Fig. 1. The transmembrane domain (S1–S6 helixes) and the pore region of cryo-EM TRPV1 structures in (a) closed state (PDB ID: 3J5P) and (b) open state (PDB ID: 3J5Q). The pore is drawn using the HOLE program.^[53] The two gates (upper and lower gates) and the corresponding residues, G643 (upper gate) and I679 (lower gate), are shown. (c) The root-mean-square deviations of four 100 ns molecular dynamics simulations at 310 K, 340 K, 360 K and 410 K. (d) A presentative open conformation of the pore obtained from molecular dynamic simulations.

Fig. 2. Time course of upper gate’s open events in the calculated temperatures. The points with the value of 1 (denoted in red) represent the open state, and those with the value of 0 (denoted in blue) represent the closed state. Because of the size of the drawing points, at some moment it appears that the upper gate is in both closed and open state, which indicates the oscillation of the upper gate between closed and open states.

Fig. 3. Sodium ion blocks the pore at the upper gate and interacts with the carbonyl oxygen atoms of G643s. The distances between the sodium ion and the oxygens of G643s of the tetramer in the simulation at 410 K temperature are shown. The inner figure shows the blocking conformation of the sodium ion.

Fig. 4. Location of M644’s side chain. (a) Conformation of side chain of M644 locates in the hydrophobic pocket formed by I642, K639 of the same monomer and F640, Y671, V667, L647 of the adjacent chain. (b) Distributions of side chain of M644 at different temperatures. The abscissa is the distance between the methyl group of M644 and C_α of Y671. The green arrows shown in the abscissa are the distances between M644 and Y671 in the closed (∼ 14 Å) and open (∼ 8 Å) state of cryo-EM structures.

Fig. 5. Time course of lower gate’s open events in the calculated temperatures. The open probability of the lower gate shows temperature-dependent character.

Fig. 6. Temperature dependency of lower-gate opening (back line), formation probability of HB between E570 and S510 (red line) and that of HB between E570 and S512 (blue line).

Fig. 7. (a) Outward movement of the N-terminal half of S5 helix. S6 helix at the position of I679 has the same outward movement. (b) Hydrophobic contact between S5 and S6 helixes. The hydrophobic residues are shown in licorice mode. I679 and D576 are explicitly shown using the VDW mode. The hydrogen atoms of the hydrophobic residues are omitted. (c) Principle component analysis shows that S5 helixes of the tetramer show the oscillation movement of S5 helixes. (d) Hydrogen bonds between E570 of S5 helix and S512/S510 of S3 helix.