+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-8235 | |||||||||
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Title | Negative stain structure of Vps15/Vps34 complex | |||||||||
Map data | Vps15/34 complex | |||||||||
Sample |
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Keywords | autophagy / phosphatidylinositol 3-kinase (PtdIns3K) / endocytosis | |||||||||
Function / homology | Function and homology information Synthesis of PIPs at the early endosome membrane / Synthesis of PIPs at the late endosome membrane / RHO GTPases Activate NADPH Oxidases / nucleus-vacuole junction / Synthesis of PIPs at the Golgi membrane / vacuole-isolation membrane contact site / phosphatidylinositol 3-kinase complex, class III, type II / cellular response to potassium ion starvation / phosphatidylinositol 3-kinase complex, class III, type I / vacuole inheritance ...Synthesis of PIPs at the early endosome membrane / Synthesis of PIPs at the late endosome membrane / RHO GTPases Activate NADPH Oxidases / nucleus-vacuole junction / Synthesis of PIPs at the Golgi membrane / vacuole-isolation membrane contact site / phosphatidylinositol 3-kinase complex, class III, type II / cellular response to potassium ion starvation / phosphatidylinositol 3-kinase complex, class III, type I / vacuole inheritance / Macroautophagy / protein retention in Golgi apparatus / autophagy of peroxisome / pexophagy / protein targeting to vacuole / phagophore assembly site membrane / late endosome to vacuole transport / phagophore assembly site / fungal-type vacuole membrane / phosphatidylinositol 3-kinase / phosphatidylinositol-3-phosphate biosynthetic process / 1-phosphatidylinositol-3-kinase activity / phosphatidylinositol-mediated signaling / phosphatidylinositol phosphate biosynthetic process / autophagosome assembly / ubiquitin binding / positive regulation of transcription elongation by RNA polymerase II / macroautophagy / autophagy / endocytosis / protein transport / late endosome / peroxisome / non-specific serine/threonine protein kinase / endosome membrane / protein kinase activity / endosome / Golgi membrane / protein serine kinase activity / protein serine/threonine kinase activity / mitochondrion / ATP binding / membrane / cytosol / cytoplasm Similarity search - Function | |||||||||
Biological species | Saccharomyces cerevisiae (brewer's yeast) | |||||||||
Method | single particle reconstruction / negative staining / Resolution: 28.0 Å | |||||||||
Authors | Kirsten ML / Zhang L | |||||||||
Citation | Journal: Autophagy / Year: 2016 Title: Characterization of Atg38 and NRBF2, a fifth subunit of the autophagic Vps34/PIK3C3 complex. Authors: Yohei Ohashi / Nicolas Soler / Miguel García Ortegón / Lufei Zhang / Marie L Kirsten / Olga Perisic / Glenn R Masson / John E Burke / Arjen J Jakobi / Apostolos A Apostolakis / Christopher ...Authors: Yohei Ohashi / Nicolas Soler / Miguel García Ortegón / Lufei Zhang / Marie L Kirsten / Olga Perisic / Glenn R Masson / John E Burke / Arjen J Jakobi / Apostolos A Apostolakis / Christopher M Johnson / Maki Ohashi / Nicholas T Ktistakis / Carsten Sachse / Roger L Williams / Abstract: The phosphatidylinositol 3-kinase Vps34 is part of several protein complexes. The structural organization of heterotetrameric complexes is starting to emerge, but little is known about organization ...The phosphatidylinositol 3-kinase Vps34 is part of several protein complexes. The structural organization of heterotetrameric complexes is starting to emerge, but little is known about organization of additional accessory subunits that interact with these assemblies. Combining hydrogen-deuterium exchange mass spectrometry (HDX-MS), X-ray crystallography and electron microscopy (EM), we have characterized Atg38 and its human ortholog NRBF2, accessory components of complex I consisting of Vps15-Vps34-Vps30/Atg6-Atg14 (yeast) and PIK3R4/VPS15-PIK3C3/VPS34-BECN1/Beclin 1-ATG14 (human). HDX-MS shows that Atg38 binds the Vps30-Atg14 subcomplex of complex I, using mainly its N-terminal MIT domain and bridges the coiled-coil I regions of Atg14 and Vps30 in the base of complex I. The Atg38 C-terminal domain is important for localization to the phagophore assembly site (PAS) and homodimerization. Our 2.2 Å resolution crystal structure of the Atg38 C-terminal homodimerization domain shows 2 segments of α-helices assembling into a mushroom-like asymmetric homodimer with a 4-helix cap and a parallel coiled-coil stalk. One Atg38 homodimer engages a single complex I. This is in sharp contrast to human NRBF2, which also forms a homodimer, but this homodimer can bridge 2 complex I assemblies. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_8235.map.gz | 3.9 MB | EMDB map data format | |
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Header (meta data) | emd-8235-v30.xml emd-8235.xml | 15.3 KB 15.3 KB | Display Display | EMDB header |
Images | emd_8235.png | 361.5 KB | ||
Filedesc metadata | emd-8235.cif.gz | 6.8 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-8235 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-8235 | HTTPS FTP |
-Validation report
Summary document | emd_8235_validation.pdf.gz | 191.1 KB | Display | EMDB validaton report |
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Full document | emd_8235_full_validation.pdf.gz | 190.2 KB | Display | |
Data in XML | emd_8235_validation.xml.gz | 5.1 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-8235 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-8235 | HTTPS FTP |
-Related structure data
Related structure data | 5kc2MC 5kc1C M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_8235.map.gz / Format: CCP4 / Size: 4.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Vps15/34 complex | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 3.82 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : Vps15/34
Entire | Name: Vps15/34 |
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Components |
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-Supramolecule #1: Vps15/34
Supramolecule | Name: Vps15/34 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all Details: Subcomplex of Vps34 phosphatidylinositol 3-kinase (PtdIns3K) complex I (yeast) |
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Source (natural) | Organism: Saccharomyces cerevisiae (brewer's yeast) |
-Macromolecule #1: Phosphatidylinositol 3-kinase VPS34
Macromolecule | Name: Phosphatidylinositol 3-kinase VPS34 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO / EC number: phosphatidylinositol 3-kinase |
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Source (natural) | Organism: Saccharomyces cerevisiae (brewer's yeast) |
Molecular weight | Theoretical: 101.04718 KDa |
Recombinant expression | Organism: Saccharomyces cerevisiae (brewer's yeast) |
Sequence | String: MSLNNITFCV SQDLDVPLKV KIKSLEGHKP LLKPSQKILN PELMLIGSNV FPSSDLIVSL QVFDKERNRN LTLPIYTPYI PFRNSRTWD YWLTLPIRIK QLTFSSHLRI ILWEYNGSKQ IPFFNLETSI FNLKDCTLKR GFESLKFRYD VIDHCEVVTD N KDQENLNK ...String: MSLNNITFCV SQDLDVPLKV KIKSLEGHKP LLKPSQKILN PELMLIGSNV FPSSDLIVSL QVFDKERNRN LTLPIYTPYI PFRNSRTWD YWLTLPIRIK QLTFSSHLRI ILWEYNGSKQ IPFFNLETSI FNLKDCTLKR GFESLKFRYD VIDHCEVVTD N KDQENLNK YFQGEFTRLP WLDEITISKL RKQRENRTWP QGTFVLNLEF PMLELPVVFI EREIMNTQMN IPTLKNNPGL ST DLREPNR NDPQIKISLG DKYHSTLKFY DPDQPNNDPI EEKYRRLERA SKNANLDKQV KPDIKKRDYL NKIINYPPGT KLT AHEKGS IWKYRYYLMN NKKALTKLLQ STNLREESER VEVLELMDSW AEIDIDDALE LLGSTFKNLS VRSYAVNRLK KASD KELEL YLLQLVEAVC FENLSTFSDK SNSEFTIVDA VSSQKLSGDS MLLSTSHANQ KLLKSISSES ETSGTESLPI VISPL AEFL IRRALVNPRL GSFFYWYLKS ESEDKPYLDQ ILSSFWSRLD KKSRNILNDQ VRLINVLREC CETIKRLKDT TAKKME LLV HLLETKVRPL VKVRPIALPL DPDVLICDVC PETSKVFKSS LSPLKITFKT TLNQPYHLMF KVGDDLRQDQ LVVQIIS LM NELLKNENVD LKLTPYKILA TGPQEGAIEF IPNDTLASIL SKYHGILGYL KLHYPDENAT LGVQGWVLDN FVKSCAGY C VITYILGVGD RHLDNLLVTP DGHFFHADFG YILGQDPKPF PPLMKLPPQI IEAFGGAESS NYDKFRSYCF VAYSILRRN AGLILNLFEL MKTSNIPDIR IDPNGAILRV RERFNLNMSE EDATVHFQNL INDSVNALLP IVIDHLHNLA QYWRT UniProtKB: Phosphatidylinositol 3-kinase VPS34 |
-Macromolecule #2: Serine/threonine-protein kinase VPS15
Macromolecule | Name: Serine/threonine-protein kinase VPS15 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO / EC number: non-specific serine/threonine protein kinase |
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Source (natural) | Organism: Saccharomyces cerevisiae (brewer's yeast) |
Molecular weight | Theoretical: 166.55 KDa |
Recombinant expression | Organism: Saccharomyces cerevisiae (brewer's yeast) |
Sequence | String: MGAQLSLVVQ ASPSIAIFSY IDVLEEVHYV SQLNSSRFLK TCKALDPNGE IVIKVFIKPK DQYSLRPFLQ RIRAQSFKLG QLPHVLNYS KLIETNRAGY MIRQHLKNNL YDRLSLRPYL QDIELKFIAF QLLNALKDIH NLNIVHGDIK TENILVTSWN W CILTDFAA ...String: MGAQLSLVVQ ASPSIAIFSY IDVLEEVHYV SQLNSSRFLK TCKALDPNGE IVIKVFIKPK DQYSLRPFLQ RIRAQSFKLG QLPHVLNYS KLIETNRAGY MIRQHLKNNL YDRLSLRPYL QDIELKFIAF QLLNALKDIH NLNIVHGDIK TENILVTSWN W CILTDFAA FIKPVYLPED NPGEFLFYFD TSKRRTCYLA PERFNSKLYQ DGKSNNGRLT KEMDIFSLGC VIAEIFAEGR PI FNLSQLF KYKSNSYDVN REFLMEEMNS TDLRNLVLDM IQLDPSKRLS CDELLNKYRG IFFPDYFYTF IYDYFRNLVT MTT STPISD NTCTNSTLED NVKLLDETTE KIYRDFSQIC HCLDFPLIKD GGEIGSDPPI LESYKIEIEI SRFLNTNLYF PQNY HLVLQ QFTKVSEKIK SVKEECALLF ISYLSHSIRS IVSTATKLKN LELLAVFAQF VSDENKIDRV VPYFVCCFED SDQDV QALS LLTLIQVLTS VRKLNQLNEN IFVDYLLPRL KRLLISNRQN TNYLRIVFAN CLSDLAIIIN RFQEFTFAQH CNDNSM DNN TEIMESSTKY SAKLIQSVED LTVSFLTDND TYVKMALLQN ILPLCKFFGR ERTNDIILSH LITYLNDKDP ALRVSLI QT ISGISILLGT VTLEQYILPL LIQTITDSEE LVVISVLQSL KSLFKTGLIR KKYYIDISKT TSPLLLHPNN WIRQFTLM I IIEIINKLSK AEVYCILYPI IRPFFEFDVE FNFKSMISCC KQPVSRSVYN LLCSWSVRAS KSLFWKKIIT NHVDSFGNN RIEFITKNYS SKNYGFNKRD TKSSSSLKGI KTSSTVYSHD NKEIPLTAED RNWIDKFHII GLTEKDIWKI VALRGYVIRT ARVMAANPD FPYNNSNYRP LVQNSPPNLN LTNIMPRNIF FDVEFAEEST SEGQDSNLEN QQIYKYDESE KDSNKLNING S KQLSTVMD INGSLIFKNK SIATTTSNLK NVFVQLEPTS YHMHSPNHGL KDNANVKPER KVVVSNSYEG DVESIEKFLS TF KILPPLR DYKEFGPIQE IVRSPNMGNL RGKLIATLME NEPNSITSSA VSPGETPYLI TGSDQGVIKI WNLKEIIVGE VYS SSLTYD CSSTVTQITM IPNFDAFAVS SKDGQIIVLK VNHYQQESEV KFLNCECIRK INLKNFGKNE YAVRMRAFVN EEKS LLVAL TNLSRVIIFD IRTLERLQII ENSPRHGAVS SICIDEECCV LILGTTRGII DIWDIRFNVL IRSWSFGDHA PITHV EVCQ FYGKNSVIVV GGSSKTFLTI WNFVKGHCQY AFINSDEQPS MEHFLPIEKG LEELNFCGIR SLNALSTISV SNDKIL LTD EATSSIVMFS LNELSSSKAV ISPSRFSDVF IPTQVTANLT MLLRKMKRTS THSVDDSLYH HDIINSISTC EVDETPL LV ACDNSGLIGI FQ UniProtKB: Serine/threonine-protein kinase VPS15 |
-Experimental details
-Structure determination
Method | negative staining |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 5 mg/mL | ||||||||||
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Buffer | pH: 8.8 Component:
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Staining | Type: NEGATIVE / Material: Uranyl acetate / Details: droplet technique | ||||||||||
Grid | Model: Quantifoil R2/2 / Material: COPPER / Mesh: 300 |
-Electron microscopy
Microscope | FEI POLARA 300 |
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Image recording | Film or detector model: GATAN ULTRASCAN 4000 (4k x 4k) / Number grids imaged: 2 / Average exposure time: 1.0 sec. / Average electron dose: 20.0 e/Å2 |
Electron beam | Acceleration voltage: 120 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.5 µm / Nominal defocus min: 1.0 µm |
Sample stage | Specimen holder model: OTHER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Tecnai Polara / Image courtesy: FEI Company |