ecgltP is a bacterial glutamate transporter homologous to mammalian EAAT glutamate transporters. We here studied purified and reconstituted ecgltP using radiotracer flux measurements. Reconstituted ecgltP transporters sustain an electrogenic co-transport of H+ and of glutamate, aspartate, alanine, serine, or cysteine. Measurements of individual transport rates and their temperature dependence demonstrated a transport process that encompasses major conformational changes, consistent with alternating access models. ecgltP transports glutamate independently of Na+ and K+ concentrations and functions at a variable H+/glutamate stoichiometry. At pH 6.0, glutamate is co-transported together with 3 protons, and lowering the external pH decreases the transport stoichiometry. To further study the variable coupling coefficient, we characterized Q320E ecgltP that exhibits a glutamate residue at the homologous position to E373, that was shown to be the proton acceptor in EAAT3. Glutamate transport by mutant ecgltP was electrogenic and depended only on the proton concentrations. However, Q320E appeared to alter the transport stoichiometry of ecgltP. We conclude that prokaryotic and eukaryotic glutamate transporters differ in the transported substrates and the transport stoichiometry, but share basic mechanisms of transport. ecgltP represents a model to study the mechanisms underlying glutamate transport and coupled transport in general.