title: Mechanism of the generation of autonomous activity of Ca2+/calmodulin-dependent protein kinase IV.
abstract: Ca2+/calmodulin-dependent protein kinase IV (CaM-KIV) is phosphorylated at Thr196 by Ca2+/calmodulin-dependent protein kinase kinase (CaM-KK), resulting in induction of both autonomous activity and a high level of Ca2+/CaM-dependent activity. We have shown that the kinetics of Thr196 phosphorylation of CaM-KIV by CaM-KK is well correlated with the generation of its autonomous activity, although Thr177 phosphorylation of CaM-KI does not induce its autonomous activity. The activities of CaM-KI chimera mutants fused with C-terminal regions (residues 296-469 and 296-350) of CaM-KIV are completely dependent on Ca2+/CaM, which is also the case for CaM-KI. Unlike wild-type CaM-KI, however, phosphorylation of Thr177 in the chimera mutants by CaM-KK resulted in generation of significant autonomous activities, indicating that the phosphorylation of Thr in the activation loop is sufficient to partially release the autoinhibitory region of CaM-KIV from the catalytic core. Indeed, the CaM-KIV peptide (residues 304-325) containing minimum autoinhibitory sequences (residues 314-321) suppressed the activity of non-phosphorylated CaM-KIV with an IC50 of approximately 50 microm, and this suppression was competitive with respect to the peptide substrate; however, the CaM-KIV peptide was not capable of inhibiting Thr196-phosphorylated CaM-KIV. Taken together, these results indicated that the Thr196 phosphorylation of CaM-KIV by CaM-KK reduced the interaction of the catalytic core with the autoinhibitory region, resulting in generation of the autonomous activity.
authors.affiliation: Department of Signal Transduction Sciences, Faculty of Medicine, Kagawa University, 1750-1 Miki-cho, Kita-gun, Kagawa 761-0793, Japan. tokumit@med.kagawa-u.ac.jp
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Tokumitsu
authors.forename: Naoya
authors.initials: N
authors.lastname: Hatano
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Inuzuka
authors.forename: Shigeyuki
authors.initials: S
authors.lastname: Yokokura
authors.forename: Naohito
authors.initials: N
authors.lastname: Nozaki
authors.forename: Ryoji
authors.initials: R
authors.lastname: Kobayashi
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D015153:Blotting, Western; D001921:Brain / Q000378:metabolism; D054737:Calcium-Calmodulin-Dependent Protein Kinase Kinase; D054734:Calcium-Calmodulin-Dependent Protein Kinase Type 4; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000737:chemistry / Q000502:physiology*; D002384:Catalysis; D020134:Catalytic Domain; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D004926:Escherichia coli / Q000378:metabolism; D005982:Glutathione Transferase / Q000378:metabolism; D020128:Inhibitory Concentration 50; D007700:Kinetics; D008969:Molecular Sequence Data; D009154:Mutation; D010455:Peptides / Q000737:chemistry; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000502:physiology*; D017434:Protein Structure, Tertiary; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry; D013696:Temperature; D013912:Threonine / Q000737:chemistry; D013997:Time Factors
chemical_list: D018076:DNA, Complementary; D010455:Peptides; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D013912:Threonine; D005982:Glutathione Transferase; D017346:Protein Serine-Threonine Kinases; D054737:Calcium-Calmodulin-Dependent Protein Kinase Kinase; D054734:Calcium-Calmodulin-Dependent Protein Kinase Type 4; D017871:Calcium-Calmodulin-Dependent Protein Kinases; C517686:Camk4 protein, rat