Specific Arabidopsis thaliana malic enzyme isoforms can provide anaplerotic pyruvate carboxylation function in Saccharomyces cerevisiae.

Badia, Mariana Beatriz; Mans, Robert; Lis, Alicia V; Tronconi, Marcos Ariel; Arias, Cintia Lucía; Maurino, Verónica Graciela; Andreo, Carlos Santiago; Drincovich, María Fabiana; van Maris, Antonius J A; Gerrard Wheeler, Mariel Claudia

Specific Arabidopsis thaliana malic enzyme isoforms can provide anaplerotic pyruvate carboxylation function in Saccharomyces cerevisiae.

Badia, Mariana Beatriz; Mans, Robert; Lis, Alicia V; Tronconi, Marcos Ariel; Arias, Cintia Lucía; Maurino, Verónica Graciela; Andreo, Carlos Santiago; Drincovich, María Fabiana; van Maris, Antonius J A; Gerrard Wheeler, Mariel Claudia.

Afiliación

Badia MB; Centro de Estudios Fotosintéticos y Bioquímicos (CEFOBI), Universidad Nacional de Rosario, Argentina.
Mans R; Department of Biotechnology, Delft University of Technology, The Netherlands.
Lis AV; Department of Biotechnology, Delft University of Technology, The Netherlands.
Tronconi MA; Biochemical Engineering Institute, Saarland University, Saarbrücken, Germany.
Arias CL; Centro de Estudios Fotosintéticos y Bioquímicos (CEFOBI), Universidad Nacional de Rosario, Argentina.
Maurino VG; Centro de Estudios Fotosintéticos y Bioquímicos (CEFOBI), Universidad Nacional de Rosario, Argentina.
Andreo CS; Plant Molecular Physiology and Biotechnology Group, Institute of Developmental and Molecular Biology of Plants, Cluster of Excellence on Plant Sciences (CEPLAS), Heinrich-Heine-Universität, Düsseldorf, Germany.
Drincovich MF; Centro de Estudios Fotosintéticos y Bioquímicos (CEFOBI), Universidad Nacional de Rosario, Argentina.
van Maris AJ; Centro de Estudios Fotosintéticos y Bioquímicos (CEFOBI), Universidad Nacional de Rosario, Argentina.
Gerrard Wheeler MC; Department of Biotechnology, Delft University of Technology, The Netherlands.

FEBS J ; 284(4): 654-665, 2017 02.

Article en En | MEDLINE | ID: mdl-28075062

RESUMEN

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.

Asunto(s)

Proteínas de Arabidopsis/genética; Arabidopsis/enzimología; Malato-Deshidrogenasa (NADP+)/genética; Malatos/metabolismo; NADP/metabolismo; NAD/metabolismo; Ácido Pirúvico/metabolismo; Saccharomyces cerevisiae/enzimología; Arabidopsis/genética; Proteínas de Arabidopsis/metabolismo; Dióxido de Carbono/metabolismo; Clonación Molecular; Expresión Génica; Prueba de Complementación Genética; Ingeniería Genética; Glucosa/metabolismo; Isoenzimas/genética; Isoenzimas/metabolismo; Malato-Deshidrogenasa (NADP+)/metabolismo; Hojas de la Planta/enzimología; Hojas de la Planta/genética; Proteínas Recombinantes/genética; Proteínas Recombinantes/metabolismo; Saccharomyces cerevisiae/genética; Transformación Genética; Transgenes

Palabras clave

Saccharomyces cerevisiae; C4 organic acids; anaplerotic role; malate synthesis; plant metabolism

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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Saccharomyces cerevisiae / Arabidopsis / Ácido Pirúvico / Proteínas de Arabidopsis / Malato-Deshidrogenasa (NADP/) / Malatos / NAD / NADP Idioma: En Revista: FEBS J Asunto de la revista: BIOQUIMICA Año: 2017 Tipo del documento: Article País de afiliación: Argentina Pais de publicación: Reino Unido

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