@article{KnappBenz2020,
  author    = {Knapp, Oliver and Benz, Roland},
  title     = {Membrane activity and channel formation of the adenylate cyclase toxin (CyaA) of Bordetella pertussis in lipid bilayer membranes},
  series = {Toxins},
  volume    = {12},
  journal   = {Toxins},
  number    = {3},
  issn      = {2072-6651},
  doi       = {10.3390/toxins12030169},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-203362},
  year      = {2020},
  abstract  = {The Gram-negative bacterium Bordetella pertussis is the cause of whooping cough. One of its pathogenicity factors is the adenylate cyclase toxin (CyaA) secreted by a Type I export system. The 1706 amino acid long CyaA (177 kDa) belongs to the continuously increasing family of repeat in toxin (RTX) toxins because it contains in its C-terminal half a high number of nine-residue tandem repeats. The protein exhibits cytotoxic and hemolytic activities that target primarily myeloid phagocytic cells expressing the αMβ2 integrin receptor (CD11b/CD18). CyaA represents an exception among RTX cytolysins because the first 400 amino acids from its N-terminal end possess a calmodulin-activated adenylate cyclase (AC) activity. The entry of the AC into target cells is not dependent on the receptor-mediated endocytosis pathway and penetrates directly across the cytoplasmic membrane of a variety of epithelial and immune effector cells. The hemolytic activity of CyaA is rather low, which may have to do with its rather low induced permeability change of target cells and its low conductance in lipid bilayer membranes. CyaA forms highly cation-selective channels in lipid bilayers that show a strong dependence on aqueous pH. The pore-forming activity of CyaA but not its single channel conductance is highly dependent on Ca\(^{2+}\) concentration with a half saturation constant of about 2 to 4 mM.},
  language  = {en}
}
@article{PollittPoulterGitzetal.2014,
  author    = {Pollitt, Alice Y. and Poulter, Natalie S. and Gitz, Eelo and Navarro-Nu{\~n}ez, Leyre and Wang, Ying-Jie and Hughes, Craig E. and Thomas, Steven G. and Nieswandt, Bernhard and Douglas, Michael R. and Owen, Dylan M. and Jackson, David G. and Dustin, Michael L. and Watson, Steve P.},
  title     = {Syk and Src Family Kinases Regulate C-type Lectin Receptor 2 (CLEC-2)-mediated Clustering of Podoplanin and Platelet Adhesion to Lymphatic Endothelial Cells*},
  series = {The Journal of Biological Chemistry},
  volume    = {289},
  journal   = {The Journal of Biological Chemistry},
  number    = {52},
  doi       = {10.1074/jbc.M114.584284},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-120770},
  pages     = {35695-710},
  year      = {2014},
  abstract  = {The interaction of CLEC-2 on platelets with Podoplanin on lymphatic endothelial cells initiates platelet signalling events that are necessary for prevention of blood-lymph mixing during development. In the present study, we show that CLEC-2 signalling via Src family and Syk tyrosine kinases promotes platelet adhesion to primary mouse lymphatic endothelial cells at low shear. Using supported lipid bilayers containing mobile Podoplanin, we further show that activation of Src and Syk in platelets promotes clustering of CLEC-2 and Podoplanin. Clusters of CLEC-2-bound Podoplanin migrate rapidly to the centre of the platelet to form a single structure. Fluorescence life-time imaging demonstrates that molecules within these clusters are within 10 nm of one another and that the clusters are disrupted by inhibition of Src and Syk family kinases. CLEC-2 clusters are also seen in platelets adhered to immobilised Podoplanin using direct stochastic optical reconstruction microscopy (dSTORM). These findings provide mechanistic insight by which CLEC-2 signalling promotes adhesion to Podoplanin and regulation of Podoplanin signalling thereby contributing to lymphatic vasculature development.},
  language  = {en}
}
@article{BenzMaierBaueretal.2014,
  author    = {Benz, Roland and Maier, Elke and Bauer, Susanne and Ludwig, Albrecht},
  title     = {The Deletion of Several Amino Acid Stretches of Escherichia coli Alpha-Hemolysin (HlyA) Suggests That the Channel-Forming Domain Contains Beta-Strands},
  series = {PLOS ONE},
  volume    = {9},
  journal   = {PLOS ONE},
  number    = {12},
  issn      = {1932-6203},
  doi       = {10.1371/journal.pone.0112248},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-118115},
  pages     = {e112248},
  year      = {2014},
  abstract  = {Escherichia coli α-hemolysin (HlyA) is a pore-forming protein of 110 kDa belonging to the family of RTX toxins. A hydrophobic region between the amino acid residues 238 and 410 in the N-terminal half of HlyA has previously been suggested to form hydrophobic and/or amphipathic α-helices and has been shown to be important for hemolytic activity and pore formation in biological and artificial membranes. The structure of the HlyA transmembrane channel is, however, largely unknown. For further investigation of the channel structure, we deleted in HlyA different stretches of amino acids that could form amphipathic β-strands according to secondary structure predictions (residues 71-110, 158-167, 180-203, and 264-286). These deletions resulted in HlyA mutants with strongly reduced hemolytic activity. Lipid bilayer measurements demonstrated that HlyAΔ71-110 and HlyAΔ264-286 formed channels with much smaller single-channel conductance than wildtype HlyA, whereas their channel-forming activity was virtually as high as that of the wildtype toxin. HlyAΔ158-167 and HlyAΔ180-203 were unable to form defined channels in lipid bilayers. Calculations based on the single-channel data indicated that the channels generated by HlyAΔ71-110 and HlyAΔ264-286 had a smaller size (diameter about 1.4 to 1.8 nm) than wildtype HlyA channels (diameter about 2.0 to 2.6 nm), suggesting that in these mutants part of the channel-forming domain was removed. Osmotic protection experiments with erythrocytes confirmed that HlyA, HlyAΔ71-110, and HlyAΔ264-286 form defined transmembrane pores and suggested channel diameters that largely agreed with those estimated from the single-channel data. Taken together, these results suggest that the channel-forming domain of HlyA might contain β-strands, possibly in addition to α-helical structures.},
  language  = {en}
}
@article{BeitzingerBronnhuberDuschaetal.2013,
  author    = {Beitzinger, Christoph and Bronnhuber, Annika and Duscha, Kerstin and Riedl, Zsuzsanna and Huber-Lang, Markus and Benz, Roland and Hajos, Gy{\"o}rgy and Barth, Holger},
  title     = {Designed Azolopyridinium Salts Block Protective Antigen Pores In Vitro and Protect Cells from Anthrax Toxin},
  series = {PLoS ONE},
  volume    = {8},
  journal   = {PLoS ONE},
  number    = {6},
  doi       = {10.1371/journal.pone.0066099},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-130097},
  pages     = {e66099},
  year      = {2013},
  abstract  = {Background Several intracellular acting bacterial protein toxins of the AB-type, which are known to enter cells by endocytosis, are shown to produce channels. This holds true for protective antigen (PA), the binding component of the tripartite anthrax-toxin of Bacillus anthracis. Evidence has been presented that translocation of the enzymatic components of anthrax-toxin across the endosomal membrane of target cells and channel formation by the heptameric/octameric \(PA_{63}\) binding/translocation component are related phenomena. Chloroquine and some 4-aminoquinolones, known as potent drugs against Plasmodium falciparium infection of humans, block efficiently the \(PA_{63}\)-channel in a dose dependent way. Methodology/Principal Findings Here we demonstrate that related positively charged heterocyclic azolopyridinium salts block the \(PA_{63}\)-channel in the µM range, when both, inhibitor and \(PA_{63}\) are added to the same side of the membrane, the cis-side, which corresponds to the lumen of acidified endosomal vesicles of target cells. Noise-analysis allowed the study of the kinetics of the plug formation by the heterocycles. In vivo experiments using J774A.1 macrophages demonstrated that the inhibitors of \(PA_{63}\)-channel function also efficiently block intoxication of the cells by the combination lethal factor and \(PA_{63}\) in the same concentration range as they block the channels in vitro. Conclusions/Significance These results strongly argue in favor of a transport of lethal factor through the \(PA_{63}\)-channel and suggest that the heterocycles used in this study could represent attractive candidates for development of novel therapeutic strategies against anthrax.},
  language  = {en}
}