Specificity of receptor-mediated signal transduction by E. coli type its heat-labile toxin

Escherichia coli enterotoxin (LT) and the homologous cholera toxin (CT) are A-B toxins that cause travelers' diarrhea and cholera, respectively. So far, experimental live and killed vaccines against these diseases have been developed using only the nontoxic B portion of these toxins. The enzymatically active A subunit has not been used because it is responsible for the toxicity and it is reported to induce a negligible titer of toxin neutralizing antibodies. We used site-directed mutagenesis to inactivate the ADP-ribosyltransferase activity of the A subunit and obtained nontoxic derivatives of LT that elicited a good titer of neutralizing antibodies recognizing the A subunit. These LT mutants and equivalent mutants of CT may be used to improve live and killed vaccines against cholera and enterotoxinogenic E. coli.

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Amplification of immune responses against a DNA-delivered idiotypic lymphoma antigen by fusion to the B subunit of E. coli heat labile toxin

Vaccine ◽

10.1016/j.vaccine.2009.05.025 ◽

2009 ◽

Vol 27 (32) ◽

pp. 4289-4296 ◽

Cited By ~ 5

Author(s):

Caleb G. Chen ◽

Yen-Ta Lu ◽

Marie Lin ◽

Natalia Savelyeva ◽

Freda K. Stevenson ◽

...

Keyword(s):

Immune Responses ◽

E Coli ◽

B Subunit ◽

Heat Labile

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Biochemical and structural characterization of the novel sialic acid-binding site of Escherichia coli heat-labile enterotoxin LT-IIb

Biochemical Journal ◽

10.1042/bcj20160575 ◽

2016 ◽

Vol 473 (21) ◽

pp. 3923-3936 ◽

Cited By ~ 3

Author(s):

Dani Zalem ◽

João P. Ribeiro ◽

Annabelle Varrot ◽

Michael Lebens ◽

Anne Imberty ◽

...

Keyword(s):

Escherichia Coli ◽

Binding Site ◽

Cholera Toxin ◽

Carbohydrate Binding ◽

Type I ◽

Type Ii ◽

E Coli ◽

B Subunit ◽

Heat Labile ◽

B Subunits

The structurally related AB5-type heat-labile enterotoxins of Escherichia coli and Vibrio cholerae are classified into two major types. The type I group includes cholera toxin (CT) and E. coli LT-I, whereas the type II subfamily comprises LT-IIa, LT-IIb and LT-IIc. The carbohydrate-binding specificities of LT-IIa, LT-IIb and LT-IIc are distinctive from those of cholera toxin and E. coli LT-I. Whereas CT and LT-I bind primarily to the GM1 ganglioside, LT-IIa binds to gangliosides GD1a, GD1b and GM1, LT-IIb binds to the GD1a and GT1b gangliosides, and LT-IIc binds to GM1, GM2, GM3 and GD1a. These previous studies of the binding properties of type II B-subunits have been focused on ganglio core chain gangliosides. To further define the carbohydrate binding specificity of LT-IIb B-subunits, we have investigated its binding to a collection of gangliosides and non-acid glycosphingolipids with different core chains. A high-affinity binding of LT-IIb B-subunits to gangliosides with a neolacto core chain, such as Neu5Gcα3- and Neu5Acα3-neolactohexaosylceramide, and Neu5Gcα3- and Neu5Acα3-neolactooctaosylceramide was detected. An LT-IIb-binding ganglioside was isolated from human small intestine and characterized as Neu5Acα3-neolactohexaosylceramide. The crystal structure of the B-subunit of LT-IIb with the pentasaccharide moiety of Neu5Acα3-neolactotetraosylceramide (Neu5Ac-nLT: Neu5Acα3Galβ4GlcNAcβ3Galβ4Glc) was determined providing the first information for a sialic-binding site in this subfamily, with clear differences from that of CT and LT-I.

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Expression of B subunit of E. coli heat-labile enterotoxin in the progenies of transgenic tobacco bred by crossing nuclear- and chloroplast-transgenic lines

Protein Expression and Purification ◽

10.1016/j.pep.2018.11.005 ◽

2019 ◽

Vol 155 ◽

pp. 54-58 ◽

Cited By ~ 3

Author(s):

Bo-Mi Kim ◽

Tae-Jin Kang

Keyword(s):

Transgenic Tobacco ◽

E Coli ◽

B Subunit ◽

Heat Labile ◽

Transgenic Lines

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Enterotoxigenic Escherichia coli infection and intestinal thiamin uptake: studies with intestinal epithelial Caco-2 monolayers

AJP Cell Physiology ◽

10.1152/ajpcell.00276.2013 ◽

2013 ◽

Vol 305 (11) ◽

pp. C1185-C1191 ◽

Cited By ~ 8

Author(s):

Abhisek Ghosal ◽

Nabendu S. Chatterjee ◽

Tristan Chou ◽

Hamid M. Said

Keyword(s):

Escherichia Coli ◽

Significant Inhibition ◽

Water Soluble ◽

Enterotoxigenic Escherichia Coli ◽

Adenylate Cyclase System ◽

Mrna Levels ◽

Intestinal Epithelial ◽

E Coli ◽

Heat Labile ◽

Etec Infection

Infections with enteric pathogens like enterotoxigenic Escherichia coli ( ETEC) is a major health issue worldwide and while diarrhea is the major problem, prolonged, severe, and dual infections with multiple pathogens may also compromise the nutritional status of the infected individuals. There is almost nothing currently known about the effect of ETEC infection on intestinal absorptions of water-soluble vitamins including thiamin. We examined the effect of ETEC infection on intestinal uptake of the thiamin using as a model the human-derived intestinal epithelial Caco-2 cells. The results showed that infecting confluent Caco-2 monolayers with live ETEC (but not with boiled/killed ETEC or nonpathogenic E. coli) or treatment with bacterial culture supernatant led to a significant inhibition in thiamin uptake. This inhibition appears to be caused by a heat-labile and -secreted ETEC component and is mediated via activation of the epithelial adenylate cyclase system. The inhibition in thiamin uptake by ETEC was associated with a significant reduction in expression of human thiamin transporter-1 and -2 (hTHTR1 and hTHTR2) at the protein and mRNA levels as well as in the activity of the SLC19A2 and SLC19A3 promoters. Dual infection of Caco-2 cells with ETEC and EPEC (enteropathogenic E. coli) led to compounded inhibition in intestinal thiamin uptake. These results show for the first time that infection of human intestinal epithelial cells with ETEC causes a significant inhibition in intestinal thiamin uptake. This inhibition is mediated by a secreted heat-labile toxin and is associated with a decrease in the expression of intestinal thiamin transporters.

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Mucosal Adjuvant Properties of Mutant LT-IIa and LT-IIb Enterotoxins That Exhibit Altered Ganglioside-Binding Activities

Infection and Immunity ◽

10.1128/iai.73.3.1330-1342.2005 ◽

2005 ◽

Vol 73 (3) ◽

pp. 1330-1342 ◽

Cited By ~ 36

Author(s):

Hesham F. Nawar ◽

Sergio Arce ◽

Michael W. Russell ◽

Terry D. Connell

Keyword(s):

Single Point ◽

Binding Activity ◽

Type I ◽

Adjuvant Activity ◽

Adrenal Cells ◽

E Coli ◽

Heat Labile ◽

Adhesin Protein ◽

And Function ◽

Major Ganglioside

ABSTRACT LT-IIa and LT-IIb, the type II heat-labile enterotoxins of Escherichia coli, are closely related in structure and function to cholera toxin and LT-I, the type I heat-labile enterotoxins of Vibrio cholerae and E. coli, respectively. Recent studies from our group demonstrated that LT-IIa and LT-IIb are potent systemic and mucosal adjuvants. To determine whether binding of LT-IIa and LT-IIb to their specific ganglioside receptors is essential for adjuvant activity, LT-IIa and LT-IIb enterotoxins were compared with their respective single-point substitution mutants which have no detectable binding activity for their major ganglioside receptors [e.g., LT-IIa(T34I) and LT-IIb(T13I)]. Both mutant enterotoxins exhibited an extremely low capacity for intoxicating mouse Y1 adrenal cells and for inducing production of cyclic AMP in a macrophage cell line. BALB/c female mice were immunized by the intranasal route with the surface adhesin protein AgI/II of Streptococcus mutans alone or in combination with LT-IIa, LT-IIa(T34I), LT-IIb, or LT-IIb(T13I). Both LT-IIa and LT-IIb potentiated strong mucosal and systemic immune responses against AgI/II. Of the two mutant enterotoxins, only LT-IIb(T13I) had the capacity to strongly potentiate mucosal anti-AgI/II and systemic anti-AgI/II antibody responses. Upon boosting with AgI/II, however, both LT-IIa(T34I) and LT-IIb(T13I) enhanced humoral memory responses to AgI/II. Flow cytometry demonstrated that LT-IIa(T34I) had no affinity for cervical lymph node lymphocytes. In contrast, LT-IIb(T13I) retained binding activity for T cells, B cells, and macrophages, indicating that this immunostimulatory mutant enterotoxin interacts with one or more unknown lymphoid cell receptors.

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Characterization of Fluorescent Chimeras of Cholera Toxin and Escherichia coli Heat-Labile Enterotoxins Produced by Use of the Twin Arginine Translocation System

Infection and Immunity ◽

10.1128/iai.73.6.3627-3635.2005 ◽

2005 ◽

Vol 73 (6) ◽

pp. 3627-3635 ◽

Cited By ~ 17

Author(s):

Juliette K. Tinker ◽

Jarrod L. Erbe ◽

Randall K. Holmes

Keyword(s):

Escherichia Coli ◽

Cholera Toxin ◽

Fusion Proteins ◽

Cell Biology ◽

Fluorescent Protein ◽

Cultured Cells ◽

Secretory Diarrhea ◽

E Coli ◽

Twin Arginine Translocation ◽

Heat Labile

ABSTRACT Cholera toxin (CT) is an AB5 toxin responsible for the profuse secretory diarrhea resulting from Vibrio cholerae infection. CT consists of a pentameric, receptor-binding B subunit (CTB) and a monomeric A subunit (CTA) that has latent enzymatic activity. In addition to its enterotoxicity, CT has potent mucosal adjuvant activity and can also function as a carrier molecule with many potential applications in cell biology. In earlier studies, the toxic CTA1 domain was replaced by several other antigenic protein domains to produce holotoxin-like chimeras for use as potential mucosal vaccines. In the present study we utilized the twin arginine translocation (tat) system to produce fluorescent CT chimeras, as well as fluorescent chimeras of Escherichia coli heat-labile toxins LTI and LTIIb. Fusion proteins containing either green fluorescent protein (GFP) or monomeric red fluorescent protein (mRFP) and the A2 domain of CT, LTI, or LTIIb were transported to the periplasm of E. coli by the tat system, and the corresponding B polypeptides of CT, LTI, and LTIIb were transported to the periplasm by the sec system. The fluorescent fusion proteins were shown to assemble spontaneously and efficiently with the corresponding B polypeptides in the periplasm to form chimeric holotoxin-like molecules, and these chimeras bound to and entered cultured cells in a manner similar to native CT, LTI, or LTIIb. The GFP and mRFP derivatives of CT, LT, and LTIIb developed here are useful tools for studies on the cell biology of trafficking of the CT/LT family of bacterial enterotoxins. In addition, these constructs provide proof in principle for the development of novel chimeric CT-like or LT-like vaccine candidates containing CTA2 fusion proteins that cannot be delivered to the periplasm of E. coli by use of the sec secretion pathway.

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