MD simulations of GIRK2 6′Y reveal dynamic opening of the channel at the HBC gate. (a) The relative PIP₂ association number (see Materials and methods) is plotted as a function of simulation time for WT (6′K, blue) and 6′Y (green) channels. (b) The cross-distance diameter of the pore at the HBC, measured between the center of mass of F192 (−2′F) side chain, is plotted as a function of simulation time. (c) The water molecule density in the conduction pathway, moving in the z plane from the selectivity filter (0 Å) to the bottom of the M2 transmembrane domain (approximately −25 Å), is plotted as a function of the simulation time for WT (top) and 6′Y (bottom) channels. Note the loss of water around the region of the −2′F in WT channels (arrow). (d and e) Structural view of the HBC at 400 ns for GIRK2-WT (6′K; d) and GIRK2-6′Y (e). Note how PIP₂ is bound loosely for WT and more tightly for 6′Y. (f) Superimposition of one subunit in 6′K (gray) and 6′Y (yellow) showing movement of M2 relative to G180 in 6′Y (red). (g) Cartoon summarizes hydrogen bond interactions for PIP₂ and GIRK2 for new PIP₂-closed and PIP₂-open conformations based on MD simulations. EtOH, ethanol.