title: Crystal structure and self-interaction of the type VI secretion tail-tube protein from enteroaggregative Escherichia coli.
abstract: The type VI secretion system (T6SS) is a widespread machine used by bacteria to control their environment and kill or disable bacterial species or eukaryotes through toxin injection. The T6SS comprises a central tube formed of stacked hexamers of hemolysin co-regulated proteins (Hcp) and terminated by a trimeric valine-glycine repeat protein G (VgrG) component, the cell puncturing device. A contractile tail sheath, formed by the TssB and TssC proteins, surrounds this tube. This syringe-like machine has been compared to an inverted phage, as both Hcp and VgrG share structural homology with tail components of Caudovirales. Here we solved the crystal structure of a tryptophan-substituted double mutant of Hcp1 from enteroaggregative Escherichia coli and compared it to the structures of other Hcps. Interestingly, we observed that the purified Hcp native protein is unable to form tubes in vitro. To better understand the rationale for observation, we measured the affinity of Hcp1 hexamers with themselves by surface plasmon resonance. The intra-hexamer interaction is weak, with a KD value of 7.2 µM. However, by engineering double cysteine mutants at defined positions, tubes of Hcp1 gathering up to 15 stacked hexamers formed in oxidative conditions. These results, together with those available in the literature regarding TssB and TssC, suggest that assembly of the T6SS tube differs significantly from that of Sipho- or Myoviridae.
authors.affiliation: Aix-Marseille Université, Architecture et Fonction des Macromolécules Biologiques, UMR 7257, Campus de Luminy, Case 932, Marseille, France ; Centre National de la Recherche Scientifique, Architecture et Fonction des Macromolécules Biologiques, UMR 7257, Campus de Luminy, Case 932, Marseille, France.
authors.forename: Badreddine
authors.initials: B
authors.lastname: Douzi
authors.affiliation: Aix-Marseille Université, Architecture et Fonction des Macromolécules Biologiques, UMR 7257, Campus de Luminy, Case 932, Marseille, France ; Centre National de la Recherche Scientifique, Architecture et Fonction des Macromolécules Biologiques, UMR 7257, Campus de Luminy, Case 932, Marseille, France.
authors.forename: Silvia
authors.initials: S
authors.lastname: Spinelli
authors.affiliation: Aix-Marseille Université, Architecture et Fonction des Macromolécules Biologiques, UMR 7257, Campus de Luminy, Case 932, Marseille, France ; Centre National de la Recherche Scientifique, Architecture et Fonction des Macromolécules Biologiques, UMR 7257, Campus de Luminy, Case 932, Marseille, France.
authors.forename: Stéphanie
authors.initials: S
authors.lastname: Blangy
authors.affiliation: Aix-Marseille Université, Architecture et Fonction des Macromolécules Biologiques, UMR 7257, Campus de Luminy, Case 932, Marseille, France ; Centre National de la Recherche Scientifique, Architecture et Fonction des Macromolécules Biologiques, UMR 7257, Campus de Luminy, Case 932, Marseille, France.
authors.forename: Alain
authors.initials: A
authors.lastname: Roussel
authors.affiliation: Aix-Marseille Université, Architecture et Fonction des Macromolécules Biologiques, UMR 7257, Campus de Luminy, Case 932, Marseille, France ; Centre National de la Recherche Scientifique, Architecture et Fonction des Macromolécules Biologiques, UMR 7257, Campus de Luminy, Case 932, Marseille, France.
authors.forename: Eric
authors.initials: E
authors.lastname: Durand
authors.affiliation: Laboratoire d'Ingénierie des Systèmes Macromoléculaires, Institut de Microbiologie de la Méditerranée, Centre National de la Recherche Scientifique UMR7255, Aix-Marseille Université, Marseille, France.
authors.forename: Yannick R
authors.initials: YR
authors.lastname: Brunet
authors.affiliation: Laboratoire d'Ingénierie des Systèmes Macromoléculaires, Institut de Microbiologie de la Méditerranée, Centre National de la Recherche Scientifique UMR7255, Aix-Marseille Université, Marseille, France.
authors.forename: Eric
authors.initials: E
authors.lastname: Cascales
authors.affiliation: Aix-Marseille Université, Architecture et Fonction des Macromolécules Biologiques, UMR 7257, Campus de Luminy, Case 932, Marseille, France ; Centre National de la Recherche Scientifique, Architecture et Fonction des Macromolécules Biologiques, UMR 7257, Campus de Luminy, Case 932, Marseille, France.
authors.forename: Christian
authors.initials: C
authors.lastname: Cambillau
mesh_terms: D000595:Amino Acid Sequence; D058947:Bacterial Secretion Systems; D002850:Chromatography, Gel; D018360:Crystallography, X-Ray; D004220:Disulfides / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism* / Q000648:ultrastructure; D008027:Light; D008958:Models, Molecular; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D055503:Protein Multimerization; D012542:Scattering, Radiation; D016415:Sequence Alignment; D040681:Structural Homology, Protein; D020349:Surface Plasmon Resonance; D037521:Virulence Factors / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D058947:Bacterial Secretion Systems; D004220:Disulfides; D029968:Escherichia coli Proteins; C571201:HCP1 protein, E coli; D050505:Mutant Proteins; D037521:Virulence Factors