A Partially Closed State in Nhtmem16 Scramblase is Enabled by Lipid Tail Insertion into the Protein Groove

Abstract

Ca2+-regulated TMEM16 proteins participate in key physiological processes such as ion transport and phospholipid scrambling. Studies of various members of the TMEM16 family have supported a functional role for the membrane-exposed groove in these proteins, showing that it can assist translocation of both ions and lipids by serving as a continuous conduit connecting the two leaflets of the membrane. The proposed mechanism of permeation involves gating of various regions of the groove through conformational rearrangements. Especially intriguing is the role of transmembrane helix 4 (TM4) that was seen in structural comparisons of the TMEM16A ion channel with TMEM16 scramblases to adopt different positions relative to the groove. To investigate the molecular mechanisms underlying the transitions between these different structural states we analyzed the dynamic properties of the nhTMEM16 scramblase with extensive atomistic molecular dynamics simulations. We found that occlusion of the groove by TM4 can occur in the Ca2+-bound state and is enabled by lipid tail penetration into the groove. Based on information about the relation between the dynamics of the scramblase and the conformational changes that facilitate lipid tail insertion into the groove, we identified a mutation in the nhTMEM16 that can stabilize the Ca2+-bound scramblase in the occluded conformation that is structurally similar to the Ca2+-bound form of the TMEM16A. These findings elucidate on how lipid-protein interactions can mechanistically drive formation of a functionally relevant state in nhTMEM16.