1 H N , 13 C, and 15 N resonance assignments of the Clostridioides difficile receptor binding domain 2 (CDTb, residues 7
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ARTICLE
1 N 13
H , C, and 15N resonance assignments of the Clostridioides difficile receptor binding domain 2 (CDTb, residues 757–876) Mary E. Cook1 · Kristen M. Varney1 · Raquel Godoy‑Ruiz1 · David J. Weber1 Received: 30 July 2020 / Accepted: 18 September 2020 © The Author(s) 2020
Abstract Clostridioides difficile is a bacterial pathogen responsible for the majority of nosocomial infections in the developed world. C. difficile infection (CDI) is difficult to treat in many cases because hypervirulent strains have evolved that contain a third toxin, termed the C. difficile toxin (CDT), in addition to the two enterotoxins TcdA and TcdB. CDT is a binary toxin comprised of an enzymatic, ADP-ribosyltransferase (ART) toxin component, CDTa, and a pore-forming or delivery subunit, CDTb. In the absence of CDTa, CDTb assembles into two distinct di-heptameric states, a symmetric and an asymmetric form with both states having two surface-accessible host cell receptor-binding domains, termed RBD1 and RBD2. RBD1 has a unique amino acid sequence, when aligned to other well-studied binary toxins (i.e., anthrax), and it contains a novel Ca2+-binding site important for CDTb stability. The other receptor binding domain, RBD2, is critically important for CDT toxicity, and a domain such as this is missing altogether in other binary toxins and shows further that CDT is unique when compared to other binary toxins. In this study, the 1H, 13C, and 15N backbone and sidechain resonances of the 120 amino acid RBD2 domain of CDTb (residues 757–876) were assigned sequence-specifically and provide a framework for future NMR-based drug discovery studies directed towards targeting the most virulent strains of CDI. Keywords Clostridioides difficile · CDI · CDTb · Binary toxin
Biological context Clostridioides difficile infection (CDI) is caused by a sporeforming, Gram-positive bacterium, and it is the most commonly reported nosocomial infection in the world, accounting for 12% of all hospital-borne infections (Gerding 2015). Prior to the emergence of hypervirulent strains early in the twenty-first century, C. difficile strains produced only two large enterotoxins, TcdA and TcdB, which inhibit signaling pathways by glucosylating small GTPases. Whereas, hypervirulent strains emerging more recently, such as the NAP1 epidemic strain, encode TcdA/TcdB plus additional virulence factors, most notably a third toxin termed the C. difficile toxin (CDT) (Perelle et al. 1997). CDT is a binary toxin that kills host cells by covalent modification of * David J. Weber [email protected] 1
Department of Biochemistry and Molecular Biology, Center for Biomolecular Therapeutics (CBT), University of Maryland School of Medicine, 108 N. Greene St., Baltimore, MD 21201, USA
essential intracellular regulators of host cell function, including G-actin. While drug options are becoming available to target the large clostridial toxins in CDI, TcdA/TcdB (Yang et al. 2015), there is nothing approved by the FDA to target CDT (Secore et al. 2017). To address this
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