biostudies-literatureUnknownSuresh SIntramural NIH HHSThe lipid kinase phosphatidylinositol 4 kinase III alpha (PI4KIIIa/PI4KA) is a master regulator of the lipid composition and asymmetry of the plasma membrane. PI4KA exists primarily in a heterotrimeric complex with its regulatory proteins TTC7 and FAM126. Fundamental to PI4KA activity is its targeted recruitment to the plasma membrane by the lipidated proteins EFR3A and EFR3B. Here, we report a cryo-EM structure of the C-terminus of EFR3A bound to the PI4KA-TTC7B-FAM126A complex, with extensive validation using both hydrogen deuterium exchange mass spectrometry (HDX-MS), and mutational analysis. The EFR3A C-terminus undergoes a disorder-order transition upon binding to the PI4KA complex, with an unexpected direct interaction with both TTC7B and FAM126A. Complex disrupting mutations in TTC7bioRxiv : the preprint server for biology2024.04.30.587787https://www.ebi.ac.uk/biostudies/studies/S-EPMC11092606biostudies-literatureMolecular basis for plasma membrane recruitment of PI4KA by EFR3.PMC11092606Z01 HD000196Pemberton JGYip CKScott MKBalla TBurke JEParson MARohilla PAlvarez-Prats ASuresh SHarris NJShaw ALJenkins MLfalseMolecular basis for plasma membrane recruitment of PI4KA by EFR3.The lipid kinase phosphatidylinositol 4 kinase III alpha (PI4KIIIa/PI4KA) is a master regulator of the lipid composition and asymmetry of the plasma membrane. PI4KA exists primarily in a heterotrimeric complex with its regulatory proteins TTC7 and FAM126. Fundamental to PI4KA activity is its targeted recruitment to the plasma membrane by the lipidated proteins EFR3A and EFR3B. Here, we report a cryo-EM structure of the C-terminus of EFR3A bound to the PI4KA-TTC7B-FAM126A complex, with extensive validation using both hydrogen deuterium exchange mass spectrometry (HDX-MS), and mutational analysis. The EFR3A C-terminus undergoes a disorder-order transition upon binding to the PI4KA complex, with an unexpected direct interaction with both TTC7B and FAM126A. Complex disrupting mutations in TTC72024-01-01T00:00:00Z2024 Jul2026-07-01T03:15:50.708Z2025-08-28T03:09:05.779ZS-EPMC110926063874645310.1101/2024.04.30.587787