title: Activity Regulation by Heteromerization of Arabidopsis Allene Oxide Cyclase Family Members.
abstract: Jasmonates (JAs) are lipid-derived signals in plant stress responses and development. A crucial step in JA biosynthesis is catalyzed by allene oxide cyclase (AOC). Four genes encoding functional AOCs (AOC1, AOC2, AOC3 and AOC4) have been characterized for Arabidopsis thaliana in terms of organ- and tissue-specific expression, mutant phenotypes, promoter activities and initial in vivo protein interaction studies suggesting functional redundancy and diversification, including first hints at enzyme activity control by protein-protein interaction. Here, these analyses were extended by detailed analysis of recombinant proteins produced in Escherichia coli. Treatment of purified AOC2 with SDS at different temperatures, chemical cross-linking experiments and protein structure analysis by molecular modelling approaches were performed. Several salt bridges between monomers and a hydrophobic core within the AOC2 trimer were identified and functionally proven by site-directed mutagenesis. The data obtained showed that AOC2 acts as a trimer. Finally, AOC activity was determined in heteromers formed by pairwise combinations of the four AOC isoforms. The highest activities were found for heteromers containing AOC4 + AOC1 and AOC4 + AOC2, respectively. All data are in line with an enzyme activity control of all four AOCs by heteromerization, thereby supporting a putative fine-tuning in JA formation by various regulatory principles.
authors.affiliation: Department of Cell and Metabolic Biology, Leibniz Institute of Plant Biochemistry, Weinberg 3, D-06120 Halle (Saale), Germany. mrmarkus.otto@googlemail.com.|Department of Molecular Signal Processing, Leibniz Institute of Plant Biochemistry, Weinberg 3, D-06120 Halle (Saale), Germany. mrmarkus.otto@googlemail.com.
authors.forename: Markus
authors.initials: M
authors.lastname: Otto
authors.affiliation: Department of Cell and Metabolic Biology, Leibniz Institute of Plant Biochemistry, Weinberg 3, D-06120 Halle (Saale), Germany. christin.naumann@ipb-halle.de.
authors.forename: Christin
authors.initials: C
authors.lastname: Naumann
authors.affiliation: Department of Natural Product Chemistry, Leibniz Institute of Plant Biochemistry, Weinberg 3, D-06120 Halle (Saale), Germany. wbrandt@ipb-halle.de.
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Brandt
authors.affiliation: Department of Molecular Signal Processing, Leibniz Institute of Plant Biochemistry, Weinberg 3, D-06120 Halle (Saale), Germany. cwastern@ipb-halle.de.|Laboratory of Growth Regulators, Centre of the Region Haná for Biotechnological and Agricultural Research, Institute of Experimental Botany AS CR &amp; Palacký University, Šlechtitelů 11, CZ-78371 Olomouc, Czech Republic. cwastern@ipb-halle.de.
authors.forename: Claus
authors.initials: C
authors.lastname: Wasternack
authors.affiliation: Department of Cell and Metabolic Biology, Leibniz Institute of Plant Biochemistry, Weinberg 3, D-06120 Halle (Saale), Germany. bhause@ipb-halle.de.
authors.forename: Bettina
authors.initials: B
authors.lastname: Hause
keywords: Arabidopsis allene oxide cyclase isoforms; activity regulation; heteromerization; protein structure analysis; site-directed mutagenesis