Dynamic role of the tether helix in PIP₂-dependent gating of a G protein-gated potassium channel

G protein-gated inwardly rectifying potassium (GIRK) channels control neuronal excitability in the brain and are implicated in several different neurological diseases. The anionic phospholipid phosphatidylinositol 4,5 bisphosphate (PIP₂) is an essential cofactor for GIRK channel gating, but the precise mechanism by which PIP₂ opens GIRK channels remains poorly understood. Previous structural studies have revealed several highly conserved, positively charged residues in the "tether helix" (C-linker) that interact with the negatively charged PIP₂. However, these crystal structures of neuronal GIRK channels in complex with PIP₂ provide only snapshots of PIP₂'s interaction with the channel and thus lack details about the gating transitions triggered by PIP₂ binding. Here, our functional studies reveal that one of these conserved basic residues in GIRK2, Lys200 (6'K), supports a complex and dynamic interaction with PIP₂. When Lys200 is mutated to an uncharged amino acid, it activates the channel by enhancing the interaction with PIP₂. Atomistic molecular dynamic simulations of neuronal GIRK2 with the same 6' substitution reveal an open GIRK2 channel with PIP₂ molecules adopting novel positions. This dynamic interaction with PIP₂ may explain the intrinsic low open probability of GIRK channels and the mechanism underlying activation by G protein Gβγ subunits and ethanol.