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Membrane topologies of PEX 13 and PEX 14 provide new insights on the mechanism of protein import into peroxisomes
Author(s) -
BarrosBarbosa Aurora,
Ferreira Maria J.,
Rodrigues Tony A.,
Pedrosa Ana G.,
Grou Cláudia P.,
Pinto Manuel P.,
Fransen Marc,
Francisco Tânia,
Azevedo Jorge E.
Publication year - 2019
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/febs.14697
Subject(s) - peroxisome , organelle , peroxisomal targeting signal , transmembrane protein , microbiology and biotechnology , biology , membrane protein , protein targeting , transmembrane domain , biochemistry , chemistry , receptor , membrane
PEX 13 and PEX 14 are two core components of the so‐called peroxisomal docking/translocation module, the transmembrane hydrophilic channel through which newly synthesized peroxisomal proteins are translocated into the organelle matrix. The two proteins interact with each other and with PEX 5, the peroxisomal matrix protein shuttling receptor, through relatively well characterized domains. However, the topologies of these membrane proteins are still poorly defined. Here, we subjected proteoliposomes containing PEX 13 or PEX 14 and purified rat liver peroxisomes to protease‐protection assays and analyzed the protected protein fragments by mass spectrometry, Edman degradation and western blotting using antibodies directed to specific domains of the proteins. Our results indicate that PEX 14 is a bona fide intrinsic membrane protein with a N in ‐C out topology, and that PEX 13 adopts a N out ‐C in topology, thus exposing its carboxy‐terminal Src homology 3 [ SH 3] domain into the organelle matrix. These results reconcile several enigmatic findings previously reported on PEX 13 and PEX 14 and provide new insights into the organization of the peroxisomal protein import machinery. Enzymes Trypsin, EC3.4.21.4 ; Proteinase K, EC3.4.21.64 ; Tobacco etch virus protease, EC3.4.22.44 .