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 15 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 Å 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 20 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.One Sentence Summary: Cryo-EM reveals lipid translocation by P4-type flippase.Main Text:

Transporters Beam Signals from Cell Membranes

“…The beta subunit is generally composed of two transmembrane helices, where there is evidence showing a shared cholesterol-binding site with the alpha subunit. [38]. Recent cryo-EM studies have provided a structural glimpse on the overall archi-tecture of P4-ATPases [38][39][40][41].…”

“…In the case of Drs2p or ATP8A, while orthologs to each other, they have distinct regulatory domains at the C-termini. In yeast Drs2p, the C-terminus has an autoinhibitory effect on ATPase activity; 4 however, the human ATP8A2 mediates a regulation mode where the regulatory domain keeps the N and A domains apart in the E2P state (Fig 1C) [38,43]. Such differential regulation may be attributed to the binding of different beta subunits.…”

Physiological and Genetic Response Characteristics of Stenotrophomonas Rhizophila JC1 Under Heavy Metal Stress

“…The P-type ATPase, located on the plasma membrane, promotes the metal e ux system through hydrolysis (Arguello et al 2011). It transfers substrates from the outside of the cell or the peripheral cytoplasm to the cytoplasm, as well as from the cytoplasm to the extracellular or peripheral cytoplasm through P-type ATPases (Hiraizumi et al 2019). Simultaneously, owing to the stimulating effects of sulfhydryl compounds on the metal e ux activity, CPx-type ATPases may also use glutathione to expel metals from the cytoplasm (Meng et al 2015).…”