cell interaction was also studied by using fragments of intact toxin and mutant toxin proteins. Neither toxin fragment A nor CRM45 nor CRM197 affected spheroplasts, but CRM197 and ATP prevented the inhibitory action of intact toxin. These results suggest that toxin acts on S. cerevisiae spheroplasts in much the same manner as it acts on sensitive mammalian cells.

scholarly journals Application of Mutated Clostridium difficile Toxin A for Determination of Glucosyltransferase-Dependent Effects

2006 ◽  
Vol 74 (10) ◽  
pp. 6006-6010 ◽  
Author(s):  
Matthias Teichert ◽  
Helma Tatge ◽  
Janett Schoentaube ◽  
Ingo Just ◽  
Ralf Gerhard
Keyword(s):  
Cell Culture ◽  
Stress Response ◽  
Clostridium Difficile ◽  
Cellular Stress Response ◽  
Toxin A ◽  
Clostridium Difficile Toxin ◽  
Clostridium Difficile Toxin A ◽  
Independent Effects ◽  
Mutant Toxin

ABSTRACT Mutation of tryptophan-101 in Clostridium difficile toxin A, a 308-kDa glucosyltransferase, resulted in a 50-fold-reduced cytopathic activity in cell culture experiments. The mutant toxin A was characterized and applied to distinguish between glucosyltransferase-dependent and -independent effects with respect to RhoB up-regulation as a cellular stress response.

scholarly journals The α-Helix 4 Residue, Asn135, Is Involved in the Oligomerization of Cry1Ac1 and Cry1Ab5 Bacillus thuringiensis Toxins

2001 ◽  
Vol 67 (12) ◽  
pp. 5715-5720 ◽  
Author(s):  
Natalie J. Tigue ◽  
Juliette Jacoby ◽  
David J. Ellar
Keyword(s):  
Bacillus Thuringiensis ◽  
Membrane Vesicles ◽  
Wild Type ◽  
Cell Plasma Membrane ◽  
Structural Domains ◽  
Cell Plasma ◽  
Α Helix ◽  
Domain I ◽  
Mutant Toxin ◽  
Made In

ABSTRACT The insecticidal Cry toxins produced by the bacteriumBacillus thuringiensis are comprised of three structural domains. Domain I, a seven-helix bundle, is thought to penetrate the insect epithelial cell plasma membrane through a hairpin composed of α-helices 4 and 5, followed by the oligomerization of four hairpin monomers. The α-helix 4 has been proposed to line the lumen of the pore, whereas some residues in α-helix 5 have been shown to be responsible for oligomerization. Mutation of the Cry1Ac1 α-helix 4 amino acid Asn135 to Gln resulted in the loss of toxicity toManduca sexta, yet binding was still observed. In this study, the equivalent mutation was made in the Cry1Ab5 toxin, and the properties of both wild-type and mutant toxin counterparts were analyzed. Both mutants appeared to bind to M. sextamembrane vesicles, but they were not able to form pores. The ability of both N135Q mutants to oligomerize was also disrupted, providing the first evidence that a residue in α-helix 4 can contribute to toxin oligomerization.

Sign in / Sign up

Export Citation Format

Share Document