Cushing's syndrome mutant PKAL205R exhibits altered substrate specificity

FEBS Lett. 2017 Feb;591(3):459-467. doi: 10.1002/1873-3468.12562. Epub 2017 Feb 3.


The PKAL205R hotspot 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 PKAWT and PKAL205R substrate 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 PKAL205R loss-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.

Keywords: Cushing's syndrome; protein kinase A; substrate specificity.

Publication types

  • Letter

MeSH terms

  • Amino Acid Sequence
  • Cushing Syndrome / enzymology*
  • Cushing Syndrome / genetics*
  • Cyclic AMP-Dependent Protein Kinases / chemistry
  • Cyclic AMP-Dependent Protein Kinases / genetics*
  • Cyclic AMP-Dependent Protein Kinases / metabolism
  • Enzyme Assays
  • Escherichia coli / metabolism
  • Humans
  • Mutant Proteins / chemistry
  • Mutant Proteins / genetics
  • Mutant Proteins / metabolism
  • Mutation / genetics*
  • Protein Engineering
  • Substrate Specificity


  • Mutant Proteins
  • Cyclic AMP-Dependent Protein Kinases