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Open data
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Basic information
Entry | Database: PDB / ID: 8fwi | ||||||
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Title | Structure of dodecameric KaiC-RS-S413E/S414E solved by cryo-EM | ||||||
![]() | Circadian clock protein KaiC | ||||||
![]() | CIRCADIAN CLOCK PROTEIN / autokinase | ||||||
Function / homology | ![]() kinase activity / non-specific serine/threonine protein kinase / ATP hydrolysis activity / ATP binding / metal ion binding Similarity search - Function | ||||||
Biological species | ![]() | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.9 Å | ||||||
![]() | Padua, R.A.P. / Grant, T. / Pitsawong, W. / Hoemberger, M.S. / Otten, R. / Bradshaw, N. / Grigorieff, N. / Kern, D. | ||||||
Funding support | ![]()
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![]() | ![]() Title: From primordial clocks to circadian oscillators. Authors: Warintra Pitsawong / Ricardo A P Pádua / Timothy Grant / Marc Hoemberger / Renee Otten / Niels Bradshaw / Nikolaus Grigorieff / Dorothee Kern / ![]() Abstract: Circadian rhythms play an essential part in many biological processes, and only three prokaryotic proteins are required to constitute a true post-translational circadian oscillator. The evolutionary ...Circadian rhythms play an essential part in many biological processes, and only three prokaryotic proteins are required to constitute a true post-translational circadian oscillator. The evolutionary history of the three Kai proteins indicates that KaiC is the oldest member and a central component of the clock. Subsequent additions of KaiB and KaiA regulate the phosphorylation state of KaiC for time synchronization. The canonical KaiABC system in cyanobacteria is well understood, but little is known about more ancient systems that only possess KaiBC. However, there are reports that they might exhibit a basic, hourglass-like timekeeping mechanism. Here we investigate the primordial circadian clock in Rhodobacter sphaeroides, which contains only KaiBC, to elucidate its inner workings despite missing KaiA. Using a combination of X-ray crystallography and cryogenic electron microscopy, we find a new dodecameric fold for KaiC, in which two hexamers are held together by a coiled-coil bundle of 12 helices. This interaction is formed by the carboxy-terminal extension of KaiC and serves as an ancient regulatory moiety that is later superseded by KaiA. A coiled-coil register shift between daytime and night-time conformations is connected to phosphorylation sites through a long-range allosteric network that spans over 140 Å. Our kinetic data identify the difference in the ATP-to-ADP ratio between day and night as the environmental cue that drives the clock. They also unravel mechanistic details that shed light on the evolution of self-sustained oscillators. | ||||||
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Structure visualization
Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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Download
PDBx/mmCIF format | ![]() | 2.3 MB | Display | ![]() |
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PDB format | ![]() | Display | ![]() | |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Arichive directory | ![]() ![]() | HTTPS FTP |
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-Related structure data
Related structure data | ![]() 29505MC ![]() 8db3C ![]() 8dbaC ![]() 8fwjC M: map data used to model this data C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
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Assembly
Deposited unit | ![]()
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1 |
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Components
#1: Protein | Mass: 62666.133 Da / Num. of mol.: 12 / Mutation: S413E, S414E Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() #2: Chemical | ChemComp-ATP / #3: Chemical | ChemComp-ADP / #4: Chemical | ChemComp-MG / #5: Water | ChemComp-HOH / | Has ligand of interest | N | |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
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Sample preparation
Component | Name: Dodecamer of KaiC-RS-S413E/S414E / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT |
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Molecular weight | Value: 0.751 MDa / Experimental value: NO |
Source (natural) | Organism: ![]() |
Source (recombinant) | Organism: ![]() ![]() |
Buffer solution | pH: 6.5 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Grid type: C-flat-1.2/1.3 |
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K |
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Electron microscopy imaging
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 2500 nm / Nominal defocus min: 500 nm |
Image recording | Electron dose: 100 e/Å2 / Film or detector model: GATAN K2 BASE (4k x 4k) |
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Processing
Software | Name: UCSF ChimeraX / Version: 1.5/v9 / Classification: model building / URL: https://www.rbvi.ucsf.edu/chimerax/ / Os: Linux / Type: package | ||||||||||||||||||||||||||||||||
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C6 (6 fold cyclic) | ||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 320000 / Algorithm: FOURIER SPACE Details: Submitted map is a composite of two hexamer maps overlayed and combined by taking the absolute maximum value Symmetry type: POINT | ||||||||||||||||||||||||||||||||
Atomic model building | Protocol: FLEXIBLE FIT |