The Role of Multivalency in Inhibition of Bacillus Anthracis and Clostridium Botulinum Binary Toxins by Cationic PAMAM Dendrimers

Abstract

Multivalent interactions between oligomeric receptors and multivalent ligands not only play an important role in many biochemical processes, but are also investigated and employed for rational drug design and development. Multivalent ligands often possess a high affinity for multivalent binding sites, significantly greater than that of a single functional group interacting with a single binding site. A number of protein toxins, including binary bacterial toxins, have recently been successfully neutralized by novel synthetic multivalent blockers.This study explores the potentially universal antitoxin activity of dendrimers, which are the repeatedly branched polymers with all bonds emanating from a central core. Dendrimers are unique highly branched macromolecules, where each consecutive growth step represents a new dendrimer “generation” with an increased diameter and double the number of reactive surface functional groups. We use several generations of polyamido amine (PAMAM) intact dendrimers and their imperfect forms, known as dendrons, to investigate their blocking activity against the pore-forming components of two binary bacterial exotoxins, Clostridium Botulinum C2 and Bacillus anthracis anthrax (PA63 and C2IIa, respectively). While aiming for different cytosolic targets, these binary toxins rely on similar cellular uptake mechanisms that include formation of oligomeric cation selective pores under acidic endosomal conditions. These pores are essential for delivery of enzymatic components of the internalized toxins from endosomes into the cytosol of target cells. By single channel reconstitution and high-resolution recording in planar bilayer lipid membranes, we show that PAMAM dendrimers obstruct PA63 and C2IIa at nM concentrations. We also strive to understand the role of attractive interaction between the dendrimer/dendron blockers and the ion channel molecules.