title: Independent domain assembly in a trapped folding intermediate of multimeric outer membrane secretins.
abstract: The outer membrane portal of the Klebsiella oxytoca type II secretion system, PulD, is a prototype of a family of proteins, the secretins, which are essential components of many bacterial secretion and pilus assembly machines. PulD is a homododecamer with a periplasmic vestibule and an outer chamber on either side of a membrane-spanning region that is poorly resolved by electron microscopy. Membrane insertion involves the formation of a dodecameric membrane-embedded intermediate. Here, we describe an amino acid substitution in PulD that blocks its assembly at this intermediate "prepore" stage. Electron microscopy indicated that the prepore has an apparently normal periplasmic vestibule but a poorly organized outer chamber. A peptide loop around this amino acid appears to be important for the formation/stability of the fully folded complex. A similar assembly intermediate results from creation of the same amino acid substitution in the Pseudomonas aeruginosa secretin XcpQ.
authors.affiliation: Molecular Genetics Unit, Departments of Microbiology and of Structural Biology and Chemistry, Institut Pasteur, rue du Dr. Roux, 75724 Paris Cedex 15, France; CNRS ERL3526, rue du Dr. Roux, 75724 Paris Cedex 15, France.
authors.forename: Ingrid
authors.initials: I
authors.lastname: Guilvout
authors.affiliation: C-CINA Center for Cellular Imaging and NanoAnalytics, Biozentrum, University of Basel, 4058 Basel, Switzerland.
authors.forename: Mohamed
authors.initials: M
authors.lastname: Chami
authors.affiliation: Molecular Genetics Unit, Departments of Microbiology and of Structural Biology and Chemistry, Institut Pasteur, rue du Dr. Roux, 75724 Paris Cedex 15, France; CNRS ERL3526, rue du Dr. Roux, 75724 Paris Cedex 15, France; Institut de Biochimie et de Biophysique Moléculaire et Cellulaire, Université de Paris-Sud, 91405 Orsay, France; CNRS UMR 8619, 91405 Orsay, France.
authors.forename: Elena
authors.initials: E
authors.lastname: Disconzi
authors.affiliation: Institut de Biochimie et de Biophysique Moléculaire et Cellulaire, Université de Paris-Sud, 91405 Orsay, France; CNRS UMR 8619, 91405 Orsay, France.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Bayan
authors.affiliation: Molecular Genetics Unit, Departments of Microbiology and of Structural Biology and Chemistry, Institut Pasteur, rue du Dr. Roux, 75724 Paris Cedex 15, France; CNRS ERL3526, rue du Dr. Roux, 75724 Paris Cedex 15, France. Electronic address: max@pasteur.fr.
authors.forename: Anthony P
authors.initials: AP
authors.lastname: Pugsley
authors.affiliation: Molecular Genetics Unit, Departments of Microbiology and of Structural Biology and Chemistry, Institut Pasteur, rue du Dr. Roux, 75724 Paris Cedex 15, France; CNRS ERL3526, rue du Dr. Roux, 75724 Paris Cedex 15, France. Electronic address: ghuysman@pasteur.fr.
authors.forename: Gerard H M
authors.initials: GH
authors.lastname: Huysmans
mesh_terms: D019943:Amino Acid Substitution; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000648:ultrastructure*; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000648:ultrastructure*; D058947:Bacterial Secretion Systems / Q000502:physiology; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D041121:Klebsiella oxytoca / Q000737:chemistry* / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000648:ultrastructure*; D011485:Protein Binding; D017510:Protein Folding; D055503:Protein Multimerization; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011550:Pseudomonas aeruginosa / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000648:ultrastructure; D040681:Structural Homology, Protein; D013329:Structure-Activity Relationship
chemical_list: D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D058947:Bacterial Secretion Systems; D008565:Membrane Proteins; D011994:Recombinant Proteins; C089841:XcpQ protein, Pseudomonas aeruginosa