The ATPase Cycle of the Tail-Anchored Protein Chaperone Get3

The cytosolic ATPase Get3/TRC40 mediates targeting of tail-anchored (TA) membrane proteins to the endoplasmic reticulum (ER). Get3 functions as a molecular chaperone by binding to transmembrane domains of newly synthesized TA proteins and delivering them to a membrane docking complex in the ER. Previous work have shown that ATP binding and hydrolysis can regulate Get3s function and are essential for the targeting of TA proteins. However, the precise mechanisms by which this is accomplished remain unclear. Using a combination of mechanistic enzymology and biophysical methods, we delineate Get3s ATPase cycle and show that it is required at multiple stages during TA protein targeting. Our data complements previous structural studies and contributes to our understanding of how Get3 harnesses the energy of ATP hydrolysis to spatially and temporally coordinate the delivery of TA proteins to the ER membrane.