Two Ca2+-Binding Sites Cooperatively Couple Together in TMEM16A Channel

TMEM16A is the molecular basis of calcium-activated chloride channels and shows Ca(2+)-dependent gating. It is critical to understand how the Ca(2+) sensors dynamically control the gate of TMEM16A. However, the detailed mechanism by which the calcium ions bind and open the channel is still obscure. In this study, the authors confirmed that there are two Ca(2+) sensors which cooperatively couple together in TMEM16A. Our data show that mutations at both Ca(2+)-sensitive domains, E447Y and E702Q-E705Q, weaken the Ca(2+) affinity for TMEM16A channel. The EC50 for WT, E447Y, and E702Q-E705Q are 0.53 ± 0.11, 14.5 ± 0.3, and 26.5 ± 3.6 μM, respectively. The triple mutation, including both of the Ca(2+) sensors, E447Y-E702Q-E705Q, with EC50 as 55.6 ± 5.1 μM, results in much further right-shifted dose response curve than the single sensor's mutations (E447Y, E702Q-E705Q) do, which indicates that there is a cooperation between the two Ca(2+)-sensitive domains. We also found that the divalent cations, both Ca(2+) and Sr(2+), share common mechanism of gating the TMEM16A.