Table 2.

Na⁺ affinity of hSGLT1 mutants

	Kd, mM	KdA, mM	KdB, mM	k16/k61	τo, ms	k16, s−1	k61, s−1
hSGLT1	20	33*	10*	100	<2†	530	5
Near sugar binding site
H83C	120	—	>150	6	6	170	10
T287C	20	—	—	200	—	—	—
Y290C	57	30	110	2.4	9	80	33
Y290S	57	30	110	1.4	17	60	25
Y290F	120	30	>150	24	10	100	4
W291C	55	30	110	2	16	21	43
W291F	20	—	—	100	4	275	3
Q457C	20	—	—	250	2	530	2
Near Na2 site
D204E	>150	—	—	2.6	—	—	—
S392A	122	50	>150	10	19	48	5
Other
F101C	18	—	—	35	—	—	—
F453C	20	33*	10*	125	—	—	—
G507C	20	33*	10*	100	—	—	—

Na⁺ affinities, K_d (= √(k₂₁/k₁₂)) for the three-state and K_dA (= k_N1/k_1N) and K_dB (= k_2N/k_N2) for the four-state model (Fig. 1), were estimated from the V_0.5 vs. log [Na⁺]_o data (Figs. 5 and 6). Parameters are those obtained in individual cells, but representative of three to five experiments conducted on different batches of oocytes. For all proteins, the variation in K_d was 10% and, excluding the WT group, the variation in K_dA was 10%. τ_o was obtained from the time constants for relaxation of the transient charge movements in the absence of sodium (e.g., Fig. 7). The values given for K_dA and K_dB are for those where V_0.5 in the absence of Na⁺ fell within the experimental range (Table 1), reducing the number of unknown parameters in Eq. 9. For all proteins, the number of Na⁺ ions (n) was 2, and net charge (z) was 1. z₁ = 0.7, z₂ = 0.3, for the three-state model and z₁ = 0.7, z₂ = 0.15, z₃ = 0.15 for the four-state model. The absence of data for mutants (—) reflects those where V_0.5 in the absence of Na⁺ fell outside the experimental range (Table 1) and those not determined.

^*The Na⁺ affinities of the two sites for the WT group are similar, because the three-state model fitted the data, and K_d = √K_dAK_dB.

^†Loo et al. (24).