jasmonic-acid and cyclopentanone

In plants, oxylipins regulate developmental processes and defense responses. The first specific step in the biosynthesis of the cyclopentanone class of oxylipins is catalyzed by allene oxide cyclase (AOC) that forms cis(+)-12-oxo-phytodienoic acid. The moss Physcomitrella patens has two AOCs (PpAOC1 and PpAOC2) with different substrate specificities for C₁₈- and C₂₀-derived substrates, respectively. To better understand AOC's catalytic mechanism and to elucidate the structural properties that explain the differences in substrate specificity, we solved and analyzed the crystal structures of 36 monomers of both apo and ligand complexes of PpAOC1 and PpAOC2. From these data, we propose the following intermediates in AOC catalysis: (1) a resting state of the apo enzyme with a closed conformation, (2) a first shallow binding mode, followed by (3) a tight binding of the substrate accompanied by conformational changes in the binding pocket, and (4) initiation of the catalytic cycle by opening of the epoxide ring. As expected, the substrate dihydro analog cis-12,13S-epoxy-9Z,15Z-octadecadienoic acid did not cyclize in the presence of PpAOC1; however, when bound to the enzyme, it underwent isomerization into the corresponding trans-epoxide. By comparing complex structures of the C₁₈ substrate analog with in silico modeling of the C₂₀ substrate analog bound to the enzyme allowed us to identify three major molecular determinants responsible for the different substrate specificities (i.e. larger active site diameter, an elongated cavity of PpAOC2, and two nonidentical residues at the entrance of the active site).

Topics: Biocatalysis; Biosynthetic Pathways; Bryopsida; Catalytic Domain; Crystallography, X-Ray; Cyclopentanes; Fatty Acids, Unsaturated; Intramolecular Oxidoreductases; Isoenzymes; Isomerism; Models, Molecular; Oxylipins; Protein Structure, Secondary; Substrate Specificity

12-oxo-phytodienoic acid and several phytoprostanes are cyclopentenone oxylipins that are formed via the enzymatic jasmonate pathway and a nonenzymatic, free radical-catalyzed pathway, respectively. Both types of cyclopentenone oxylipins induce the expression of genes related to detoxification, stress responses, and secondary metabolism, a profile clearly distinct from that of the cyclopentanone jasmonic acid. Microarray analyses revealed that 60% of the induction by phytoprostanes and 30% of the induction by 12-oxo-phytodienoic acid was dependent on the TGA transcription factors TGA2, TGA5, and TGA6. Moreover, treatment with phytoprostanes and 12-oxo-phytodienoic acid inhibited cell division and root growth, a property also shared by jasmonic acid. Besides being potent signals, cyclopentenones and other lipid peroxidation products are reactive electrophiles that can covalently bind to and damage proteins. To this end, we show that at least two of the induced detoxification enzymes efficiently metabolize cyclopentenones in vitro. Accumulation of two of these metabolites was detectable during Pseudomonas infection. The cyclopentenone oxylipin gene induction profile resembles the defense response induced by a variety of lipophilic xenobiotics. Hence, oxidized lipids may activate chemosensory mechanisms of a general broad-spectrum detoxification network involving TGA transcription factors.

Topics: Arabidopsis; Arabidopsis Proteins; Basic-Leucine Zipper Transcription Factors; Cell Cycle; Cell Division; Cyclopentanes; Fatty Acids, Unsaturated; Gene Expression Regulation, Plant; Nuclear Proteins; Oligonucleotide Array Sequence Analysis; Oxylipins; Plant Roots; Pseudomonas; Reverse Transcriptase Polymerase Chain Reaction; Transcriptional Activation