Citrate metabolism in Enterococcus faecalis is repressed by PTS sugars

V.S. BLANCATO, G.D. REPIZO, AND CH. MAGNI

Mar del Plata – Buenos Aires - Argentina

Congreso; XLIII Reunión Anual de SAIB; 2007

SAIB

Enterococcus are involved in the ripening and in aroma development of diverse cheeses. These positive effects have been attributed to specific biochemical traits such as lipolytic activity and citrate utilization. The expression pattern of the cit locus showed the presence of two divergent operons, citHO and oadDBcitCDEFXoadAcitMG encoding the enzymes for citrate metabolism. Northern blot analysis showed activation of both operons by specific addition of citrate to the medium and repression in the presence of glucose. In accordance with that, cell extracts showed repression of citrate lyase activity and the level of OadD protein, determined by western blot, was also repressed when the cells were grown in the presence of citrate and glucose. Carbohydrate repression was only observed with PTS sugars suggesting a Carbon Catabolite Repression (CCR). Moreover, the repression was partially relieved when transcriptional fusions of the promoters were analyzed in an Enterococcus ccpA mutant strain. Sequence analysis of the promoter regions revealed the presence of catabolite responsive elements (cre). Band shift experiments demonstrated that BsCcpA could bind to these cre sites downregulating the expression of the operons. In order to establish which one is involved in the CCR observed, the cre sites were systematically mutated and the promoter activities were determined.