Reconstitution of Circadian Oscillation of Cyanobacterial KaiC Phosphorylation in Vitro

@article{Nakajima2005ReconstitutionOC,
  title={Reconstitution of Circadian Oscillation of Cyanobacterial KaiC Phosphorylation in Vitro},
  author={Masato Nakajima and Keiko Imai and Hiroshi Ito and Taeko Nishiwaki and Yoriko Murayama and Hideo Iwasaki and Tokitaka Oyama and Takao Kondo},
  journal={Science},
  year={2005},
  volume={308},
  pages={414 - 415}
}
Kai proteins globally regulate circadian gene expression of cyanobacteria. The KaiC phosphorylation cycle, which persists even without transcription or translation, is assumed to be a basic timing process of the circadian clock. We have reconstituted the self-sustainable oscillation of KaiC phosphorylation in vitro by incubating KaiC with KaiA, KaiB, and adenosine triphosphate. The period of the in vitro oscillation was stable despite temperature change (temperature compensation), and the… Expand
In vitro regulation of circadian phosphorylation rhythm of cyanobacterial clock protein KaiC by KaiA and KaiB
TLDR
KaiA and KaiB were found to be “parameter‐tuning” and “state‐switching” regulators of KaiC phosphorylation rhythm, respectively, suggesting a possible entrainment mechanism of the cellular circadian clock with the circadian variation of intracellular levels of Kai proteins. Expand
Autonomous synchronization of the circadian KaiC phosphorylation rhythm
TLDR
The cyanobacterial circadian oscillator can be reconstituted in vitro by mixing three purified clock proteins, KaiA, KaiB and KaiC, with ATP and it is demonstrated that synchronization is tightly linked with KaiC dephosphorylation and is mediated by monomer exchange between KaiC hexamers during the early deph phosphorylation phase. Expand
Circadian Rhythmicity by Autocatalysis
TLDR
This model, based upon autocatalysis instead of transcription-translation negative feedback, shows temperature-compensated circadian limit-cycle oscillations with KaiC phosphorylation profiles and has period lengths and rate constant values that are consistent with experimental observations. Expand
Evolution Analysis of the Circadian Clock Protein KaiB
TLDR
Here, the evolution of the KaiB protein is studied to help understand the development of the circadian clock system in cyanobacteria. Expand
Detecting KaiC phosphorylation rhythms of the cyanobacterial circadian oscillator in vitro and in vivo.
TLDR
Here, a detailed protocol to obtain pure recombinant proteins from Escherichia coli to construct a robust cyanobacterial circadian oscillator in vitro is provided and a protocol that facilitates analysis of phosphorylation states of KaiC and other phosphorylated proteins from in vivo samples is presented. Expand
A Molecular Dynamics Study of the Cyanobacterial Clock Protein KaiA
Regulation of daily physiological functions with a ~24-hour periodicity, or circadian rhythms, exists in both eukaryotes and prokaryotes. So far, cyanobacteria are only known prokaryotes proved toExpand
Synchronization of circadian oscillation of phosphorylation level of KaiC in vitro.
TLDR
This study extends the previous model of oscillation by explicitly taking two phosphorylation sites of KaiC into account and applies the extended model to the problem of synchrony of two oscillatory samples mixed at different phases, suggesting that the combined mechanism of the allosteric transition of Kai C hexamers and the monomer shuffling between them plays a key role in synchronization among KaiC hexamer and hence underlies the population-level oscillation of the ensemble of Kai proteins. Expand
Expression and Purification of Cyanobacterial Circadian Clock Protein KaiC and Determination of Its Auto-phosphatase Activity.
TLDR
Circadian clocks of cyanobacteria are the simplest and one of the best studied models and can be used for in vitro reconstitution experiments and determination of the auto-phosphatase activity of KaiC as described in this protocol. Expand
Cyanobacterial circadian pacemaker: Kai protein complex dynamics in the KaiC phosphorylation cycle in vitro.
TLDR
The molecular model proposed here provides mechanisms for circadian timing systems and confirmed that a similar model can be applied in cyanobacterial cells. Expand
A model for circadian rhythm of cyanobacteria , which maintains oscillation without gene expression
An intriguing property of the cyanobacterial circadian clock is that endogenous rhythm persists when protein abundances are kept constant either in the presence of translation and transcriptionExpand
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TLDR
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TLDR
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