PER3, a Gene Required for Peroxisome Biogenesis in Pichia pastoris, Encodes a Peroxisomal Membrane Protein Involved in Protein Import

Henry Liu,Xuqiu Tan,Kimberly A Russell,Marten Veenhuis,James M Cregg

doi:10.1074/jbc.270.18.10940

Henry Liu, Xuqiu Tan + Show 3 more

Open Access

https://doi.org/10.1074/jbc.270.18.10940

Copy DOI

Abstract

PER genes are essential for the biogenesis of peroxisomes in the yeast Pichia pastoris. Here we describe the cloning of PER3 and functional characterization of its product Per3p. The PER3 sequence predicts that Per3p is a 713-amino acid (81-kDa) hydrophobic protein with at least three potential membrane-spanning domains. We show that Per3p is a membrane protein of the peroxisome. Methanol- or oleate-induced cells of per3-1, a mutant strain generated by chemical mutagenesis, lack normal peroxisomes but contain numerous abnormal vesicular structures. The vesicles contain thiolase, a PTS2 protein, but only a small portion of several other peroxisomal enzymes, including heterologously expressed luciferase, a PTS1 protein. These results suggest that the vesicles in per3-1 cells are peroxisomal remnants similar to those observed in cells of patients with the peroxisomal disorder Zellweger syndrome, and that the mutant is deficient in PTS1 but not PTS2 import. In a strain in which most of PER3 was deleted, peroxisomes as well as peroxisomal remnants appeared to be completely absent, and both PTS1- and PTS2-containing enzymes were located in the cytosol. We propose that Per3p is an essential component of the machinery required for import of all peroxisomal matrix proteins and is composed of independent domains involved in the import of specific PTS groups.

Highlights

Ing signals (PTSS)l have been characterized in detail [9]
Alcohol oxidase, an abundant peroxisomal enzyme required for methanol metabolism, is present at greatly reduced levels in methanol-induced per mutant cells, a phenomenon that is observed with certain peroxisomal enzymes in Zellweger cell lines
We report that pe r3 (PER3) encodes a peroxisomal membrane protein that appears to be involved in peroxisomal protein import

Summary

Introduction

Ing signals (PTSS)l have been characterized in detail [9]. The first, PTS1, is a tripeptide motif, Ser-Lys-Leu (and conservative variants), that is located at the carboxyl terminus of many peroxisomal matrix proteins [10, 11]. Cells ind uced with eit he r meth anol or oleate , ca ta lase (CAT) activity wa s present at a pproxima te ly t he sa me level as wild-type cells (Fig. 2A), In contrast, activity for alcohol oxidase (AOX), th e fir st e nzy me in meth a nol catabolism and a major cons t it ue nt of th e peroxisomal matrix of methanol-grown cell s , was cons iste nt ly pr esen t at levels that were less than 3'# of th ose in wild -typ e P. pastoris .

Results

Conclusion