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- PDB-8ow1: Cryo-EM structure of the yeast Inner kinetochore bound to a CENP-... -

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Entry
Database: PDB / ID: 8ow1
TitleCryo-EM structure of the yeast Inner kinetochore bound to a CENP-A nucleosome.
Components
  • (C0N3) x 2
  • (Centromere DNA-binding protein complex CBF3 subunit ...) x 2
  • (Inner kinetochore subunit ...) x 13
  • Centromere-binding protein 1
  • Histone H2A.1
  • Histone H2B.1
  • Histone H3-like centromeric protein CSE4
  • Histone H4
  • Suppressor of kinetochore protein 1
KeywordsCELL CYCLE / kinetochore / point centromere / CENP-A nucleosome / topological entrapment / centromeric DNA
Function / homology
Function and homology information


Cbf1-Met4-Met28 complex / regulation of sulfur metabolic process / negative regulation of kinetochore assembly / 2-micrometer circle DNA / 2-micrometer plasmid partitioning / negative regulation of meiotic DNA double-strand break formation involved in reciprocal meiotic recombination / COMA complex / maintenance of meiotic sister chromatid cohesion / RAVE complex / Iron uptake and transport ...Cbf1-Met4-Met28 complex / regulation of sulfur metabolic process / negative regulation of kinetochore assembly / 2-micrometer circle DNA / 2-micrometer plasmid partitioning / negative regulation of meiotic DNA double-strand break formation involved in reciprocal meiotic recombination / COMA complex / maintenance of meiotic sister chromatid cohesion / RAVE complex / Iron uptake and transport / CBF3 complex / regulation of transcription by galactose / regulation of sulfur amino acid metabolic process / cellular response to methylmercury / meiotic sister chromatid segregation / Mis6-Sim4 complex / vacuolar proton-transporting V-type ATPase complex assembly / FBXL7 down-regulates AURKA during mitotic entry and in early mitosis / centromere complex assembly / establishment of meiotic sister chromatid cohesion / septin ring assembly / HDMs demethylate histones / ascospore formation / HATs acetylate histones / Condensation of Prophase Chromosomes / RNA polymerase I upstream activating factor complex / attachment of spindle microtubules to kinetochore / centromeric DNA binding / CENP-A containing chromatin assembly / regulation of exit from mitosis / Antigen processing: Ubiquitination & Proteasome degradation / outer kinetochore / SUMOylation of chromatin organization proteins / protein localization to chromosome, centromeric region / establishment of mitotic sister chromatid cohesion / vacuolar acidification / kinetochore assembly / condensed chromosome, centromeric region / regulation of metabolic process / exit from mitosis / replication fork protection complex / spindle pole body / protein localization to kinetochore / RMTs methylate histone arginines / positive regulation of glucose transmembrane transport / protein neddylation / postreplication repair / mitotic intra-S DNA damage checkpoint signaling / silent mating-type cassette heterochromatin formation / mitochondrial fusion / DNA binding, bending / SCF-dependent proteasomal ubiquitin-dependent protein catabolic process / SCF ubiquitin ligase complex / mitotic spindle assembly checkpoint signaling / positive regulation of transcription by RNA polymerase I / DNA replication origin binding / nucleolar large rRNA transcription by RNA polymerase I / mitotic sister chromatid segregation / cullin family protein binding / subtelomeric heterochromatin formation / rRNA transcription / chromosome, centromeric region / DNA replication initiation / regulation of protein-containing complex assembly / endomembrane system / protein localization to CENP-A containing chromatin / negative regulation of cytoplasmic translation / CENP-A containing nucleosome / regulation of mitotic cell cycle / meiotic cell cycle / chromosome segregation / G1/S transition of mitotic cell cycle / kinetochore / DNA-binding transcription repressor activity, RNA polymerase II-specific / nucleosome assembly / structural constituent of chromatin / G2/M transition of mitotic cell cycle / nucleosome / mitotic cell cycle / chromatin organization / ubiquitin-dependent protein catabolic process / DNA-binding transcription activator activity, RNA polymerase II-specific / protein-containing complex assembly / RNA polymerase II-specific DNA-binding transcription factor binding / sequence-specific DNA binding / chromosome, telomeric region / protein dimerization activity / protein ubiquitination / DNA-binding transcription factor activity, RNA polymerase II-specific / chromatin remodeling / RNA polymerase II cis-regulatory region sequence-specific DNA binding / DNA-binding transcription factor activity / protein heterodimerization activity / cell division / DNA repair / protein-containing complex binding / regulation of DNA-templated transcription / regulation of transcription by RNA polymerase II / structural molecule activity / negative regulation of transcription by RNA polymerase II
Similarity search - Function
: / Kinetochore subunit Nkp2/Cnl2 / Cnl2/NKP2 family protein / : / : / CENPH protein / CENPU protein / Centromere DNA-binding protein complex CBF3 subunit B, C-terminal / Centromere DNA-binding protein complex CBF3 subunit B / Centromere protein O ...: / Kinetochore subunit Nkp2/Cnl2 / Cnl2/NKP2 family protein / : / : / CENPH protein / CENPU protein / Centromere DNA-binding protein complex CBF3 subunit B, C-terminal / Centromere DNA-binding protein complex CBF3 subunit B / Centromere protein O / Cenp-O kinetochore centromere component / Centromere protein Chl4/mis15/CENP-N / Kinetochore protein CHL4 like / Zn(2)-C6 fungal-type DNA-binding domain signature. / Fungal Zn(2)-Cys(6) binuclear cluster domain / Zn(2)-C6 fungal-type DNA-binding domain superfamily / Zn(2)-C6 fungal-type DNA-binding domain profile. / GAL4-like Zn(II)2Cys6 (or C6 zinc) binuclear cluster DNA-binding domain / Zn(2)-C6 fungal-type DNA-binding domain / SKP1 component, dimerisation / S-phase kinase-associated protein 1 / SKP1-like, dimerisation domain superfamily / Skp1 family, dimerisation domain / Helix-loop-helix DNA-binding domain / S-phase kinase-associated protein 1-like / SKP1 component, POZ domain / Skp1 family, tetramerisation domain / Found in Skp1 protein family / Helix-loop-helix DNA-binding domain superfamily / helix loop helix domain / Myc-type, basic helix-loop-helix (bHLH) domain / Myc-type, basic helix-loop-helix (bHLH) domain profile. / SKP1/BTB/POZ domain superfamily / Histone H2B signature. / Histone H2B / Histone H2B / Histone H2A conserved site / Histone H2A signature. / Histone H2A, C-terminal domain / C-terminus of histone H2A / Histone H2A / Histone 2A / Histone H4, conserved site / Histone H4 signature. / Histone H4 / Histone H4 / CENP-T/Histone H4, histone fold / Centromere kinetochore component CENP-T histone fold / Leucine-rich repeat domain superfamily / Histone H3 signature 2. / Histone H3 / Histone H3/CENP-A / Histone H2A/H2B/H3 / Core histone H2A/H2B/H3/H4 / Histone-fold
Similarity search - Domain/homology
DNA / DNA (> 10) / DNA (> 100) / Histone H2B.1 / Histone H4 / Histone H2A.1 / Centromere-binding protein 1 / Centromere DNA-binding protein complex CBF3 subunit C / Histone H3-like centromeric protein CSE4 / Inner kinetochore subunit IML3 ...DNA / DNA (> 10) / DNA (> 100) / Histone H2B.1 / Histone H4 / Histone H2A.1 / Centromere-binding protein 1 / Centromere DNA-binding protein complex CBF3 subunit C / Histone H3-like centromeric protein CSE4 / Inner kinetochore subunit IML3 / Inner kinetochore subunit AME1 / Inner kinetochore subunit CHL4 / Centromere DNA-binding protein complex CBF3 subunit B / Inner kinetochore subunit CNN1 / Inner kinetochore subunit MCM22 / Suppressor of kinetochore protein 1 / Inner kinetochore subunit OKP1 / Inner kinetochore subunit CTF19 / Inner kinetochore subunit NKP2 / Inner kinetochore subunit MCM21 / Inner kinetochore subunit MCM16 / Inner kinetochore subunit NKP1 / Inner kinetochore subunit CTF3 / Inner kinetochore subunit WIP1
Similarity search - Component
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsDendooven, T.D. / Zhang, Z. / Yang, J. / McLaughlin, S. / Schwabb, J. / Scheres, S. / Yatskevich, S. / Barford, D.
Funding support United Kingdom, Germany, 4items
OrganizationGrant numberCountry
UK Research and Innovation (UKRI)MC_UP_1201/6 United Kingdom
Cancer Research UKC576/A14109 United Kingdom
UK Research and Innovation (UKRI)MC_UP_ A025_1013 United Kingdom
Boehringer Ingelheim Fonds (BIF) Germany
CitationJournal: Sci Adv / Year: 2023
Title: Cryo-EM structure of the complete inner kinetochore of the budding yeast point centromere.
Authors: Tom Dendooven / Ziguo Zhang / Jing Yang / Stephen H McLaughlin / Johannes Schwab / Sjors H W Scheres / Stanislau Yatskevich / David Barford /
Abstract: The point centromere of budding yeast specifies assembly of the large kinetochore complex to mediate chromatid segregation. Kinetochores comprise the centromere-associated inner kinetochore (CCAN) ...The point centromere of budding yeast specifies assembly of the large kinetochore complex to mediate chromatid segregation. Kinetochores comprise the centromere-associated inner kinetochore (CCAN) complex and the microtubule-binding outer kinetochore KNL1-MIS12-NDC80 (KMN) network. The budding yeast inner kinetochore also contains the DNA binding centromere-binding factor 1 (CBF1) and CBF3 complexes. We determined the cryo-electron microscopy structure of the yeast inner kinetochore assembled onto the centromere-specific centromere protein A nucleosomes (CENP-A). This revealed a central CENP-A with extensively unwrapped DNA ends. These free DNA duplexes bind two CCAN protomers, one of which entraps DNA topologically, positioned on the centromere DNA element I (CDEI) motif by CBF1. The two CCAN protomers are linked through CBF3 forming an arch-like configuration. With a structural mechanism for how CENP-A can also be linked to KMN involving only CENP-QU, we present a model for inner kinetochore assembly onto a point centromere and how it organizes the outer kinetochore for chromosome attachment to the mitotic spindle.
History
DepositionApr 26, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Aug 9, 2023Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Centromere-binding protein 1
B: Centromere-binding protein 1
I: Inner kinetochore subunit CTF3
K: Inner kinetochore subunit MCM22
H: Inner kinetochore subunit MCM16
LL: Inner kinetochore subunit IML3
NN: Inner kinetochore subunit CHL4
CT: Centromere DNA-binding protein complex CBF3 subunit C
ce: Centromere DNA-binding protein complex CBF3 subunit B
L: Inner kinetochore subunit IML3
SK: Suppressor of kinetochore protein 1
CE: Centromere DNA-binding protein complex CBF3 subunit B
P: Inner kinetochore subunit CTF19
Q: Inner kinetochore subunit OKP1
OO: Inner kinetochore subunit MCM21
N: Inner kinetochore subunit CHL4
T: Inner kinetochore subunit CNN1
U: Inner kinetochore subunit AME1
PP: Inner kinetochore subunit CTF19
W: Inner kinetochore subunit WIP1
O: Inner kinetochore subunit MCM21
Y: Inner kinetochore subunit NKP1
Z: Inner kinetochore subunit NKP2
UU: Inner kinetochore subunit AME1
YY: Inner kinetochore subunit NKP1
ZZ: Inner kinetochore subunit NKP2
II: Inner kinetochore subunit CTF3
KK: Inner kinetochore subunit MCM22
HH: Inner kinetochore subunit MCM16
QQ: Inner kinetochore subunit OKP1
D: C0N3
E: C0N3
b: Histone H4
d: Histone H2B.1
e: Histone H3-like centromeric protein CSE4
f: Histone H4
g: Histone H2A.1
h: Histone H2B.1
c: Histone H2A.1
a: Histone H3-like centromeric protein CSE4
TT: Inner kinetochore subunit CNN1
WW: Inner kinetochore subunit WIP1


Theoretical massNumber of molelcules
Total (without water)1,490,63342
Polymers1,490,63342
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: light scattering
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area236670 Å2
ΔGint-1564 kcal/mol
Surface area452540 Å2
MethodPISA

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Components

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Protein , 6 types, 11 molecules ABSKbfdheagc

#1: Protein Centromere-binding protein 1 / CBP-1 / Centromere promoter factor 1 / Centromere-binding factor 1


Mass: 39444.715 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: CBF1, CEP1, CP1, CPF1, YJR060W, J1730 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P17106
#9: Protein Suppressor of kinetochore protein 1 / Centromere DNA-binding protein complex CBF3 subunit D / E3 ubiquitin ligase complex SCF subunit SKP1


Mass: 22357.270 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: SKP1, CBF3D, YDR328C, D9798.14 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P52286
#20: Protein Histone H4 /


Mass: 11395.390 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: HHF1, YBR009C, YBR0122, HHF2, YNL030W, N2752 / Production host: Escherichia coli (E. coli) / References: UniProt: P02309
#21: Protein Histone H2B.1 / Suppressor of Ty protein 12


Mass: 14280.362 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: HTB1, H2B1, SPT12, YDR224C, YD9934.09C / Production host: Escherichia coli (E. coli) / References: UniProt: P02293
#22: Protein Histone H3-like centromeric protein CSE4 / CENP-A homolog / CENPA homolog / Chromosome segregation protein 4


Mass: 26885.434 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: CSE4, CSL2, YKL049C, YKL262 / Production host: Escherichia coli (E. coli) / References: UniProt: P36012
#23: Protein Histone H2A.1


Mass: 14013.177 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: HTA1, H2A1, SPT11, YDR225W, YD9934.10 / Production host: Escherichia coli (E. coli) / References: UniProt: P04911

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Inner kinetochore subunit ... , 13 types, 26 molecules IIIKKKHHHLLLNNNPPPQQQOOOTTTUUUWWWYYYZZZ

#2: Protein Inner kinetochore subunit CTF3 / CENP-I homolog / Chromosome loss protein 3 / Chromosome transmission fidelity protein 3 / ...CENP-I homolog / Chromosome loss protein 3 / Chromosome transmission fidelity protein 3 / Constitutive centromere-associated network protein CTF3


Mass: 84345.633 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: CTF3, CHL3, YLR381W / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q12748
#3: Protein Inner kinetochore subunit MCM22 / CENP-K homolog / Constitutive centromere-associated network protein MCM22 / Minichromosome ...CENP-K homolog / Constitutive centromere-associated network protein MCM22 / Minichromosome maintenance protein 22


Mass: 27602.541 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: MCM22, YJR135C, J2122 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P47167
#4: Protein Inner kinetochore subunit MCM16 / CENP-H homolog / Constitutive centromere-associated network protein MCM16 / Minichromosome ...CENP-H homolog / Constitutive centromere-associated network protein MCM16 / Minichromosome maintenance protein 16


Mass: 21166.100 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: MCM16, YPR046W / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q12262
#5: Protein Inner kinetochore subunit IML3 / CENP-L homolog / Constitutive centromere-associated network protein IML3 / Increased minichromosome ...CENP-L homolog / Constitutive centromere-associated network protein IML3 / Increased minichromosome loss protein 3 / Minichromosome maintenance protein 19


Mass: 28093.223 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: IML3, MCM19, YBR107C, YBR0836 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P38265
#6: Protein Inner kinetochore subunit CHL4 / CENP-N homolog / Chromosome loss protein 4 / Chromosome transmission fidelity protein 17 / ...CENP-N homolog / Chromosome loss protein 4 / Chromosome transmission fidelity protein 17 / Constitutive centromere-associated network protein CHL4 / Minichromosome maintenance protein 17


Mass: 52743.723 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: CHL4, CTF17, MCM17, YDR254W, YD9320A.04 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P38907
#10: Protein Inner kinetochore subunit CTF19 / CENP-P homolog / Chromosome transmission fidelity protein 19 / Constitutive centromere-associated ...CENP-P homolog / Chromosome transmission fidelity protein 19 / Constitutive centromere-associated network protein CTF19 / Minichromosome maintenance protein 18


Mass: 42841.113 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: CTF19, MCM18, YPL018W, LPB13W / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q02732
#11: Protein Inner kinetochore subunit OKP1 / CENP-Q homolog / Constitutive centromere-associated network protein OKP1 / Outer kinetochore protein 1


Mass: 47427.246 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: OKP1, YGR179C / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P53298
#12: Protein Inner kinetochore subunit MCM21 / CENP-O homolog / Chromosome transmission fidelity protein 5 / Constitutive centromere-associated ...CENP-O homolog / Chromosome transmission fidelity protein 5 / Constitutive centromere-associated network protein MCM21 / Minichromosome maintenance protein 21


Mass: 43028.879 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: MCM21, CTF5, YDR318W / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q06675
#13: Protein Inner kinetochore subunit CNN1 / CENP-T homolog / Co-purified with NNF1 protein 1 / Constitutive centromere-associated network protein CNN1


Mass: 41359.785 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: CNN1, YFR046C / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P43618
#14: Protein Inner kinetochore subunit AME1 / Associated with microtubules and essential protein 1 / CENP-U homolog / Constitutive centromere- ...Associated with microtubules and essential protein 1 / CENP-U homolog / Constitutive centromere-associated network protein AME1


Mass: 37506.723 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: AME1, ARP100, YBR211C, YBR1458 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P38313
#15: Protein Inner kinetochore subunit WIP1 / CENP-W homolog / Constitutive centromere-associated network protein WIP1 / W-like protein 1


Mass: 10255.458 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: WIP1, YDR374W-A / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q2V2P8
#16: Protein Inner kinetochore subunit NKP1 / Constitutive centromere-associated network protein NKP1 / Non-essential kinetochore protein 1


Mass: 27006.451 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: NKP1, YDR383C / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q12493
#17: Protein Inner kinetochore subunit NKP2 / Constitutive centromere-associated network protein NKP2 / Non-essential kinetochore protein 2


Mass: 17877.033 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: NKP2, YLR315W / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q06162

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Centromere DNA-binding protein complex CBF3 subunit ... , 2 types, 3 molecules CTceCE

#7: Protein Centromere DNA-binding protein complex CBF3 subunit C / Chromosome transmission fidelity protein 13 / Kinetochore protein CTF13


Mass: 56416.863 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: CTF13, CBF3C, YMR094W, YM6543.01, YM9582.19 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P35203
#8: Protein Centromere DNA-binding protein complex CBF3 subunit B / Centromere protein 3


Mass: 71439.891 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: CEP3, CBF3, CBF3B, CSL1, YMR168C, YM8520.17C / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P40969

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DNA chain , 2 types, 2 molecules DE

#18: DNA chain C0N3


Mass: 47239.277 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Saccharomyces cerevisiae (brewer's yeast)
#19: DNA chain C0N3


Mass: 47194.199 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Saccharomyces cerevisiae (brewer's yeast)

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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: A complex of CBF1-CCAN bound to centromeric C0N3 DNA / Type: COMPLEX
Entity ID: #1-#5, #7, #9, #8, #10-#11, #6, #13-#15, #12, #16-#23
Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
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: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: SPOT SCAN
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2600 nm / Nominal defocus min: 1000 nm
Image recordingElectron dose: 40 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 108672 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00380835
ELECTRON MICROSCOPYf_angle_d0.698110508
ELECTRON MICROSCOPYf_dihedral_angle_d22.66812800
ELECTRON MICROSCOPYf_chiral_restr0.03812616
ELECTRON MICROSCOPYf_plane_restr0.00512969

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