Table 1 Relative effects of alanine mutations on the ion-exchange capacity in a given pair of inverted residues.
From: Asymmetric Preorganization of Inverted Pair Residues in the Sodium-Calcium Exchanger
Mutated pair residues | RA values (V″max1/V″max2) Na/Ca exchange | RA values (V″max1/V″max2) Ca/Ca exchange |
|---|---|---|
S236A/S77A | 31.3 ± 3.6 | 32.8 ± 2.3 |
N81A/D240A | 3.5 ± 0.4 | 4.5 ± 0.3 |
T50A/T209A | 2.5 ± 0.3 | 3.0 ± 0.3 |
S210A/S51A | 2.2 ± 0.3 | 1.9 ± 0.3 |
E213A/E54A | 1.8 ± 0.4 | 1.6 ± 0.3 |
N73A/N232A | 2.1 ± 0.3 | 1.7 ± 0.3 |
- The V″max1 and V″max2 ratios of the Na+/Ca2+ and Ca2+/Ca2+ exchange reactions were measured in mutants of matching pair residues as described in Figs 3 and 4 (see also Methods). The V″max1/V″max2 ratios are presented as the index of asymmetry (RA) for a given pair of residues. The numbers represent the mean ± SE.
Figure 4 
Identification of distinct residues stabilizing the Ca2+ transition state.
(A) The Km and Vmax values of the Ca2+/Ca2+ exchange reaction were measured by varying the extravesicular Ca2+ concentrations (20–2000 μM) at saturating intravesicular Ca2+ (250 μM) (see Methods and Fig. S1). The kcat values were calculated as described in the legend of Fig. 3D (see also Methods). The bars represent the mean ± SE. (B) The kcat/Km values were derived from experimentally observed kcat and Km values, as outlined in panel A. The bars represent the mean ± SE. (C) Experimentally derived kcat and Km values of the Ca2+/Ca2+ exchange were used for calculating the ∆∆G≠app values according to the equation ∆∆G≠app = −2.303RT•log[(kcat/Km)mut/(kcat/Km)wt. The bars represent the mean ± SE.
