[English] 日本語
Yorodumi- EMDB-27701: Focused map (monomer A) for Arabidopsis SPY in complex with GDP-fucose -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-27701 | |||||||||
---|---|---|---|---|---|---|---|---|---|---|
Title | Focused map (monomer A) for Arabidopsis SPY in complex with GDP-fucose | |||||||||
Map data | ||||||||||
Sample |
| |||||||||
Keywords | O-Fucosyltransferase / PLANT PROTEIN | |||||||||
Biological species | Arabidopsis thaliana (thale cress) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.1 Å | |||||||||
Authors | Kumar S / Zhou Y / Dillard L / Borgnia MJ / Bartesaghi A / Zhou P | |||||||||
Funding support | United States, 1 items
| |||||||||
Citation | Journal: Nat Commun / Year: 2023 Title: Structure and dynamics of the Arabidopsis O-fucosyltransferase SPINDLY. Authors: Shivesh Kumar / Yan Wang / Ye Zhou / Lucas Dillard / Fay-Wei Li / Carly A Sciandra / Ning Sui / Rodolfo Zentella / Emily Zahn / Jeffrey Shabanowitz / Donald F Hunt / Mario J Borgnia / ...Authors: Shivesh Kumar / Yan Wang / Ye Zhou / Lucas Dillard / Fay-Wei Li / Carly A Sciandra / Ning Sui / Rodolfo Zentella / Emily Zahn / Jeffrey Shabanowitz / Donald F Hunt / Mario J Borgnia / Alberto Bartesaghi / Tai-Ping Sun / Pei Zhou / Abstract: SPINDLY (SPY) in Arabidopsis thaliana is a novel nucleocytoplasmic protein O-fucosyltransferase (POFUT), which regulates diverse developmental processes. Sequence analysis indicates that SPY is ...SPINDLY (SPY) in Arabidopsis thaliana is a novel nucleocytoplasmic protein O-fucosyltransferase (POFUT), which regulates diverse developmental processes. Sequence analysis indicates that SPY is distinct from ER-localized POFUTs and contains N-terminal tetratricopeptide repeats (TPRs) and a C-terminal catalytic domain resembling the O-linked-N-acetylglucosamine (GlcNAc) transferases (OGTs). However, the structural feature that determines the distinct enzymatic selectivity of SPY remains unknown. Here we report the cryo-electron microscopy (cryo-EM) structure of SPY and its complex with GDP-fucose, revealing distinct active-site features enabling GDP-fucose instead of UDP-GlcNAc binding. SPY forms an antiparallel dimer instead of the X-shaped dimer in human OGT, and its catalytic domain interconverts among multiple conformations. Analysis of mass spectrometry, co-IP, fucosylation activity, and cryo-EM data further demonstrates that the N-terminal disordered peptide in SPY contains trans auto-fucosylation sites and inhibits the POFUT activity, whereas TPRs 1-5 dynamically regulate SPY activity by interfering with protein substrate binding. | |||||||||
History |
|
-Structure visualization
Supplemental images |
---|
-Downloads & links
-EMDB archive
Map data | emd_27701.map.gz | 118 MB | EMDB map data format | |
---|---|---|---|---|
Header (meta data) | emd-27701-v30.xml emd-27701.xml | 17.3 KB 17.3 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_27701_fsc.xml | 11.3 KB | Display | FSC data file |
Images | emd_27701.png | 49.3 KB | ||
Masks | emd_27701_msk_1.map | 125 MB | Mask map | |
Others | emd_27701_half_map_1.map.gz emd_27701_half_map_2.map.gz | 115.9 MB 115.9 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-27701 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-27701 | HTTPS FTP |
-Validation report
Summary document | emd_27701_validation.pdf.gz | 759.5 KB | Display | EMDB validaton report |
---|---|---|---|---|
Full document | emd_27701_full_validation.pdf.gz | 759.1 KB | Display | |
Data in XML | emd_27701_validation.xml.gz | 18.1 KB | Display | |
Data in CIF | emd_27701_validation.cif.gz | 23.9 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-27701 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-27701 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
---|
-Map
File | Download / File: emd_27701.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Voxel size | X=Y=Z: 1.08 Å | ||||||||||||||||||||
Density |
| ||||||||||||||||||||
Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
|
-Supplemental data
-Mask #1
File | emd_27701_msk_1.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Projections & Slices |
| ||||||||||||
Density Histograms |
-Half map: #2
File | emd_27701_half_map_1.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Projections & Slices |
| ||||||||||||
Density Histograms |
-Half map: #1
File | emd_27701_half_map_2.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Projections & Slices |
| ||||||||||||
Density Histograms |
-Sample components
-Entire : SPY GDP-fucose complex
Entire | Name: SPY GDP-fucose complex |
---|---|
Components |
|
-Supramolecule #1: SPY GDP-fucose complex
Supramolecule | Name: SPY GDP-fucose complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
---|---|
Source (natural) | Organism: Arabidopsis thaliana (thale cress) |
-Macromolecule #1: SPY GDP-fucose complex
Macromolecule | Name: SPY GDP-fucose complex / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO |
---|---|
Source (natural) | Organism: Arabidopsis thaliana (thale cress) |
Sequence | String: MVGLEDDTER ERSPVVENGF SNGSRSSSSS AGVLSPSRKV TQGNDTLSYA NILRARNKFA DALALYEAML EKDSKNVEAH IGKGICLQTQ NKGNLAFDCF SEAIRLDPHN ACALTHCGIL HKEEGRLVEA AESYQKALMA DASYKPAAEC LAIVLTDLGT SLKLAGNTQE ...String: MVGLEDDTER ERSPVVENGF SNGSRSSSSS AGVLSPSRKV TQGNDTLSYA NILRARNKFA DALALYEAML EKDSKNVEAH IGKGICLQTQ NKGNLAFDCF SEAIRLDPHN ACALTHCGIL HKEEGRLVEA AESYQKALMA DASYKPAAEC LAIVLTDLGT SLKLAGNTQE GIQKYYEALK IDPHYAPAYY NLGVVYSEMM QYDNALSCYE KAALERPMYA EAYCNMGVIY KNRGDLEMAI TCYERCLAVS PNFEIAKNNM AIALTDLGTK VKLEGDVTQG VAYYKKALYY NWHYADAMYN LGVAYGEMLK FDMAIVFYEL AFHFNPHCAE ACNNLGVLYK DRDNLDKAVE CYQMALSIKP NFAQSLNNLG VVYTVQGKMD AAASMIEKAI LANPTYAEAF NNLGVLYRDA GNITMAIDAY EECLKIDPDS RNAGQNRLLA MNYINEGLDD KLFEAHRDWG WRFTRLHPQY TSWDNLKDPE RPITIGYISP DFFTHSVSYF IEAPLTHHDY TKYKVVVYSA VVKADAKTYR FRDKVLKKGG VWKDIYGIDE KKIASMVRED KIDILVELTG HTANNKLGTM ACRPAPVQVT WIGYPNTTGL PTVDYRITDS LADPPDTKQK QVEELVRLPD CFLCYTPSPE AGPVCPTPAL SNGFVTFGSF NNLAKITPKV LQVWARILCA VPNSRLVVKC KPFCCDSIRQ RFLTTLEQLG LESKRVDLLP LILFNHDHMQ AYSLMDISLD TFPYAGTTTT CESLYMGVPC VTMAGSVHAH NVGVSLLTKV GLGHLVAKNE DEYVQLSVDL ASDVTALSKL RMSLRDLMAG SPVCNGPSFA VGLESAYRNM WKKYCKGEVP SLRRMEMLQK EVHDDPLISK DLGPSRVSVT GEATPSLKAN GSAPVPSSLP TQSPQLSKRM DSTSGGSENL YFQGGSHHHH HHHHHHGGWS HPQFEK |
-Experimental details
-Structure determination
Method | cryo EM |
---|---|
Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.8 mg/mL | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Buffer | pH: 7.5 Component:
| ||||||||||||
Grid | Model: C-flat-1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: GOLD / Support film - topology: HOLEY / Support film - Film thickness: 40 / Pretreatment - Type: PLASMA CLEANING | ||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 289.15 K / Instrument: LEICA EM GP |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
---|---|
Specialist optics | Energy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV |
Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 1 / Average electron dose: 52.4 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 81000 |
Sample stage | Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |