title: The Role of Regulatory Domains in Maintaining Autoinhibition in the Multidomain Kinase PKCα.
abstract: Resolving the conformational dynamics of large multidomain proteins has proven to be a significant challenge. Here we use a variety of techniques to dissect the roles of individual protein kinase Cα (PKCα) regulatory domains in maintaining catalytic autoinhibition. We find that whereas the pseudosubstrate domain is necessary for autoinhibition it is not sufficient. Instead, each regulatory domain (C1a, C1b, and C2) appears to strengthen the pseudosubstrate-catalytic domain interaction in a nucleotide-dependent manner. The pseudosubstrate and C1a domains, however, are minimally essential for maintaining the inactivated state. Furthermore, disrupting known interactions between the C1a and other regulatory domains releases the autoinhibited interaction and increases basal activity. Modulating this interaction between the catalytic and regulatory domains reveals a direct correlation between autoinhibition and membrane translocation following PKC activation.
authors.affiliation: From the Department of Genetics, Cell Biology, and Development, University of Minnesota, Minneapolis, Minnesota 55455 and.
authors.forename: Ruth F
authors.initials: RF
authors.lastname: Sommese
authors.affiliation: From the Department of Genetics, Cell Biology, and Development, University of Minnesota, Minneapolis, Minnesota 55455 and.
authors.forename: Michael
authors.initials: M
authors.lastname: Ritt
authors.affiliation: the Biophysics Program, University of Michigan, Ann Arbor, Michigan 48109.
authors.forename: Carter J
authors.initials: CJ
authors.lastname: Swanson
authors.affiliation: From the Department of Genetics, Cell Biology, and Development, University of Minnesota, Minneapolis, Minnesota 55455 and sivaraj@umn.edu.
authors.forename: Sivaraj
authors.initials: S
authors.lastname: Sivaramakrishnan
mesh_terms: D000818:Animals; D002384:Catalysis; D020134:Catalytic Domain; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans; D009154:Mutation; D051571:Protein Kinase C-alpha / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D021381:Protein Transport; D061987:Sf9 Cells; D013379:Substrate Specificity
chemical_list: D051571:Protein Kinase C-alpha
keywords: fluorescence resonance energy transfer (FRET); inhibition mechanism; protein domain; protein kinase C (PKC); protein translocation; signal transduction