Mutations in the hydrophobic core of ubiquitin differentially affect its recognition by receptor proteins

J Mol Biol. 2008 Jan 25;375(4):979-96. doi: 10.1016/j.jmb.2007.11.016. Epub 2007 Nov 13.


Ubiquitin (Ub) is one of the most highly conserved signaling proteins in eukaryotes. In carrying out its myriad functions, Ub conjugated to substrate proteins interacts with dozens of receptor proteins that link the Ub signal to various biological outcomes. Here we report mutations in conserved residues of Ub's hydrophobic core that have surprisingly potent and specific effects on molecular recognition. Mutant Ubs bind tightly to the Ub-associated domain of the receptor proteins Rad23 and hHR23A but fail to bind the Ub-interacting motif present in the receptors Rpn10 and S5a. Moreover, chains assembled on target substrates with mutant Ubs are unable to support substrate degradation by the proteasome in vitro or sustain viability of yeast cells. The mutations have relatively little effect on Ub's overall structure but reduce its rigidity and cause a slight displacement of the C-terminal beta-sheet, thereby compromising association with Ub-interacting motif but not with Ub-associated domains. These studies emphasize an unexpected role for Ub's core in molecular recognition and suggest that the diversity of protein-protein interactions in which Ub engages placed enormous constraints on its evolvability.

Publication types

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

MeSH terms

  • Alanine / metabolism
  • Amino Acid Motifs
  • Amino Acid Sequence
  • Amino Acid Substitution
  • Carrier Proteins / chemistry
  • Carrier Proteins / genetics
  • Carrier Proteins / metabolism*
  • DNA-Binding Proteins / chemistry
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism*
  • Deuterium / metabolism
  • Dose-Response Relationship, Drug
  • Glutathione Transferase / metabolism
  • Glycine / metabolism
  • Guanidine / pharmacology
  • Humans
  • Hydrogen / metabolism
  • Hydrogen Bonding
  • Hydrogen-Ion Concentration
  • Hydrophobic and Hydrophilic Interactions
  • Ligands
  • Lysine / metabolism
  • Models, Molecular
  • Mutation*
  • Proteasome Endopeptidase Complex / metabolism
  • Protein Binding
  • Protein Conformation
  • Protein Denaturation
  • Protein Folding
  • Protein Structure, Secondary
  • Protein Structure, Tertiary
  • Saccharomyces cerevisiae Proteins / chemistry
  • Saccharomyces cerevisiae Proteins / genetics
  • Saccharomyces cerevisiae Proteins / metabolism*
  • Structure-Activity Relationship
  • Substrate Specificity
  • Temperature
  • Thermodynamics
  • Ubiquitin / chemistry
  • Ubiquitin / genetics*
  • Ubiquitin / metabolism*
  • Ubiquitination


  • Carrier Proteins
  • DNA-Binding Proteins
  • Ligands
  • RAD23 protein, S cerevisiae
  • RPN10 protein, S cerevisiae
  • Saccharomyces cerevisiae Proteins
  • Ubiquitin
  • Hydrogen
  • Deuterium
  • Glutathione Transferase
  • Proteasome Endopeptidase Complex
  • Guanidine
  • Lysine
  • Alanine
  • Glycine

Associated data

  • PDB/2JWZ