The peroxisomal membrane protein Pex13p shows a novel mode of SH3 interaction

Src homology 3 (SH3) domains are small non-catalytic protein modules capabl e of mediating protein-protein interactions by binding to proline-X-X-proli ne (P-X-Y-P) moths. Here we demonstrate that the SH3 domain of the integral peroxisomal membrane protein Pex13p is able to bind two proteins, one of w hich, Pex5p, represents a novel non-P-X-X-P ligand. Using alanine scanning, two-hybrid and irt vitro interaction analysis, we show that an alpha -heli cal element in Pex5p is necessary and sufficient for SH3 interaction. Suppr essor analysis using Pex5p mutants located in this alpha -helical element a llowed the identification of a unique site of interaction for Pex5p on the Pex13pSH3 domain that is distinct from the classical P-X-X-P binding pocket . On the basis of a structural model of the Pex13p-SH3 domain we show that this interaction probably takes place between the RT- and distal loops. Thu s, the Pes13p-SH3-Pex5p interaction establishes a novel mode of SH3 interac tion.