title: The N-terminal beta-barrel structure of lipid body lipoxygenase mediates its binding to liposomes and lipid bodies.
abstract: Phospholipase A2 and a particular isoform of lipoxygenase are synthesized and transferred to lipid bodies during the stage of triacylglycerol mobilization in germinating cucumber seedlings. Lipid body lipoxygenase (LBLOX) is post-translationally transported to lipid bodies without proteolytic modification. Fractionation of homogenates from cucumber cotyledons or transgenic tobacco leaves expressing LBLOX showed that a small but significant amount was detectable in the microsomal fraction. A beta-barrel-forming N-terminal domain in the structure of LBLOX, as deduced from sequence data, was shown to be crucial for selective intracellular transport from the cytosol to lipid bodies. Although a specific signal sequence for targeting protein domains to the lipid bodies could not be established, it was evident that the beta-barrel represents a membrane-binding domain that is functionally comparable with the C2 domains of mammalian phospholipases. The intact beta-barrel of LBLOX was demonstrated to be sufficient to target in vitro a fusion protein of LBLOX beta-barrel with glutathione S-transferase (GST) to lipid bodies. In addition, binding experiments on liposomes using lipoxygenase isoforms, LBLOX deletions and the GST-fusion protein confirmed the role of the beta-barrel as the membrane-targeting domain. In this respect, the cucumber LBLOX differs from cytosolic isoforms in cucumber and from the soybean LOX-1. When the beta-barrel of LBLOX was destroyed by insertion of an additional peptide sequence, its ability to target proteins to membranes was abolished.
authors.affiliation: Fachbereich Chemie der Philipps-Universität, Marburg, Germany.
authors.forename: C
authors.initials: C
authors.lastname: May
authors.forename: M
authors.initials: M
authors.lastname: Höhne
authors.forename: P
authors.initials: P
authors.lastname: Gnau
authors.forename: K
authors.initials: K
authors.lastname: Schwennesen
authors.forename: H
authors.initials: H
authors.lastname: Kindl
mesh_terms: D001665:Binding Sites; D001692:Biological Transport; D018553:Cucumis sativus / Q000166:cytology / Q000201:enzymology; D003600:Cytosol / Q000201:enzymology / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000201:enzymology / Q000378:metabolism; D007425:Intracellular Membranes / Q000201:enzymology / Q000378:metabolism; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008081:Liposomes / Q000378:metabolism*; D008084:Lipoxygenase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008861:Microsomes / Q000201:enzymology / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D010741:Phospholipases A / Q000378:metabolism; D054467:Phospholipases A2; D018514:Plant Structures / Q000166:cytology / Q000201:enzymology* / Q000378:metabolism*; D030821:Plants, Genetically Modified; D010947:Plants, Toxic; D011485:Protein Binding; D014176:Protein Biosynthesis / Q000502:physiology; D021382:Protein Sorting Signals / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014026:Nicotiana / Q000166:cytology / Q000201:enzymology / Q000235:genetics
chemical_list: D007527:Isoenzymes; D008081:Liposomes; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D008084:Lipoxygenase; D010741:Phospholipases A; D054467:Phospholipases A2