Journal: Elife / Year: 2019 Title: Structural basis of Ca-dependent activation and lipid transport by a TMEM16 scramblase. Authors: Maria E Falzone / Jan Rheinberger / Byoung-Cheol Lee / Thasin Peyear / Linda Sasset / Ashleigh M Raczkowski / Edward T Eng / Annarita Di Lorenzo / Olaf S Andersen / Crina M Nimigean / Alessio Accardi Abstract: The lipid distribution of plasma membranes of eukaryotic cells is asymmetric and phospholipid scramblases disrupt this asymmetry by mediating the rapid, nonselective transport of lipids down their ...The lipid distribution of plasma membranes of eukaryotic cells is asymmetric and phospholipid scramblases disrupt this asymmetry by mediating the rapid, nonselective transport of lipids down their concentration gradients. As a result, phosphatidylserine is exposed to the outer leaflet of membrane, an important step in extracellular signaling networks controlling processes such as apoptosis, blood coagulation, membrane fusion and repair. Several TMEM16 family members have been identified as Ca-activated scramblases, but the mechanisms underlying their Ca-dependent gating and their effects on the surrounding lipid bilayer remain poorly understood. Here, we describe three high-resolution cryo-electron microscopy structures of a fungal scramblase from , afTMEM16, reconstituted in lipid nanodiscs. These structures reveal that Ca-dependent activation of the scramblase entails global rearrangement of the transmembrane and cytosolic domains. These structures, together with functional experiments, suggest that activation of the protein thins the membrane near the transport pathway to facilitate rapid transbilayer lipid movement.
Conc.: 7 mg/ml Details: afTMEM16 reconstituted in nanodiscs in the absence of Ca2+ Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Vitrification
Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 288 kelvins
-
Electron microscopy imaging
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
Microscopy
Microscope model: FEI TITAN KRIOS
Electron gun
Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: OTHER
Electron lens
Mode: BRIGHT FIELDBright-field microscopy
Image recording
Electron dose: 62.61 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Number of grids imaged: 1
-
Processing
Software
Name
Version
Classification
phenix.real_space_refine
1.13_2998
refinement
PHENIX
1.13_2998
refinement
EM software
ID
Name
Version
Category
2
Leginon
imageacquisition
4
CTFFIND
4
CTFcorrection
9
PHENIX
modelrefinement
CTF correction
Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Symmetry
Point symmetry: C2
3D reconstruction
Resolution: 3.89 Å / Resolution method: FSC 0.143 CUT-OFF / Number of particles: 70356 / Symmetry type: POINT
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