title: Expression and functional analysis of the subtilin immunity genes spaIFEG in the subtilin-sensitive host Bacillus subtilis MO1099.
abstract: Bacillus subtilis ATCC 6633 produces the cationic pore-forming lantibiotic subtilin, which preferentially acts on gram-positive microorganisms; self protection of the producer cells is mediated by the four genes spaIFEG. To elucidate the mechanism of subtilin autoimmunity, we transferred different combinations of subtilin immunity genes under the control of an inducible promoter into the genome of subtilin-sensitive host strain B. subtilis MO1099. Recipient cells acquired subtilin tolerance through expression of either spaI or spaFEG, which shows that subtilin immunity is based on two independently acting systems. Cells coordinately expressing all four immunity genes acquired the strongest subtilin protection level. Quantitative in vivo peptide release assays demonstrated that SpaFEG diminished the quantity of cell-associated subtilin, suggesting that SpaFEG transports subtilin molecules from the membrane into the extracellular space. Homology and secondary structure analyses define SpaFEG as a prototype of lantibiotic immunity transporters that fall into the ABC-2 subfamily of multidrug resistance proteins. Membrane localization of the lipoprotein SpaI and specific interaction of SpaI with the cognate lantibiotic subtilin suggest a function of SpaI as a subtilin-intercepting protein. This interpretation was supported by hexahistidine-mediated 0-A cross-linking between hexahistidine-tagged SpaI and subtilin.
authors.affiliation: Johann Wolfgang Goethe-Universität, Institut für Mikrobiologie, Marie-Curie-Strasse 9, 60439 Frankfurt am Main, Germany. T.Stein@em.uni-frankfurt.de
authors.forename: Torsten
authors.initials: T
authors.lastname: Stein
authors.forename: Stefan
authors.initials: S
authors.lastname: Heinzmann
authors.forename: Stefanie
authors.initials: S
authors.lastname: Düsterhus
authors.forename: Stefan
authors.initials: S
authors.lastname: Borchert
authors.forename: Karl-Dieter
authors.initials: KD
authors.lastname: Entian
mesh_terms: D000595:Amino Acid Sequence; D001412:Bacillus subtilis / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics* / Q000276:immunology; D001430:Bacteriocins; D001483:Base Sequence; D017931:DNA Primers; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D010455:Peptides / Q000235:genetics* / Q000276:immunology; D010957:Plasmids; D016133:Polymerase Chain Reaction; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
chemical_list: D001426:Bacterial Proteins; D001430:Bacteriocins; D017931:DNA Primers; D010455:Peptides; C002836:subtilin