ATPase activity and its temperature compensation of the cyanobacterial clock protein KaiC.
KaiA, KaiB and KaiC constitute the circadian clock machinery in cyanobacteria. KaiC is a homohexamer; its subunit contains duplicated halves, each with a set of ATPase motifs. Here, using highly purified KaiC preparations of the thermophilic cyanobacterium Thermosynechococcus elongatus BP-1 produced in Escherichia coli, we found that the N- and C-terminal domains of KaiC had extremely weak ATPase activity. ATPase activity showed temperature compensation in wild-type KaiC, but not in KaiC(S431A/T432A), a mutant that lacks two phosphorylation sites. We concluded that KaiC phosphorylation is involved in the ATPase temperature-compensation mechanism-which is probably critical to the stability of the circadian clock in cyanobacteria-and we hypothesized the following temperature-compensation mechanism: (i) The C-terminal phosphorylation sites of a KaiC hexamer subunit are phosphorylated by the C-terminal domain of an adjacent KaiC subunit; (ii) the phosphorylation suppresses the ATPase activity of the C-terminal domain; and (iii) the phosphorylated KaiC spontaneously dephosphorylates, resulting in the recover of ATPase activity.
Murakami
Reiko
R
Center for Gene Research, Nagoya University, Furocho, Chikusa, Nagoya 464-8602, Japan.
Miyake
Ayumi
A
Iwase
Ryo
R
Hayashi
Fumio
F
Uzumaki
Tatsuya
T
Ishiura
Masahiro
M
D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002940:Circadian Rhythm; D056925:Circadian Rhythm Signaling Peptides and Proteins; D000458:Cyanobacteria / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D017434:Protein Structure, Tertiary; D021122:Protein Subunits; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013696:Temperature
D001426:Bacterial Proteins; D056925:Circadian Rhythm Signaling Peptides and Proteins; C114269:KaiA protein, cyanobacteria; C114271:KaiC protein, cyanobacteria; D021122:Protein Subunits; D011994:Recombinant Proteins; D000251:Adenosine Triphosphatases