Tomato allene oxide synthase and fatty acid hydroperoxide lyase, two cytochrome P450s involved in oxylipin metabolism, are targeted to different membranes of chloroplast envelope.
Allene oxide synthase (AOS) and hydroperoxide lyase (HPL) are related cytochrome P450s that metabolize a common fatty acid hydroperoxide substrate to different classes of bioactive oxylipins within chloroplasts. Here, we report the use of in vitro import assays to investigate the targeting of tomato (Lycopersicon esculentum) AOS (LeAOS) and HPL (LeHPL) to isolated chloroplasts. LeAOS, which contains a typical N-terminal transit peptide, was targeted to the inner envelope membrane by a route that requires both ATP and proteinase-sensitive components on the surface of chloroplasts. Imported LeAOS was peripherally associated with the inner envelope; the bulk of the protein facing the stroma. LeHPL, which lacks a typical chloroplast-targeting sequence, was targeted to the outer envelope by an ATP-independent and protease-insensitive pathway. Imported LeHPL was integrated into the outer envelope with most of the protein exposed to the inter-membrane space. We conclude that LeAOS and LeHPL are routed to different envelope membranes by distinct targeting pathways. Partitioning of AOS and HPL to different envelope membranes suggests differences in the spatial organization of these two branches of oxylipin metabolism.
Froehlich
J E
JE
Department of Energy-Plant Research Laboratory, Michigan State University, East Lansing, Michigan 48824, USA.
Itoh
A
A
Howe
G A
GA
D000446:Aldehyde-Lyases / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D001483:Base Sequence; D002736:Chloroplasts / Q000201:enzymology*; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017931:DNA Primers; D007425:Intracellular Membranes / Q000201:enzymology; D019746:Intramolecular Oxidoreductases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018551:Solanum lycopersicum / Q000201:enzymology*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D018532:Pisum sativum / Q000201:enzymology*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
D017931:DNA Primers; D011994:Recombinant Proteins; D003577:Cytochrome P-450 Enzyme System; D000446:Aldehyde-Lyases; C054540:hydroperoxide lyase; D019746:Intramolecular Oxidoreductases; C021377:hydroperoxide isomerase