+
Open data
-
Basic information
Entry | ![]() | |||||||||
---|---|---|---|---|---|---|---|---|---|---|
Title | Consensus map of Sec7 dimer autoinhibited conformation | |||||||||
![]() | ||||||||||
![]() |
| |||||||||
![]() | GEF / Exchange Factor / Golgi / PROTEIN TRANSPORT | |||||||||
Biological species | ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 5.3 Å | |||||||||
![]() | Brownfield BA / Fromme JC | |||||||||
Funding support | ![]()
| |||||||||
![]() | ![]() Title: Sec7 regulatory domains scaffold autoinhibited and active conformations. Authors: Bryce A Brownfield / Brian C Richardson / Steve L Halaby / J Christopher Fromme / ![]() Abstract: The late stages of Golgi maturation involve a series of sequential trafficking events in which cargo-laden vesicles are produced and targeted to multiple distinct subcellular destinations. Each of ...The late stages of Golgi maturation involve a series of sequential trafficking events in which cargo-laden vesicles are produced and targeted to multiple distinct subcellular destinations. Each of these vesicle biogenesis events requires activation of an Arf GTPase by the Sec7/BIG guanine nucleotide exchange factor (GEF). Sec7 localization and activity is regulated by autoinhibition, positive feedback, and interaction with other GTPases. Although these mechanisms have been characterized biochemically, we lack a clear picture of how GEF localization and activity is modulated by these signals. Here, we report the cryogenic electron microscopy structure of full-length Sec7 in its autoinhibited form, revealing the architecture of its multiple regulatory domains. We use functional experiments to determine the basis for autoinhibition and use structural predictions to produce a model for an active conformation of the GEF that is supported empirically. This study therefore elucidates the conformational transition that Sec7 undergoes to become active on the organelle membrane surface. | |||||||||
History |
|
-
Structure visualization
Supplemental images |
---|
-
Downloads & links
-EMDB archive
Map data | ![]() | 1.8 MB | ![]() | |
---|---|---|---|---|
Header (meta data) | ![]() ![]() | 18.2 KB 18.2 KB | Display Display | ![]() |
Images | ![]() | 24.5 KB | ||
Filedesc metadata | ![]() | 6.5 KB | ||
Others | ![]() ![]() | 337.4 MB 337.4 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
-
Links
EMDB pages | ![]() ![]() |
---|
-
Map
File | ![]() | ||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Voxel size | X=Y=Z: 1.7415 Å | ||||||||||||||||||||
Density |
| ||||||||||||||||||||
Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
|
-Supplemental data
-Half map: #1
File | emd_42183_half_map_1.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Projections & Slices |
| ||||||||||||
Density Histograms |
-Half map: #2
File | emd_42183_half_map_2.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Projections & Slices |
| ||||||||||||
Density Histograms |
-
Sample components
-Entire : Sec7 homodimer
Entire | Name: Sec7 homodimer |
---|---|
Components |
|
-Supramolecule #1: Sec7 homodimer
Supramolecule | Name: Sec7 homodimer / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
---|---|
Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 392.14 KDa |
-Macromolecule #1: SEC7 domain-containing protein
Macromolecule | Name: SEC7 domain-containing protein / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO |
---|---|
Source (natural) | Organism: ![]() |
Recombinant expression | Organism: ![]() |
Sequence | String: MEQKLISEED LNSAVDHHHH HHRIPGLINS SLKFVVSSLD IIAAQAGRNK QLAELAEKAL AAIKENDQQL PDPEVVFAPL QLATKSGTIP LTTTALDCIG KLISYSYFSA PSSSATQDGT EQTPLIERAI DTICDCFQGE TTLVEIQLQI VKSLLAAVLN DKIIVHGAGL ...String: MEQKLISEED LNSAVDHHHH HHRIPGLINS SLKFVVSSLD IIAAQAGRNK QLAELAEKAL AAIKENDQQL PDPEVVFAPL QLATKSGTIP LTTTALDCIG KLISYSYFSA PSSSATQDGT EQTPLIERAI DTICDCFQGE TTLVEIQLQI VKSLLAAVLN DKIIVHGAGL LKAVRQVYNI FLLSRSTANQ QVAQGTLTQM VGTVFERVKT RLHMKEARAN LGRLKASRSS LAVDRSDDQD SQAGKVDGED ATVETVSDAT PSESVDKAGG GKLTLKDLEH RKSFDDSHMG DGPTMVSQVK PMKKASRSVS EQSLQESPQD ETPESLDAED EAYIRDAYLV FRSFCNLSTK ILPPDQLYDL RGQPMRSKLI SLHLIHTLLN NHITVFTSPL CTIRNTKNNE PTNFLQAIKY YLCLSITRNG ASSVDRVFDI CCEIFWLMLK YMRSSFKNEI EVFLNEIYLA LLARRNAPLS QKLTFVGILK RLCEDPRALV ELYLNYDCNQ NVDNIFQTIV EDLSRFATAS VPITPTQEQQ YEESRSKSAT AGEWQIKSVL PPPLSVALIA TNHEADTELP KEYVMKRTAL DSLVETLRSL VHWSQPGRPE LNGASGDVQR RTSSDDLGDS IDPSMSETAS RMEVPIAPAT PVIDDDPDQL EKEKARKTAM TNAIKVFNFK PKHGIKLLIK EGFIPSDKPE DIARFLLREE RLDKAQIGEY LGEGDQKNVD IMHAFVDMMD FSKKRFVDAL REFLQAFRLP GEAQKIDRFM LKFAHRYVTG NPNAFANADT PYVLAYSVIM LNTDLHSSKV VKRMSKAEFI KNNRGINDNA DLPDEYLIGI YDDIASNEIV LKSEREAAAA AGTLPAQSTG LAGLGQAFSN VGRDLQREAY VQQSEEISLR SEQLFRDLYR SQRKSATKGG VKFISATSFK HVGPMFDATW MSFFSTLSSL VQKTHNLDVN KLCLEGMKLA TKIACLFDLS TPREAFISML KNTANLNNPR EMQAKNVEAL KVLLDLAQTE GNYLKESWKD VLLCISQLDR LQLISGGVDE SAVPDVSRAR FVPPPRTETG ESRKSTSSAR RTRPRAHTGP QGVSLEIALE SRSDEVIKSV DRIFTNTANL SRDAIIHFAR ALTEVSWDEI KVSGSNDSPR TYSLQKIVEI SYYNMTRVRF EWSHIWDVLG EHFNRVGCHA NTAIVFFALD SLRQLSMRFM EIEELAGFKF QKDFLKPFEH VMSNSSNVTV KDMVLRCLIQ MIQARGENIR SGWRTMFGVF TVAAREPYES IVNLAYENVT QVYKTRFGVV ISQGAFTDLI VCLTEFSKNM RFQKKSLQAM ETLKSVIPTM LKTPECPLSQ HKPTATTASG SESHSKKAAV QQTRTSVEEG FWFPVLFAFH DVLMTGEDLE VRSNALNYFF ETLLRYGGDF PPEFWDILWR QQLYPIFMVL RSRPEMTNAL NHEELSVWLS TTMIQALRNM ITLFTHYFDA LEYMLDRFLE LLALCICQEN DTIARIGSNC LQQLILQNVT KFTAEHWAKI VGAFCELFER TTAYQLFSAT TINSTASLSP PPSGLELGGP LSPTSATAPV DGKSLKINGV ETNGQTPGAE PANGDADGNG TAAAAADASA PAATPQPQQG PAQQLEEFKP NNPLQQQPVV VTAARRRFFN RIISRCVLQL LMIETVNELF SNDAVYAQIP SAELLRLMAL LKKSFLFAKR FNADKDLRMR LWREGFMKQP PNLLKQESGS AATYVAILFR MFGDTAPDRR GSRADVEAAL VPLCRDIIRG YTALDDESQH RNIVAWRPVV VDVLEGYAAF PRDAFAAHIR SFYPLVVELL GKDLGQDLRA ALLLVLRRVG EVGLGIEGMG SGGAAAAAAA GAAAASSGQG NGNGAAAAAA DSERRSSVLS VPSGPRHTPS MDSLNDDPSR QVMGKAEQKL ISEEDLNSAV DHHHHHH |
-Experimental details
-Structure determination
Method | cryo EM |
---|---|
![]() | single particle reconstruction |
Aggregation state | particle |
-
Sample preparation
Concentration | 8.0 mg/mL | |||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Buffer | pH: 7.4 Component:
| |||||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV |
-
Electron microscopy
Microscope | FEI TALOS ARCTICA |
---|---|
Specialist optics | Energy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV |
Image recording | Film or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Number real images: 4474 / Average exposure time: 3.092 sec. / Average electron dose: 55.6 e/Å2 |
Electron beam | Acceleration voltage: 200 kV / Electron source: ![]() |
Electron optics | C2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.2 µm / Nominal defocus min: 0.6 µm / Nominal magnification: 63000 |
Sample stage | Cooling holder cryogen: NITROGEN |
Experimental equipment | ![]() Model: Talos Arctica / Image courtesy: FEI Company |
+
Image processing
-Atomic model buiding 1
Initial model | Chain - Source name: AlphaFold / Chain - Initial model type: in silico model |
---|---|
Details | Phenix Real Space Refinement |
Refinement | Space: REAL / Protocol: OTHER |