Mammalian Pex14p: membrane topology and characterisation of the Pex14p–Pex14p interaction

Oliveira, Márcia E.; Reguenga, Carlos; Gouveia, Alexandra; Guimarães, Carla P.; Schliebs, Wolfgang; Kunau, Wolf‐H.; Silva, Manuel T.; Sá-Miranda, Clara; Azevedo, Jorge E.

doi:10.1016/s0005-2736(02)00635-1

Cited by 46 publications

(142 citation statements)

References 43 publications

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“…Mammalian PEX14 is a transmembrane protein and forms homodimers or larger oligomers in the transmembrane domain. The N-terminus is protected from chemical or proteolytic cleavage Oliveira et al, 2002). This portion of PEX14 has been shown to interact directly with PEX5 via the W-X 3 -F/Y pentapeptide repeat units of the Nterminal half of PEX5 (Schliebs et al, 1999;Nito et al, 2002;Otera et al, 2002).…”

Section: Docking Complexmentioning

confidence: 99%

Getting a camel through the eye of a needle: the import of folded proteins by peroxisomes

Lanyon‐Hogg

Warriner

Baker

2010

Biology of the Cell

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Peroxisomes are a family of organelles which have many unusual features. They can arise de novo from the endoplasmic reticulum by a still poorly characterized process, yet possess a unique machinery for the import of their matrix proteins. As peroxisomes lack DNA, their function, which is highly variable and dependent on developmental and/or environmental conditions, is determined by the post-translational import of specific metabolic enzymes in folded or oligomeric states. The two classes of matrix targeting signals for peroxisomal proteins [PTS1 (peroxisomal targeting signal 1) and PTS2] are recognized by cytosolic receptors [PEX5 (peroxin 5) and PEX7 respectively] which escort their cargo proteins to, or possibly across, the peroxisome membrane. Although the membrane translocation mechanism remains unclear, it appears to be driven by thermodynamically favourable binding interactions. Recycling of the receptors from the peroxisome membrane requires ATP hydrolysis for two linked processes: ubiquitination of PEX5 (and the PEX7 co-receptors in yeast) and the function of two peroxisome-associated AAA (ATPase associated with various cellular activities) ATPases, which play a role in recycling or turnover of the ubiquitinated receptors. This review summarizes and integrates recent findings on peroxisome matrix protein import from yeast, plant and mammalian model systems, and discusses some of the gaps in our understanding of this remarkable protein transport system.

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Section: Docking Complexmentioning

confidence: 99%

Getting a camel through the eye of a needle: the import of folded proteins by peroxisomes

Lanyon‐Hogg

Warriner

Baker

2010

Biology of the Cell

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“…The amplified fragments were digested with BamHI and KpnI, and inserted into pQE-30 (QIAGEN) according to the manufacturer's instructions. The cDNA encoding the first 110 amino acid residues of Pex5p was obtained from pGEM-4-Pex5p 62 by PCR using the forward primer 5 0 -CCGGCATGCGCAATGCGG-GAGCTGGTGG and the reverse primer 5 0 -CCGGTC-GACGCCACACCAGGGCTCTCTGG-3 0 . The fragment was cloned into the pGEM w -T easy vector according to the manufacturer's instructions (Promega).…”

Section: Recombinant Proteinsmentioning

confidence: 99%

The N-terminal Half of the Peroxisomal Cycling Receptor Pex5p is a Natively Unfolded Domain

Carvalho

Costa‐Rodrigues

Correia

et al. 2006

Journal of Molecular Biology

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Targeting of most newly synthesised peroxisomal matrix proteins to the organelle requires Pex5p, the so-called PTS1 receptor. According to current models of peroxisomal biogenesis, Pex5p interacts with these proteins in the cytosol, transports them to the peroxisomal membrane and catalyses their translocation across the membrane. Presently, our knowledge on the structural details behind the interaction of Pex5p with the cargo proteins is reasonably complete. In contrast, information regarding the structure of the Pex5p N-terminal half (a region containing its peroxisomal targeting domain) is still limited. We have recently observed that the Stokes radius of this Pex5p domain is anomalously large, suggesting that this portion of the protein is either a structured elongated domain or that it adopts a low compactness conformation. Here, we address this issue using a combination of biophysical and biochemical approaches. Our results indicate that the N-terminal half of Pex5p is best described as a natively unfolded premolten globule-like domain. The implications of these findings on the mechanism of protein import into the peroxisome are discussed.

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“…Total IgGs were purified from rabbit sera using protein A-Sepharose beads according to the manufacturer's protocol (Sigma). PEX5 in HeLa cell lysates was detected by blot overlay using 35 S-labeled PEX14, as described previously (56). Preparation of samples for SDS-PAGE under nonreducing conditions (necessary to preserve the Ub-PEX5 thioester conjugate) and autoradiography following Western blot were described previously (33,38).…”

mentioning

confidence: 99%

Identification of Ubiquitin-specific Protease 9X (USP9X) as a Deubiquitinase Acting on Ubiquitin-Peroxin 5 (PEX5) Thioester Conjugate

Grou

Francisco

Rodrigues

et al. 2012

Journal of Biological Chemistry

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Background:The mammalian deubiquitinase that hydrolyzes the ubiquitin-PEX5 thioester conjugate was unknown. Results: USP9X was found to be the most active deubiquitinase acting on ubiquitin-PEX5. Conclusion:We propose that USP9X participates in the PEX5-mediated peroxisomal protein import pathway. Significance: The unbiased biochemical strategy described here will be useful to identify deubiquitinases acting on other substrates.

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Mammalian Pex14p: membrane topology and characterisation of the Pex14p–Pex14p interaction

Cited by 46 publications

References 43 publications

Getting a camel through the eye of a needle: the import of folded proteins by peroxisomes

Getting a camel through the eye of a needle: the import of folded proteins by peroxisomes

The N-terminal Half of the Peroxisomal Cycling Receptor Pex5p is a Natively Unfolded Domain

Identification of Ubiquitin-specific Protease 9X (USP9X) as a Deubiquitinase Acting on Ubiquitin-Peroxin 5 (PEX5) Thioester Conjugate

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