+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-20221 | |||||||||
---|---|---|---|---|---|---|---|---|---|---|
Title | nhTMEM16 L302A +Ca2+ in nanodiscs | |||||||||
Map data | Final EM map for nhTMEM16 L302A/nanodisc complex with C2 symmetry. Used for model buidling. | |||||||||
Sample |
| |||||||||
Keywords | TMEM16 / scramblase / lipid transport / anoactamin | |||||||||
Function / homology | Function and homology information cortical endoplasmic reticulum / chloride channel activity / identical protein binding / metal ion binding / plasma membrane Similarity search - Function | |||||||||
Biological species | Nectria haematococca mpVI (fungus) / Nectria haematococca (strain 77-13-4 / ATCC MYA-4622 / FGSC 9596 / MPVI) (fungus) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 4.0 Å | |||||||||
Authors | Falzone M / Lee BC | |||||||||
Funding support | United States, 1 items
| |||||||||
Citation | Journal: Nat Commun / Year: 2019 Title: Dynamic modulation of the lipid translocation groove generates a conductive ion channel in Ca-bound nhTMEM16. Authors: George Khelashvili / Maria E Falzone / Xiaolu Cheng / Byoung-Cheol Lee / Alessio Accardi / Harel Weinstein / Abstract: Both lipid and ion translocation by Ca-regulated TMEM16 transmembrane proteins utilizes a membrane-exposed hydrophilic groove. Several conformations of the groove are observed in TMEM16 protein ...Both lipid and ion translocation by Ca-regulated TMEM16 transmembrane proteins utilizes a membrane-exposed hydrophilic groove. Several conformations of the groove are observed in TMEM16 protein structures, but how these conformations form, and what functions they support, remains unknown. From analyses of atomistic molecular dynamics simulations of Ca-bound nhTMEM16 we find that the mechanism of a conformational transition of the groove from membrane-exposed to occluded from the membrane involves the repositioning of transmembrane helix 4 (TM4) following its disengagement from a TM3/TM4 interaction interface. Residue L302 is a key element in the hydrophobic TM3/TM4 interaction patch that braces the open-groove conformation, which should be changed by an L302A mutation. The structure of the L302A mutant determined by cryogenic electron microscopy (cryo-EM) reveals a partially closed groove that could translocate ions, but not lipids. This is corroborated with functional assays showing severely impaired lipid scrambling, but robust channel activity by L302A. | |||||||||
History |
|
-Structure visualization
Movie |
Movie viewer |
---|---|
Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_20221.map.gz | 58 MB | EMDB map data format | |
---|---|---|---|---|
Header (meta data) | emd-20221-v30.xml emd-20221.xml | 15.5 KB 15.5 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_20221_fsc.xml | 9.2 KB | Display | FSC data file |
Images | emd_20221.png | 224.1 KB | ||
Filedesc metadata | emd-20221.cif.gz | 5.9 KB | ||
Others | emd_20221_additional_1.map.gz emd_20221_additional_2.map.gz | 59.8 MB 55.3 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-20221 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-20221 | HTTPS FTP |
-Related structure data
Related structure data | 6oy3MC M: atomic model generated by this map C: citing same article (ref.) |
---|---|
Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
---|
-Map
File | Download / File: emd_20221.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | Final EM map for nhTMEM16 L302A/nanodisc complex with C2 symmetry. Used for model buidling. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.0961 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
|
-Supplemental data
-Additional map: Final EM map for nhTMEM16 L302A/nanodisc complex with...
File | emd_20221_additional_1.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | Final EM map for nhTMEM16 L302A/nanodisc complex with C1 symmetry. Used to analyze effects of nhTMEM16 | ||||||||||||
Projections & Slices |
| ||||||||||||
Density Histograms |
-Additional map: Unfiltered map from refinement of the nhTMEM16 L302A/nanodisc....
File | emd_20221_additional_2.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | Unfiltered map from refinement of the nhTMEM16 L302A/nanodisc. Used to assist model building. | ||||||||||||
Projections & Slices |
| ||||||||||||
Density Histograms |
-Sample components
-Entire : nhTMEM16 L302A reconstituted in nanodiscs in the presence of Ca2+
Entire | Name: nhTMEM16 L302A reconstituted in nanodiscs in the presence of Ca2+ |
---|---|
Components |
|
-Supramolecule #1: nhTMEM16 L302A reconstituted in nanodiscs in the presence of Ca2+
Supramolecule | Name: nhTMEM16 L302A reconstituted in nanodiscs in the presence of Ca2+ type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 |
---|---|
Source (natural) | Organism: Nectria haematococca mpVI (fungus) |
Molecular weight | Theoretical: 166 kDa/nm |
-Macromolecule #1: nhTMEM16
Macromolecule | Name: nhTMEM16 / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO |
---|---|
Source (natural) | Organism: Nectria haematococca (strain 77-13-4 / ATCC MYA-4622 / FGSC 9596 / MPVI) (fungus) Strain: 77-13-4 / ATCC MYA-4622 / FGSC 9596 / MPVI |
Molecular weight | Theoretical: 83.15793 KDa |
Recombinant expression | Organism: Saccharomyces cerevisiae (brewer's yeast) |
Sequence | String: MSNLKDFSQP GSGQESNFGV DFVIHYKVPA AERDEAEAGF VQLIRALTTV GLATEVRHGE NESLLVFVKV ASPDLFAKQV YRARLGDWL HGVRVSAPHN DIAQALQDEP VVEAERLRLI YLMITKPHNE GGAGVTPTNA KWKHVESIFP LHSHSFNKEW I KKWSSKYT ...String: MSNLKDFSQP GSGQESNFGV DFVIHYKVPA AERDEAEAGF VQLIRALTTV GLATEVRHGE NESLLVFVKV ASPDLFAKQV YRARLGDWL HGVRVSAPHN DIAQALQDEP VVEAERLRLI YLMITKPHNE GGAGVTPTNA KWKHVESIFP LHSHSFNKEW I KKWSSKYT LEQTDIDNIR DKFGESVAFY FAFLRSYFRF LVIPSAFGFG AWLLLGQFSY LYALLCGLWS VVFFEYWKKQ EV DLAVQWG VRGVSSIQQS RPEFEWEHEA EDPITGEPVK VYPPMKRVKT QLLQIPFALA CVVAAGALIV TCNSLEVFIN EVY SGPGKQ YLGFLPTIFL VIGTPTISGV LMGAAEKLNA MENYATVDAH DAALIQKQFV LNFMTSYMAL FFTAFVYIPF GHIL HPFLN FWRATAQTLT FSEKELPTRE FQINPARISN QMFYFTVTAQ IVNFATEVVV PYIKQQAFQK AKQLKSGSKV QEDHE EEAE FLQRVREECT LEEYDVSGDY REMVMQFGYV AMFSVAWPLA ACCFLVNNWV ELRSDALKIA ISSRRPIPWR TDSIGP WLT ALSFLSWLGS ITSSAIVYLC SNSKNGTQGE ASPLKAWGLL LSILFAEHFY LVVQLAVRFV LSKLDSPGLQ KERKERF QT KKRLLQENLG QDAAEEAAAP GIEHSEKITR EALEEEARQA SIRGHGTPEE MFWQRQRGMQ ETIEIGRRMI EQQLAAGK N GKKSAPAVPS EKAS UniProtKB: Plasma membrane channel protein |
-Macromolecule #2: CALCIUM ION
Macromolecule | Name: CALCIUM ION / type: ligand / ID: 2 / Number of copies: 4 / Formula: CA |
---|---|
Molecular weight | Theoretical: 40.078 Da |
-Experimental details
-Structure determination
Method | cryo EM |
---|---|
Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 6.5 mg/mL |
---|---|
Buffer | pH: 8 |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 288 K / Instrument: FEI VITROBOT MARK IV |
Details | nhTMEM16 L302A reconstituted in nanodiscs in the absence of Ca2+ |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
---|---|
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Average electron dose: 70.7 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: OTHER / Imaging mode: BRIGHT FIELD |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |