title: Inhibition of type VI secretion by an anti-TssM llama nanobody.
abstract: The type VI secretion system (T6SS) is a secretion pathway widespread in Gram-negative bacteria that targets toxins in both prokaryotic and eukaryotic cells. Although most T6SSs identified so far are involved in inter-bacterial competition, a few are directly required for full virulence of pathogens. The T6SS comprises 13 core proteins that assemble a large complex structurally and functionally similar to a phage contractile tail structure anchored to the cell envelope by a trans-membrane spanning stator. The central part of this stator, TssM, is a 1129-amino-acid protein anchored in the inner membrane that binds to the TssJ outer membrane lipoprotein. In this study, we have raised camelid antibodies against the purified TssM periplasmic domain. We report the crystal structure of two specific nanobodies that bind to TssM in the nanomolar range. Interestingly, the most potent nanobody, nb25, competes with the TssJ lipoprotein for TssM binding in vitro suggesting that TssJ and the nb25 CDR3 loop share the same TssM binding site or causes a steric hindrance preventing TssM-TssJ complex formation. Indeed, periplasmic production of the nanobodies displacing the TssM-TssJ interaction inhibits the T6SS function in vivo. This study illustrates the power of nanobodies to specifically target and inhibit bacterial secretion systems.
authors: Architecture et Fonction des Macromolécules Biologiques, Centre National de la Recherche Scientifique (CNRS)-UMR 7257, Marseille, France; Architecture et Fonction des Macromolécules Biologiques, Aix-Marseille Université, Campus de Luminy, Case 932, Marseille, France.
authors: Van Son
authors: VS
authors: Nguyen
authors: Laboratoire d'Ingénierie des Systèmes Macromoléculaires, Institut de Microbiologie de la Méditerranée, Aix-Marseille Université, CNRS-UMR 7255, 31 chemin Joseph Aiguier, Marseille, France.
authors: Laureen
authors: L
authors: Logger
authors: Architecture et Fonction des Macromolécules Biologiques, Centre National de la Recherche Scientifique (CNRS)-UMR 7257, Marseille, France; Architecture et Fonction des Macromolécules Biologiques, Aix-Marseille Université, Campus de Luminy, Case 932, Marseille, France.
authors: Silvia
authors: S
authors: Spinelli
authors: Architecture et Fonction des Macromolécules Biologiques, Centre National de la Recherche Scientifique (CNRS)-UMR 7257, Marseille, France; Architecture et Fonction des Macromolécules Biologiques, Aix-Marseille Université, Campus de Luminy, Case 932, Marseille, France.
authors: Aline
authors: A
authors: Desmyter
authors: Architecture et Fonction des Macromolécules Biologiques, Centre National de la Recherche Scientifique (CNRS)-UMR 7257, Marseille, France; Architecture et Fonction des Macromolécules Biologiques, Aix-Marseille Université, Campus de Luminy, Case 932, Marseille, France.
authors: Thi Thu Hang
authors: TT
authors: Le
authors: Architecture et Fonction des Macromolécules Biologiques, Centre National de la Recherche Scientifique (CNRS)-UMR 7257, Marseille, France; Architecture et Fonction des Macromolécules Biologiques, Aix-Marseille Université, Campus de Luminy, Case 932, Marseille, France.
authors: Christine
authors: C
authors: Kellenberger
authors: Architecture et Fonction des Macromolécules Biologiques, Centre National de la Recherche Scientifique (CNRS)-UMR 7257, Marseille, France; Architecture et Fonction des Macromolécules Biologiques, Aix-Marseille Université, Campus de Luminy, Case 932, Marseille, France.
authors: Badreddine
authors: B
authors: Douzi
authors: Architecture et Fonction des Macromolécules Biologiques, Centre National de la Recherche Scientifique (CNRS)-UMR 7257, Marseille, France; Architecture et Fonction des Macromolécules Biologiques, Aix-Marseille Université, Campus de Luminy, Case 932, Marseille, France.
authors: Eric
authors: E
authors: Durand
authors: Architecture et Fonction des Macromolécules Biologiques, Centre National de la Recherche Scientifique (CNRS)-UMR 7257, Marseille, France; Architecture et Fonction des Macromolécules Biologiques, Aix-Marseille Université, Campus de Luminy, Case 932, Marseille, France.
authors: Alain
authors: A
authors: Roussel
authors: Laboratoire d'Ingénierie des Systèmes Macromoléculaires, Institut de Microbiologie de la Méditerranée, Aix-Marseille Université, CNRS-UMR 7255, 31 chemin Joseph Aiguier, Marseille, France.
authors: Eric
authors: E
authors: Cascales
authors: Architecture et Fonction des Macromolécules Biologiques, Centre National de la Recherche Scientifique (CNRS)-UMR 7257, Marseille, France; Architecture et Fonction des Macromolécules Biologiques, Aix-Marseille Université, Campus de Luminy, Case 932, Marseille, France.
authors: Christian
authors: C
authors: Cambillau
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000915:Antibody Affinity; D000918:Antibody Specificity / Q000276:immunology; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism; D002161:Camelids, New World; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011485:Protein Binding; D011487:Protein Conformation; D016415:Sequence Alignment; D061905:Single-Domain Antibodies / Q000737:chemistry / Q000276:immunology / Q000494:pharmacology*; D013816:Thermodynamics; D000069376:Type VI Secretion Systems / Q000037:antagonists & inhibitors* / Q000276:immunology
chemical_list: D001426:Bacterial Proteins; D061905:Single-Domain Antibodies; D000069376:Type VI Secretion Systems