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- PDB-4kso: Crystal Structure of Circadian clock protein KaiB from S.Elongatus -

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Basic information

Entry
Database: PDB / ID: 4kso
TitleCrystal Structure of Circadian clock protein KaiB from S.Elongatus
ComponentsCircadian clock protein KaiB
KeywordsCIRCADIAN CLOCK PROTEIN / CYANOBACTERIAL CIRCADIAN CLOCK PROTEIN KaiB / CIRCADIAN CLOCK / KaiC Protein / Soluble
Function / homology
Function and homology information


negative regulation of phosphorylation / entrainment of circadian clock / regulation of circadian rhythm / circadian rhythm / identical protein binding / plasma membrane / cytoplasm
Similarity search - Function
Circadian clock protein KaiB / Circadian clock protein KaiB-like / KaiB domain / KaiB domain / KaiB / Glutaredoxin / Glutaredoxin / Thioredoxin-like superfamily / 3-Layer(aba) Sandwich / Alpha Beta
Similarity search - Domain/homology
Circadian clock oscillator protein KaiB
Similarity search - Component
Biological speciesSynechococcus elongatus (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.622 Å
AuthorsPattanayek, R. / Egli, M.
CitationJournal: J Mol Biol / Year: 2013
Title: CryoEM and molecular dynamics of the circadian KaiB-KaiC complex indicates that KaiB monomers interact with KaiC and block ATP binding clefts.
Authors: Seth A Villarreal / Rekha Pattanayek / Dewight R Williams / Tetsuya Mori / Ximing Qin / Carl H Johnson / Martin Egli / Phoebe L Stewart /
Abstract: The circadian control of cellular processes in cyanobacteria is regulated by a posttranslational oscillator formed by three Kai proteins. During the oscillator cycle, KaiA serves to promote ...The circadian control of cellular processes in cyanobacteria is regulated by a posttranslational oscillator formed by three Kai proteins. During the oscillator cycle, KaiA serves to promote autophosphorylation of KaiC while KaiB counteracts this effect. Here, we present a crystallographic structure of the wild-type Synechococcus elongatus KaiB and a cryo-electron microscopy (cryoEM) structure of a KaiBC complex. The crystal structure shows the expected dimer core structure and significant conformational variations of the KaiB C-terminal region, which is functionally important in maintaining rhythmicity. The KaiBC sample was formed with a C-terminally truncated form of KaiC, KaiC-Δ489, which is persistently phosphorylated. The KaiB-KaiC-Δ489 structure reveals that the KaiC hexamer can bind six monomers of KaiB, which form a continuous ring of density in the KaiBC complex. We performed cryoEM-guided molecular dynamics flexible fitting simulations with crystal structures of KaiB and KaiC to probe the KaiBC protein-protein interface. This analysis indicated a favorable binding mode for the KaiB monomer on the CII end of KaiC, involving two adjacent KaiC subunits and spanning an ATP binding cleft. A KaiC mutation, R468C, which has been shown to affect the affinity of KaiB for KaiC and lengthen the period in a bioluminescence rhythm assay, is found within the middle of the predicted KaiBC interface. The proposed KaiB binding mode blocks access to the ATP binding cleft in the CII ring of KaiC, which provides insight into how KaiB might influence the phosphorylation status of KaiC.
History
DepositionMay 17, 2013Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jul 10, 2013Provider: repository / Type: Initial release
Revision 1.1Sep 18, 2013Group: Database references
Revision 1.2Sep 20, 2023Group: Data collection / Database references / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
C: Circadian clock protein KaiB
D: Circadian clock protein KaiB
A: Circadian clock protein KaiB
B: Circadian clock protein KaiB


Theoretical massNumber of molelcules
Total (without water)45,8024
Polymers45,8024
Non-polymers00
Water1,856103
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area6690 Å2
ΔGint-41 kcal/mol
Surface area20690 Å2
MethodPISA
Unit cell
Length a, b, c (Å)69.821, 116.124, 53.228
Angle α, β, γ (deg.)90.00, 98.28, 90.00
Int Tables number5
Space group name H-MC121

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Components

#1: Protein
Circadian clock protein KaiB


Mass: 11450.387 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Synechococcus elongatus (bacteria) / Strain: PCC 7942 / Gene: kaiB / Plasmid: pGex vector / Production host: Escherichia coli (E. coli) / Strain (production host): BL21 / References: UniProt: Q79PF5
#2: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 103 / Source method: isolated from a natural source / Formula: H2O

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 2.33 Å3/Da / Density % sol: 47.23 %
Crystal growTemperature: 292 K / Method: vapor diffusion / pH: 5.7
Details: KaiB were grown from droplets containing 6.5 mg/mL KaiB-KaiC complex, 20 mM Tris (pH 7.8), 100 mM NaCl, 5 mM MgCl2,1 mM ATP and 2 mM BME. The reservoir solution was 100 mM sodium acetate, ...Details: KaiB were grown from droplets containing 6.5 mg/mL KaiB-KaiC complex, 20 mM Tris (pH 7.8), 100 mM NaCl, 5 mM MgCl2,1 mM ATP and 2 mM BME. The reservoir solution was 100 mM sodium acetate, 500 mM sodium formate and 5% glycerol (v/v), VAPOR DIFFUSION, temperature 292K

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Data collection

DiffractionMean temperature: 113 K
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 21-ID-G / Wavelength: 0.98 Å
DetectorType: MARMOSAIC 300 mm CCD / Detector: CCD / Date: Dec 14, 2012 / Details: Mirrors
RadiationMonochromator: C(111) / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.98 Å / Relative weight: 1
ReflectionResolution: 2.6→30 Å / Num. obs: 12670 / % possible obs: 100 % / Observed criterion σ(F): 2 / Rmerge(I) obs: 0.06 / Net I/σ(I): 14.6
Reflection shellResolution: 2.6→2.69 Å / Rmerge(I) obs: 0.267 / Mean I/σ(I) obs: 2.1 / % possible all: 99.9

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Processing

Software
NameVersionClassification
HKL-2000data collection
CCP4model building
PHENIX(phenix.refine: 1.8.2_1309)refinement
HKL-2000data reduction
HKL-2000data scaling
CCP4phasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 2QKE

2qke


Resolution: 2.622→29.689 Å / SU ML: 0.45 / σ(F): 1.39 / Phase error: 41.33 / Stereochemistry target values: ML
RfactorNum. reflection% reflection
Rfree0.3104 424 4.73 %
Rwork0.2793 --
obs0.2809 8967 71.01 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Refinement stepCycle: LAST / Resolution: 2.622→29.689 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms3160 0 0 103 3263
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.0043202
X-RAY DIFFRACTIONf_angle_d1.0064341
X-RAY DIFFRACTIONf_dihedral_angle_d13.8821245
X-RAY DIFFRACTIONf_chiral_restr0.069536
X-RAY DIFFRACTIONf_plane_restr0.005546
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
2.6215-3.00060.404850.31881591X-RAY DIFFRACTION40
3.0006-3.77920.38331330.31732943X-RAY DIFFRACTION73
3.7792-29.69070.272060.25814009X-RAY DIFFRACTION99

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