Allosteric modulation of peroxisomal membrane protein recognition by farnesylation of the peroxisomal import receptor PEX19

Leonidas Emmanouilidis,Konstantinos Tripsianes,Matthias Wilmanns,Ralf Erdmann,Ulrike Schütz,Juliane Radke,Tobias Madl,Wolfgang Schliebs,Robert Rucktäschel,Michael Sattler

doi:10.1038/ncomms14635

Abstract

The transport of peroxisomal membrane proteins (PMPs) requires the soluble PEX19 protein as chaperone and import receptor. Recognition of cargo PMPs by the C-terminal domain (CTD) of PEX19 is required for peroxisome biogenesis in vivo. Farnesylation at a C-terminal CaaX motif in PEX19 enhances the PMP interaction, but the underlying molecular mechanisms are unknown. Here, we report the NMR-derived structure of the farnesylated human PEX19 CTD, which reveals that the farnesyl moiety is buried in an internal hydrophobic cavity. This induces substantial conformational changes that allosterically reshape the PEX19 surface to form two hydrophobic pockets for the recognition of conserved aromatic/aliphatic side chains in PMPs. Mutations of PEX19 residues that either mediate farnesyl contacts or are directly involved in PMP recognition abolish cargo binding and cannot complement a ΔPEX19 phenotype in human Zellweger patient fibroblasts. Our results demonstrate an allosteric mechanism for the modulation of protein function by farnesylation.

Highlights

The transport of peroxisomal membrane proteins (PMPs) requires the soluble Peroxisomal biogenesis factor 19 (PEX19) protein as chaperone and import receptor
To investigate the effects of farnesylation, we studied the human PEX19 C-terminal domain (CTD) that includes the native C-terminus with the CaaX box of the protein (Fig. 1a)
Our structural analysis shows that the farnesyl group in the PEX19 CTD is buried within a hydrophobic cavity

Summary

Introduction

The transport of peroxisomal membrane proteins (PMPs) requires the soluble PEX19 protein as chaperone and import receptor. Recognition of cargo PMPs by the C-terminal domain (CTD) of PEX19 is required for peroxisome biogenesis in vivo. PEX19 directs the cargo to the peroxisomal membrane, where it docks to the transmembrane protein PEX3 thereby acting as a shuttling receptor[6] It could be involved in membrane insertion of PMPs (refs 7,8). The crystallized PEX19 fragment lacks the C-terminal 16 amino acids, which include the farnesylation site, and the structural impact of farnesylation and its role in the modulation of PMP binding remain unknown. We present the solution structure of the farnesylated C-terminal PMP binding domain of human PEX19 and report molecular details for the recognition of hydrophobic residues in PMPs determined by NMR spectroscopy. Mutations interfering with either farnesyl recognition or PMP interactions affect the PMP binding in vitro and the biological activity of PEX19 during peroxisome biogenesis in vivo. These findings reveal a novel mechanism for the modulation of protein function by farnesylation

Methods

Results

Conclusion