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A conserved 20S proteasome assembly factor requires a C-terminal HbYX motif for proteasomal precursor binding

Nature Structural & Molecular Biology volume 18, pages 622–629 (2011)Cite this article

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Abstract

Dedicated chaperones facilitate the assembly of the eukaryotic proteasome, but how they function remains largely unknown. Here we show that a yeast 20S proteasome assembly factor, Pba1–Pba2, requires a previously overlooked C-terminal hydrophobic-tyrosine-X (HbYX) motif for function. HbYX motifs in proteasome activators open the 20S proteasome entry pore, but Pba1–Pba2 instead binds inactive proteasomal precursors. We discovered an archaeal ortholog of this factor, here named PbaA, that also binds preferentially to proteasomal precursors in a HbYX motif–dependent fashion using the same proteasomal α-ring surface pockets as are bound by activators. PbaA and the related PbaB protein can be induced to bind mature 20S proteasomes if the active sites in the central chamber are occupied by inhibitors. Our data are consistent with an allosteric mechanism in which the maturation of the proteasome active sites determines the binding of assembly chaperones, potentially shielding assembly intermediates or misassembled complexes from nonproductive associations until assembly is complete.

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Figure 1: An HbYX motif in the yeast Pba1–Pba2 assembly factor and its archaeal orthologs.
Figure 2: The HbYX motifs of yeast Pba1–Pba2 are functionally important in vivo.
Figure 3: The HbYX motifs of yeast Pba1–Pba2 are necessary for proteasome precursor binding.
Figure 4: Archaeal PbaA binds preferentially to a 20S proteasome intermediate.
Figure 5: PbaA HbYX motif mediates binding to the α-ring of 20S proteasome intermediates.
Figure 6: Binding of archaeal Pba proteins to inhibitor-treated mature 20S proteasomes.
Figure 7: Binding of PbaA to PHP requires the α-subunit N termini.

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Acknowledgements

We thank R.J. Tomko Jr. for comments on the manuscript and J. Leigh (University of Washington) for the archaeal genomic DNA. This work was supported by US National Institutes of Health grant GM083050 to M.H.

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  1. Andrew R Kusmierczyk

    Present address: Present address: Department of Biology, Indiana University–Purdue University Indianapolis, Indianapolis, Indiana, USA.,

Authors and Affiliations

  1. Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut, USA

    Andrew R Kusmierczyk, Roger Y Kim & Mark Hochstrasser

  2. Department of Molecular, Cellular and Developmental Biology, Yale University, New Haven, Connecticut, USA

    Mary J Kunjappu

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Contributions

A.R.K. and M.H. developed the experimental approach. A.R.K., M.J.K. and R.Y.K. carried out experiments. A.R.K. and M.H. wrote the paper.

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Correspondence to Mark Hochstrasser.

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Supplementary Figures 1–12, Supplementary Tables 1 and 2, Supplementary Notes and Supplementary Methods (PDF 543 kb)

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Kusmierczyk, A., Kunjappu, M., Kim, R. et al. A conserved 20S proteasome assembly factor requires a C-terminal HbYX motif for proteasomal precursor binding. Nat Struct Mol Biol 18, 622–629 (2011). https://doi.org/10.1038/nsmb.2027

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