Receptor interactions of protoxin and activated Vip3Aa structural conformations in Spodoptera exigua

Abstract

AbstractBACKGROUNDThe Vip3Aa insecticidal protein, produced by Bacillus thuringiensis, has been effectively used in commercial Bt‐crops to manage lepidopteran pests. Upon ingestion by larvae, the protoxin is processed by midgut proteases into the activated protein and binds specifically to its receptors in the midgut, leading to insect mortality. Cryo‐EM resolution of the trypsin‐processed Vip3Aa protein unveiled structural remodelling of the N‐terminal region during the transition from protoxin to activated protein. This conformational change has been demonstrated to be crucial for toxicity against Spodoptera exigua larvae, a major global lepidopteran pest. In this study, we investigated the relevance of the structural remodelling for the specific binding to midgut receptors.RESULTSWe conducted in vitro binding assays with radiolabelled proteins and brush border membrane vesicles (BBMV) from S. exigua, employing structural mutants that lock the protein in either its protoxin or its activated conformation. Our results indicate that both structural stages of the protein share binding sites in the midgut epithelium. Moreover, in vivo competition assays revealed that Vip3Aa is able to bind to functional receptors in S. exigua larvae both as protoxin and as activated protein.CONCLUSIONAltogether, our findings point to both structural conformations contributing to receptor binding. In vivo, either spontaneous structural shift upon proteolytic cleavage or receptor‐mediated remodelling could be occurring. However, the timing and context in which the conformational change occurs could influence membrane insertion and toxicity. Our results show the complex interplay between proteolytic processing, protein structure and receptor interactions in Vip3Aa's toxicity. © 2024 The Author(s). Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.