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Fumarate and cytosolic pH as modulators of the synthesis or consumption of C(4) organic acids through NADP-malic enzyme in Arabidopsis thaliana.
- Source :
-
Plant molecular biology [Plant Mol Biol] 2013 Feb; Vol. 81 (3), pp. 297-307. Date of Electronic Publication: 2012 Dec 16. - Publication Year :
- 2013
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Abstract
- Arabidopsis thaliana is a plant species that accumulates high levels of organic acids and uses them as carbon, energy and reducing power sources. Among the enzymes that metabolize these compounds, one of the most important ones is malic enzyme (ME). A. thaliana contains four malic enzymes (NADP-ME 1-4) to catalyze the reversible oxidative decarboxylation of malate in the presence of NADP. NADP-ME2 is the only one located in the cell cytosol of all Arabidopsis organs providing most of the total NADP-ME activity. In the present work, the regulation of this key enzyme by fumarate was investigated by kinetic assays, structural analysis and a site-directed mutagenesis approach. The final effect of this metabolite on NADP-ME2 forward activity not only depends on fumarate and substrate concentrations but also on the pH of the reaction medium. Fumarate produced an increase in NADP-ME2 activity by binding to an allosteric site. However at higher concentrations, fumarate caused a competitive inhibition, excluding the substrate malate from binding to the active site. The characterization of ME2-R115A mutant, which is not activated by fumarate, confirms this hypothesis. In addition, the reverse reaction (reductive carboxylation of pyruvate) is also modulated by fumarate, but in a different way. The results indicate pH-dependence of the fumarate modulation with opposite behavior on the two activities analyzed. Thereby, the coordinated action of fumarate over the direct and reverse reactions would allow a precise and specific modulation of the metabolic flux through this enzyme, leading to the synthesis or degradation of C(4) compounds under certain conditions. Thus, the physiological context might be exerting an accurate control of ME activity in planta, through changes in metabolite and substrate concentrations and cytosolic pH.
- Subjects :
- Allosteric Regulation drug effects
Allosteric Site
Amino Acid Substitution
Arabidopsis drug effects
Arabidopsis genetics
Arabidopsis Proteins drug effects
Arabidopsis Proteins genetics
Arabidopsis Proteins metabolism
Cytosol enzymology
Enzyme Activation drug effects
Hydrogen-Ion Concentration
Kinetics
Malate Dehydrogenase drug effects
Malate Dehydrogenase genetics
Malates metabolism
Mutagenesis, Site-Directed
NADP metabolism
Protein Structure, Tertiary
Recombinant Fusion Proteins
Arabidopsis enzymology
Carboxylic Acids metabolism
Fumarates pharmacology
Malate Dehydrogenase metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 1573-5028
- Volume :
- 81
- Issue :
- 3
- Database :
- MEDLINE
- Journal :
- Plant molecular biology
- Publication Type :
- Academic Journal
- Accession number :
- 23242919
- Full Text :
- https://doi.org/10.1007/s11103-012-9999-6