Figure 5: Recognition of a PMP-derived peptide by the farnesylated PEX19 CTD.

(a) Amino acid sequences of peroxisomal membrane proteins. Aromatic residues are highlighted in red; regions known to be involved in PEX19 binding are shown in bold. (b) Docking models showing two binding pockets for aromatic side chains of the PMP-derived peptide (green stick and ball representation). Top: The PEX19 lid and helix α1 are coloured dark blue and cyan, respectively. Side chains of M179, Leu182, Pro272 and Pro273 are shown as sticks. Bottom: Surface representation of the PEX19 CTD indicating the hydrophobic cavities that accommodate the aromatic side chain of Phe71 in the ALDP peptide. (c) In vivo effects of mutations of the PMP binding site of PEX19. Immunofluorescence microscopy images of PEX19-deficient human fibroblasts transfected with bicistronic vectors encoding for eGFP-PTS1 and different PEX19 variants. The same plasmid lacking PEX19 was used as a negative control (ΔPEX19) showing diffuse staining due to mitochondrial mislocalization for both peroxisomal marker proteins, eGFP-PTS1 (matrix protein) and PEX14 (PMP). The same plasmid lacking PEX19 was used as a negative control (ΔPEX19) showing cytosolic and mitochondrial mislocalization for the peroxisomal marker proteins eGFP-PTS1 (matrix protein) and PEX14 (PMP), respectively. A congruent punctate pattern of eGFP-SKL and PEX14 indicates formation of import-competent peroxisomes 72 h after transfection. Scale bar: 10 μm. (d) Statistical analysis of number of transfected cells containing import-competent peroxisomes. Values were obtained from 100 cells each of three independent transfected experiments. Data shown represent mean±s.d. (e) Immunoblot analysis of mutants shows that all variants are present in an amount similar to the wild type protein.