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- EMDB-32952: Structure of the phosphorylation-site double mutant S431A/T432A o... -

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Basic information

Entry
Database: EMDB / ID: EMD-32952
TitleStructure of the phosphorylation-site double mutant S431A/T432A of the KaiC circadian clock protein
Map data
Sample
  • Complex: KaiC hexamer
    • Protein or peptide: Circadian clock oscillator protein KaiC
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE
  • Ligand: MAGNESIUM ION
Keywordsmutant / CIRCADIAN CLOCK PROTEIN
Function / homology
Function and homology information


regulation of phosphorelay signal transduction system / negative regulation of circadian rhythm / entrainment of circadian clock / protein serine/threonine/tyrosine kinase activity / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / circadian rhythm / non-specific serine/threonine protein kinase / protein serine kinase activity / protein serine/threonine kinase activity / regulation of DNA-templated transcription ...regulation of phosphorelay signal transduction system / negative regulation of circadian rhythm / entrainment of circadian clock / protein serine/threonine/tyrosine kinase activity / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / circadian rhythm / non-specific serine/threonine protein kinase / protein serine kinase activity / protein serine/threonine kinase activity / regulation of DNA-templated transcription / magnesium ion binding / ATP hydrolysis activity / DNA binding / ATP binding / identical protein binding
Similarity search - Function
Circadian clock KaiC, bacteria / : / Circadian clock protein kinase KaiC / : / : / KaiC domain / KaiC domain profile. / KaiC-like domain / KaiC / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Circadian clock oscillator protein KaiC
Similarity search - Component
Biological speciesSynechococcus elongatus PCC 7942 = FACHB-805 (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsHan X / Zhang DL / Hong L / Yu DQ / Wu ZL / Yang T / Rust MJ / Tu YH / Ouyang Q
Funding support China, 1 items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)12090054 China
CitationJournal: Nat Commun / Year: 2023
Title: Determining subunit-subunit interaction from statistics of cryo-EM images: observation of nearest-neighbor coupling in a circadian clock protein complex
Authors: Han X / Zhang D / Hong L / Yu D / Wu Z / Yang T / Rust M / Tu Y / Ouyang Q
History
DepositionFeb 25, 2022-
Header (metadata) releaseApr 26, 2023-
Map releaseApr 26, 2023-
UpdateNov 8, 2023-
Current statusNov 8, 2023Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_32952.png

Downloads & links

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Map

FileDownload / File: emd_32952.map.gz / Format: CCP4 / Size: 91.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.69 Å/pix.
x 288 pix.
= 197.28 Å		0.69 Å/pix.
x 288 pix.
= 197.28 Å		0.69 Å/pix.
x 288 pix.
= 197.28 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.685 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.01
Minimum - Maximum-0.031022402 - 0.05920836
Average (Standard dev.)0.000020200572 (±0.0029225615)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions288288288
Spacing288288288
CellA=B=C: 197.28 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_32952_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_32952_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : KaiC hexamer

EntireName: KaiC hexamer
Components
  • Complex: KaiC hexamer
    • Protein or peptide: Circadian clock oscillator protein KaiC
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE
  • Ligand: MAGNESIUM ION

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Supramolecule #1: KaiC hexamer

SupramoleculeName: KaiC hexamer / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Synechococcus elongatus PCC 7942 = FACHB-805 (bacteria)
Strain: PCC 7942

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Macromolecule #1: Circadian clock oscillator protein KaiC

MacromoleculeName: Circadian clock oscillator protein KaiC / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Enantiomer: LEVO
Source (natural)Organism: Synechococcus elongatus PCC 7942 = FACHB-805 (bacteria)
Strain: PCC 7942
Molecular weightTheoretical: 52.757922 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: EHQAIAKMRT MIEGFDDISH GGLPIGRSTL VSGTSGTGKT LFSIQFLYNG IIEFDEPGVF VTFEETPQDI IKNARSFGWD LAKLVDEGK LFILDASPDP EGQEVVGGFD LSALIERINY AIQKYRARRV SIDSVTSVFQ QYDASSVVRR ELFRLVARLK Q IGATTVMT ...String:
EHQAIAKMRT MIEGFDDISH GGLPIGRSTL VSGTSGTGKT LFSIQFLYNG IIEFDEPGVF VTFEETPQDI IKNARSFGWD LAKLVDEGK LFILDASPDP EGQEVVGGFD LSALIERINY AIQKYRARRV SIDSVTSVFQ QYDASSVVRR ELFRLVARLK Q IGATTVMT TERIEEYGPI ARYGVEEFVS DNVVILRNVL EGERRRRTLE ILKLRGTSHM KGEYPFTITD HGINIFPLGA MR LTQRSSN VRVSSGVVRL DEMCGGGFFK DSIILATGAT GTGKTLLVSR FVENACANKE RAILFAYEES RAQLLRNAYS WGM DFEEME RQNLLKIVCA YPESAGLEDH LQIIKSEIND FKPARIAIDS LSALARGVSN NAFRQFVIGV TGYAKQEEIT GLFT NTSDQ FMGAHSITDS HIAAITDTII LLQYVEIRGE MSRAINVFKM RGSWHDKAIR EFMISDKGPD IKDSFR

UniProtKB: Circadian clock oscillator protein KaiC

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Macromolecule #2: ADENOSINE-5'-TRIPHOSPHATE

MacromoleculeName: ADENOSINE-5'-TRIPHOSPHATE / type: ligand / ID: 2 / Number of copies: 12 / Formula: ATP
Molecular weightTheoretical: 507.181 Da
Chemical component information

ChemComp-ATP:
ADENOSINE-5'-TRIPHOSPHATE / ATP, energy-carrying molecule*YM

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Macromolecule #3: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 3 / Number of copies: 12 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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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

Concentration0.35 mg/mL
BufferpH: 8
GridModel: Quantifoil R2/1 / Material: COPPER / Mesh: 300
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 5.0 µm / Nominal defocus min: 0.4 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 140475
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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