Unique structural features govern the activity of a human mitochondrial AAA+ disaggregase, Skd3

Ryan R Cupo; Alexandrea N Rizo; Gabriel A Braun; Eric Tse; Edward Chuang; Kushol Gupta; Daniel R Southworth; James Shorter

doi:10.1016/j.celrep.2022.111408

Unique structural features govern the activity of a human mitochondrial AAA+ disaggregase, Skd3

Cell Rep. 2022 Sep 27;40(13):111408. doi: 10.1016/j.celrep.2022.111408.

Authors

Ryan R Cupo¹, Alexandrea N Rizo², Gabriel A Braun³, Eric Tse⁴, Edward Chuang¹, Kushol Gupta⁵, Daniel R Southworth⁶, James Shorter⁷

Affiliations

¹ Department of Biochemistry and Biophysics, University of Pennsylvania, Philadelphia, PA, USA; Pharmacology Graduate Group, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA.
² Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, CA, USA; Graduate Program in Chemical Biology, University of Michigan, Ann Arbor, MI, USA.
³ Chemistry and Chemical Biology Graduate Program, Institute for Neurodegenerative Diseases, University of California, San Francisco, San Francisco, CA, USA.
⁴ Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, CA, USA.
⁵ Department of Biochemistry and Biophysics, University of Pennsylvania, Philadelphia, PA, USA.
⁶ Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, CA, USA. Electronic address: daniel.southworth@ucsf.edu.
⁷ Department of Biochemistry and Biophysics, University of Pennsylvania, Philadelphia, PA, USA; Pharmacology Graduate Group, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA. Electronic address: jshorter@pennmedicine.upenn.edu.

Abstract

The AAA+ protein, Skd3 (human CLPB), solubilizes proteins in the mitochondrial intermembrane space, which is critical for human health. Skd3 variants with defective protein-disaggregase activity cause severe congenital neutropenia (SCN) and 3-methylglutaconic aciduria type 7 (MGCA7). How Skd3 disaggregates proteins remains poorly understood. Here, we report a high-resolution structure of a Skd3-substrate complex. Skd3 adopts a spiral hexameric arrangement that engages substrate via pore-loop interactions in the nucleotide-binding domain (NBD). Substrate-bound Skd3 hexamers stack head-to-head via unique, adaptable ankyrin-repeat domain (ANK)-mediated interactions to form dodecamers. Deleting the ANK linker region reduces dodecamerization and disaggregase activity. We elucidate apomorphic features of the Skd3 NBD and C-terminal domain that regulate disaggregase activity. We also define how Skd3 subunits collaborate to disaggregate proteins. Importantly, SCN-linked subunits sharply inhibit disaggregase activity, whereas MGCA7-linked subunits do not. These advances illuminate Skd3 structure and mechanism, explain SCN and MGCA7 inheritance patterns, and suggest therapeutic strategies.

Keywords: AAA+ proteins, protein aggregation; CP: Cell biology; chaperone; disaggregase; mitochondria; mitochondrial disorders; therapeutics.

Publication types

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

MeSH terms

Adenosine Triphosphate / metabolism
Ankyrins* / metabolism
Heat-Shock Proteins* / metabolism
Humans
Models, Molecular
Nucleotides / metabolism
Protein Transport

Substances

Ankyrins
Heat-Shock Proteins
Nucleotides
Adenosine Triphosphate

Abstract

Publication types

MeSH terms

Substances

Grants and funding