title: Aggregates of nisin with various bactoprenol-containing cell wall precursors differ in size and membrane permeation capacity.
abstract: Many lantibiotics use the membrane bound cell wall precursor Lipid II as a specific target for killing Gram-positive bacteria. Binding of Lipid II usually impedes cell wall biosynthesis, however, some elongated lantibiotics such as nisin, use Lipid II also as a docking molecule for pore formation in bacterial membranes. Although the unique nisin pore formation can be analyzed in Lipid II-doped vesicles, mechanistic details remain elusive. We used optical sectioning microscopy to directly visualize the interaction of fluorescently labeled nisin with membranes of giant unilamellar vesicles containing Lipid II and its various bactoprenol precursors. We quantitatively analyzed the binding and permeation capacity of nisin when applied at nanomolar concentrations. Specific interactions with Lipid I, Lipid II and bactoprenol-diphosphate (C55-PP), but not bactoprenol-phosphate (C55-P), resulted in the formation of large molecular aggregates. For Lipid II, we demonstrated the presence of both nisin and Lipid II in these aggregates. Membrane permeation induced by nisin was observed in the presence of Lipid I and Lipid II, but not in the presence of C55-PP. Notably, the size of the C55-PP-nisin aggregates was significantly smaller than that of the aggregates formed with Lipid I and Lipid II. We conclude that the membrane permeation capacity of nisin is determined by the size of the bactoprenol-containing aggregates in the membrane. Notably, transmitted light images indicated that the formation of large aggregates led to a pinch-off of small vesicles, a mechanism, which probably limits the growth of aggregates and induces membrane leakage.
authors.affiliation: Institute for Physical and Theoretical Chemistry, Wegeler Str. 12, Rheinische Friedrich-Wilhelms-University Bonn, 53115 Bonn, Germany. Electronic address: kscherer@uni-bonn.de.
authors.forename: Katharina
authors.initials: K
authors.lastname: Scherer
authors.forename: Imke
authors.initials: I
authors.lastname: Wiedemann
authors.forename: Corina
authors.initials: C
authors.lastname: Ciobanasu
authors.forename: Hans-Georg
authors.initials: HG
authors.lastname: Sahl
authors.forename: Ulrich
authors.initials: U
authors.lastname: Kubitscheck
mesh_terms: D002463:Cell Membrane Permeability / Q000187:drug effects*; D002473:Cell Wall / Q000737:chemistry*; D005456:Fluorescent Dyes / Q000737:chemistry; D018613:Microscopy, Confocal; D009561:Nisin / Q000737:chemistry* / Q000494:pharmacology; D013729:Terpenes / Q000737:chemistry*
chemical_list: D005456:Fluorescent Dyes; D013729:Terpenes; C000146:bactoprenol; D009561:Nisin
keywords: 1,2-dioleoyl-sn-glycero-3-phospho-(1′-rac-glycerol) (sodium salt); 1,2-dioleoyl-sn-glycero-3-phosphocholine; 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide; 2-(N-morpholino)ethanesulfonic acid; 5(6)-carboxyfluorescein; AF647; Alexa Fluor 647 N-hydroxysuccinimide hydrazide; Atto 488 N-hydroxysuccinimide; Atto488-NHS; C(55)-P; C(55)-PP; CF; CLSM; Cell wall precursor; Confocal microscopy; DOPC; DOPG; EDC; GUV; Giant unilamellar vesicles; GlcNAc; LI; LII; LSM; LUV; LY; Lipid I; Lipid II; Lucifer yellow; MES; MIC; Membrane pore formation; MurNAc; N-acetylglucosamine; N-acetylmuramic acid; Nisin; SUV; TFA; UDP; confocal laser scanning microscopy; giant unilamellar vesicle; large unilamellar vesicle; laser scanning microscope; minimal inhibitory concentration; nisin coupled to Alexa Fluor 647; nisin–AF647; small unilamellar vesicle; trifluoroacetic acid; undecaprenyl-diphosphate (bactoprenol-diphosphate); undecaprenyl-phosphate (bactoprenol-phosphate); uridine diphosphate