NAD(P)-malic enzyme (NAD(P)-ME) catalyzes the reversible oxidative decarboxylation of malate to pyruvate, CO₂ , and NAD(P)H and is present as a multigene family in Arabidopsis thaliana. The carboxylation reaction catalyzed by purified recombinant Arabidopsis NADP-ME proteins is faster than those reported for other animal or plant isoforms. In contrast, no carboxylation activity could be detected in vitro for the NAD-dependent counterparts. In order to further investigate their putative carboxylating role in vivo, Arabidopsis NAD(P)-ME isoforms, as well as the NADP-ME2del2 (with a decreased ability to carboxylate pyruvate) and NADP-ME2R115A (lacking fumarate activation) versions, were functionally expressed in the cytosol of pyruvate carboxylase-negative (Pyc^- ) Saccharomyces cerevisiae strains. The heterologous expression of NADP-ME1, NADP-ME2 (and its mutant proteins), and NADP-ME3 restored the growth of Pyc^- S. cerevisiae on glucose, and this capacity was dependent on the availability of CO₂ . On the other hand, NADP-ME4, NAD-ME1, and NAD-ME2 could not rescue the Pyc^- strains from C₄ auxotrophy. NADP-ME carboxylation activity could be measured in leaf crude extracts of knockout and overexpressing Arabidopsis lines with modified levels of NADP-ME, where this activity was correlated with the amount of NADP-ME2 transcript. These results indicate that specific A. thaliana NADP-ME isoforms are able to play an anaplerotic role in vivo and provide a basis for the study on the carboxylating activity of NADP-ME, which may contribute to the synthesis of C₄ compounds and redox shuttling in plant cells.

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Journal Article

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Badia MB, Mans R, Lis AV, Tronconi MA, Arias CL, Maurino VG, Andreo CS, Drincovich MF, van Maris AJ, Gerrard Wheeler MC

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