title: The role of conserved PEX3 regions in PEX19-binding and peroxisome biogenesis.
abstract: The human peroxins PEX3 and PEX19 are essential for peroxisome biogenesis. They mediate the import of membrane proteins as well as the de novo formation of peroxisomes. PEX19 binds newly synthesized peroxisomal membrane proteins post-translationally and directs them to peroxisomes by engaging PEX3, a protein anchored in the peroxisomal membrane. After protein insertion into the lipid bilayer, PEX19 is released back to the cytosol. Crystallographic analysis provided detailed insights into the PEX3-PEX19 interaction and identified three highly conserved regions, the PEX19-binding region, a hydrophobic groove and an acidic cluster, on the surface of PEX3. Here, we used site-directed mutagenesis and biochemical and functional assays to determine the role of these regions in PEX19-binding and peroxisome biogenesis. Mutations in the PEX19-binding region reduce the affinity for PEX19 and destabilize PEX3. Furthermore, we provide evidence for a crucial function of the PEX3-PEX19 complex during de novo formation of peroxisomes in peroxisome-deficient cells, pointing to a dual function of the PEX3-PEX19 interaction in peroxisome biogenesis. The maturation of preperoxisomes appears to require the hydrophobic groove near the base of PEX3, presumably by its involvement in peroxisomal membrane protein insertion, while the acidic cluster does not appear to be functionally relevant.
authors.affiliation: Interfaculty Institute of Biochemistry, University of Tübingen, D-72076, Tübingen, Germany.
authors.forename: Friederike
authors.initials: F
authors.lastname: Schmidt
authors.forename: Denise
authors.initials: D
authors.lastname: Dietrich
authors.forename: Roy
authors.initials: R
authors.lastname: Eylenstein
authors.forename: Yvonne
authors.initials: Y
authors.lastname: Groemping
authors.forename: Thilo
authors.initials: T
authors.lastname: Stehle
authors.forename: Gabriele
authors.initials: G
authors.lastname: Dodt
mesh_terms: D000595:Amino Acid Sequence; D002460:Cell Line; D017353:Gene Deletion; D006801:Humans; D007425:Intracellular Membranes / Q000378:metabolism; D008074:Lipoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D000074401:Peroxins; D020675:Peroxisomes / Q000378:metabolism*; D054730:Protein Interaction Domains and Motifs; D034741:RNA, Small Interfering
chemical_list: D008074:Lipoproteins; D008565:Membrane Proteins; D000074401:Peroxins; C113138:Pex3 protein, human; D034741:RNA, Small Interfering; C087343:PEX19 protein, human