Two novel D-2-Hydroxyacid dehydrogenases in Arabidopsis thaliana with catalyric capacity to participate in the last reactions of the methylglyoxal and ß-oxidation pathways

MARTIN ENGQVIST; MARIA FABIANA DRINCOVICH; FLUGGE, U.; MAURINO, VERÓNICA G

JOURNAL OF BIOLOGICAL CHEMISTRY

AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC

Año: 2009 vol. 284 p. 25026 - 25037

0021-9258

The Arabidopsis thaliana locus At5g06580 encodes an ortholog to Saccharomyces cerevisiae D-lactate dehydrogenase (AtD-LDH). The recombinant protein is a homodimer of 59 kDa subunits with one FAD per monomer. A substrate screen indicated that AtD-LDH catalyzes the oxidation of D- and L-lactate, D-2-hydroxybutyrate, glycerate and glycolate using cytochrome c as electron acceptor. AtD-LDH shows a clear preference for D-lactate, with a catalytic efficiency 200- and 2000-fold higher than that for L-lactate and glycolate, respectively, and a Km value for D-lactate of ~160 ìM. Knock-out mutants showed impaired growth in the presence of D-lactate or methylglyoxal. Collectively, the data indicated that the protein is a D-LDH that participates in planta in the methylglyoxal pathway. Webbased bioinformatic tools revealed the existence of a paralogous protein encoded by locus At4g36400. The recombinant protein is a homodimer of 61 kDa subunits with one FAD per monomer. A substrate screening revealed highly specific D-2-hydroxyglutarate (D-2HG) conversion in the presence of an organic cofactor with a Km value of ~580 μM. Thus, the enzyme was characterized as a D-2HG dehydrogenase (AtD-2HGDH). Analysis of knock-out mutants demonstrated that AtD- 2HGDH is responsible for the total D-2HGDH activity present in A. thaliana. Gene coexpression analysis indicated that AtD- 2HGDH is in the same network as several genes involved in ß-oxidation and degradation of branched-chain amino acids and chlorophyll. It is proposed that AtD-2HGDH participates in the catabolism of D-2HG most probably during the mobilization of alternate substrates from proteolysis and/or lipid degradation