title: Functional assembly of two membrane-binding domains in listeriolysin O, the cytolysin of Listeria monocytogenes.
abstract: Listeriolysin O (LLO) is a major virulence factor secreted by the pathogenic Listeria monocytogenes and acts as pore-forming cytolysin. Based on sequence similarities between LLO and perfringolysin (PFO), the cytolysin from Clostridium perfringens of known crystallographic structure, two truncated LLO proteins were produced: LLO-d123, comprising the first three predicted domains, and LLO-d4, the last C-terminal domain. The two proteins were efficiently secreted into the culture supernatant of L. monocytogenes and were able to bind to cell membranes. Strikingly, when expressed simultaneously, the two secreted domains LLO-d123 and LLO-d4 reassembled into a haemolytically active form. Two in-frame linker insertions were generated in the hinge region between the d123 and d4 domains. In both cases, the insertion created a major cleavage site for proteolytic degradation and abolished cytolytic activity, which might suggest that the region connecting d123 and d4 participates in the interaction between the two portions of the monomer.
authors.affiliation: Laboratoire de Microbiologie, INSERM U-411, Faculté de Médecine Necker-Enfants Malades, 156 rue de Vaugirard, 75730 Paris Cedex 15, France1.
authors.forename: Iharilalao
authors.initials: I
authors.lastname: Dubail
authors.affiliation: Laboratoire de Microbiologie, INSERM U-411, Faculté de Médecine Necker-Enfants Malades, 156 rue de Vaugirard, 75730 Paris Cedex 15, France1.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Autret
authors.affiliation: Laboratoire de Microbiologie, INSERM U-411, Faculté de Médecine Necker-Enfants Malades, 156 rue de Vaugirard, 75730 Paris Cedex 15, France1.
authors.forename: Jean-Luc
authors.initials: JL
authors.lastname: Beretti
authors.affiliation: Laboratoire de Microbiologie, INSERM U-411, Faculté de Médecine Necker-Enfants Malades, 156 rue de Vaugirard, 75730 Paris Cedex 15, France1.
authors.forename: Samer
authors.initials: S
authors.lastname: Kayal
authors.affiliation: Laboratoire de Microbiologie, INSERM U-411, Faculté de Médecine Necker-Enfants Malades, 156 rue de Vaugirard, 75730 Paris Cedex 15, France1.
authors.forename: Patrick
authors.initials: P
authors.lastname: Berche
authors.affiliation: Laboratoire de Microbiologie, INSERM U-411, Faculté de Médecine Necker-Enfants Malades, 156 rue de Vaugirard, 75730 Paris Cedex 15, France1.
authors.forename: Alain
authors.initials: A
authors.lastname: Charbit
mesh_terms: D000818:Animals; D001427:Bacterial Toxins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism*; D003603:Cytotoxins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004910:Erythrocyte Membrane / Q000378:metabolism; D005057:Eukaryotic Cells / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006460:Hemolysin Proteins; D006461:Hemolysis; D006801:Humans; D008089:Listeria monocytogenes / Q000737:chemistry / Q000378:metabolism*; D008958:Models, Molecular; D016254:Mutagenesis, Insertional; D017510:Protein Folding; D017422:Sequence Analysis, DNA; D013329:Structure-Activity Relationship
chemical_list: D001427:Bacterial Toxins; D003603:Cytotoxins; D006360:Heat-Shock Proteins; D006460:Hemolysin Proteins; C020452:Clostridium perfringens theta-toxin; C038705:hlyA protein, Listeria monocytogenes