Structural basis of substrate recognition and specificity in the N-end rule pathway

Nat Struct Mol Biol. 2010 Oct;17(10):1182-7. doi: 10.1038/nsmb.1894. Epub 2010 Sep 12.

Abstract

The N-end rule links the half-life of a protein to the identity of its N-terminal residue. Destabilizing N-terminal residues are recognized by E3 ubiquitin ligases, termed N-recognins. A conserved structural domain called the UBR box is responsible for their specificity. Here we report the crystal structures of the UBR boxes of the human N-recognins UBR1 and UBR2, alone and in complex with an N-end rule peptide, Arg-Ile-Phe-Ser. These structures show that the UBR box adopts a previously undescribed fold stabilized through the binding of three zinc ions to form a binding pocket for type 1 N-degrons. NMR experiments reveal a preference for N-terminal arginine. Peptide binding is abrogated by N-terminal acetylation of the peptide or loss of the positive charge of the N-terminal residue. These results rationalize and refine the empirical rules for the classification of type 1 N-degrons. We also confirm that a missense mutation in UBR1 that is responsible for Johanson-Blizzard syndrome leads to UBR box unfolding and loss of function.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Acetylation
  • Amino Acid Sequence
  • Amino Acids / chemistry
  • Binding Sites
  • Crystallography, X-Ray
  • Exocrine Pancreatic Insufficiency / genetics
  • Humans
  • Models, Molecular
  • Molecular Sequence Data
  • Mutation, Missense
  • Oligopeptides / chemistry
  • Oligopeptides / metabolism*
  • Protein Binding
  • Protein Conformation
  • Protein Folding
  • Protein Interaction Mapping
  • Protein Processing, Post-Translational
  • Protein Stability
  • Protein Structure, Tertiary
  • Structure-Activity Relationship
  • Substrate Specificity
  • Syndrome
  • Ubiquitin-Protein Ligases / chemistry
  • Ubiquitin-Protein Ligases / genetics
  • Ubiquitin-Protein Ligases / metabolism*
  • Zinc Fingers / physiology

Substances

  • Amino Acids
  • Oligopeptides
  • UBR1 protein, human
  • UBR2 protein, human
  • Ubiquitin-Protein Ligases

Associated data

  • PDB/3NY1
  • PDB/3NY2
  • PDB/3NY3
  • PDB/UBR1
  • PDB/UBR2