title: Identification of domains essential for the assembly of calcium/calmodulin-dependent protein kinase II holoenzymes.
abstract: Ca2+/calmodulin-dependent protein kinase II (CaM kinase II), as isolated from brain, is a multimeric complex composed predominantly of two subunits, alpha and beta, products of unique genes. Little is known about how subunit composition influences holoenzyme structure or how the domain(s) of each subunit interact to form holoenzymes. We show here that holoenzymes composed of only alpha or only beta subunits exhibit different biophysical properties. The S values of alpha and beta are 17.2 and 14.5 S while the Stokes's radii are 85 and 111 A, respectively, indicating their structures are different. C-terminal truncations of the alpha subunit show that amino acids 382-478 are necessary for holoenzyme formation and that amino acids 427-478 contribute to holoenzyme stability. Additionally, the C-terminal domains of both the alpha subunit, alpha315-478, and beta subunit, beta314-542, formed oligomers indicating the sufficiency of the C-terminal domain for multimer formation. Using the yeast two-hybrid system we show, in vivo, that full-length subunits, alpha1-478 and beta1-542, interact with themselves or each other interchangeably. Additionally, the C-terminal domains of the alpha subunit, alpha315-478 and beta subunit, beta314-542 associated with themselves in a manner indistinguishable from their association with full-length alpha or beta subunits. Further studies revealed that the C-terminal domains of the alpha and beta subunits contain information necessary for interaction with beta but not alpha. These data are summarized into a model describing the assembly of CaM kinase II holoenzymes.
authors.affiliation: Department of Neurobiology and Anatomy, The University of Texas Medical School, Houston, Texas 77030, USA.
authors.forename: S J
authors.initials: SJ
authors.lastname: Kolb
authors.forename: A
authors.initials: A
authors.lastname: Hudmon
authors.forename: T R
authors.initials: TR
authors.lastname: Ginsberg
authors.forename: M N
authors.initials: MN
authors.lastname: Waxham
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004795:Enzyme Stability; D020035:Holoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008956:Models, Chemical; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D010766:Phosphorylation; D011485:Protein Binding; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000378:metabolism
chemical_list: D020035:Holoenzymes; D007527:Isoenzymes; D010446:Peptide Fragments; D011994:Recombinant Proteins; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases