Visualising the membrane insertion mechanism of the bio-insecticide Vip3Aa by cryo-ET

Vip3Aa is an important insecticidal protein used in agriculture as both a biopesticide and a transgenic crop trait for control of lepidopteran pests. After ingestion by larvae, Vip3Aa acts in the midgut by binding to specific receptors on brush border epithelial cell membranes, where it inserts into and permeabilises the membrane. However, its membrane-inserted conformation and the details of the membrane association, insertion and permeabilization process remain unclear. Here, we use cryo-EM and cryo-ET to visualise Vip3Aa inserted into synthetic liposomes and into enriched preparations of brush border membrane vesicles for the first time. Our structures demonstrate that Vip3Aa retains a consistent overall conformation in both synthetic and native membranes. These structures provide insights into the interaction between the membrane and the Vip3Aa N-terminal region, and how Vip3Aa orients itself within the bilayer. These results further our understanding of Vip3Aa’s mechanism of action, contributing to the knowledge required to develop more powerful Vip3Aa-based crop defences.