Characterization of recombinant REGα, REGβ, and REGγ proteasome activators

Claudio Realini, Christopher C. Jensen, Zhi Guo Zhang, Steven C. Johnston, J. Randalph Knowlton, Christopher P. Hill, Martin Rechsteiner

Research output: Contribution to journalArticle

144 Scopus citations

Abstract

Full-length cDNAs for three human proteasome activator subunits, called REGα, REGβ, and REGγ, have been expressed in Escherichia coli, and the purified recombinant proteins have been characterized. Recombinant α or γ subunits form heptameric species; recombinant β subunits are found largely as monomers or small multimers. Each recombinant REG stimulates cleavage of fluorogenic peptides by human red cell proteasomes. The pattern of activated peptide hydrolysis is virtually identical for REGα and REGβ. These two subunits, alone or in combination, stimulate cleavage after basic, acidic, and most hydrophobic residues in many peptides. Recombinant a and β subunits bind each other with high affinity, and the REGα/β heteromeric complex activates hydrolysis of LLVY-methylcoumaryl-7-amide (LLVY-MCA) and LLE-β- nitroanilide (LLE-βNA) more than REGα or REGβ alone. Using filter binding and gel filtration assays, recombinant REGγ subunits were shown to bind themselves but not α or β subunits. REGγ differs from REGα and REGβ in that it markedly stimulates hydrolysis of peptides with basic residues in the P1 position but only modestly activates cleavage of LLVY-MCA or LLE-βNA by the proteasome. REGγ binds the proteasome with higher affinity than REGα or REGβ yet with lower affinity than complexes containing both REGα and REGβ. In summary, each of the three PEG homologs is a proteasome activator with unique biochemical properties.

Original languageEnglish (US)
Pages (from-to)25483-25492
Number of pages10
JournalJournal of Biological Chemistry
Volume272
Issue number41
DOIs
StatePublished - Oct 10 1997

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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    Realini, C., Jensen, C. C., Zhang, Z. G., Johnston, S. C., Knowlton, J. R., Hill, C. P., & Rechsteiner, M. (1997). Characterization of recombinant REGα, REGβ, and REGγ proteasome activators. Journal of Biological Chemistry, 272(41), 25483-25492. https://doi.org/10.1074/jbc.272.41.25483