Protein Structure Facilitates High-Resolution Immunological Mapping

Abstract

Select agents (SA) pose unique challenges for licensing vaccines and therapies. In the case of toxin-mediated diseases, HHS assigns guidelines for SA use, oversees vaccine and therapy development, and approves animal models and approaches to identify mechanisms for toxin neutralization. In this commentary, we discuss next-generation vaccines and therapies against ricin toxin and botulinum toxin, which are regulated SA toxins that utilize structure-based approaches for countermeasures to guide rapid response to future biothreats.KEYWORDS botulinum toxin, RTA, RVEc, RiVax, antibodies, mass spectrometry, ricin T his commentary addresses studies by Mantis and coworkers, who in this issue of Clinical and Vaccine Immunology describe the continued mapping of the neutralizing epitopes within ricin toxin (RTX) and relate these studies to current research on the botulinum neurotoxins (BoNTs) (1, 2). Our goal is to provide a perspective on how protein structure has facilitated development of vaccines and therapies against regulated select agent (SA) toxins. An overview of the properties of ricin toxin and botulinum neurotoxin is presented in Fig. 1. Ricin toxin and the botulinum neurotoxins are HHS and USDA Select Agents and Toxins (7 CFR part 331, 9 CFR part 121, and 42 CFR part 73).AB toxin structure/function. AB toxins contain modular domains with distinct functional activities (3). The A domain encodes an enzymatic activity which targets a host protein to modulate host cell physiology, often inactivating a biological process within cells, leading to cell death or occasionally enhancing the action of a cellular process. The B domain often binds a host receptor, such as a protein or glycolipid, via interactions that bind directly to a specific host receptor or that bind a carbohydrate present on a glycosylated host receptor. AB toxins are often synthesized as single-chain proteins in an inactive form and are converted to an active di-chain by proteolysis between the A and B domains, which remain connected by an interchain disulfide (4, 5).RTX is a category B bioterrorism toxigenic lectin enriched in seeds of the castor bean plant Ricinus communis. RTX is posttranslationally processed by host glycosylation and activated by cleavage between A and B domains (6, 7). The ricin toxin A domain (RTA) is a type II ribosome-inactivating protein with N-glycosidase activity, which depurinates 28S rRNA within the 60S host ribosome (8). Depurination stalls protein synthesis and induces a ribotoxic stress response (9). Ricin toxin B domain (RTB) contains at least two homologous lectin binding sites specific for galactose and binds galactose-containing lipids and glycoproteins (10). While binding of galactose results in nonproductive uptake of RTX, binding to mannose moieties on host mannose-containing receptors results in RTX retrograde trafficking to the endoplasmic reticulum, where RTA is delivered into the cytosol to target the ribosome (11). RTX exploits receptors on multiple cell types; therefore, the symptoms vary based ...

Fine-Specificity Epitope Analysis Identifies Contact Points on Ricin Toxin Recognized by Protective Monoclonal Antibodies

“…Establishing comprehensive, high-resolution B cell epitope maps ofpathogen-associated Ags and toxins provides a basis for rational vaccine design and reveals possible targets for drug candidates (24, 25). Ricin toxin’s enzymatic subunit, RTA, is of particular interest in this regard because it is the foundation for two recombinant subunit vaccines under development (i.e., RiVax and RVEc), as well as a prime target for candidate immunotherapeutics (8).…”