+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-28782 | |||||||||
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Title | Cryo-EM structure of Kap114 bound to H2A-H2B | |||||||||
Map data | sharpened | |||||||||
Sample |
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Keywords | Karyopherin Beta Nuclear Transport Histone Histone Chaperone / PROTEIN TRANSPORT-STRUCTURAL PROTEIN complex | |||||||||
Function / homology | Function and homology information HATs acetylate histones / Condensation of Prophase Chromosomes / SIRT1 negatively regulates rRNA expression / Activated PKN1 stimulates transcription of AR (androgen receptor) regulated genes KLK2 and KLK3 / Assembly of the ORC complex at the origin of replication / HDACs deacetylate histones / Recruitment and ATM-mediated phosphorylation of repair and signaling proteins at DNA double strand breaks / replication fork protection complex / Oxidative Stress Induced Senescence / RMTs methylate histone arginines ...HATs acetylate histones / Condensation of Prophase Chromosomes / SIRT1 negatively regulates rRNA expression / Activated PKN1 stimulates transcription of AR (androgen receptor) regulated genes KLK2 and KLK3 / Assembly of the ORC complex at the origin of replication / HDACs deacetylate histones / Recruitment and ATM-mediated phosphorylation of repair and signaling proteins at DNA double strand breaks / replication fork protection complex / Oxidative Stress Induced Senescence / RMTs methylate histone arginines / nuclear import signal receptor activity / NLS-bearing protein import into nucleus / RNA Polymerase I Promoter Escape / Estrogen-dependent gene expression / mRNA transport / Ub-specific processing proteases / nuclear pore / nucleosomal DNA binding / heterochromatin formation / small GTPase binding / protein import into nucleus / structural constituent of chromatin / nucleosome / nuclear envelope / chromatin organization / protein heterodimerization activity / DNA repair / regulation of DNA-templated transcription / DNA binding / nucleus / cytosol / cytoplasm Similarity search - Function | |||||||||
Biological species | Saccharomyces cerevisiae S288C (yeast) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.21 Å | |||||||||
Authors | Jiou J / Chook YM | |||||||||
Funding support | United States, 1 items
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Citation | Journal: Proc Natl Acad Sci U S A / Year: 2023 Title: Mechanism of RanGTP priming H2A-H2B release from Kap114 in an atypical RanGTP•Kap114•H2A-H2B complex. Authors: Jenny Jiou / Joy M Shaffer / Natalia E Bernades / Ho Yee Joyce Fung / Juliana Kikumoto Dias / Sheena D'Arcy / Yuh Min Chook / Abstract: Previously, we showed that the nuclear import receptor Importin-9 wraps around the H2A-H2B core to chaperone and transport it from the cytoplasm to the nucleus. However, unlike most nuclear import ...Previously, we showed that the nuclear import receptor Importin-9 wraps around the H2A-H2B core to chaperone and transport it from the cytoplasm to the nucleus. However, unlike most nuclear import systems where RanGTP dissociates cargoes from their importins, RanGTP binds stably to the Importin-9•H2A-H2B complex, and formation of the ternary RanGTP•Importin-9•H2A-H2B complex facilitates H2A-H2B release to the assembling nucleosome. It was unclear how RanGTP and the cargo H2A-H2B can bind simultaneously to an importin, and how interactions of the three components position H2A-H2B for release. Here, we show cryo-EM structures of Importin-9•RanGTP and of its yeast homolog Kap114, including Kap114•RanGTP, Kap114•H2A-H2B, and RanGTP•Kap114•H2A-H2B, to explain how the conserved Kap114 binds H2A-H2B and RanGTP simultaneously and how the GTPase primes histone transfer to the nucleosome. In the ternary complex, RanGTP binds to the N-terminal repeats of Kap114 in the same manner as in the Kap114/Importin-9•RanGTP complex, and H2A-H2B binds via its acidic patch to the Kap114 C-terminal repeats much like in the Kap114/Importin-9•H2A-H2B complex. Ran binds to a different conformation of Kap114 in the ternary RanGTP•Kap114•H2A-H2B complex. Here, Kap114 no longer contacts the H2A-H2B surface proximal to the H2A docking domain that drives nucleosome assembly, positioning it for transfer to the assembling nucleosome or to dedicated H2A-H2B chaperones in the nucleus. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_28782.map.gz | 143.9 MB | EMDB map data format | |
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Header (meta data) | emd-28782-v30.xml emd-28782.xml | 19.2 KB 19.2 KB | Display Display | EMDB header |
Images | emd_28782.png | 65 KB | ||
Filedesc metadata | emd-28782.cif.gz | 6.5 KB | ||
Others | emd_28782_half_map_1.map.gz emd_28782_half_map_2.map.gz | 262.7 MB 262.7 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-28782 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-28782 | HTTPS FTP |
-Validation report
Summary document | emd_28782_validation.pdf.gz | 914 KB | Display | EMDB validaton report |
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Full document | emd_28782_full_validation.pdf.gz | 913.6 KB | Display | |
Data in XML | emd_28782_validation.xml.gz | 16.3 KB | Display | |
Data in CIF | emd_28782_validation.cif.gz | 19.2 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-28782 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-28782 | HTTPS FTP |
-Related structure data
Related structure data | 8f0xMC 8f19C 8f1eC 8f7aC C: citing same article (ref.) M: atomic model generated by this map |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_28782.map.gz / Format: CCP4 / Size: 282.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||
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Annotation | sharpened | ||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 0.5295 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Half map: half-A
File | emd_28782_half_map_1.map | ||||||||||||
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Annotation | half-A | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: half-B
File | emd_28782_half_map_2.map | ||||||||||||
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Annotation | half-B | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Complex of Kap114 bound to H2A-H2B dimer
Entire | Name: Complex of Kap114 bound to H2A-H2B dimer |
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Components |
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-Supramolecule #1: Complex of Kap114 bound to H2A-H2B dimer
Supramolecule | Name: Complex of Kap114 bound to H2A-H2B dimer / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
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Source (natural) | Organism: Saccharomyces cerevisiae S288C (yeast) |
-Macromolecule #1: Importin subunit beta-5
Macromolecule | Name: Importin subunit beta-5 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Saccharomyces cerevisiae S288C (yeast) / Strain: ATCC 204508 / S288c |
Molecular weight | Theoretical: 114.019695 KDa |
Recombinant expression | Organism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) |
Sequence | String: MDINELIIGA QSADKHTREV AETQLLQWCD SDASQVFKAL ANVALQHEAS LESRQFALLS LRKLITMYWS PGFESYRSTS NVEIDVKDF IREVLLKLCL NDNENTKIKN GASYCIVQIS AVDFPDQWPQ LLTVIYDAIS HQHSLNAMSL LNEIYDDVVS E EMFFEGGI ...String: MDINELIIGA QSADKHTREV AETQLLQWCD SDASQVFKAL ANVALQHEAS LESRQFALLS LRKLITMYWS PGFESYRSTS NVEIDVKDF IREVLLKLCL NDNENTKIKN GASYCIVQIS AVDFPDQWPQ LLTVIYDAIS HQHSLNAMSL LNEIYDDVVS E EMFFEGGI GLATMEIVFK VLNTETSTLI AKIAALKLLK ACLLQMSSHN EYDEASRKSF VSQCLATSLQ ILGQLLTLNF GN VDVISQL KFKSIIYENL VFIKNDFSRK HFSSELQKQF KIMAIQDLEN VTHINANVET TESEPLLETV HDCSIYIVEF LTS VCTLQF SVEEMNKIIT SLTILCQLSS ETREIWTSDF NTFVSKETGL AASYNVRDQA NEFFTSLPNP QLSLIFKVVS NDIE HSTCN YSTLESLLYL LQCILLNDDE ITGENIDQSL QILIKTLENI LVSQEIPELI LARAILTIPR VLDKFIDALP DIKPL TSAF LAKSLNLALK SDKELIKSAT LIAFTYYCYF AELDSVLGPE VCSETQEKVI RIINQVSSDA EEDTNGALME VLSQVI SYN PKEPHSRKEI LQAEFHLVFT ISSEDPANVQ VVVQSQECLE KLLDNINMDN YKNYIELCLP SFINVLDSNN ANNYRYS PL LSLVLEFITV FLKKKPNDGF LPDEINQYLF EPLAKVLAFS TEDETLQLAT EAFSYLIFNT DTRAMEPRLM DIMKVLER L LSLEVSDSAA MNVGPLVVAI FTRFSKEIQP LIGRILEAVV VRLIKTQNIS TEQNLLSVLC FLTCNDPKQT VDFLSSFQI DNTDALTLVM RKWIEAFEVI RGEKRIKENI VALSNLFFLN DKRLQKVVVN GNLIPYEGDL IITRSMAKKM PDRYVQVPLY TKIIKLFVS ELSFQSKQPN PEQLITSDIK QEVVNANKDD DNDDWEDVDD VLDYDKLKEY IDDDVDEEAD DDSDDITGLM D VKESVVQL LVRFFKEVAS KDVSGFHCIY ETLSDSERKV LSEALL UniProtKB: Importin subunit beta-5 |
-Macromolecule #2: Histone H2A.2
Macromolecule | Name: Histone H2A.2 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Saccharomyces cerevisiae S288C (yeast) / Strain: ATCC 204508 / S288c |
Molecular weight | Theoretical: 13.88198 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: SGGKGGKAGS AAKASQSRSA KAGLTFPVGR VHRLLRRGNY AQRIGSGAPV YLTAVLEYLA AEILELAGNA ARDNKKTRII PRHLQLAIR NDDELNKLLG NVTIAQGGVL PNIHQNLLPK KSAKTAKASQ EL UniProtKB: Histone H2A.2 |
-Macromolecule #3: Histone H2B.2
Macromolecule | Name: Histone H2B.2 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Saccharomyces cerevisiae S288C (yeast) / Strain: ATCC 204508 / S288c |
Molecular weight | Theoretical: 14.133145 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: SSAAEKKPAS KAPAEKKPAA KKTSTSVDGK KRSKVRKETY SSYIYKVLKQ THPDTGISQK SMSILNSFVN DIFERIATEA SKLAAYNKK STISAREIQT AVRLILPGEL AKHAVSEGTR AVTKYSSSTQ A UniProtKB: Histone H2B.2 |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 4.0 mg/mL | ||||||||||||||||||
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Buffer | pH: 7.5 Component:
Details: 20 mM Tris pH 7.5, 300 mM NaCl, 2 mM MgCl2, 1 mM TCEP, and 0.1% NP-40 | ||||||||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 4 K / Instrument: FEI VITROBOT MARK IV | ||||||||||||||||||
Details | 1 Kap114 to 1.2 H2A-H2B ratio |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K3 (6k x 4k) / Average electron dose: 50.0 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: 2.5 µm / Nominal defocus min: 1.5 µm |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
Startup model | Type of model: PDB ENTRY PDB model - PDB ID: |
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Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 3.21 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 554529 |
Initial angle assignment | Type: MAXIMUM LIKELIHOOD |
Final angle assignment | Type: MAXIMUM LIKELIHOOD |