title: Cushing's syndrome mutant PKAL205Rexhibits altered substrate specificity.
abstract: The PKAL205Rhotspot mutation has been implicated in Cushing's syndrome through hyperactive gain-of-function PKA signaling; however, its influence on substrate specificity has not been investigated. Here, we employ the Proteomic Peptide Library (ProPeL) approach to create high-resolution models for PKAWTand PKAL205Rsubstrate specificity. We reveal that the L205R mutation reduces canonical hydrophobic preference at the substrate P + 1 position, and increases acidic preference in downstream positions. Using these models, we designed peptide substrates that exhibit altered selectivity for specific PKA variants, and demonstrate the feasibility of selective PKAL205Rloss-of-function signaling. Through these results, we suggest that substrate rewiring may contribute to Cushing's syndrome disease etiology, and introduce a powerful new paradigm for investigating mutation-induced kinase substrate rewiring in human disease.
authors.affiliation: Department of Physiology and Neurobiology, University of Connecticut, Storrs, CT, USA.
authors.forename: Joshua M
authors.initials: JM
authors.lastname: Lubner
authors.affiliation: Pat and Jim Calhoun Center for Cardiology, Department of Cell Biology, University of Connecticut Health Center, Farmington, CT, USA.
authors.forename: Kimberly L
authors.initials: KL
authors.lastname: Dodge-Kafka
authors.affiliation: Institute for Experimental Medical Research, Oslo University Hospital and University of Oslo, Norway.
authors.forename: Cathrine R
authors.initials: CR
authors.lastname: Carlson
authors.affiliation: Department of Genetics, Harvard Medical School, Boston, MA, USA.|Wyss Institute for Biologically Inspired Engineering, Harvard University, Boston, MA, USA.
authors.forename: George M
authors.initials: GM
authors.lastname: Church
authors.affiliation: Department of Genetics, Harvard Medical School, Boston, MA, USA.|Wyss Institute for Biologically Inspired Engineering, Harvard University, Boston, MA, USA.
authors.forename: Michael F
authors.initials: MF
authors.lastname: Chou
authors.affiliation: Department of Physiology and Neurobiology, University of Connecticut, Storrs, CT, USA.
authors.forename: Daniel
authors.initials: D
authors.lastname: Schwartz
mesh_terms: D000595:Amino Acid Sequence; D003480:Cushing Syndrome / Q000201:enzymology* / Q000235:genetics*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D057075:Enzyme Assays; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D009154:Mutation / Q000235:genetics*; D015202:Protein Engineering; D013379:Substrate Specificity
chemical_list: D050505:Mutant Proteins; D017868:Cyclic AMP-Dependent Protein Kinases
keywords: Cushing's syndrome; protein kinase A; substrate specificity