Analysis of the Carboxyl-terminal Peroxisomal Targeting Signal 1 in a Homologous Context in Saccharomyces cerevisiae

Abstract

Most peroxisomal matrix proteins contain a carboxylterminal tripeptide that directs them to peroxisomes. Within limits, these amino acids may be varied, without loss of function. The specificity of this peroxisomal targeting signal (PTS1) is remarkable considering its small size and its relaxed consensus sequence. Moreover, several peroxisomal proteins have a PTS1-like signal that does not fit the reported consensus sequence. Because many of these PTS1 variants seem to be functional in a species-dependent or protein context-dependent manner, we investigated the PTS1 requirements in a homologous context, using Saccharomyces cerevisiae and endogenous peroxisomal malate dehydrogenase (MDH3). Peroxisomal import of the MDH3-PTS1 variants was tested qualitatively by the ability to complement the ⌬mdh3 mutant and quantitatively by subcellular fractionation. We observed efficient import of MDH3 into peroxisomes with a large variety of PTS1 tripeptides. Many of these variants do not fit the observed PTS1 requirements for heterologously expressed proteins, which suggests that additional domains in the protein may be of decisive importance whether or not a certain PTS1 variant is recognized by the components of the peroxisomal import machinery. Because we show that dimerization of MDH3 precedes import into the organelle, these domains are most likely conformational domains.

The methylotrophic yeast Hansenula polymorpha: a versatile cell factory

“…The first identified PTS1, -SKL-COOH), was found in firefly luciferase. Later studies revealed that several amino acid substitutions are allowed in this sequence [37]. In H. polymorpha the enzymes AOX (-ARF), DHAS (-NKL), catalase (-SKI), and the peroxisomal matrix protein Pex8p (-AKL; see Fig.…”

Yarrowia lipolyticaCells Mutant for the Peroxisomal Peroxin Pex19p Contain Structures Resembling Wild-Type Peroxisomes

“…Most soluble matrix proteins are targeted to peroxisomes by one of two types of peroxisomal targeting signals (PTS). PTS1 is a carboxylterminal tripeptide (SKL and conserved variants; Gould et al, 1987Gould et al, , 1989Aitchison et al, 1991;Elgersma et al, 1996) found in the majority of matrix proteins, whereas PTS2 is an amino-terminal nonapeptide found in a very limited subset of matrix proteins (Swinkels et al, 1991;Glover et al, 1994;Waterham et al, 1994). Pex5p and Pex7p are the receptors for PTS1 and PTS2, respectively, and various peroxins, including Pex13p and Pex14p, form a docking complex at the peroxisomal membrane for these receptors (reviewed by Erdmann et al, 1997;Subramani, 1998Titorenko and Rachubinski, 2001).…”

Mutants of theYarrowia lipolytica PEX23Gene Encoding an Integral Peroxisomal Membrane Peroxin Mislocalize Matrix Proteins and Accumulate Vesicles Containing Peroxisomal Matrix and Membrane Proteins

“…Most soluble proteins of the matrix are targeted by one of two types of peroxisomal targeting signal (PTS). PTS1 is a carboxyl-terminal tripeptide (SKL and conserved variants) (Gould et al, 1987(Gould et al, , 1989 found in a large number of matrix proteins (Gould et al, 1989;Motley et al, 1995;Elgersma et al, 1996b), whereas PTS2 is a sometimes cleaved amino-terminal nonapeptide found in a smaller subset of matrix proteins (Swinkels et al, 1991;Glover et al, 1994;Waterham et al, 1994). Pex5p and Pex7p are the receptors for PTS1 and PTS2, respectively, and various peroxins, including Pex13p and Pex14p, form a docking complex at the peroxisomal membrane for these receptors (reviewed by Erdmann et al, 1997;Subramani, 1998).…”