Dataset Information

Structural Basis for the Specific Neutralization of Stx2a with a Camelid Single Domain Antibody Fragment.

ABSTRACT: BACKGROUND:Shiga toxin-producing Escherichia coli (STEC) are a subset of pathogens leading to illnesses such as diarrhea, hemolytic uremic syndrome and even death. The Shiga toxins are the main virulence factors and divided in two groups: Stx1 and Stx2, of which the latter is more frequently associated with severe pathologies in humans. RESULTS:An immune library of nanobodies (Nbs) was constructed after immunizing an alpaca with recombinant Shiga toxin-2a B subunit (rStx2aB), to retrieve multiple rStx2aB-specific Nbs. The specificity of five Nbs towards rStx2aB was confirmed in ELISA and Western blot. Nb113 had the highest affinity (9.6 nM) and its bivalent construct exhibited a 100-fold higher functional affinity. The structure of the Nb113 in complex with rStx2aB was determined via X-ray crystallography. The crystal structure of the Nb113-rStx2aB complex revealed that five copies of Nb113 bind to the rStx2aB pentamer and that the Nb113 epitope overlaps with the Gb3 binding site, thereby providing a structural basis for the neutralization of Stx2a by Nb113 that was observed on Vero cells. Finally, the tandem-repeated, bivalent Nb113? exhibits a higher toxin neutralization capacity compared to monovalent Nb113. CONCLUSIONS:The Nb of highest affinity for rStx2aB is also the best Stx2a and Stx2c toxin neutralizing Nb, especially in a bivalent format. This lead Nb neutralizes Stx2a by competing for the Gb3 receptor. The fusion of the bivalent Nb113? with a serum albumin specific Nb is expected to combine high toxin neutralization potential with prolonged blood circulation.

SUBMITTER: Bernedo-Navarro RA

PROVIDER: S-EPMC5869396 | biostudies-literature | 2018 Mar

REPOSITORIES: biostudies-literature

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Structural Basis for the Specific Neutralization of Stx2a with a Camelid Single Domain Antibody Fragment.

Bernedo-Navarro Robert Alvin RA Romão Ema E Yano Tomomasa T Pinto Joar J De Greve Henri H Sterckx Yann G-J YG Muyldermans Serge S

Toxins 20180301 3

<h4>Background</h4>Shiga toxin-producing <i>Escherichia coli</i> (STEC) are a subset of pathogens leading to illnesses such as diarrhea, hemolytic uremic syndrome and even death. The Shiga toxins are the main virulence factors and divided in two groups: Stx1 and Stx2, of which the latter is more frequently associated with severe pathologies in humans.<h4>Results</h4>An immune library of nanobodies (Nbs) was constructed after immunizing an alpaca with recombinant Shiga toxin-2a B subunit (rStx2aB ...[more]

PMID: 29494518

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Dataset Information

Structural Basis for the Specific Neutralization of Stx2a with a Camelid Single Domain Antibody Fragment.

Publications

Structural Basis for the Specific Neutralization of Stx2a with a Camelid Single Domain Antibody Fragment.

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