title: Identification of malic and soluble oxaloacetate decarboxylase enzymes in Enterococcus faecalis.
abstract: Two paralogous genes, maeE and citM, that encode putative malic enzyme family members were identified in the Enterococcus faecalis genome. MaeE (41 kDa) and CitM (42 kDa) share a high degree of homology between them (47% identities and 68% conservative substitutions). However, the genetic context of each gene suggested that maeE is associated with malate utilization whereas citM is linked to the citrate fermentation pathway. In the present work, we focus on the biochemical characterization and physiological contribution of these enzymes in E. faecalis. With this aim, the recombinant versions of the two proteins were expressed in Escherichia coli, affinity purified and finally their kinetic parameters were determined. This approach allowed us to establish that MaeE is a malate oxidative decarboxylating enzyme and CitM is a soluble oxaloacetate decarboxylase. Moreover, our genetic studies in E. faecalis showed that the citrate fermentation phenotype is not affected by citM deletion. On the other hand, maeE gene disruption resulted in a malate fermentation deficient strain indicating that MaeE is responsible for malate metabolism in E. faecalis. Lastly, it was demonstrated that malate fermentation in E. faecalis is associated with cytoplasmic and extracellular alkalinization which clearly contributes to pH homeostasis in neutral or mild acidic conditions.
authors.affiliation: Instituto de Biología Molecular y Celular de Rosario (IBR-CONICET), Universidad Nacional de Rosario, Argentina.
authors.forename: Martín
authors.initials: M
authors.lastname: Espariz
authors.forename: Guillermo
authors.initials: G
authors.lastname: Repizo
authors.forename: Víctor
authors.initials: V
authors.lastname: Blancato
authors.forename: Pablo
authors.initials: P
authors.lastname: Mortera
authors.forename: Sergio
authors.initials: S
authors.lastname: Alarcón
authors.forename: Christian
authors.initials: C
authors.lastname: Magni
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D002262:Carboxy-Lyases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D019343:Citric Acid / Q000378:metabolism; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000235:genetics; D013293:Enterococcus faecalis / Q000201:enzymology* / Q000235:genetics*; D005285:Fermentation; D005798:Genes, Bacterial; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008291:Malate Dehydrogenase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D008293:Malates / Q000378:metabolism; D008969:Molecular Sequence Data; D010802:Phylogeny; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012995:Solubility
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; D008293:Malates; D011994:Recombinant Proteins; D019343:Citric Acid; C030298:malic acid; D008291:Malate Dehydrogenase; D002262:Carboxy-Lyases; C021916:oxaloacetate decarboxylase