Cryo-EM structures capturing the entire transport cycle of the P4-ATPase flippase

Abstract

In eukaryotic membranes, P4-ATPases mediate the translocation of phospholipids from the outer to inner leaflet and maintain lipid asymmetry, which is critical for protein trafficking and signaling pathways. Here we report the cryo-EM structures of six distinct intermediates of the human ATP8A1-CDC50a hetero-complex, at 2.6-3.3 [A] resolutions, revealing the entire lipid translocation cycle of this P4-ATPase. ATP-dependent phosphorylation induces a large rotational movement of the actuator domain around the phosphorylation site, accompanied by lateral shifts of the first and second transmembrane helices, thereby allowing phosphatidylserine binding. The phospholipid head group passes through the hydrophilic cleft, while the acyl chain is exposed toward the lipid environment. These findings advance our understanding of the flippase mechanism and the disease-associated mutants of P4-ATPases.\n\nOne Sentence SummaryCryo-EM reveals lipid translocation by P4-type flippase.