CLONING OF NADP DEPENDENT MALIC ENZYME FROM C4, C3 AND C3-C4 FLAVERIA SPECIES

SAAVEDRA DAMIAN DARÍO; AMERISO MARÍA CARMEN; DRINCOVICH MARÍA FABIANA; ANDREO CARLOS SANTIAGO

ROSARIO

Congreso; XLII Reunión Anual SOCIEDAD ARGENTINA DE INVESTIGACIÓN BIOQUÍMICA Y BIOLOGÍA MOLECULAR; 2006

CLONING OF NADP DEPENDENT MALIC ENZYME FROM C4, C3 AND C3-C4 FLAVERIA SPECIESSaavedra, Damián D.; Ameriso, María Carmen; Drincovich, María F. and Andreo, Carlos S. CEFOBI. Facultad de Cs Bioquímicas y Farmacéuticas. UNR. Suipacha 531. 2000. Rosario. Argentina. e-mail: dsaavedr@fbioyf.unr.edu.arC4 plants have evolved independently from C3 species many times during the evolution. NADP-malic enzyme (NADP-ME) is a widely distributed enzyme involved in different metabolic pathways. The photosynthetic isoform of this enzyme has evolved from non-photosynthetic isoforms. In order to analyze the origin of the NADP-ME C4-specific isoform, cDNAs encoding this enzyme were isolated in the genus Flaveria. This genus is well suited for studying the evolution of photosynthesis because it contains a more or less continuous range of species between C3 and C4, including C3-C4 and C4-like species. In the present work, five cDNAs codifying NADP-ME isoforms were isolated by RT-PCR. Four of these cDNAs correspond to plastidic isoforms from C3 F. pringlei, C3-C4 F. floridana and C4 F. bidentis and F. trinervia species. The remainig cDNA correspond to a cytosolic isoform from F. pringlei. The amplified products were cloned into pGEM-T vector, sequenced and the complete codifying sequences corresponding to mature proteins subcloned into the pET 32 expression vector. These recombinant proteins will be use for kinetic and structural characterizations. In addition, small portions to 3? ends to cDNAs encoding NADP-ME from intermediate species F. anomala, F. palmeri and F. sonorensis were amplified by RT-PCR. These sequences (previously unknown) will be used to obtain completes cDNAs with RACE method.