Loss of cell viability by histidine substitution of leucine 325 of the glutamate transporter EAAT1

Although glutamate transporters and neutral amino acid transporters have 55% amino acid identity in the transmembrane domains, many residues are still unique to individual transporters, providing for structural stability or substrate binding. In this study, the mutant protein L325H, which replaced a leucine 325 of the glutamate transporter EAAT1 by a histidine, was evaluated. When expressed in Xenopus oocytes, L325H caused oocytes to weaken pigmentation in the animal pole, accompanied by patches of colorless spots. Oocytes finally oozed cytoplasm. The resting membrane potential in L325H oocytes was -18.9 +/- 2.5 mV, significantly more positive than -37.3 +/- 2.5 mV of oocytes expressing EAAT1. The holding current at -60 mV was 283.1 +/- 48.3 nA in L325H oocytes and 92.2 +/- 12.6 nA in EAAT1 oocytes. These results suggest that even though glutamate and neutral amino acid transporters have strong overall homology, the local structure in the transmembrane domains may be different. (C) 2000 Academic Press.