Recruiting Mechanism of the AAA Peroxins, Pex1p and Pex6p, to Pex26p on the Peroxisomal Membrane

Chika Nashiro, Astuko Kashiwagi, Takashi Matsuzaki, Shigehiko Tamura, Yukio Fujiki

Research output: Contribution to journalArticle

20 Citations (Scopus)

Abstract

A peroxisomal C-tail-anchored type-II membrane protein, Pex26p, recruits AAA ATPase Pex1p-Pex6p complexes to peroxisomes. We herein attempted to gain mechanistic insight into Pex26p function. Pex26pΔ33-40 truncated in amino-acid residues at 33-40 abolishes the recruiting of Pex1p-Pex6p complex to peroxisomes and fails to complement the impaired phenotype of pex26 CHO cell mutant ZP167, thereby suggesting that peroxisomal localization of Pex1p and Pex6p is indispensable for the transport of matrix proteins. In in vitro transport assay using semipermeabilized CHO cells, Pex1p is targeted to peroxisomes in a manner dependent on ATP hydrolysis, while Pex6p targeting requires ATP but not its hydrolysis. This finding is confirmed by the assay using Walker-motif mutants. Transport of Pex1p and Pex6p is temperature-dependent. In vitro binding assays with glutathione-S-transferase-fused Pex26p, Pex1p and Pex6p bind to Pex26p in a manner dependent on ATP binding but not ATP hydrolysis. These results suggest that ATP hydrolysis is required for stable localization of Pex1p to peroxisomes, but not for binding to Pex26p. Moreover, Pex1p and Pex6p are altered to a more compact conformation upon binding to ATP, as verified by limited proteolysis. Taken together, Pex1p and Pex6p are most likely regulated in their peroxisomal localization onto Pex26p via conformational changes by the ATPase cycle.

Original languageEnglish
Pages (from-to)774-788
Number of pages15
JournalTraffic
Volume12
Issue number6
DOIs
Publication statusPublished - Jun 1 2011

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Peroxisomes
Adenosine Triphosphate
Membranes
Hydrolysis
Assays
CHO Cells
Adenosine Triphosphatases
Walkers
Proteolysis
Glutathione Transferase
Conformations
Carrier Proteins
Membrane Proteins
Phenotype
Amino Acids
Temperature
Proteins
In Vitro Techniques

All Science Journal Classification (ASJC) codes

  • Structural Biology
  • Biochemistry
  • Molecular Biology
  • Genetics
  • Cell Biology

Cite this

Recruiting Mechanism of the AAA Peroxins, Pex1p and Pex6p, to Pex26p on the Peroxisomal Membrane. / Nashiro, Chika; Kashiwagi, Astuko; Matsuzaki, Takashi; Tamura, Shigehiko; Fujiki, Yukio.

In: Traffic, Vol. 12, No. 6, 01.06.2011, p. 774-788.

Research output: Contribution to journalArticle

Nashiro, Chika ; Kashiwagi, Astuko ; Matsuzaki, Takashi ; Tamura, Shigehiko ; Fujiki, Yukio. / Recruiting Mechanism of the AAA Peroxins, Pex1p and Pex6p, to Pex26p on the Peroxisomal Membrane. In: Traffic. 2011 ; Vol. 12, No. 6. pp. 774-788.
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