Abstract
The lipid scramblase TMEM16F initiates blood coagulation by catalyzing the exposure of phosphatidylserine in platelets. The protein is part of a family of membrane proteins, which encompasses calcium-activated channels for ions and lipids. Here, we reveal features of murine TMEM16F (mTMEM16F) that underlie its function as a lipid scramblase and an ion channel. The cryo-EM data of mTMEM16F in absence and presence of Ca2+ define the ligand-free closed conformation of the protein and the structure of a Ca2+-bound intermediate. Both conformations resemble their counterparts of the scrambling-incompetent anion channel mTMEM16A, yet with distinct differences in the region of ion and lipid permeation. In conjunction with functional data, we demonstrate the relationship between ion conduction and lipid scrambling. Although activated by a common mechanism, both functions appear to be mediated by alternate protein conformations that are at equilibrium in the ligand-bound state.
Highlights
Lipid scramblases facilitate the movement of lipids between both leaflets of the bilayer (Bevers and Williamson, 2016; Nagata et al, 2016; Williamson, 2015), thereby dissipating the asymmetry present in the membrane
We explored the relationship between the structure of mTMEM16F, which shares a sequence identity of over 90% with its human orthologue, to its diverse functional properties
To confirm that the purified protein has retained its function as a Ca2+-activated lipid scramblase, we investigated lipid transport with proteoliposomes using an assay that was previously established for fungal TMEM16 scramblases (Malvezzi et al, 2013; Ploier and Menon, 2016)
Summary
Lipid scramblases facilitate the movement of lipids between both leaflets of the bilayer (Bevers and Williamson, 2016; Nagata et al, 2016; Williamson, 2015), thereby dissipating the asymmetry present in the membrane. In mTMEM16A, the rearrangement of an a-helix that lines one edge of the ‘subunit cavity’ in nhTMEM16 seals the membrane-accessible furrow, resulting in the formation of a protein-enclosed aqueous pore that is for a large part shielded from the bilayer. In both proteins, binding of Ca2+ mediates the activation of the permeation region contained in each subunit, which in mTMEM16A was shown to act as an independent entity (Jeng et al, 2016; Lim et al, 2016). In the most plausible scenario, both transport processes are activated by the same mechanism and mediated by distinct protein conformations, which are at equilibrium in a calcium-bound state
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