Contribution of cysteines C59 and C61 in the first transmembrane helix (TM1) to the stabilization of the ASIC1a complex by BMOE.
A: ASIC1a oligomeric states, resolved by SDS-PAGE under reducing conditions and detected by anti-His-tag western blotting (same experimental procedure as in Fig 3A), of the His8-tagged forms of ASIC1a-ΔCCt as control (Ctrl), and the cysteine substitution mutant G433C-ΔCCt (433), G433C-C59V-ΔCCt (433–59) or G433C-C59V-C61S-ΔCCt (433-59-61). Numbers I to IV have the same meaning as in Fig 3. B: Relative intensities (mean ±SEM, n = 4) of each of the 4 bands (I to IV) for ASIC1a identified on SDS-PAGE from cells expressing ASIC1a-ΔCCt, (C), G433C-ΔCCt (433) G433C-C59V-ΔCCt (433–59) and G433C-C59V-C61S-ΔCCt (433-59-61). The average molecular weight estimated for the four bands I, II, III, IV are respectively 80±10, 160±6, 230±9 and 300±19 kDa (Mw±SD, n = 4) for the G433C-ΔCCt, G433C-C59V-ΔCCt, and G433C-C59V-C61S-ΔCCt constructs. * denotes p<0.01