title: Inhibition of Bacterial Toxin Activity by the Nuclear Stain, DRAQ5™.
abstract: The repeats-in-toxin family of toxins includes proteins produced by Gram negative bacteria such as Escherichia coli (α-hemolysin), Bordetella pertussis (adenylate cyclase toxin), and Aggregatibacter actinomycetemcomitans (LtxA), which contribute to the pathogenesis of these organisms by killing host cells. In the case of LtxA produced by A. actinomycetemcomitans, white blood cells are targeted, allowing the bacteria to avoid clearance by the host immune system. In its association with target cells, LtxA binds to a receptor, lymphocyte function-associated antigen-1, as well as membrane lipids and cholesterol, before being internalized via a lysosomal-mediated pathway. The motivation for this project comes from our discovery that DRAQ5™, a membrane-permeable nuclear stain, prevents the internalization of LtxA in a Jurkat T cell line. We hypothesized that DRAQ5™, in crossing the plasma membrane, alters the properties of the membrane to inhibit LtxA internalization. To investigate how DRAQ5™ interacts with the lipid membrane to prevent LtxA internalization, we used studied DRAQ5™-mediated membrane changes in model membranes using a variety of techniques, including differential scanning calorimetry and fluorescence spectroscopy. Our results suggest that DRAQ5™ inhibits the activity of LtxA by decreasing the fluidity of the cellular lipid membrane, which decreases LtxA binding. These results present an interesting possible anti-virulence strategy; by altering bacterial toxin activity by modifying membrane fluidity, it may be possible to inhibit the pathogenicity of A. actinomycetemcomitans.
authors.affiliation: Program in Bioengineering, Lehigh University, Bethlehem, PA, 18015, USA.
authors.forename: Joshua N
authors.initials: JN
authors.lastname: Webb
authors.affiliation: Department of Chemical and Biomolecular Engineering, Lehigh University, B323 Iacocca Hall, 111 Research Drive, Bethlehem, PA, 18015, USA.
authors.forename: Evan
authors.initials: E
authors.lastname: Koufos
authors.affiliation: Department of Chemical and Biomolecular Engineering, Lehigh University, B323 Iacocca Hall, 111 Research Drive, Bethlehem, PA, 18015, USA. acb313@lehigh.edu.
authors.forename: Angela C
authors.initials: AC
authors.lastname: Brown
mesh_terms: D000880:Anthraquinones / Q000378:metabolism / Q000494:pharmacology*; D000992:Antitoxins / Q000378:metabolism / Q000494:pharmacology*; D001427:Bacterial Toxins / Q000037:antagonists & inhibitors* / Q000378:metabolism* / Q000633:toxicity; D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D002463:Cell Membrane Permeability / Q000187:drug effects; D005098:Exotoxins / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000633:toxicity; D006801:Humans; D008560:Membrane Fluidity / Q000187:drug effects; D013696:Temperature
chemical_list: C520537:1,5-bis((2-(methylamino)ethyl)amino)-4,8-dihydroxyanthracene-9,10-dione; D000880:Anthraquinones; D000992:Antitoxins; D001427:Bacterial Toxins; D005098:Exotoxins; C018579:leukotoxin
keywords: Anthracycline; Bacterial toxin; Membrane fluidity; Virulence