The MrpA, MrpB and MrpD subunits of the Mrp antiporter complex in Bacillus subtilis contain membrane-embedded and essential acidic residues.

Bacillus subtilis Mrp is a unique Na+/H+ antiporter with a multicomponent structure consisting of the mrpABCDEFG gene products. We have previously reported that the conserved and putative membrane-embedded Glu-113, Glu-657, Asp-743 and Glu-747 of MrpA (ShaA) are essential for the transport function. In this study, we further investigated the functional involvement of the equivalent conserved acidic residues of other Mrp proteins in heterologous Escherichia coli and natural B. subtilis backgrounds. Asp-121 of MrpB and Glu-137 of MrpD were additionally identified to be essential for the transport function in both systems. Glu-137 of MrpD and Glu-113 of MrpA were found to be conserved in the homologous MrpD/MrpA proteins as well as in the homologous subunits of H+-translocating primary active transporters such as Nuo and Mbh, suggesting their critical role in ion binding. The remaining essential acidic residues clustered in the C-terminal domain of MrpA (Glu-657, Asp-743 and Glu-747) and MrpB (Asp-121); these subunits are fused in some Gram-negative species. It is possible that the MrpA, MrpB and MrpD subunits, which contain essential transmembrane acidic residues, form the ion translocation site(s) of the Mrp antiporter complex.