| MD run . | Simulation time (µs) . | Applied electric field (mV) . | Ion sites occupied at starting state . | HBC gate permeation events . |
|---|---|---|---|---|
| 200ns_run1 | 0.2 | — | S2, S4 | 0 |
| 200ns_run2 | 0.2 | — | S2, S4 | 0 |
| 200ns_run3 | 0.2 | — | S2, S4 | 0 |
| 200ns_run4 | 0.2 | — | S2, S4 | 0 |
| 200ns_run5 | 0.2 | — | S2, S4 | 0 |
| 200ns_run6 | 0.2 | — | S2, S4 | 1 |
| 200ns_run7 | 0.2 | — | S2, S4 | 1 |
| 200ns_run8 | 0.2 | — | S2, S4 | 1 |
| 200ns_run9 | 0.2 | — | S2, S4 | 0 |
| 200ns_run10 | 0.2 | — | S2, S4 | 0 |
| Full SF permeation events | ||||
| 1µs_run1 | 1 | 580 | S1, S2, S4 | 30 |
| 1µs_run2 | 1 | 580 | S1, S2, S4 | 9 |
| 1µs_run3 | 1 | 580 | S1, S2, S4 | 0 |
| 1µs_run4 | 1 | 580 | S2, S3 | 8 |
| 1µs_run5 | 1 | 580 | S2, S3 | 27 |
| 1µs_run6 | 1 | 580 | S2, S3 | 20 |
| 1µs_run7 | 1 | 580 | S2, S3 | 15 |
| 1µs_run8a | 1 | 580 | S2, S3 | 24 |
| 1µs_run9a | 1 | 580 | S2, S3 | 19 |
| 1µs_run10a | 1 | 580 | S2, S3 | 27 |
| 1µs_run11 | 1 | 290 | S2, S3 | 0 |
| 1µs_run12 | 1 | 290 | S2, S3 | 19 |
| 1µs_run13a | 1 | 290 | S2, S3 | 27 |
| 1µs_run14a | 1 | 290 | S2, S3 | 3 |
| Total number | 16 | 228 |
| MD run . | Simulation time (µs) . | Applied electric field (mV) . | Ion sites occupied at starting state . | HBC gate permeation events . |
|---|---|---|---|---|
| 200ns_run1 | 0.2 | — | S2, S4 | 0 |
| 200ns_run2 | 0.2 | — | S2, S4 | 0 |
| 200ns_run3 | 0.2 | — | S2, S4 | 0 |
| 200ns_run4 | 0.2 | — | S2, S4 | 0 |
| 200ns_run5 | 0.2 | — | S2, S4 | 0 |
| 200ns_run6 | 0.2 | — | S2, S4 | 1 |
| 200ns_run7 | 0.2 | — | S2, S4 | 1 |
| 200ns_run8 | 0.2 | — | S2, S4 | 1 |
| 200ns_run9 | 0.2 | — | S2, S4 | 0 |
| 200ns_run10 | 0.2 | — | S2, S4 | 0 |
| Full SF permeation events | ||||
| 1µs_run1 | 1 | 580 | S1, S2, S4 | 30 |
| 1µs_run2 | 1 | 580 | S1, S2, S4 | 9 |
| 1µs_run3 | 1 | 580 | S1, S2, S4 | 0 |
| 1µs_run4 | 1 | 580 | S2, S3 | 8 |
| 1µs_run5 | 1 | 580 | S2, S3 | 27 |
| 1µs_run6 | 1 | 580 | S2, S3 | 20 |
| 1µs_run7 | 1 | 580 | S2, S3 | 15 |
| 1µs_run8a | 1 | 580 | S2, S3 | 24 |
| 1µs_run9a | 1 | 580 | S2, S3 | 19 |
| 1µs_run10a | 1 | 580 | S2, S3 | 27 |
| 1µs_run11 | 1 | 290 | S2, S3 | 0 |
| 1µs_run12 | 1 | 290 | S2, S3 | 19 |
| 1µs_run13a | 1 | 290 | S2, S3 | 27 |
| 1µs_run14a | 1 | 290 | S2, S3 | 3 |
| Total number | 16 | 228 |
10 replicas of 0.2-µs free MD simulations were performed using the equilibrated system of the crystal structure with bound PIP2 (PDB accession no. 3SYA). 14 1-µs simulations were performed with an applied electric field. 1µs_run1, 1µs_run2 and 1µs_run3 started from a snapshot at 50ns of 200ns_run8. All the other MD runs with applied electric field were initiated from a snapshot of 1µs_run1 (t = 400 ns), where the HBC gate reached a steady state of a relatively wide and solvated conformation (see Figs. S4 and S5 B). For all 1-µs simulations, either 40 mV nm−1 or 20 mV nm−1 was applied; this equals ∼580 mV or ∼290 mV, respectively, when calculated over the whole box (z axis; Roux, 2008; Bjelkmar et al., 2009; Gumbart et al., 2012).
Harmonic restraints on G-loop gate–forming residues G318 and M319.