title: Discoupling the Ca(2+)-activation from the pore-forming function of the bi-component Panton-Valentine leucocidin in human PMNs.
abstract: The consecutive cell activation, including Ca(2+)-channel opening, and pore formation leading to human neutrophil lysis were the two functions of the staphylococcal Panton-Valentine leucocidin attempted to be discoupled by site-directed mutagenesis. In a first approach consisting in deletions of the cytoplasmic extremity of the transmembranous domain, we produced a LukF-PV DeltaSer125-Leu128 with a slightly reduced Ca(2+) induction but with a significantly lowered lytic activity when combined with its synergistic protein LukS-PV. The second approach consisted in the modification of charges and/or introduction of a steric hindrance inside the pore, which also led to interesting mutated proteins: LukF-PV G131D, G131W and G130D. The latter had an intact Ca(2+) induction ability while the lytic one was 20-fold diminished. Binding properties and intrinsic pore diameters of these discoupled toxins remained comparable to the wild-type protein. The mutated proteins promoted interleukin-8 secretion, but they were rather inactive in an experimental model. New insights are brought concerning the role of the two functions in the virulence of this bi-component leucotoxin.
authors.affiliation: UPRES EA-1318, Institut de Bactériologie de la Faculté de Médecine, Université Louis Pasteur-Hôpitaux Universitaires de Strasbourg, 3 rue Koeberlé, F-67000, Strasbourg, France.
authors.forename: L
authors.initials: L
authors.lastname: Baba Moussa
authors.forename: S
authors.initials: S
authors.lastname: Werner
authors.forename: D A
authors.initials: DA
authors.lastname: Colin
authors.forename: L
authors.initials: L
authors.lastname: Mourey
authors.forename: J D
authors.initials: JD
authors.lastname: Pédelacq
authors.forename: J P
authors.initials: JP
authors.lastname: Samama
authors.forename: A
authors.initials: A
authors.lastname: Sanni
authors.forename: H
authors.initials: H
authors.lastname: Monteil
authors.forename: G
authors.initials: G
authors.lastname: Prévost
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D001427:Bacterial Toxins; D015220:Calcium Channels / Q000502:physiology*; D020013:Calcium Signaling / Q000187:drug effects*; D002462:Cell Membrane / Q000187:drug effects / Q000648:ultrastructure; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D004926:Escherichia coli; D005098:Exotoxins; D006801:Humans; D016209:Interleukin-8 / Q000378:metabolism; D017136:Ion Transport; D007956:Leukocidins / Q000737:chemistry / Q000235:genetics / Q000633:toxicity*; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D009504:Neutrophils / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000633:toxicity; D013211:Staphylococcus aureus / Q000472:pathogenicity*; D013329:Structure-Activity Relationship; D014774:Virulence
chemical_list: D001427:Bacterial Toxins; D015220:Calcium Channels; D005098:Exotoxins; D016209:Interleukin-8; D007956:Leukocidins; C078284:Panton-Valentine leukocidin; D011993:Recombinant Fusion Proteins