Biochemical and genetic characterization of the Enterococcus faecalis oxaloacetate decarboxylase complex

REPIZO G. D.; BLANCATO V. S.; MORTERA P.; LOLKEMA J. S.; MAGNI C.; REPIZO G. D.; BLANCATO V. S.; MORTERA P.; LOLKEMA J. S.; MAGNI C.

APPLIED AND ENVIRONMENTAL MICROBIOLOGY

AMER SOC MICROBIOLOGY

Lugar: Washington; Año: 2013 p. 2882 - 2890

0099-2240

Enterococcus faecalis encodes a biotin-dependent oxaloacetate decarboxylase (OAD), which is constituted by four subunits: E.faecalis carboxyltransferase subunit OadA (termed Ef-A), membrane pump Ef-B, biotin acceptor protein Ef-D, and the novelsubunit Ef-H. Our results show that in E. faecalis, subunits Ef-A, Ef-D, and Ef-H form a cytoplasmic soluble complex (termedEf-AHD) which is also associated with the membrane. In order to characterize the role of the novel Ef-H subunit, coexpression ofoad genes was performed in Escherichia coli, showing that this subunit is vital for Ef-A and Ef-D interaction. Diminished growthof the oadA and oadD single deletion mutants in citrate-supplemented medium indicated that the activity of the complex is essentialfor citrate utilization. Remarkably, the oadB-deficient strain was still capable of growing to wild-type levels but with adelay during the citrate-consuming phase, suggesting that the soluble Ef-AHD complex is functional in E. faecalis. These resultssuggest that the Ef-AHD complex is active in its soluble form, and that it is capable of interacting in a dynamic way with themembrane-bound Ef-B subunit to achieve its maximal alkalinization capacity during citrate fermentation.