Expression and characterization of recombinant cytosolicNAD+-dependent malate dehydrogenase from Mesembyanthemum crystallinum |
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| Authors: | Karina E J Trípodi María E Gómez Casati Antonio D Uttaro Florencio E Podestá |
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| Institution: | Centro de Estudios Fotosintéticos y Bioquímicos (CONICET), Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario, Suipacha 531 2000 Rosario, Argentina;IBR, Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario, Suipacha 531 2000 Rosario, Argentina |
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| Abstract: | An NAD+-dependent cytosolic malate dehydrogenase (MDH, EC 1.1.1.37) from leaves of Mesembryanthemum crystallinum in the Crassulacean Acid Metabolism (CAM) mode was cloned, expressed in E. coli and characterized. The recombinant enzyme had a subunit molecular mass of 39.5 kDa and was recognized by antibodies raised against the cytosolic MDH from Ananas comosus . Its activity showed a maximum in the pH range of 7.5–9.5. The purified MDH is highly but not completely specific for oxaloacetate, as indicated by a low activity using various other α-ketoacids as substrates. The sequence data, subunit mass and immunoreactivity suggest that the MDH that has been cloned and characterized corresponds to the cytosolic isoform. Yet, the biochemistry of this enzyme comparative with the only other CAM plant cytosolic MDH characterized so far (that of pineapple) hints at a distinct isoform being expressed in M. crystallinum leaves. |
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