Biochemical properties of CikA, an unusual phytochrome-like histidine protein kinase that resets the circadian clock in Synechococcus elongatus PCC 7942.
We recently described the cikA (circadian input kinase A) gene, whose product supplies environmental information to the circadian oscillator in the cyanobacterium Synechococcus elongatus PCC 7942. CikA possesses three distinct domains: a GAF, a histidine protein kinase (HPK), and a receiver domain similar to those of the response regulator family. To determine how CikA functions in providing circadian input, we constructed modified alleles to tag and truncate the protein, allowing analysis of each domain individually. CikA covalently bound bilin chromophores in vitro, even though it lacks the expected ligand residues, and the GAF domain influenced but did not entirely account for this function. Full-length CikA and truncated variants that carry the HPK domain showed autophosphorylation activity. Deletion of the GAF domain or the N-terminal region adjacent to GAF dramatically reduced autophosphorylation, whereas elimination of the receiver domain increased activity 10-fold. Assays to test phosphorelay from the HPK to the cryptic receiver domain, which lacks the conserved aspartyl residue that serves as a phosphoryl acceptor in response regulators, were negative. We propose that the cryptic receiver is a regulatory domain that interacts with an unknown protein partner to modulate the autokinase activity of CikA but does not work as bona fide receiver domain in a phosphorelay.
Mutsuda
Michinori
M
Department of Biology, Texas A & M University, College Station, Texas 77843-3258, USA.
Michel
Klaus-Peter
KP
Zhang
Xiaofan
X
Montgomery
Beronda L
BL
Golden
Susan S
SS
D000255:Adenosine Triphosphate / Q000378:metabolism; D000483:Alleles; D001426:Bacterial Proteins; D001654:Bile Pigments / Q000378:metabolism; D001664:Biliverdine / Q000235:genetics / Q000378:metabolism; D002940:Circadian Rhythm / Q000502:physiology*; D000458:Cyanobacteria / Q000201:enzymology*; D004926:Escherichia coli / Q000235:genetics; D017353:Gene Deletion; D015870:Gene Expression; D000071677:Histidine Kinase; D008163:Luminescent Measurements; D016296:Mutagenesis; D010766:Phosphorylation; D052980:Phycobilins; D010798:Phycocyanin / Q000235:genetics / Q000378:metabolism; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D011494:Protein Kinases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D011758:Pyrroles / Q000378:metabolism; D011994:Recombinant Proteins; D013329:Structure-Activity Relationship; D045725:Tetrapyrroles
D001426:Bacterial Proteins; D001654:Bile Pigments; D052980:Phycobilins; D011758:Pyrroles; D011994:Recombinant Proteins; D045725:Tetrapyrroles; D010798:Phycocyanin; C011069:phycocyanobilin; D000255:Adenosine Triphosphate; D011494:Protein Kinases; D000071677:Histidine Kinase; C411989:CikA protein, bacteria; D001664:Biliverdine