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- EMDB-31812: masked histone region in the catalytic core of human telomerase h... -

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Basic information

Entry
Database: EMDB / ID: EMD-31812
Titlemasked histone region in the catalytic core of human telomerase holoenzyme
Map data
Sample
  • Complex: Catalytic core
    • Protein or peptide: Telomerase reverse transcriptase
    • Protein or peptide: Histone H2A type 1-B/E
    • Protein or peptide: Histone H2B type 1-K
    • RNA: Telomerase RNA component
    • DNA: Primer DNA
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.97 Å
AuthorsWan F / Ding Y / Yang L / Wu Z / Wu J / Lei M
Funding support China, 1 items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC) China
CitationJournal: Cell Res / Year: 2021
Title: Zipper head mechanism of telomere synthesis by human telomerase.
Authors: Futang Wan / Yongbo Ding / Yuebin Zhang / Zhenfang Wu / Shaobai Li / Lin Yang / Xiangyu Yan / Pengfei Lan / Guohui Li / Jian Wu / Ming Lei /
Abstract: Telomerase, a multi-subunit ribonucleoprotein complex, is a unique reverse transcriptase that catalyzes the processive addition of a repeat sequence to extend the telomere end using a short fragment ...Telomerase, a multi-subunit ribonucleoprotein complex, is a unique reverse transcriptase that catalyzes the processive addition of a repeat sequence to extend the telomere end using a short fragment of its own RNA component as the template. Despite recent structural characterizations of human and Tetrahymena telomerase, it is still a mystery how telomerase repeatedly uses its RNA template to synthesize telomeric DNA. Here, we report the cryo-EM structure of human telomerase holoenzyme bound with telomeric DNA at resolutions of 3.5 Å and 3.9 Å for the catalytic core and biogenesis module, respectively. The structure reveals that a leucine residue Leu980 in telomerase reverse transcriptase (TERT) catalytic subunit functions as a zipper head to limit the length of the short primer-template duplex in the active center. Moreover, our structural and computational analyses suggest that TERT and telomerase RNA (hTR) are organized to harbor a preformed active site that can accommodate short primer-template duplex substrates for catalysis. Furthermore, our findings unveil a double-fingers architecture in TERT that ensures nucleotide addition processivity of human telomerase. We propose that the zipper head Leu980 is a structural determinant for the sequence-based pausing signal of DNA synthesis that coincides with the RNA element-based physical template boundary. Functional analyses unveil that the non-glycine zipper head plays an essential role in both telomerase repeat addition processivity and telomere length homeostasis. In addition, we also demonstrate that this zipper head mechanism is conserved in all eukaryotic telomerases. Together, our study provides an integrated model for telomerase-mediated telomere synthesis.
History
DepositionAug 24, 2021-
Header (metadata) releaseMar 30, 2022-
Map releaseMar 30, 2022-
UpdateMar 30, 2022-
Current statusMar 30, 2022Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_31812.png

Downloads & links

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Map

FileDownload / File: emd_31812.map.gz / Format: CCP4 / Size: 274.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.1 Å
Density
Contour LevelBy AUTHOR: 3.0
Minimum - Maximum-9.654326 - 15.548514
Average (Standard dev.)-0.0061614523 (±0.2957092)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions416416416
Spacing416416416
CellA=B=C: 457.6 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : Catalytic core

EntireName: Catalytic core
Components
  • Complex: Catalytic core
    • Protein or peptide: Telomerase reverse transcriptase
    • Protein or peptide: Histone H2A type 1-B/E
    • Protein or peptide: Histone H2B type 1-K
    • RNA: Telomerase RNA component
    • DNA: Primer DNA

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Supramolecule #1: Catalytic core

SupramoleculeName: Catalytic core / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Telomerase reverse transcriptase

MacromoleculeName: Telomerase reverse transcriptase / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
SequenceString: MPRAPRCRAV RSLLRSHYRE VLPLATFVRR LGPQGWRLVQ RGDPAAFRAL VAQCLVCVPW DARPPPAAPS FRQVSCLKE LVARVLQRLC ERGAKNVLAF GFALLDGARG GPPEAFTTSV RSYLPNTVTD ALRGSGAWGL L LRRVGDDV LVHLLARCAL FVLVAPSCAY ...String:
MPRAPRCRAV RSLLRSHYRE VLPLATFVRR LGPQGWRLVQ RGDPAAFRAL VAQCLVCVPW DARPPPAAPS FRQVSCLKE LVARVLQRLC ERGAKNVLAF GFALLDGARG GPPEAFTTSV RSYLPNTVTD ALRGSGAWGL L LRRVGDDV LVHLLARCAL FVLVAPSCAY QVCGPPLYQL GAATQARPPP HASGPRRRLG CERAWNHSVR EA GVPLGLP APGARRRGGS ASRSLPLPKR PRRGAAPEPE RTPVGQGSWA HPGRTRGPSD RGFCVVSPAR PAE EATSLE GALSGTRHSH PSVGRQHHAG PPSTSRPPRP WDTPCPPVYA ETKHFLYSSG DKEQLRPSFL LSSL RPSLT GARRLVETIF LGSRPWMPGT PRRLPRLPQR YWQMRPLFLE LLGNHAQCPY GVLLKTHCPL RAAVT PAAG VCAREKPQGS VAAPEEEDTD PRRLVQLLRQ HSSPWQVYGF VRACLRRLVP PGLWGSRHNE RRFLRN TKK FISLGKHAKL SLQELTWKMS VRDCAWLRRS PGVGCVPAAE HRLREEILAK FLHWLMSVYV VELLRSF FY VTETTFQKNR LFFYRKSVWS KLQSIGIRQH LKRVQLRELS EAEVRQHREA RPALLTSRLR FIPKPDGL R PIVNMDYVVG ARTFRREKRA ERLTSRVKAL FSVLNYERAR RPGLLGASVL GLDDIHRAWR TFVLRVRAQ DPPPELYFVK VDVTGAYDTI PQDRLTEVIA SIIKPQNTYC VRRYAVVQKA AHGHVRKAFK SHVSTLTDLQ PYMRQFVAH LQETSPLRDA VVIEQSSSLN EASSGLFDVF LRFMCHHAVR IRGKSYVQCQ GIPQGSILST L LCSLCYGD MENKLFAGIR RDGLLLRLVD DFLLVTPHLT HAKTFLRTLV RGVPEYGCVV NLRKTVVNFP VE DEALGGT AFVQMPAHGL FPWCGLLLDT RTLEVQSDYS SYARTSIRAS LTFNRGFKAG RNMRRKLFGV LRL KCHSLF LDLQVNSLQT VCTNIYKILL LQAYRFHACV LQLPFHQQVW KNPTFFLRVI SDTASLCYSI LKAK NAGMS LGAKGAAGPL PSEAVQWLCH QAFLLKLTRH RVTYVPLLGS LRTAQTQLSR KLPGTTLTAL EAAAN PALP SDFKTILD

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Macromolecule #2: Histone H2A type 1-B/E

MacromoleculeName: Histone H2A type 1-B/E / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
SequenceString:
SGRGKQGGKA RAKAKTRSSR AGLQFPVGRV HRLLRKGNYS ERVGAGAPVY LAAVLEYLTA EILELAGNA ARDNKKTRII PRHLQLAIRN DEELNKLLGR VTIAQGGVLP NIQAVLLPKK TESHHKAKGK

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Macromolecule #3: Histone H2B type 1-K

MacromoleculeName: Histone H2B type 1-K / type: protein_or_peptide / ID: 3 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
SequenceString:
PEPAKSAPAP KKGSKKAVTK AQKKDGKKRK RSRKESYSVY VYKVLKQVHP DTGISSKAMG IMNSFVNDI FERIAGEASR LAHYNKRSTI TSREIQTAVR LLLPGELAKH AVSEGTKAVT KYTSAK

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Macromolecule #4: Telomerase RNA component

MacromoleculeName: Telomerase RNA component / type: rna / ID: 4
Source (natural)Organism: Homo sapiens (human)
SequenceString: GGGUUGCGGA GGGUGGGCCU GGGAGGGGUG GUGGCCAUUU UUUGUCUAAC CCUAACUGAG AAGGGCGUAG GCGCCGUGC UUUUGCUCCC CGCGCGCUGU UUUUCUCGCU GACUUUCAGC GGGCGGAAAA GCCUCGGCCU G CCGCCUUC CACCGUUCAU UCUAGAGCAA ...String:
GGGUUGCGGA GGGUGGGCCU GGGAGGGGUG GUGGCCAUUU UUUGUCUAAC CCUAACUGAG AAGGGCGUAG GCGCCGUGC UUUUGCUCCC CGCGCGCUGU UUUUCUCGCU GACUUUCAGC GGGCGGAAAA GCCUCGGCCU G CCGCCUUC CACCGUUCAU UCUAGAGCAA ACAAAAAAUG UCAGCUGCUG GCCCGUUCGC CCCUCCCGGG GA CCUGCGG CGGGUCGCCU GCCCAGCCCC CGAACCCCGC CUGGAGGCCG CGGUCGGCCC GGGGCUUCUC CGG AGGCAC CCACUGCCAC CGCGAAGAGU UGGGCUCUGU CAGCCGCGGG UCUCUCGGGG GCGAGGGCGA GGUU CAGGC CUUUCAGGCC GCAGGAAGAG GAACGGAGCG AGUCCCCGCG CGCGGCGCGA UUCCCUGAGC UGUGG GACG UGCACCCAGG ACUCGGCUCA CACAUGC

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Macromolecule #5: Primer DNA

MacromoleculeName: Primer DNA / type: dna / ID: 5 / Classification: DNA
Source (natural)Organism: Homo sapiens (human)
SequenceString:
TTTTTTTTTT TTTTTTTTTT AGGG

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 7.9
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 62.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionResolution.type: BY AUTHOR / Resolution: 4.97 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 50600
Initial angle assignmentType: COMMON LINE
Final angle assignmentType: MAXIMUM LIKELIHOOD

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