Dynamic change in hydrogen bonding with PIP₂ in GIRK2-6′K and GIRK2-6′Y MD simulations. (a) Examples of GIRK2/PIP₂ structures at the start and after 200 ns of MD simulations for GIRK2-6′K (WT) and GIRK2-6′Y channels. Note the 5′-PO₄ of PIP₂ moves away from starting position in the WT simulation. In contrast, the 5′-PO₄ of PIP₂ appears to engage PIP₂ more deeply in the pocket in the 6′Y channel. One of four subunits is shown. (b) Each graph plots the number of hydrogen bonds during the 400-ns simulation time for the indicated pairs for WT GIRK2-6′K (blue) and GIRK2-′Y (green) for the PIP₂ binding pockets shown in a. (c and d) Bar graphs show the mean relative probability of hydrogen bonds for all four subunits during the two 400-ns simulations for GIRK2-6′K (blue) and GIRK2-6′Y (green) channels. (c) Probability of hydrogen bonding between 9′E and 6′K/Y. (d) Probability of hydrogen bonding for K64, K90, 0′K, 5′K, and 6′K/Y with 5′-PO₄ of PIP₂ (of two maximal bonds). Note the number of hydrogen bonds increases with 0′K and 5′K but decreases for K64 with the 6′Y channel simulations. Bars represent mean ± SEM. *, P < 0.05; ns, not significant; Student’s t test.