A calmodulin-dependent protein kinase from lower eukaryote Physarum polycephalum.
A full-length cDNA coding a calmodulin (CaM)-dependent protein kinase gene was cloned from Physarum plasmodia poly(A)-RNA by polymerase chain reaction with the oligonucleotide primers that were designed after the amino acid sequence of highly conserved regions of myosin light-chain kinase. Sequence analysis of the cDNA revealed that this Physarum kinase was a 42,519-Da protein with an ATP-binding domain, Ser/Thr kinase active site signature, and CaM-binding domain. Expression of the cDNA in Escherichia coli demonstrated that the Physarum kinase in the presence of Ca2+ and CaM phosphorylated the recombinant phosphorylatable light chain (PLc) of Physarum myosin II. The peptide analysis after proteolysis of the phosphorylated PLc indicated that Ser 18 was phosphorylated. The site was confirmed by the failure of phosphorylation of PLc, the Ser 18 of which was replaced by Ala. The physiological role of the kinase will be discussed with special reference to the 55-kDa kinase, which had been previously purified from Physarum plasmodia for phosphorylated PLc.
Nakamura
Akio
A
Department of Molecular and Cellular Pharmacology, Faculty of Medicine, Gunma University Graduate School of Medicine, Gunma 371-8511, Japan. gacho@med.gunma-u.ac.jp
Hanyuda
Yuki
Y
Okagaki
Tuyoshi
T
Takagi
Takashi
T
Kohama
Kazuhiro
K
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D017124:Conserved Sequence; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010766:Phosphorylation; D016839:Physarum polycephalum / Q000201:enzymology* / Q000235:genetics; D011485:Protein Binding; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D020539:Sequence Analysis, Protein; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship
D011994:Recombinant Proteins; D017871:Calcium-Calmodulin-Dependent Protein Kinases