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J Biol Chem. 2016 Jan 22;291(4):1991-2003. doi: 10.1074/jbc.M115.677047. Epub 2015 Dec 1.

Distinct Elements in the Proteasomal β5 Subunit Propeptide Required for Autocatalytic Processing and Proteasome Assembly.

Author information

1
From the Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut 06520 and.
2
the Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, Illinois 60637.
3
From the Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut 06520 and mark.hochstrasser@yale.edu.

Abstract

Eukaryotic 20S proteasome assembly remains poorly understood. The subunits stack into four heteroheptameric rings; three inner-ring subunits (β1, β2, and β5) bear the protease catalytic residues and are synthesized with N-terminal propeptides. These propeptides are removed autocatalytically late in assembly. In Saccharomyces cerevisiae, β5 (Doa3/Pre2) has a 75-residue propeptide, β5pro, that is essential for proteasome assembly and can work in trans. We show that deletion of the poorly conserved N-terminal half of the β5 propeptide nonetheless causes substantial defects in proteasome maturation. Sequences closer to the cleavage site have critical but redundant roles in both assembly and self-cleavage. A conserved histidine two residues upstream of the autocleavage site strongly promotes processing. Surprisingly, although β5pro is functionally linked to the Ump1 assembly factor, trans-expressed β5pro associates only weakly with Ump1-containing precursors. Several genes were identified as dosage suppressors of trans-expressed β5pro mutants; the strongest encoded the β7 proteasome subunit. Previous data suggested that β7 and β5pro have overlapping roles in bringing together two half-proteasomes, but the timing of β7 addition relative to half-mer joining was unclear. Here we report conditions where dimerization lags behind β7 incorporation into the half-mer. Our results suggest that β7 insertion precedes half-mer dimerization, and the β7 tail and β5 propeptide have unequal roles in half-mer joining.

KEYWORDS:

chaperone; propeptide; proteasome; protein assembly; protein degradation; ubiquitin

PMID:
26627836
PMCID:
PMC4722473
DOI:
10.1074/jbc.M115.677047
[Indexed for MEDLINE]
Free PMC Article

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