The Crystal Structure of Tetanus Toxin Hc Fragment Complexed with a Synthetic GT1b Analogue Suggests Cross-linking between Ganglioside Receptors and the Toxin

Fotinou, Constantina; Emsley, Paul; Black, Isobel; Ando, Hiromune; Ishida, Hideyuki; Kiso, Makoto; Sinha, Katharine A.; Fairweather, Neil F.; Isaacs, Neil W.

doi:10.1074/jbc.m103285200

Cited by 175 publications

(199 citation statements)

References 49 publications

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“…The critical importance of the last 34 residues from Hc-C, in particular His 1293 of TeNT, for binding the oligosaccharide portion of polysialogangliosides was demonstrated by photoaffinity labelling (Shapiro et al, 1997). In BoNT/B and BoNT/A, one ganglioside binding site was mapped within the Hc-C, whereas TeNT Hc-C binds simultaneously to two ganglioside molecules (Swaminathan and Eswaramoorthy, 2000;Fotinou et al, 2001;Rummel et al, 2003Rummel et al, , 2004b. These data support a doublereceptor model for binding of clostridial NTs to the presynaptic membrane.…”

Section: Transcytotic Pathway and Gangliosides: The Case Of Clostridimentioning

confidence: 99%

Bacterial protein toxins and lipids: role in toxin targeting and activity

Geny

Popoff

2006

Biology of the Cell

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All bacterial toxins, which globally are hydrophilic proteins, interact first with their target cells by recognizing a surface receptor, which is either a lipid or a lipid derivative, or another compound but in a lipid environment. Intracellular active toxins follow various trafficking pathways, the sorting of which is greatly dependent on the nature of the receptor, notably lipidic receptor or receptor embedded into a distinct environment such as lipid microdomains. Numerous other toxins act locally on cell membrane. Indeed, phospholipase activity is a common mechanism shared by several membrane‐damaging toxins. In addition, many toxins active intracellularly or on cell membrane modulate host cell phospholipid pathways. Unusually, a few bacterial toxins require a lipid post‐translational modification to be active. Thereby, lipids are obligate partners of bacterial toxins.

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Section: Transcytotic Pathway and Gangliosides: The Case Of Clostridimentioning

confidence: 99%

Bacterial protein toxins and lipids: role in toxin targeting and activity

Geny

Popoff

2006

Biology of the Cell

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“…As expected for booster vaccination, all antibodies bound the vaccine antigen TT with high affinities (K d ∼ 2.2 × 10 −8 − 1 × 10 −10 M). Neutralizing epitopes of the 150-kDa holotoxin have been reported to reside within rTT.C (24). Five serum IgGs (HD1-2, HD1-5, HD2-49, HD2-88, and HD2-89) were shown to bind to rTT.C with subnanomolar K d values.…”

Section: Identities and Dynamics Of The Serum Antibody Response To Vamentioning

confidence: 99%

Identification and characterization of the constituent human serum antibodies elicited by vaccination

Lavinder

Wine

Giesecke

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

239

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Most vaccines confer protection via the elicitation of serum antibodies, yet more than 100 y after the discovery of antibodies, the molecular composition of the human serum antibody repertoire to an antigen remains unknown. Using high-resolution liquid chromatography tandem MS proteomic analyses of serum antibodies coupled with next-generation sequencing of the V gene repertoire in peripheral B cells, we have delineated the human serum IgG and B-cell receptor repertoires following tetanus toxoid (TT) booster vaccination. We show that the TT + serum IgG repertoire comprises ∼100 antibody clonotypes, with three clonotypes accounting for >40% of the response. All 13 recombinant IgGs examined bound to vaccine antigen with K d ∼ 10 −8 -10 −10 M. Five of 13 IgGs recognized the same linear epitope on TT, occluding the binding site used by the toxin for cell entry, suggesting a possible explanation for the mechanism of protection conferred by the vaccine. Importantly, only a small fraction (<5%) of peripheral blood plasmablast clonotypes (CD3 − CD14 − CD19 + CD27 ++ CD38 ++ CD20 − TT + ) at the peak of the response (day 7), and an even smaller fraction of memory B cells, were found to encode antibodies that could be detected in the serological memory response 9 mo postvaccination. This suggests that only a small fraction of responding peripheral B cells give rise to the bone marrow long-lived plasma cells responsible for the production of biologically relevant amounts of vaccine-specific antibodies (near or above the K d ). Collectively, our results reveal the nature and dynamics of the serological response to vaccination with direct implications for vaccine design and evaluation.B-cell repertoire | proteomics M ost approved vaccines confer protection against infectious diseases by the induction of long-lived plasma cells (LLPCs), which secrete antibodies that serve to neutralize and opsonize the pathogen for many years or decades (1-3). Additionally, the generation of memory B cells (mBCs) provides both a mechanism for the rapid synthesis of affinity matured, antigenspecific antibodies following rechallenge and a means to diversify the humoral immune response to confer protection against rapidly evolving viruses or bacteria (4). Although some vaccines elicit antibody titers that remain virtually constant for many decades, for others, including the tetanus toxoid (TT) vaccine, antibody titers wane monotonically over time (5). Booster immunization triggers the rapid expansion and differentiation of cognate B cells, generating antigen-specific plasmablasts that peak in concentration in peripheral blood after 6-7 d and subsequently rapidly decline to nearly undetectable levels (6, 7). Some, but not all, of these peak-wave plasmablasts migrate to specialized niches overwhelmingly located in the bone marrow (BM) and survive as LLPCs (8), which constitute the major source of all classes of Ig in the serum (9).The establishment of serological memory following either primary or booster vaccination is not understood well (10-14)...

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“…1B). Strikingly, pocket 4 matches the position of TeNT's second carbohydrate binding site, the sialic acid binding site (24,27). Fig.…”

Section: Computer-based Surface Analysis Of the Hcc Of Bont/b Revealsmentioning

confidence: 99%

“…However, a second carbohydrate binding site, the sialic acid binding site, was described only for TeNT (26,27). The sialic acid binding site in TeNT could be used to interact with the sialic acid-containing glycoprotein receptor (21,24).…”

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confidence: 99%

Identification of the protein receptor binding site of botulinum neurotoxins B and G proves the double-receptor concept

Rummel

Eichner

Weil

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

171

216

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Botulinum neurotoxins (BoNTs) cause muscle paralysis by selectively cleaving core components of the vesicular fusion machinery within motoneurons. Complex gangliosides initially bind into a pocket that is conserved among the seven BoNTs and tetanus neurotoxin. Productive neurotoxin uptake also requires protein receptors. The interaction site of the protein receptor within the neurotoxin is currently unknown. We report the identification and characterization of the protein receptor binding site of BoNT/B and BoNT/G. Their protein receptors, synaptotagmins I and II, bind to a pocket at the tip of their HCC (C-terminal domain of the C-terminal fragment of the heavy chain) that corresponds to the unique second carbohydrate binding site of tetanus neurotoxin, the sialic acid binding site. Substitution of amino acids in this region impaired binding to synaptotagmins and drastically decreased toxicity at mouse phrenic nerve preparations; CD-spectroscopic analyses evidenced that the secondary structure of the mutated neurotoxins was unaltered. Deactivation of the synaptotagmin binding site by single mutations led to virtually inactive BoNT/B and BoNT/G when assayed at phrenic nerve preparations of complex-ganglioside-deficient mice. Analogously, a BoNT B mutant with deactivated ganglioside and synaptotagmin binding sites lacked appreciable activity at wild-type mouse phrenic nerve preparations. Thus, these data exclude relevant contributions of any cell surface molecule other than one ganglioside and one protein receptor to the entry process of BoNTs, which substantiates the double-receptor concept. The molecular characterization of the synaptotagmin binding site provides the basis for designing a novel class of potent binding inhibitors.synaptotagmin ͉ tetanus B otulinum neurotoxins (BoNTs) (serotypes A-G) are the causative agents of the disease botulism. The neurotoxins are produced as Ϸ150-kDa single-chain proteins in Clostridium botulinum and subsequently cleaved by proteases, yielding an Ϸ100-kDa heavy chain (HC) and an Ϸ50-kDa light chain (LC). These chains remain connected via a single disulfide bridge, noncovalent interactions, and a HC-derived peptide loop wrapped around the LC. The LCs act as zinc metallopeptidases, which solely hydrolyze one of three SNARE (soluble N-ethylmaleimide-sensitive fusion protein attachment protein receptor) proteins: vesicle associated membrane protein/synaptobrevin, synaptosomal-associated protein of 25 kDa, or syntaxin. Together, these substrate molecules constitute the core of the vesicular fusion machinery. Thus, cleavage of one of these proteins inhibits the release of neurotransmitters from synaptic vesicles into the synaptic cleft. The HCs mediate the neurospecific binding, uptake by receptor-mediated endocytosis, and transport of the LC across the endosomal membrane into the cytosol, where the LCs encounter their substrates. The Ϸ50-kDa

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The Crystal Structure of Tetanus Toxin Hc Fragment Complexed with a Synthetic GT1b Analogue Suggests Cross-linking between Ganglioside Receptors and the Toxin

Cited by 175 publications

References 49 publications

Bacterial protein toxins and lipids: role in toxin targeting and activity

Bacterial protein toxins and lipids: role in toxin targeting and activity

Identification and characterization of the constituent human serum antibodies elicited by vaccination

Identification of the protein receptor binding site of botulinum neurotoxins B and G proves the double-receptor concept

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