Fig 5. Identification of AimA as a novel low-affinity glutamate transporter.

(A) E. coli JM109 was transformed with pGP2987 (AimA) and pWH844 (empty vector) and growth of E. coli JM109 was assessed on MSSM minimal medium with glucose and ammonium or glutamate as carbon and nitrogen source. (B) Growth assay of B. subtilis glutamate transporter mutants. B. subtilis strains were cultivated in MSSM minimal medium with 0.1 mM KCl and ammonium. Cells were harvested, washed, and the OD₆₀₀ was adjusted to 1.0. Serial dilutions were dropped onto MSSM minimal plates with the indicated potassium concentration and ammonium or glutamate (1, 5, or 20 mM). (C) Determination of glutamate transporter kinetics. The growth rates of B. subtilis strains expressing either AimA (upper panel) or GltT (lower panel) as the only glutamate transporter were used to determine the apparent glutamate transporter kinetics of AimA and GltT. A B. subtilis strain lacking all glutamate transporters (ΔgltT ΔaimA ΔgltP; light grey) served as a control. The strains were grown in MSSM minimal medium with 0.1 mM or 5 mM KCl and various glutamate concentrations. The growth rate was plotted against the glutamate concentration. (D) Kinetic parameters (K_S and v_max values) for AimA and GltT. The numerical data are presented in S4 Table. (E) Expression of gltT, aimA, and gltP. The expression levels of the genes encoding the glutamate transporters GltT, AimA, and GltP were extracted from the transcriptomic analysis.