Table 1.

Alanine mutations in M4 of each subunit that led to statistically significant changes in pEC₅₀

Mutant	Dose responsea,b				Fold change
	EC50 (μM)	pEC50 (M)	Hill slope	n	$[\frac{{\text{EC}}_{50}(\text{Mut})}{{\text{EC}}_{50}(\text{WT})}]$
WT	7.61	5.12 ± 0.07	1.70 ± 0.47	50	—
α subunit
αL433A	10.3	4.99 ± 0.04	1.93 ± 0.14	12	1.35
αR429A	40.0	4.41 ± 0.11	1.09 ± 0.12	10	5.25
αF426A	2.02	5.71 ± 0.14	1.80 ± 0.76	11	0.27
αV425A	4.30	5.37 ± 0.05	1.89 ± 0.28	10	0.57
αL423A	5.80	5.24 ± 0.07	2.54 ± 0.66	10	0.76
αT422A	31.2	4.52 ± 0.13	1.40 ± 0.12	10	4.10
αG421A	10.0	5.00 ± 0.05	1.96 ± 0.26	9	1.32
αI420A	5.97	5.23 ± 0.07	2.71 ± 0.33	10	0.78
αC418A	10.6	4.99 ± 0.13	1.82 ± 0.33	9	1.40
αM415A	12.5	4.92 ± 0.13	1.61 ± 0.27	10	1.65
αF414A	4.47	5.36 ± 0.09	1.76 ± 0.34	10	0.59
αL411A	9.86	5.02 ± 0.10	2.35 ± 0.40	13	1.30
αL410A	5.21	5.27 ± 0.06	2.06 ± 0.59	21	0.68
αH408A	10.1	5.00 ± 0.05	1.80 ± 0.27	10	1.33
αD407A	4.97	5.31 ± 0.04	3.12 ± 0.78	11	0.65
αY401A	9.86	5.01 ± 0.03	1.75 ± 0.48	11	1.30
αK400A	12.9	4.90 ± 0.09	1.89 ± 0.63	15	1.70
β subunit
βP476A	4.98	5.33 ± 0.20	1.49 ± 0.34	13	0.65
βD475A	4.15	5.39 ± 0.11	1.27 ± 0.31	8	0.54
βH470A	13.0	4.90 ± 0.11	1.24 ± 0.19	13	1.71
βD466A	5.08	5.31 ± 0.14	1.57 ± 0.48	11	0.67
βI463A	3.67	5.44 ± 0.16b	1.78 ± 0.11	8	0.48
βG459A	14.4	4.90 ± 0.28	1.12 ± 0.30	10	1.89
βS457A	13.2	4.93 ± 0.22	1.10 ± 0.24	11	1.74
βI453A	4.57	5.36 ± 0.12	1.46 ± 0.24	8	0.60
βW450A	12.7	4.94 ± 0.19	1.27 ± 0.20	14	1.66
βL449A	15.9	4.82 ± 0.15	1.45 ± 0.19	8	2.09
βF448A	4.36	5.38 ± 0.15	1.63 ± 0.48	8	0.57
βV444A	2.22	5.66 ± 0.07	1.89 ± 0.30	11	0.29
δ subunit
δP478A	4.72	5.34 ± 0.11b	1.43 ± 0.56	7	0.62
δP477A	11.7	4.94 ± 0.09b	1.26 ± 0.14	8	1.51
δG471A	14.1	4.86 ± 0.11b	1.57 ± 0.25	4	1.85
δL469A	5.13	5.30 ± 0.09b	1.71 ± 0.38	9	0.67
δI467A	4.63	5.34 ± 0.07b	1.91 ± 0.79	7	0.61
δW466A	5.49	5.28 ± 0.14b	1.57 ± 0.27	8	0.72
δG463A	4.88	5.32 ± 0.10b	1.71 ± 0.38	8	0.64
δP458A	4.08	5.40 ± 0.09b	2.06 ± 0.75	7	0.54
δV456A	5.06	5.33 ± 0.19b	1.58 ± 0.41	8	0.66
δC452A	4.86	5.32 ± 0.07b	1.56 ± 0.16	11	0.64
δR450A	5.15	5.29 ± 0.04b	1.72 ± 0.17	7	0.68
δD449A	5.03	5.34 ± 0.20b	2.00 ± 0.19	8	0.66
δV444A	4.35	5.36 ± 0.03b	2.02 ± 0.17	8	0.57
ε subunit
εI471A	15.5	4.81 ± 0.07b	1.61 ± 0.24	10	2.04
εC470A	13.4	4.88 ± 0.05b	1.54 ± 0.08	12	1.76
εP463A	12.1	4.93 ± 0.08b	1.77 ± 0.10	9	1.59
εY458A	5.10	5.30 ± 0.07b	1.43 ± 0.20	8	0.67
εF454A	3.90	5.42 ± 0.10b	1.80 ± 0.25	8	0.51
εI453A	4.50	5.35 ± 0.06b	1.83 ± 0.27	8	0.59
εG449A	5.62	5.26 ± 0.12b	1.54 ± 0.52	9	0.74
εC438A	10.1	5.00 ± 0.06b	1.67 ± 0.20	9	1.33
εN436A	12.9	4.90 ± 0.09b	1.76 ± 0.22	11	1.70
εG431A	10.6	4.99 ± 0.11b	1.60 ± 0.17	8	1.40
εM430A	No currentc	-	-	8	-
εV428A	5.62	5.25 ± 0.06b	1.56 ± 0.19	8	0.74
εW427A	5.85	5.25 ± 0.14b	1.37 ± 0.39	9	0.77

^aMeasurements performed 1 to 4 days after cRNA injection (V_hold ranging from −20 to −80 mV). Error values are represented as standard deviation.

^bp < 0.001 relative to WT via one-way ANOVA followed by Dunnet’s post hoc test.

^cNo significant current observed up to 4 days after cRNA injection.