Selection of nanobodies that block the enzymatic and cytotoxic activities of the binary Clostridium difficile toxin CDT

Authors
Unger, Mandy; Eichhoff, Anna Marei; Schumacher, Lucas; Strysio, Moritz; Menzel, Stephan; Schwan, Carsten; Alzogaray, Vanina Andrea; Zylberman, Vanesa; Seman, Michel; Brandner, Johanna; Rohde, Holger; Zhu, Kai; Haag, Friedrich; Mittrucker, Hans Willi; Goldbaum, Fernando Alberto; Aktories, Klaus; Koch Nolte, Friedrich
Publication Year
2015
Language
English
Format
article
Status
Published version
Description
The spore-forming gut bacterium Clostridium difficile is the leading cause of antibiotic-associated diarrhea in hospitalized patients. The major virulence factors are two large glucosylating cytotoxins. Hypervirulent strains (e.g. ribotype 027) with higher morbidity and mortality additionally produce the binary CDT toxin (Clostridium difficile transferase) that ADP-ribosylates actin and induces microtubule-based cell protrusions. Nanobodies are robust single domain antibodies derived from camelid heavy chain antibodies. Here we report the generation of functional nanobodies against the enzymatic CDTa and the heptameric receptor binding subunit CDTb. The nanobodies were obtained from a variable-domain repertoire library isolated from llamas immunized with recombinant CDTa or CDTb. Five CDTa-specific nanobodies blocked CDTa-mediated ADP-ribosylation of actin. Three CDTa-specific and two CDTb-specific nanobodies neutralized the cytotoxicity of CDTa+b. These nanobodies hold promise as new tools for research, diagnosis and therapy of C. difficile associated disease.
Fil: Unger, Mandy. Universitat Hamburg; Alemania
Fil: Eichhoff, Anna Marei. Universitat Hamburg; Alemania
Fil: Schumacher, Lucas. Universitat Hamburg; Alemania
Fil: Strysio, Moritz. Universitat Hamburg; Alemania
Fil: Menzel, Stephan. Universitat Hamburg; Alemania
Fil: Schwan, Carsten. University of Freiburg; Alemania
Fil: Alzogaray, Vanina Andrea. Fundación Instituto Leloir; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina
Fil: Zylberman, Vanesa. Fundación Instituto Leloir; Argentina
Fil: Seman, Michel. University Henry Poincare; Francia
Fil: Brandner, Johanna. Universitat Hamburg; Alemania
Fil: Rohde, Holger. Universitat Hamburg; Alemania
Fil: Zhu, Kai. State University Of New York; Estados Unidos
Fil: Haag, Friedrich. Universitat Hamburg; Alemania
Fil: Mittrucker, Hans Willi. Universitat Hamburg; Alemania
Fil: Goldbaum, Fernando Alberto. Fundación Instituto Leloir; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina
Fil: Aktories, Klaus. Universitat Hamburg; Alemania
Fil: Koch Nolte, Friedrich. Universitat Hamburg; Alemania
Subject
nanobodies
CDT toxin
Clostridium difficile
Bioquímica y Biología Molecular
Ciencias Biológicas
CIENCIAS NATURALES Y EXACTAS
Access level
Open access
License
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
Repository
CONICET Digital (CONICET)
Institution
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identifier
oai:ri.conicet.gov.ar:11336/9651