title: The allene oxide cyclase family of Arabidopsis thaliana: localization and cyclization.
abstract: Jasmonates are derived from oxygenated fatty acids (oxylipins) via the octadecanoid pathway and are characterized by a pentacyclic ring structure. They have regulatory functions as signaling molecules in plant development and adaptation to environmental stress. Recently, we solved the structure of allene oxide cyclase 2 (AOC2) of Arabidopsis thaliana, which is, together with the other three AOCs, a key enzyme in the biosynthesis of jasmonates, in that it releases the first cyclic and biologically active metabolite -- 12-oxo-phytodienoic acid (OPDA). On the basis of models for the bound substrate, 12,13(S)-epoxy-9(Z),11,15(Z)-octadecatrienoic acid, and the product, OPDA, we proposed that a conserved Glu promotes the reaction by anchimeric assistance. According to this hypothesis, the transition state with a pentadienyl carbocation and an oxyanion is stabilized by a strongly bound water molecule and favorable pi-pi interactions with aromatic residues in the cavity. Stereoselectivity results from steric restrictions to the necessary substrate isomerizations imposed by the protein environment. Here, site-directed mutagenesis was used to explore and verify the proposed reaction mechanism. In a comparative analysis of the AOC family from A. thaliana involving enzymatic characterization, in vitro import, and transient expression of AOC-enhanced green fluorescent protein fusion proteins for analysis of subcellular targeting, we demonstrate that all four AOC isoenzymes may contribute to jasmonate biosynthesis, as they are all located in chloroplasts and, in concert with the allene oxide synthase, they are all able to convert 13(S)-hydroperoxy-9(Z),11(E),15(Z)-octadecatrienoic acid into enantiomerically pure cis(+)-OPDA.
authors.affiliation: Lehrstuhl für Pflanzenphysiologie, Ruhr-Universität Bochum, Germany.
authors.forename: Florian
authors.initials: F
authors.lastname: Schaller
authors.forename: Philipp
authors.initials: P
authors.lastname: Zerbe
authors.forename: Steffen
authors.initials: S
authors.lastname: Reinbothe
authors.forename: Christiane
authors.initials: C
authors.lastname: Reinbothe
authors.forename: Eckhard
authors.initials: E
authors.lastname: Hofmann
authors.forename: Stephan
authors.initials: S
authors.lastname: Pollmann
mesh_terms: D000818:Animals; D017360:Arabidopsis / Q000201:enzymology*; D029681:Arabidopsis Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D003500:Cyclization; D003517:Cyclopentanes / Q000737:chemistry / Q000378:metabolism; D005231:Fatty Acids, Unsaturated / Q000737:chemistry / Q000378:metabolism; D019746:Intramolecular Oxidoreductases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D007527:Isoenzymes / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D015394:Molecular Structure; D054883:Oxylipins / Q000737:chemistry / Q000378:metabolism; D017354:Point Mutation; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity; D014018:Tissue Distribution; D003313:Zea mays / Q000201:enzymology
chemical_list: D029681:Arabidopsis Proteins; D003517:Cyclopentanes; D005231:Fatty Acids, Unsaturated; D007527:Isoenzymes; D054883:Oxylipins; D011993:Recombinant Fusion Proteins; C025999:12-oxophytodienoic acid; C011006:jasmonic acid; D019746:Intramolecular Oxidoreductases; C021377:hydroperoxide isomerase