title: Influence of a mutation in the ATP-binding region of Ca2+/calmodulin-dependent protein kinase II on its interaction with peptide substrates.
abstract: CaMKII (Ca2+/calmodulin-dependent protein kinase II) is expressed in high concentrations in the brain and is found enriched in the postsynaptic densities. The enzyme is activated by the binding of calmodulin to the autoregulatory domain in the presence of high levels of intracellular Ca2+, which causes removal of auto-inhibition from the N-terminal catalytic domain. Knowledge of the 3D (three-dimensional) structure of this enzyme at atomic resolution is restricted to the association domain, a region at the extreme C-terminus. The catalytic domain of CaMKII shares high sequence similarity with CaMKI. The 3D structure of the catalytic core of CaMKI comprises ATP- and substrate-binding regions in a cleft between two distinct lobes, similar to the structures of all protein kinases solved to date. Mutation of Glu-60, a residue in the ATP-binding region of CaMKII, to glycine exerts different effects on phosphorylation of two peptide substrates, syntide and NR2B ( N -methyl-D-aspartate receptor subunit 2B) 17-mer. Although the mutation caused increases in the Km values for phosphorylation for both the peptide substrates, the effect on the kcat values for each was different. The kcat value decreased in the case of syntide, whereas it increased in the case of the NR2B peptide as a result of the mutation. This resulted in a significant decrease in the apparent kcat/Km value for syntide, but the change was minimal for the NR2B peptide. These results indicate that different catalytic mechanisms are employed by the kinase for the two peptides. Molecular modelling suggests structural changes are likely to occur at the peptide-binding pocket in the active state of the enzyme as a consequence of the Glu-60-->Gly mutation.
authors.affiliation: Rajiv Gandhi Centre for Biotechnology, Thycaud P.O., Thiruvananthapuram, Kerala 695014, India.
authors.forename: Mullasseril
authors.initials: M
authors.lastname: Praseeda
authors.forename: Kurup K
authors.initials: KK
authors.lastname: Pradeep
authors.forename: Ananth
authors.initials: A
authors.lastname: Krupa
authors.forename: S Sri
authors.initials: SS
authors.lastname: Krishna
authors.forename: Suseela
authors.initials: S
authors.lastname: Leena
authors.forename: R Rajeev
authors.initials: RR
authors.lastname: Kumar
authors.forename: John
authors.initials: J
authors.lastname: Cheriyan
authors.forename: Madhavan
authors.initials: M
authors.lastname: Mayadevi
authors.forename: Narayanaswamy
authors.initials: N
authors.lastname: Srinivasan
authors.forename: Ramakrishnapillai V
authors.initials: RV
authors.lastname: Omkumar
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D036341:Intercellular Signaling Peptides and Proteins; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D010455:Peptides / Q000737:chemistry / Q000378:metabolism*; D016194:Receptors, N-Methyl-D-Aspartate / Q000737:chemistry / Q000378:metabolism; D018411:Spodoptera / Q000166:cytology
chemical_list: D036341:Intercellular Signaling Peptides and Proteins; C485096:NR2B protein (1292-1308); D010446:Peptide Fragments; D010455:Peptides; D016194:Receptors, N-Methyl-D-Aspartate; C063391:syntide-2; D000255:Adenosine Triphosphate; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases