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          Institute: MPI für molekulare Pflanzenphysiologie     Collection: Publikationen Pflanzenphysiologie     Display Documents



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ID: 224231.0, MPI für molekulare Pflanzenphysiologie / Publikationen Pflanzenphysiologie
AtDGK2, a novel diacylglycerol kinase from Arabidopsis thaliana, phosphorylates 1-stearoyl-2-arachidonoyl-sn-glycerol and 1,2-dioleoyl-sn-glycerol and exhibits cold-inducible gene expression
Authors:Gomez-Merino, F. C.; Brearley, C. A.; Ornatowska, M.; Abdel-Haliem, M. E. F.; Zanor, M. I.; Mueller-Roeber, B.
Language:English
Date of Publication (YYYY-MM-DD):2004-02-27
Title of Journal:Journal of Biological Chemistry
Journal Abbrev.:J Biol Chem
Volume:279
Issue / Number:9
Start Page:8230
End Page:8241
Review Status:not specified
Audience:Not Specified
Abstract / Description:Diacylglycerol kinase (DGK) phosphorylates diacylglycerol (DAG) to generate phosphatidic acid (PA). Both DAG and PA are implicated in signal transduction pathways. DGKs have been widely studied in animals, but their analysis in plants is fragmentary. Here, we report the cloning and biochemical characterization of AtDGK2, encoding DGK from Arabidopsis thaliana. AtDGK2 has a predicted molecular mass of 79.4 kDa and, like AtDGK1 previously reported, harbors two copies of a phorbol ester/DAG-binding domain in its N-terminal region. AtDGK2 belongs to a family of seven DGK genes in A. thaliana. AtDGK3 to AtDGK7 encode similar to55-kDa DGKs that lack a typical phorbol ester/DAG-binding domain. Phylogenetically, plant DGKs fall into three clusters. Members of all three clusters are widely expressed in vascular plants. Recombinant AtDGK2 was expressed in Escherichia coli and biochemically characterized. The enzyme phosphorylated 1,2-dioleoyl-sn-glycerol to yield PA, exhibiting Michaelis-Menten type kinetics. Estimated K-m and V-max values were 125 muM for DAG and 0.25 pmol of PA min(-1) mug(-1), respectively. The enzyme was maximally active at pH 7.2. Its activity was Mg2+-dependent and affected by the presence of detergents, salts, and the DGK inhibitor R59022, but not by Ca2+. AtDGK2 exhibited substrate preference for unsaturated DAG analogues (i.e. 1-stearoyl-2-arachidonoyl-sn-glycerol and 1,2-dioleoyl-sn-glycerol). The AtDGK2 gene is expressed in various tissues of the Arabidopsis plant, including leaves, roots, and flowers, as shown by Northern blot analysis and promoter-reporter gene fusions. We found that AtDGK2 is induced by exposure to low temperature (4degreesC), pointing to a role in cold signal transduction.
Free Keywords:cysteine-rich domains
; protein-kinase
; phorbol ester
; phosphatidic-acid
; phospholipase-c
; beta-glucuronidase
; molecular-cloning
; psaf gene
; cells
; activation
External Publication Status:published
Document Type:Article
Communicated by:N. N.
Affiliations:MPI für molekulare Pflanzenphysiologie/Universitätsgastgruppen/AG Mueller-Roeber
External Affiliations:Univ Potsdam, Inst Biochem & Biol, Karl Liebknecht Str 24-25,Haus 20, D-14476 Golm Potsdam, Germany
Univ Potsdam, Inst Biochem & Biol, D-14476 Golm Potsdam, Germany
Univ E Anglia, Sch Biol Sci, Norwich NR4 7TJ, Norfolk, England
Max Planck Inst Mol Plant Physiol, Cooperat Res Grp, D-14476 Golm Potsdam, Germany
Identifiers:ISI:000189103300105 [ID No:1]
ISI:000189103300105 [ID No:2]
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