EAAT1 | EAAT2 | |||
---|---|---|---|---|
Km | Imax | Km /Kb | Imax | |
μm | μm | |||
l-Glutamate | 20 ± 3 | 1 | 18 ± 3 | 1 |
l-Kainate | >3000 | 17.4 ± 3.4 | 0 | |
tPDC | 26.3 ± 0.8 | 0.74 ± 0.03 | 2.3 ± 0.5 | 0.23 ± 0.02 |
T3MG | No activity (300) | 18.3 ± 1.0 | 0 | |
E3MG | No activity (100) | No activity (100) | ||
2S4R4MG | 54 ± 17 | 0.80 ± 0.05 | 3.4 ± 0.2 | 0 |
(2S,4S)-4-Methylglutamate | No activity (100) | No activity (100) | ||
(±)-4-Methylene-glutamate | 391 ± 51 | 0.43 ± 0.02 | 39 ± 7 | 0 |
LT4HG | 61 ± 14 | 0.78 ± 0.06 | 48 ± 5 | 0.90 ± 0.04 |
E4HG | > 1000 | ∼1.0 | > 1000 | ∼1.0 |
(S)-α-Methyl glutamate | No activity (100) | No activity (100) |
Km and Imax values for the various competitive substrates were determined as described in Materials and Methods. Imax values are relative to the maximal current generated by glutamate. T3MG, 2S4R4MG, and kainate blocked glutamate transport by EAAT2; Schild analysis (22) was used to establish competitive antagonism and also to determine theKb for binding of the compound to the transporter (see Fig. 3). The Ki for (±)-4-methyleneglutamate block of glutamate transport by EAAT2 was derived from the IC50 using the Cheng-Prusoff equation, assuming a competitive antagonism mechanism (23). The number of cells tested for each compound on both EAAT1 and EAAT2 was between three and eight, except for E4HG, in which only one cell expressing EAAT1 and one cell expressing EAAT2 were tested.