Identification and Characterization of malic enzymes from Streptomyces coelicolor

E. RODRÍGUEZ, P. CASATI Y H. GRAMAJO

Rosario, Argentina

Congreso; XLII Reunión Anual de la Sociedad Argentina de Investigadores en Bioquímica y Biología Molecular (SAIB); 2006

S. coelicolor has been adopted as a model system to study secondary metabolism, which is regulated through a complex cascade of regulatory factors. Primary metabolism, which supplies the building blocks for antibiotic biosynthesis, has received little attention. Since anaplerotic enzymes give rise to branch points between energy-producing and biosynthetic pathways, their activity is likely to be regulated at the transition phase between primary and secondary metabolism. For this reason, we studied anaplerotic malic enzymes from S. coelicolor. Malic enzymes catalyze the oxidative decarboxylation of L-malate into piruvate with reduction of NAD or NADP. Here we describe the characterization of two genes encoding putative malic enzymes that where found in the chromosome sequence project of S. coelicolor. In vitro enzymatic assays using recombinant proteins allowed validate Sco2951 protein as a NAD- and Sco5261 as a NADP-dependent malic enzyme. Single and double mutant strains were constructed by genetic manipulation. Individual mutations in either Sco2951 or Sco5261 genes resulted in no significant growth phenotype, but the double mutant strain showed severe differences in growth and antibiotic production under different carbon sources tested, indicating a main role of the activity of malic enzyme in primary and secondary metabolism of S. coelicolor.