Plant Cell Advance Online Publication
Published on October 19, 2007; 10.1105/tpc.107.050989
Received February 7, 2007
Returned for revision September 12, 2007
Accepted September 24, 2007
Proteome Analysis of Arabidopsis Leaf Peroxisomes Reveals Novel Targeting Peptides, Metabolic Pathways, and Defense Mechanisms
Sigrun Reumann 1*, Lavanya Babujee 1, Changle Ma 1, Stephanie Wienkoop 2, Tanja Siemsen 1, Gerardo E. Antonicelli 1, Nicolas Rasche 1, Franziska Lüder 1, Wolfram Weckwerth 3, and Olaf Jahn 4
1 Department of Plant Biochemistry, Georg-August-University of Goettingen, Albrecht-von-Haller-Institute for Plant Sciences, D-37077 Goettingen, Germany
2 Max-Planck-Institute of Molecular Plant Physiology, Metabolic Networks, D-14424 Potsdam, Germany
3 Max-Planck-Institute of Molecular Plant Physiology, Metabolic Networks, D-14424 Potsdam, Germany; University of Potsdam, Institute of Biochemistry and Biology, 14469 Potsdam, Germany
4 Max-Planck-Institute of Experimental Medicine, Proteomics Group, D-37075 Goettingen, Germany; Deutsche Forschungsgemeinschaft Research Center for Molecular Physiology of the Brain, D-37073 Goettingen, Germany
* To whom correspondence should be addressed. E-mail: sreuman{at}gwdg.de.
We have established a protocol for the isolation of highly purified peroxisomes from mature Arabidopsis thaliana leaves and analyzed the proteome by complementary gel-based and gel-free approaches. Seventy-eight nonredundant proteins were identified, of which 42 novel proteins had previously not been associated with plant peroxisomes. Seventeen novel proteins carried predicted peroxisomal targeting signals (PTS) type 1 or type 2; 11 proteins contained PTS-related peptides. Peroxisome targeting was supported for many novel proteins by in silico analyses and confirmed for 11 representative full-length fusion proteins by fluorescence microscopy. The targeting function of predicted and unpredicted signals was investigated and SSL>, SSI>, and ASL> were established as novel functional PTS1 peptides. In contrast with the generally accepted confinement of PTS2 peptides to the N-terminal domain, the bifunctional transthyretin-like protein was demonstrated to carry internally a functional PTS2. The novel enzymes include numerous enoyl-CoA hydratases, short-chain dehydrogenases, and several enzymes involved in NADP and glutathione metabolism. Seven proteins, including  -glucosidases and myrosinases, support the currently emerging evidence for an important role of leaf peroxisomes in defense against pathogens and herbivores. The data provide new insights into the biology of plant peroxisomes and improve the prediction accuracy of peroxisome-targeted proteins from genome sequences.
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