A theoretical approach towards the identification of cleavage-determining amino acid motifs of the 20 S proteasome

J Mol Biol. 1999 Mar 5;286(4):1251-65. doi: 10.1006/jmbi.1998.2530.


Hitherto the mechanisms controlling the selective cleavage of peptide bonds by the 20 S proteasome have been poorly understood. The observation that peptide bond cleavage may eventually occur at the carboxyl site of either amino acid residue rules out a simple control of cleavage preferences by the P1 residue alone. Here, we follow the rationale that the presence of specific cleavage-determining amino acids motifs (CDAAMs) around the scissile peptide bond are required for the attainment of substrate conformations susceptible to cleavage. We present an exploratory search for these putative motifs based on empirical regression functions relating the cleavage probability for a given peptide bond to some selected side-chain properties of the flanking amino acid residues. Identification of the sequence locations of cleavage-determining residues relative to the scissile bond and of their optimal side-chain properties was carried out by fitting the cleavage probability to (binary) experimental observations on peptide bond cleavage gathered among a set of seven different peptide substrates with known patterns of proteolytic degradation products. In this analysis, all peptide bonds containing the same residue in the P1 position were assumed to be cleaved by the same active sites of the proteasome, and thus to be under control of the same CDAAMs. We arrived at a final set of ten different CDAAMs, accounting for the cleavage of one to five different groups of peptide bonds with an overall predictive correctness of 93 %. The CDAAM is composed of two to four "anchor" positions preferentially located between P5 and P5' around the scissile bond. This implies a length constraint for the usage of cleavage sites, which could considerably suppress the excision of shorter fragments and thus partially explain for the observed preponderance of medium-size cleavage products.

MeSH terms

  • Amino Acid Sequence
  • Binding Sites
  • Cysteine Endopeptidases / chemistry*
  • Cysteine Endopeptidases / metabolism
  • Eukaryotic Cells
  • Histocompatibility Antigens Class I / metabolism
  • Kinetics
  • Models, Theoretical
  • Molecular Sequence Data
  • Multienzyme Complexes / chemistry*
  • Multienzyme Complexes / metabolism
  • Proteasome Endopeptidase Complex
  • Protein Conformation
  • Regression Analysis
  • Sequence Homology, Amino Acid


  • Histocompatibility Antigens Class I
  • Multienzyme Complexes
  • Cysteine Endopeptidases
  • Proteasome Endopeptidase Complex