Recombinant Human Peroxisomal Targeting Signal Receptor PEX5

Schliebs, Wolfgang; Saidowsky, Jürgen; Agianian, Bogos; Dodt, Gabriele; Herberg, Friedrich W.; Kunau, Wolf‐H.

doi:10.1074/jbc.274.9.5666

Cited by 164 publications

(92 citation statements)

References 34 publications

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“…The purified preparation of Pex5p exhibited a major band of approximately 80 kDa on SDS-PAGE (Fig. 1A), being consistent with a previous report (23). Minor bands were seen in the lower molecular weight region.…”

Section: Purification Of Recombinant Proteins-humansupporting

confidence: 79%

Formation of Peroxisomes from Peroxisomal Ghosts in a Peroxisome-deficient Mammalian Cell Mutant upon Complementation by Protein Microinjection

Yamasaki

Hashiguchi

Fujiwara

et al. 1999

Journal of Biological Chemistry

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Most mammalian cell strains genetically deficient in peroxisome biogenesis have abnormal membrane structures called ghosts, containing integral peroxisomal membrane protein, PMP70, but lacking the peroxisomal matrix proteins. Upon genetic complementation, these mutants regain the ability of peroxisome biogenesis. It is postulated that, in this process, the ghosts act as the precursors of peroxisomes, but there has been no evidence to support this. In the present study, we investigated this issue by protein microinjection to a mutant Chinese hamster ovary cell line defective of PEX5, encoding a peroxisome-targeting signal receptor. When recombinant Pex5p and green fluorescent protein (GFP) carrying a peroxisome-targeting signal were co-injected into the mutant cells, the GFP fluorescence gathered over time to particulate structures where PMP70 was co-localized. This process was dependent on both Pex5p and the targeting signal, and, most importantly, occurred even in the presence of cycloheximide, a protein synthesis inhibitor. These findings suggest that the ghosts act as acceptors of matrix proteins in the peroxisome recovery process at least in the PEX5 mutant, and support the view that peroxisomes can grow by incorporating newly synthesized matrix proteins.

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Section: Purification Of Recombinant Proteins-humansupporting

confidence: 79%

Formation of Peroxisomes from Peroxisomal Ghosts in a Peroxisome-deficient Mammalian Cell Mutant upon Complementation by Protein Microinjection

Yamasaki

Hashiguchi

Fujiwara

et al. 1999

Journal of Biological Chemistry

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“…Presently, our knowledge on the structure/function of amino acid residues 118 -639 of mammalian Pex5p is quite vast. Indeed, a variety of biochemical and structural studies have shown that this region of the protein contains the binding site for PTS1-containing cargo proteins (7,(22)(23)(24)(25)(26)(27), seven Pex14p-binding domains (43)(44)(45), and binding sites for Pex7p (11)(12)(13)(14), Pex12p (46), and Pex13p (45). In sharp contrast, not much is known regarding the role of the first 117 amino acid residues of Pex5p.…”

Section: Pex5p Lacking the First 110 Amino Acid Residues Is Efficientmentioning

confidence: 99%

The N Terminus of the Peroxisomal Cycling Receptor, Pex5p, Is Required for Redirecting the Peroxisome-associated Peroxin Back to the Cytosol

Costa‐Rodrigues

Carvalho

Gouveia³

et al. 2004

Journal of Biological Chemistry

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Most newly synthesized peroxisomal matrix proteins are transported to the organelle by Pex5p, a remarkable multidomain protein involved in an intricate network of transient protein-protein interactions. Presently, our knowledge regarding the structure/function of amino acid residues 118 to the very last residue of mammalian Pex5p is quite vast. Indeed, the cargo-protein receptor domain as well as the binding sites for several peroxins have all been mapped to this region of Pex5p. In contrast, structural/functional data regarding the first 117 amino acid residues of Pex5p are still scarce. Here we show that a truncated Pex5p lacking the first 110 amino acid residues (⌬N110-Pex5p) displays exactly the peroxisomal import properties of the full-length peroxin implying that this N-terminal domain is involved neither in cargo-protein binding nor in the docking/translocation step of the Pex5p-cargo protein complex at the peroxisomal membrane. However, the ATP-dependent export step of ⌬N110-Pex5p from the peroxisomal membrane is completely blocked, a phenomenon that was also observed for a Pex5p version lacking just the first 17 amino acid residues but not for a truncated protein comprising amino acid residues 1-324 of Pex5p. By exploring the unique properties of ⌬N110-Pex5p, the effect of temperature on the import/export kinetics of Pex5p was characterized. Our data indicate that the export step of Pex5p from the peroxisomal compartment (in contrast with its insertion into the organelle membrane) is highly dependent on the temperature.

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“…Pex5p recognizes matrix proteins via their C-terminal tripeptide motif (PTS1); Pex7p recognizes an N-terminally located amino acid motif (PTS2). The main components onto which cargo and receptors dock are Pex13p (an integral peroxisomal membrane protein) and Pex14p (a peroxisomal membrane-associated protein), as established by a combination of techniques: yeast two-hybrid interactions, co-immunoprecipitations and, recently, surface plasmon resonance analysis using purified proteins 9,22 . Well-defined (sub)domains exist in many of the Pex proteins (peroxins) for which the three-dimensional structure is known from other protein family members (see Table 1).…”

Section: Figurementioning

confidence: 99%