7kdm

Publication Abstract from PubMed

Ricin toxin kills mammalian cells with notorious efficiency. The toxin's B subunit (RTB) is a Gal/GalNAc-specific lectin that attaches to cell surfaces and promotes retrograde transport of ricin's A subunit (RTA) to the trans Golgi network (TGN) and endoplasmic reticulum (ER). RTA is liberated from RTB in the ER and translocated into the cell cytoplasm, where it functions as a ribosome-inactivating protein. While antibodies against ricin's individual subunits have been reported, we now describe seven alpaca-derived, single-domain antibodies (V(H)Hs) that span the RTA-RTB interface, including four Tier 1 V(H)Hs with IC(50) values <1 nM. Crystal structures of each V(H)H bound to native ricin holotoxin revealed three different binding modes, based on contact with RTA's F-G loop (mode 1), RTB's subdomain 2gamma (mode 2) or both (mode 3). V(H)Hs in modes 2 and 3 were highly effective at blocking ricin attachment to HeLa cells and immobilized asialofetuin, due to framework residues (FR3) that occupied the 2gamma Gal/GalNAc-binding pocket and mimic ligand. The four Tier 1 V(H)Hs also interfered with intracellular functions of RTB, as they neutralized ricin in a post-attachment cytotoxicity assay (e.g., the toxin was bound to cell surfaces before antibody addition) and reduced the efficiency of toxin transport to the TGN. We conclude that the RTA-RTB interface is a target of potent toxin-neutralizing antibodies that interfere with both extracellular and intracellular events in ricin's cytotoxic pathway.

Structural Analysis of Toxin-Neutralizing, Single-Domain Antibodies that Bridge Ricin's A-B Subunit Interface.,Rudolph MJ, Poon AY, Kavaliauskiene S, Myrann AG, Reynolds-Peterson C, Davis SA, Sandvig K, Vance DJ, Mantis NJ J Mol Biol. 2021 Jul 23;433(15):167086. doi: 10.1016/j.jmb.2021.167086. Epub 2021 , Jun 3. PMID:34089718^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.