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- PDB-6tlj: Cryo-EM structure of the Anaphase-promoting complex/Cyclosome, in... -

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Entry
Database: PDB / ID: 6tlj
TitleCryo-EM structure of the Anaphase-promoting complex/Cyclosome, in complex with the Mitotic checkpoint complex (APC/C-MCC) at 3.8 angstrom resolution
Components
  • (Anaphase-promoting complex subunit ...) x 11
  • (Cell division cycle protein ...) x 5
  • Mitotic checkpoint serine/threonine-protein kinase BUB1 beta
  • Mitotic spindle assembly checkpoint protein MAD2A
KeywordsCELL CYCLE / E3 ligase / Complex / Mitotic checkpoint complex
Function / homology
Function and homology information


mitotic spindle assembly checkpoint MAD1-MAD2 complex / metaphase/anaphase transition of cell cycle / metaphase/anaphase transition of meiosis I / Inhibition of the proteolytic activity of APC/C required for the onset of anaphase by mitotic spindle checkpoint components / positive regulation of ubiquitin protein ligase activity / negative regulation of ubiquitin protein ligase activity / positive regulation of anaphase-promoting complex-dependent catabolic process / positive regulation of synapse maturation / regulation of mitotic cell cycle spindle assembly checkpoint / Conversion from APC/C:Cdc20 to APC/C:Cdh1 in late anaphase ...mitotic spindle assembly checkpoint MAD1-MAD2 complex / metaphase/anaphase transition of cell cycle / metaphase/anaphase transition of meiosis I / Inhibition of the proteolytic activity of APC/C required for the onset of anaphase by mitotic spindle checkpoint components / positive regulation of ubiquitin protein ligase activity / negative regulation of ubiquitin protein ligase activity / positive regulation of anaphase-promoting complex-dependent catabolic process / positive regulation of synapse maturation / regulation of mitotic cell cycle spindle assembly checkpoint / Conversion from APC/C:Cdc20 to APC/C:Cdh1 in late anaphase / meiotic sister chromatid cohesion, centromeric / Inactivation of APC/C via direct inhibition of the APC/C complex / APC/C:Cdc20 mediated degradation of mitotic proteins / regulation of dendrite development / anaphase-promoting complex / Phosphorylation of Emi1 / anaphase-promoting complex-dependent catabolic process / metaphase/anaphase transition of mitotic cell cycle / Aberrant regulation of mitotic exit in cancer due to RB1 defects / regulation of meiotic cell cycle / protein branched polyubiquitination / regulation of exit from mitosis / anaphase-promoting complex binding / Phosphorylation of the APC/C / negative regulation of mitotic cell cycle / nuclear pore nuclear basket / outer kinetochore / positive regulation of synaptic plasticity / positive regulation of dendrite morphogenesis / positive regulation of mitotic metaphase/anaphase transition / positive regulation of mitotic cell cycle spindle assembly checkpoint / protein K11-linked ubiquitination / ubiquitin ligase activator activity / regulation of mitotic metaphase/anaphase transition / ubiquitin-ubiquitin ligase activity / mitotic spindle assembly checkpoint signaling / intercellular bridge / Regulation of APC/C activators between G1/S and early anaphase / Transcriptional Regulation by VENTX / cullin family protein binding / mitotic metaphase chromosome alignment / enzyme-substrate adaptor activity / positive regulation of axon extension / ubiquitin-like ligase-substrate adaptor activity / heterochromatin / protein K48-linked ubiquitination / Amplification of signal from unattached kinetochores via a MAD2 inhibitory signal / nuclear periphery / Mitotic Prometaphase / EML4 and NUDC in mitotic spindle formation / regulation of mitotic cell cycle / APC/C:Cdc20 mediated degradation of Cyclin B / APC-Cdc20 mediated degradation of Nek2A / Resolution of Sister Chromatid Cohesion / brain development / Autodegradation of Cdh1 by Cdh1:APC/C / APC/C:Cdc20 mediated degradation of Securin / negative regulation of protein catabolic process / SCF-beta-TrCP mediated degradation of Emi1 / Assembly of the pre-replicative complex / Cdc20:Phospho-APC/C mediated degradation of Cyclin A / RHO GTPases Activate Formins / APC/C:Cdh1 mediated degradation of Cdc20 and other APC/C:Cdh1 targeted proteins in late mitosis/early G1 / G protein-coupled receptor binding / CDK-mediated phosphorylation and removal of Cdc6 / mitotic spindle / spindle / histone deacetylase binding / neuron projection development / kinetochore / spindle pole / ubiquitin-protein transferase activity / Separation of Sister Chromatids / microtubule cytoskeleton / ubiquitin protein ligase activity / mitotic cell cycle / nervous system development / Antigen processing: Ubiquitination & Proteasome degradation / Senescence-Associated Secretory Phenotype (SASP) / protein phosphatase binding / molecular adaptor activity / ubiquitin-dependent protein catabolic process / cell differentiation / protein kinase activity / non-specific serine/threonine protein kinase / Ub-specific processing proteases / protein ubiquitination / negative regulation of gene expression / protein serine kinase activity / cell division / protein serine/threonine kinase activity / centrosome / positive regulation of cell population proliferation / ubiquitin protein ligase binding / apoptotic process / nucleolus / perinuclear region of cytoplasm / protein homodimerization activity / nucleoplasm / zinc ion binding
Similarity search - Function
: / Mad3/Bub1 homology region 1 / Mitotic spindle checkpoint protein Bub1/Mad3 / Mad3/BUB1 homology region 1 / BUB1 N-terminal domain profile. / Mad3/BUB1 hoMad3/BUB1 homology region 1 / Anaphase-promoting complex subunit 15 / Anaphase-promoting complex subunit 15 / : / CDC20/Fizzy WD40 domain ...: / Mad3/Bub1 homology region 1 / Mitotic spindle checkpoint protein Bub1/Mad3 / Mad3/BUB1 homology region 1 / BUB1 N-terminal domain profile. / Mad3/BUB1 hoMad3/BUB1 homology region 1 / Anaphase-promoting complex subunit 15 / Anaphase-promoting complex subunit 15 / : / CDC20/Fizzy WD40 domain / Cell Cycle, Spindle Assembly Checkpoint Protein; Chain A / HORMA domain / The WD repeat Cdc20/Fizzy family / : / APC4, WD40 domain C-terminal half / Anaphase-promoting complex subunit 4, metazoa / Anaphase-promoting complex subunit 1, C-terminal / Anaphase-promoting complex subunit 1, middle domain / : / : / : / Anaphase-promoting complex subunit 1 WD40 beta-propeller domain / Anaphase-promoting complex sub unit 1 C-terminal domain / Anaphase-promoting complex subunit 1 middle domain / APC1 beta sandwich domain / Anaphase-promoting complex subunit 5, N-terminal domain / Anaphase-promoting complex subunit 16 / Anaphase-promoting complex, subunit 16 / Cdc23 / Apc13 / Anaphase promoting complex subunit 8 / Cdc23 / Apc13p protein / Anaphase-promoting complex subunit 4 / Anaphase-promoting complex subunit 4 long domain / Anaphase-promoting complex subunit 1 / Anaphase-promoting complex subunit 5 / Anaphase-promoting complex, cyclosome, subunit 4 / Anaphase-promoting complex subunit APC10/Doc1 / Anaphase-promoting complex, subunit CDC26 / Anaphase-promoting complex APC subunit CDC26 / Anaphase-promoting complex subunit 11, RING-H2 finger / Anaphase-promoting complex subunit 5 domain / Anaphase-promoting complex subunit 11 RING-H2 finger / Anaphase-promoting complex subunit 5 / Anaphase-promoting complex subunit 2, C-terminal / Anaphase-promoting complex subunit 2 / : / Anaphase promoting complex (APC) subunit 2 / ANAPC2-like, TPR repeats / Anaphase promoting complex (APC) subunit 2 / Mad2-like / HORMA domain / HORMA domain / HORMA domain profile. / APC10/DOC domain / Anaphase-promoting complex, subunit 10 (APC10) / DOC domain profile. / Anaphase-promoting complex, subunit 10 (APC10) / HORMA domain superfamily / Anaphase-promoting complex, cyclosome, subunit 3 / TPR repeat / Anaphase-promoting complex subunit 4, WD40 domain / Anaphase-promoting complex subunit 4 WD40 domain / Tetratricopeptide repeat / : / Tetratricopeptide repeat / Cullin alpha solenoid domain / Cullin / Tetratricopeptide repeat / : / Cullin alpha+beta domain / Cullin homology domain / Cullin homology domain superfamily / Cullin family profile. / Tetratricopeptide repeat domain / Zinc/RING finger domain, C3HC4 (zinc finger) / Herpes Virus-1 / Tetratricopeptide repeat / Galactose-binding domain-like / TPR repeat region circular profile. / TPR repeat profile. / Tetratricopeptide repeats / YVTN repeat-like/Quinoprotein amine dehydrogenase / Tetratricopeptide repeat / 7 Propeller / Methylamine Dehydrogenase; Chain H / Galactose-binding-like domain superfamily / Zinc finger RING-type profile. / Serine Threonine Protein Phosphatase 5, Tetratricopeptide repeat / Zinc finger, RING-type / Alpha Horseshoe / Armadillo-like helical / Tetratricopeptide-like helical domain superfamily / WD domain, G-beta repeat / Zinc finger, RING/FYVE/PHD-type / Winged helix DNA-binding domain superfamily / Jelly Rolls / Trp-Asp (WD) repeats signature. / Trp-Asp (WD) repeats profile. / Trp-Asp (WD) repeats circular profile.
Similarity search - Domain/homology
Mitotic checkpoint serine/threonine-protein kinase BUB1 beta / Cell division cycle protein 27 homolog / Anaphase-promoting complex subunit 15 / Cell division cycle protein 20 homolog / Cell division cycle protein 16 homolog / Mitotic spindle assembly checkpoint protein MAD2A / Anaphase-promoting complex subunit CDC26 / Anaphase-promoting complex subunit 16 / Anaphase-promoting complex subunit 13 / Anaphase-promoting complex subunit 1 ...Mitotic checkpoint serine/threonine-protein kinase BUB1 beta / Cell division cycle protein 27 homolog / Anaphase-promoting complex subunit 15 / Cell division cycle protein 20 homolog / Cell division cycle protein 16 homolog / Mitotic spindle assembly checkpoint protein MAD2A / Anaphase-promoting complex subunit CDC26 / Anaphase-promoting complex subunit 16 / Anaphase-promoting complex subunit 13 / Anaphase-promoting complex subunit 1 / Anaphase-promoting complex subunit 11 / Cell division cycle protein 23 homolog / Anaphase-promoting complex subunit 7 / Anaphase-promoting complex subunit 5 / Anaphase-promoting complex subunit 4 / Anaphase-promoting complex subunit 2 / Anaphase-promoting complex subunit 10
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.8 Å
AuthorsAlfieri, C. / Barford, D.
Funding support United Kingdom, 1items
OrganizationGrant numberCountry
Cancer Research UK United Kingdom
CitationJournal: EMBO Rep / Year: 2020
Title: A unique binding mode of Nek2A to the APC/C allows its ubiquitination during prometaphase.
Authors: Claudio Alfieri / Thomas Tischer / David Barford /
Abstract: The anaphase-promoting complex (APC/C) is the key E3 ubiquitin ligase which directs mitotic progression and exit by catalysing the sequential ubiquitination of specific substrates. The activity of ...The anaphase-promoting complex (APC/C) is the key E3 ubiquitin ligase which directs mitotic progression and exit by catalysing the sequential ubiquitination of specific substrates. The activity of the APC/C in mitosis is restrained by the spindle assembly checkpoint (SAC), which coordinates chromosome segregation with the assembly of the mitotic spindle. The SAC effector is the mitotic checkpoint complex (MCC), which binds and inhibits the APC/C. It is incompletely understood how the APC/C switches substrate specificity in a cell cycle-specific manner. For instance, it is unclear how in prometaphase, when APC/C activity towards cyclin B and securin is repressed by the MCC, the kinase Nek2A is ubiquitinated. Here, we combine biochemical and structural analysis with functional studies in cells to show that Nek2A is a conformational-specific binder of the APC/C-MCC complex (APC/C ) and that, in contrast to cyclin A, Nek2A can be ubiquitinated efficiently by the APC/C in conjunction with both the E2 enzymes UbcH10 and UbcH5. We propose that these special features of Nek2A allow its prometaphase-specific ubiquitination.
History
DepositionDec 2, 2019Deposition site: PDBE / Processing site: PDBE
Revision 1.0Feb 19, 2020Provider: repository / Type: Initial release
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Revision 1.1Apr 29, 2020Group: Database references / Category: citation / citation_author
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Revision 1.3Oct 9, 2024Group: Data collection / Database references / Structure summary
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Assembly

Deposited unit
A: Anaphase-promoting complex subunit 1
B: Anaphase-promoting complex subunit 11
C: Cell division cycle protein 23 homolog
D: Anaphase-promoting complex subunit 15
E: Anaphase-promoting complex subunit 16
F: Cell division cycle protein 27 homolog
G: Anaphase-promoting complex subunit CDC26
H: Cell division cycle protein 27 homolog
I: Anaphase-promoting complex subunit 4
J: Cell division cycle protein 16 homolog
K: Cell division cycle protein 16 homolog
L: Anaphase-promoting complex subunit 10
M: Anaphase-promoting complex subunit 13
N: Anaphase-promoting complex subunit 2
O: Anaphase-promoting complex subunit 5
P: Cell division cycle protein 23 homolog
Q: Cell division cycle protein 20 homolog
R: Cell division cycle protein 20 homolog
S: Mitotic checkpoint serine/threonine-protein kinase BUB1 beta
W: Anaphase-promoting complex subunit CDC26
X: Anaphase-promoting complex subunit 7
Y: Anaphase-promoting complex subunit 7
Z: Mitotic spindle assembly checkpoint protein MAD2A


Theoretical massNumber of molelcules
Total (without water)1,411,71223
Polymers1,411,71223
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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Anaphase-promoting complex subunit ... , 11 types, 13 molecules ABDEGWILMNOXY

#1: Protein Anaphase-promoting complex subunit 1 / APC1 / Cyclosome subunit 1 / Mitotic checkpoint regulator / Testis-specific gene 24 protein


Mass: 216775.516 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: ANAPC1, TSG24 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9H1A4
#2: Protein Anaphase-promoting complex subunit 11 / APC11 / Cyclosome subunit 11 / Hepatocellular carcinoma-associated RING finger protein


Mass: 9854.647 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: ANAPC11, HSPC214 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9NYG5
#4: Protein Anaphase-promoting complex subunit 15 / APC15


Mass: 14286.727 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: ANAPC15, C11orf51, HSPC020 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P60006
#5: Protein Anaphase-promoting complex subunit 16 / APC16 / Cyclosome subunit 16


Mass: 11677.995 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: ANAPC16, C10orf104, CENP-27 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q96DE5
#7: Protein Anaphase-promoting complex subunit CDC26 / Anaphase-promoting complex subunit 12 / APC12 / Cell division cycle protein 26 homolog


Mass: 9793.999 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CDC26, ANAPC12, C9orf17 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q8NHZ8
#8: Protein Anaphase-promoting complex subunit 4 / APC4 / Cyclosome subunit 4


Mass: 92219.227 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: ANAPC4, APC4 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9UJX5
#10: Protein Anaphase-promoting complex subunit 10 / APC10 / Cyclosome subunit 10


Mass: 21282.143 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: ANAPC10, APC10 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9UM13
#11: Protein Anaphase-promoting complex subunit 13 / APC13 / Cyclosome subunit 13


Mass: 8528.309 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: ANAPC13 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9BS18
#12: Protein Anaphase-promoting complex subunit 2 / APC2 / Cyclosome subunit 2


Mass: 93938.977 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: ANAPC2, APC2, KIAA1406 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9UJX6
#13: Protein Anaphase-promoting complex subunit 5 / APC5 / Cyclosome subunit 5


Mass: 85179.766 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: ANAPC5, APC5 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9UJX4
#17: Protein Anaphase-promoting complex subunit 7 / APC7 / Cyclosome subunit 7


Mass: 66929.367 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: ANAPC7, APC7 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9UJX3

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Cell division cycle protein ... , 5 types, 8 molecules CPFHJKQR

#3: Protein Cell division cycle protein 23 homolog / Anaphase-promoting complex subunit 8 / APC8 / Cyclosome subunit 8


Mass: 68921.031 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CDC23, ANAPC8 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9UJX2
#6: Protein Cell division cycle protein 27 homolog / Anaphase-promoting complex subunit 3 / APC3 / CDC27 homolog / CDC27Hs / H-NUC


Mass: 91973.125 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CDC27, ANAPC3, D0S1430E, D17S978E / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P30260
#9: Protein Cell division cycle protein 16 homolog / Anaphase-promoting complex subunit 6 / APC6 / CDC16 homolog / CDC16Hs / Cyclosome subunit 6


Mass: 71747.516 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CDC16, ANAPC6 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q13042
#14: Protein Cell division cycle protein 20 homolog / p55CDC


Mass: 41226.332 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CDC20 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q12834
#15: Protein Cell division cycle protein 20 homolog / p55CDC


Mass: 54796.508 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CDC20 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q12834

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Protein , 2 types, 2 molecules SZ

#16: Protein Mitotic checkpoint serine/threonine-protein kinase BUB1 beta / MAD3/BUB1-related protein kinase / hBUBR1 / Mitotic checkpoint kinase MAD3L / Protein SSK1


Mass: 119681.977 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: BUB1B, BUBR1, MAD3L, SSK1 / Production host: Trichoplusia ni (cabbage looper)
References: UniProt: O60566, non-specific serine/threonine protein kinase
#18: Protein Mitotic spindle assembly checkpoint protein MAD2A / HsMAD2 / Mitotic arrest deficient 2-like protein 1 / MAD2-like protein 1


Mass: 23533.883 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: MAD2L1, MAD2 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q13257

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Details

Has protein modificationY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Anaphase-promoting complex/Cyclosome, in complex with the Mitotic checkpoint complex (APC/C-MCC)
Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Trichoplusia ni (cabbage looper)
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company
MicroscopyModel: FEI POLARA 300
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD
Image recordingElectron dose: 27 e/Å2 / Film or detector model: FEI FALCON III (4k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.17.1_3660: / Classification: refinement
EM softwareName: PHENIX / Category: model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 343551 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00573849
ELECTRON MICROSCOPYf_angle_d0.759100076
ELECTRON MICROSCOPYf_dihedral_angle_d12.2359974
ELECTRON MICROSCOPYf_chiral_restr0.04311236
ELECTRON MICROSCOPYf_plane_restr0.00412734

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Related info.:EMDB / PDB / SASBDB / Comparison of 3 databanks / Yorodumi Search / Aug 31, 2016. New EM Navigator & Yorodumi / Yorodumi Papers / Jmol/JSmol / Function and homology information / Changes in new EM Navigator and Yorodumi

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