Evidence of in vivo pyruvate carboxylase activity for an Arabidopsis NADP-malic enzyme

BADIA, M. B.; MANS, R.; LIS, A.; TRONCONI, M. A.; ARIAS, C. L.; ANDREO, C. S.; DRINCOVICH, M. F.; VAN MARIS, A. J. A.; GERRARD WHEELER, M. C.

Rosario

Congreso; L Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología MolecularFisiología Vegetal; 2014

Sociedad Argentina de Investigación en Bioquímica y Biología Molecular

NADP-malic enzyme (NADP-ME) catalyzes the reversible oxidative decarboxylation of malate to pyruvate, CO2 and NADPH and is present as a multigene family in A. thaliana. The reverse reaction catalyzed by recombinant AtNADP-MEs was faster than reported for other animal or plant isoforms. Particularly, AtNADP-ME2, the cytosolic isoform that mostly contributes to the ME activity in all organs of the plant, presented the higher catalytic efficiency of the family. In this work, AtNADP-ME2 as well as its N-truncated version NADP-MEdel2 (unable to catalyze the carboxylation of pyruvate in vitro) were functionally expressed in pyruvate carboxylase-negative (Pyc-) Saccharomyces cerevisiae strains, which are incapable of growing on glucose as the sole carbon source. Experiments conducted on agar plates as well as in batch cultivation under ambient and thermodynamically favorable conditions showed that heterologous expression of both NADP-ME2 and NADP-MEdel2 enabled growth of Pyc- S. cerevisiae on solely glucose. This capacity of rescuing Pyc- strains from C4 auxotrophy was dependent on the provision of the reaction substrates, particularly CO2. These results indicate that AtNADP-ME2 is able to play an anaplerotic role in vivo and provide a basis for the study of the carboxylase activity of malic enzyme, which may contribute to the synthesis of C4 compounds in plant cells.