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- EMDB-13189: Homology model of the full-length AP-3 complex in a stretched ope... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-13189 | |||||||||
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Title | Homology model of the full-length AP-3 complex in a stretched open conformation | |||||||||
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Function / homology | ![]() AP-3 adaptor complex / clathrin adaptor complex / Golgi to vacuole transport / protein targeting to vacuole / membrane coat / vesicle-mediated transport / intracellular protein transport / cytoplasmic vesicle membrane / endocytosis / cytoplasmic vesicle / Golgi apparatus Similarity search - Function | |||||||||
Biological species | ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 10.5 Å | |||||||||
![]() | Schubert E / Raunser S | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Flexible open conformation of the AP-3 complex explains its role in cargo recruitment at the Golgi. Authors: Jannis Schoppe / Evelyn Schubert / Amir Apelbaum / Erdal Yavavli / Oliver Birkholz / Heike Stephanowitz / Yaping Han / Angela Perz / Oliver Hofnagel / Fan Liu / Jacob Piehler / Stefan ...Authors: Jannis Schoppe / Evelyn Schubert / Amir Apelbaum / Erdal Yavavli / Oliver Birkholz / Heike Stephanowitz / Yaping Han / Angela Perz / Oliver Hofnagel / Fan Liu / Jacob Piehler / Stefan Raunser / Christian Ungermann / ![]() Abstract: Vesicle formation at endomembranes requires the selective concentration of cargo by coat proteins. Conserved adapter protein complexes at the Golgi (AP-3), the endosome (AP-1), or the plasma membrane ...Vesicle formation at endomembranes requires the selective concentration of cargo by coat proteins. Conserved adapter protein complexes at the Golgi (AP-3), the endosome (AP-1), or the plasma membrane (AP-2) with their conserved core domain and flexible ear domains mediate this function. These complexes also rely on the small GTPase Arf1 and/or specific phosphoinositides for membrane binding. The structural details that influence these processes, however, are still poorly understood. Here we present cryo-EM structures of the full-length stable 300 kDa yeast AP-3 complex. The structures reveal that AP-3 adopts an open conformation in solution, comparable to the membrane-bound conformations of AP-1 or AP-2. This open conformation appears to be far more flexible than AP-1 or AP-2, resulting in compact, intermediate, and stretched subconformations. Mass spectrometrical analysis of the cross-linked AP-3 complex further indicates that the ear domains are flexibly attached to the surface of the complex. Using biochemical reconstitution assays, we also show that efficient AP-3 recruitment to the membrane depends primarily on cargo binding. Once bound to cargo, AP-3 clustered and immobilized cargo molecules, as revealed by single-molecule imaging on polymer-supported membranes. We conclude that its flexible open state may enable AP-3 to bind and collect cargo at the Golgi and could thus allow coordinated vesicle formation at the trans-Golgi upon Arf1 activation. | |||||||||
History |
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Structure visualization
Movie |
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 12.9 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 17.9 KB 17.9 KB | Display Display | ![]() |
Images | ![]() | 52.7 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Validation report
Summary document | ![]() | 295.6 KB | Display | ![]() |
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Full document | ![]() | 295.2 KB | Display | |
Data in XML | ![]() | 6.2 KB | Display | |
Data in CIF | ![]() | 7 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 7p3zMC ![]() 7p3xC ![]() 7p3yC M: atomic model generated by this map C: citing same article ( |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Voxel size | X=Y=Z: 1.07 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
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Sample components
-Entire : Full-length AP-3 complex from Saccharomyces cerevisiae
Entire | Name: Full-length AP-3 complex from Saccharomyces cerevisiae |
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Components |
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-Supramolecule #1: Full-length AP-3 complex from Saccharomyces cerevisiae
Supramolecule | Name: Full-length AP-3 complex from Saccharomyces cerevisiae type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
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Source (natural) | Organism: ![]() ![]() |
Recombinant expression | Organism: ![]() ![]() |
-Macromolecule #1: AP-3 complex subunit delta
Macromolecule | Name: AP-3 complex subunit delta / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 110.903016 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MTSLYAPGAE DIRQRLRPFG FFFEKSLKDL IKGIRSHNET PEKLDQFFKQ VLSECREEVN SPDLNSKTNA VLKLTYLEMY GFDMAWCNF HILEVMSSNK LQQKRVGYLA ASQSFYKDSD ILMLATNLLK KDLKYDGNND VVKVGIALSG LSTIITPSLA R DIADDLFT ...String: MTSLYAPGAE DIRQRLRPFG FFFEKSLKDL IKGIRSHNET PEKLDQFFKQ VLSECREEVN SPDLNSKTNA VLKLTYLEMY GFDMAWCNF HILEVMSSNK LQQKRVGYLA ASQSFYKDSD ILMLATNLLK KDLKYDGNND VVKVGIALSG LSTIITPSLA R DIADDLFT MLNSTRPYIR KKAITALFKV FLQYPEALRD NFDKFVSKLD DDDISVVSAA VSVICELSKK NPQPFIQLSP LL YEILVTI DNNWIIIRLL KLFTNLSQVE PKLRAKLLPK ILELMESTVA TSVIYESVNC IVKGNMLEED DFETAMACLE RLH TFCDSQ DPNLRYISCI LFYKIGKINT DFISRFDQLI IRLLSDVDVS IRSKAIELVE GIVDEDNLKA IVQTLMKQFV DEDV VILQT GSIVYEKSKR IPIIIPENYK IKMVNVIISI CSADNYSSVN DFEWYNAVIM DLAMLCQDIS DKSLGSKIGE QFRNL MIKV PSMREVTIAN IIKLISNDNI NKQLPTVLRE CIWCLGEFST LVENGNDLIK IMTENISYYS HSVQEVLILA LVKVFS NWC NNFQEDKRFE IKMVLKELIE FFENLSYSST FEVQERSVEV LEFLRLSLEA LEEDTEGLPM LLSEVLPSFF NAYELAP IA RGTQLKLAVD ENLDLETPFL TKEAADELLD EQKSDAISDL MSDISMDEQV ELKFVDDSDT SYEEKEKLDD FENPFEIE R EKERMSNPYY LGEEDEERTK NSKDLLDLNE EESSDKKPET IRLNRTDNSL NSLSLSTTEI SRKKKKGKKK NRVQVLSDE PVIEAAPKRK DAFQKPHDNH STQNPLKKDK INLRMHSQLE NFDFSNFGQS SNAGRGSQEE GNLRKEDELE LSRLEANLIV KDEKDNLSD TEEVIVIKKK KKGKKSKSKN KLKTKAKNSP EPNEFLRDQS TDIRTLQVDG SDYKDDDDKD YKDDDDKDYK D DDDK |
-Macromolecule #2: Y55_G0035830.mRNA.1.CDS.1
Macromolecule | Name: Y55_G0035830.mRNA.1.CDS.1 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 91.712016 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MVDSIHRIAS ALDTAKVITR EAAAVATSKL GESSYTYYSQ NINPQQLVTL LNSRNSREVR DAMKRIISIM ASDDDSIDVQ LYFADVVKN ITTNDTKVKR LIHLYLLRFA ENDPNLTLLS INSLQKSLSD SNSELRCFAL SALSDMKMSS LAPIILHTVK K LVTDPSAM ...String: MVDSIHRIAS ALDTAKVITR EAAAVATSKL GESSYTYYSQ NINPQQLVTL LNSRNSREVR DAMKRIISIM ASDDDSIDVQ LYFADVVKN ITTNDTKVKR LIHLYLLRFA ENDPNLTLLS INSLQKSLSD SNSELRCFAL SALSDMKMSS LAPIILHTVK K LVTDPSAM VRGEVALAII KLYRAGKNDY HEELLDILKE LMADTDPKVI SCAVLAYKEC YADHLELLHG HFRRYCRIIK QL DSWSQSY LIELLIKYCK QYLPKPTVVD KSSEGSPRSC PLPDKYNEIE YPSYEVVNDP DLDLFLQSLN CLIYSSNPTV ILS CCNALY QLASPLQMKN TKFIEALVRT VTMTENQGNK EMLLQAIHFL SILDQTLFLP YTKKFYVFPK DPIVASIWKI QILS TLINE SNVKEIFKEL KYYVASAHFP ENVVIMAVKS LSRCGQLSTS WESHVMKWLI DHMESHNLSA SVLDAYVNVI RMLVQ KNPT KHLRIIFKLA DLLTVQTSLA DNARAGIVWL FGEIASIEFK ICPDVLRRLI QNFSNEGPET RCQILVLSAK LLSYDI DNF KQAQVTGSEE NNQNPPYYDF SGSRISQMYN AVLYLAKYDD EFDIRDRARM ISSLFDSGKY EIVSLLLQAP KPTARSD DF IVSARLETHT PEIKEFFRML PWNTEITEVG ETGNDIREGA ELKDYNKYKK SFSSQSFITN NSARSFTSSS NAKLTGIN D GDSNSISGKG NVNTFTSQNG KKYRLQSLDE FFSDIPERKS KPRKIIKVVE ESSDEDEDES EESSDDDEYS DSSLGTSSS GTSSSHLEL |
-Macromolecule #3: AP-3 complex subunit mu
Macromolecule | Name: AP-3 complex subunit mu / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 54.899062 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MYLSFYITDT KNKLIFQYLL GATAPSFKHL WTRVQSTCPQ LLEDSSSDDY LDHSMVGRDL EVYKYFSVIN KLNYWCLAST SKSKGPLDC FTFLETIDRI LLEYFDKDKL SIKKIVNNYD RISLIFNCCV EAGEPNVSDM LYVNKIKEAV PERSDLSKFI S STAHNLQQ ...String: MYLSFYITDT KNKLIFQYLL GATAPSFKHL WTRVQSTCPQ LLEDSSSDDY LDHSMVGRDL EVYKYFSVIN KLNYWCLAST SKSKGPLDC FTFLETIDRI LLEYFDKDKL SIKKIVNNYD RISLIFNCCV EAGEPNVSDM LYVNKIKEAV PERSDLSKFI S STAHNLQQ AVQLPQQRQQ QLQQNQISRG SNSLIENEEI VPWRTSRASK HENNELYVDL LETFHVVFEK KKSHLRLLTG SI HGIVDVR SYLNDNPLVA VKLNTMGNDI GIPSLHDCVE INDGVFSPSN ITFIPPDGKF RLLEYSVDLS SQVKQSGVRM NSI GLMSLH FQNGLGKDSD EFELSLNIEN FKKVSQVDDL KIDLQFNVEN ADPNEIAYKI KILRNTHGRF ENSIIMGQGQ WIFD KSTAT GTVPVLRGCI EYENTGPNFT KKVDLQTVSL EYSYIGQSAS GIYVEAIDIV SGLTIGKNTK LYKGAKYKTQ TGNFQ VRL |
-Macromolecule #4: AP complex subunit sigma
Macromolecule | Name: AP complex subunit sigma / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 21.951646 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MIHAVLIFNK KCQPRLVKFY TPVDLPKQKL LLEQVYELIS QRNSDFQSSF LVTPPSLLLS NENNNDEVNN EDIQIIYKNY ATLYFTFIV DDQESELAIL DLIQTFVESL DRCFTEVNEL DLIFNWQTLE SVLEEIVQGG MVIETNVNRI VASVDELNKA A ESTDSKIG ...String: MIHAVLIFNK KCQPRLVKFY TPVDLPKQKL LLEQVYELIS QRNSDFQSSF LVTPPSLLLS NENNNDEVNN EDIQIIYKNY ATLYFTFIV DDQESELAIL DLIQTFVESL DRCFTEVNEL DLIFNWQTLE SVLEEIVQGG MVIETNVNRI VASVDELNKA A ESTDSKIG RLTSTGFGSA LQAFAQGGFA QWATGQ |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 0.25 mg/mL |
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Buffer | pH: 7.4 / Details: 20 mM Hepes pH 7.4 150 mM NaCl 1.5 mM MgCl2 |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK III Details: The sample was blotted using a 2.5 s blotting time and 0 blotting force with 100% humidity at 277.15 K. |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average electron dose: 81.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELD / Nominal defocus max: 3.6 µm / Nominal defocus min: 1.5 µm / Nominal magnification: 130000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Image processing
-Atomic model buiding 1
Refinement | Protocol: FLEXIBLE FIT |
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Output model | ![]() PDB-7p3z: |