Mechanism of Translocation by the Bacterial ATPase SecA

Abstract

Fully synthesized bacterial secretory proteins are transported across the plasma membrane post-translationally by the ATPase SecA. SecA is a member of the large AAA ATPase family and uses the energy from ATP hydrolysis to push nascent polypeptides through the SecY channel. However, the mechanism by which SecA couples its energy consumption to this translocation activity is unclear. Here, we use single molecule FRET to observe the conformational changes of SecA as it translocates a model polypeptide substrate and ensemble biochemistry experiments, including protease protection assays, to monitor the corresponding movements of the substrate itself. Upon ATP-binding, SecA’s two-helix finger inserts into the SecY channel and pushes the translocating polypeptide chain forward. Subsequently, a clamp domain closes around the substrate and holds it in place as the two-helix finger resets during ATP hydrolysis. The clamp domain re-opens after phosphate release, freeing the polypeptide to passively slide through the SecY channel until the subsequent ATP binding event. The alternating movements of these two domains ensure that translocation is directional and provide a model that may be applicable to a wide range of ATPases that work on polypeptide chains.