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http://dx.doi.org/10.14348/molcells.2021.2203

Ca2+ Sensitivity of Anoctamin 6/TMEM16F Is Regulated by the Putative Ca2+-Binding Reservoir at the N-Terminal Domain  

Roh, Jae Won (Department of Physiology, Dongguk University College of Medicine)
Hwang, Ga Eun (Department of Physiology, Dongguk University College of Medicine)
Kim, Woo Kyung (Department of Internal Medicine, Graduate School of Medicine, Dongguk University)
Nam, Joo Hyun (Department of Physiology, Dongguk University College of Medicine)
Publication Information
Molecules and Cells / v.44, no.2, 2021 , pp. 88-100 More about this Journal
Abstract
Anoctamin 6/TMEM16F (ANO6) is a dual-function protein with Ca2+-activated ion channel and Ca2+-activated phospholipid scramblase activities, requiring a high intracellular Ca2+ concentration (e.g., half-maximal effective Ca2+ concentration [EC50] of [Ca2+]i > 10 μM), and strong and sustained depolarization above 0 mV. Structural comparison with Anoctamin 1/TMEM16A (ANO1), a canonical Ca2+-activated chloride channel exhibiting higher Ca2+ sensitivity (EC50 of 1 μM) than ANO6, suggested that a homologous Ca2+-transferring site in the N-terminal domain (Nt) might be responsible for the differential Ca2+ sensitivity and kinetics of activation between ANO6 and ANO1. To elucidate the role of the putative Ca2+-transferring reservoir in the Nt (Nt-CaRes), we constructed an ANO6-1-6 chimera in which Nt-CaRes was replaced with the corresponding domain of ANO1. ANO6-1-6 showed higher sensitivity to Ca2+ than ANO6. However, neither the speed of activation nor the voltage-dependence differed between ANO6 and ANO6-1-6. Molecular dynamics simulation revealed a reduced Ca2+ interaction with Nt-CaRes in ANO6 than ANO6-1-6. Moreover, mutations on potentially Ca2+-interacting acidic amino acids in ANO6 Nt-CaRes resulted in reduced Ca2+ sensitivity, implying direct interactions of Ca2+ with these residues. Based on these results, we cautiously suggest that the net charge of Nt-CaRes is responsible for the difference in Ca2+ sensitivity between ANO1 and ANO6.
Keywords
anoctamin 6; calcium-binding domain; calcium sensitivity; TMEM16F;
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