caa a22 4500 528785125 20190923032428.0 cr unu---uuuuu 180924e20130729xx s 000 0 eng 10.3929/ethz-b-000070973 doi 10.1371/journal.pone.0069901 doi (ETHRESEARCH)oai:www.research-collection.ethz.ch:20.500.11850/70973 Miki Tsuyoshi Outer Membrane Permeabilization Is an Essential Step in the Killing of Gram-Negative Bacteria by the Lectin RegIIIβ [Elektronische Daten] [Tsuyoshi Miki, Wolf-Dietrich; id_orcid 0000-0002-9892-6420 Hardt] PLoS ONE Open access ethresearch The C-type lectin RegIIIβ can kill certain Gram-positive and Gram-negative bacteria. The susceptibility of S. Typhimurium depends on the bacterial growth phase, i.e., bacteria from the logarithmic growth phase do bind RegIIIβ and are subsequently killed. Lipid A is one of the bacterial targets for RegIIIβ. However, at the molecular level, it is not understood how RegIIIβ interacts with and kills Gram-negative bacteria. Here, we show that RegIIIβ interacts with Gram-negative bacteria in two distinct steps. Initially, it binds to surface-exposed lipid A. The lipid A can be shielded by the O-antigen of lipopolysaccharide (LPS), as indicated by the exquisite susceptibility of wbaP mutants to RegIIIβ-mediated killing. Increased cell viability after incubation with an anti-lipid A antibody also supports this conclusion. This RegIIIβ-binding permeabilizes the outer membrane to hydrophobic dyes like Ethidium bromide or to bulky bacteriolytic enzymes like lysozyme. Conversely, compromising the outer membrane integrity by the mild detergent Triton X-100 enhances the antibacterial effect of RegIIIβ. Based on our observations, we conclude that RegIIIβ interacts with Gram-negative bacteria in two subsequent steps. Initially, it binds to the outer membrane thus leading to outer membrane permeabilization. This initial step is necessary for RegIIIβ to reach a second, still not well understood target site (presumably localized in the periplasm or the cytoplasmic membrane), thereby triggering bacterial death. This provides novel insights into the outer membrane-step of the bactericidal mechanism of RegIIIβ. Creative Commons Attribution 3.0 Unported http://creativecommons.org/licenses/by/3.0 ethresearch Hardt Wolf-Dietrich; id_orcid 0000-0002-9892-6420 joint author PLoS ONE San Francisco, CA, USA : Public Library of Science 8 (7), p. e69901 1932-6203 http://hdl.handle.net/20.500.11850/70973 text/html WWW-Backlink auf das Repository (Open access) 1 Journal Article ethresearch ETHRESEARCH 856 40 http://hdl.handle.net/20.500.11850/70973 text/html WWW-Backlink auf das Repository (Open access) ETHRESEARCH 100 1- Miki Tsuyoshi ETHRESEARCH 700 1- Hardt Wolf-Dietrich; id_orcid 0000-0002-9892-6420 joint author ETHRESEARCH 773 0- PLoS ONE San Francisco, CA, USA : Public Library of Science 8 (7), p. e69901 1932-6203 BK010053 XK010053 XK010000 ETHRESEARCH ETHRESEARCH ETHRESEARCH Journal Article Open access