|Entry||Database: EMDB / ID: 8993|
|Title||Class III PI3K Complex 2 with inhibitor Rubicon|
|Map data||CryoEM map between 6-8 angstroms of the Class III PI3K Complex 2, bound to an inhibitor from Rubicon|
|Sample||Core scaffold of PI3K Complex 2 bound to inhibitor Rubicon:|
|Source||Homo sapiens (human)|
|Method||single particle reconstruction / cryo EM / 6.8 Å resolution|
|Authors||Young LN / Morris KL / Hurley JH|
|Citation||Journal: Mol. Cell / Year: 2019|
Title: Bidirectional Control of Autophagy by BECN1 BARA Domain Dynamics.
Authors: Chunmei Chang / Lindsey N Young / Kyle L Morris / Sören von Bülow / Johannes Schöneberg / Hitomi Yamamoto-Imoto / Yukako Oe / Kentaro Yamamoto / Shuhei Nakamura / Goran Stjepanovic / Gerhard Hummer / Tamotsu Yoshimori / James H Hurley
Abstract: Membrane targeting of the BECN1-containing class III PI 3-kinase (PI3KC3) complexes is pivotal to the regulation of autophagy. The interaction of PI3KC3 complex II and its ubiquitously expressed ...Membrane targeting of the BECN1-containing class III PI 3-kinase (PI3KC3) complexes is pivotal to the regulation of autophagy. The interaction of PI3KC3 complex II and its ubiquitously expressed inhibitor, Rubicon, was mapped to the first β sheet of the BECN1 BARA domain and the UVRAG BARA2 domain by hydrogen-deuterium exchange and cryo-EM. These data suggest that the BARA β sheet 1 unfolds to directly engage the membrane. This mechanism was confirmed using protein engineering, giant unilamellar vesicle assays, and molecular simulations. Using this mechanism, a BECN1 β sheet-1 derived peptide activates both PI3KC3 complexes I and II, while HIV-1 Nef inhibits complex II. These data reveal how BECN1 switches on and off PI3KC3 binding to membranes. The observations explain how PI3KC3 inhibition by Rubicon, activation by autophagy-inducing BECN1 peptides, and inhibition by HIV-1 Nef are mediated by the switchable ability of the BECN1 BARA domain to partially unfold and insert into membranes.
|Date||Deposition: Jul 25, 2018 / Header (metadata) release: Aug 29, 2018 / Map release: Jan 23, 2019 / Last update: Feb 6, 2019|
|Structure viewer||EM map: |
Downloads & links
|File||emd_8993.map.gz (map file in CCP4 format, 174457 KB)|
|Projections & slices|
Images are generated by Spider.
|Voxel size||X=Y=Z: 1.15 Å|
CCP4 map header:
-Entire Core scaffold of PI3K Complex 2 bound to inhibitor Rubicon
|Entire||Name: Core scaffold of PI3K Complex 2 bound to inhibitor Rubicon|
Number of components: 1
-Component #1: protein, Core scaffold of PI3K Complex 2 bound to inhibitor Rubicon
|Protein||Name: Core scaffold of PI3K Complex 2 bound to inhibitor Rubicon|
Recombinant expression: No
|Mass||Theoretical: 360 kDa|
|Source||Species: Homo sapiens (human)|
|Source (engineered)||Expression System: Homo sapiens (human) / Strain: HEK 293 GNTI|
|Specimen||Specimen state: particle / Method: cryo EM|
|Sample solution||Specimen conc.: 0.2 mg/ml / pH: 8|
|Vitrification||Instrument: FEI VITROBOT MARK IV / Cryogen name: OTHER / Temperature: 288 K / Humidity: 100 %|
-Electron microscopy imaging
Model: Titan Krios / Image courtesy: FEI Company
|Imaging||Microscope: FEI TITAN KRIOS|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 59.7 e/Å2 / Illumination mode: FLOOD BEAM|
|Lens||Imaging mode: BRIGHT FIELD|
|Specimen Holder||Model: OTHER|
|Camera||Detector: GATAN K2 SUMMIT (4k x 4k)|
-Atomic model buiding
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