2.A.81.1.1
The L-aspartate:L-alanine exchanger, AspT (Abe et al. 1996; Abe et al. 2002).  A mutant, R76K, has higher activity than the AspT-WT (R76), whereas R76D and R76E have lower activity than AspT-WT. Thus, R76 is involved in AspT substrate transport (Suzuki et al. 2016).  AspT catalyzes self-exchange of aspartate and electrogenic heterologous exchange of aspartate with alanine. Thus, the asp operon confers a proton motive metabolic cycle consisting of the electrogenic aspartate-alanine antiporter and  aspartate decarboxylase, which keeps intracellular levels of alanine, the countersubstrate for aspartate, high (Abe et al. 2002). AspT has been reported to have a unique topology; 8 TMS, a large cytoplasmic loop (183 amino acids) between TMS5 and TMS6, and N- and C-termini that both face the periplasm. This may be a unique 2D-structure of AspT, a representative of the novel AAE family (Nanatani et al. 2005). However, bioinformatic analyses suggest that this prediction may be in error, and the true topology is 12 TMSs with an internal repeat of 6 TMSs (M. Saier, unpublished observations; see 2.A.81.2.1). The K_m values = 0.35 mm for L-aspartate, 0.098 mm for D-aspartate, 26 mm for L-alanine, and 3.3 mm for D-alanine). Competitive inhibition by various amino acids of L-aspartate or L-alanine in self-exchange reactions revealed that L-cysteine selectively inhibited L-aspartate self-exchange but only weakly inhibited L-alanine self-exchange while L-serine selectively inhibited L-alanine self-exchange but barely inhibited L-aspartate self-exchange. The aspartate analogs L-cysteine sulfinic acid, L-cysteic acid, and D-cysteic acid competitively and strongly inhibited L-aspartate self-exchange compared with L-alanine self-exchange. Taken together, these kinetic data suggest that the putative binding sites of L-aspartate and L-alanine are independently located in the substrate translocation pathway of AspT (Sasahara et al. 2011). L-Ala binding yields a conformation different from the apo or L-Asp binding conformations (Suzuki et al. 2022).