Fig 1.
Schematic of the molecular fragments method.
(A) Each structural constraint (e.g., a metal ion bridge or a spin-label) is present as a molecular fragment with all atomic details in the system. (B) The residue(s) in the molecular fragment (colored in red) is/are attached to the targeting residue(s) in the wild-type protein, via harmonic restraints, with their backbone atoms (N, C, O and Cα) staying on top of each other, respectively. The residue in the molecular fragment does not have interactions with its targeting residue in the wild-type protein (residue i), and the interactions between the backbone atoms of the residue in the molecular fragment and the backbone atoms of the two nearby residues in the wild-type protein (residues i-1 and i+1; embraced by red dashed lines) are also turned off. (C) During the restrained MD simulations, the interactions within each molecular fragment are evaluated accurately triggering the conformational changes of the wild-type protein, while different molecular fragments do not have interactions with one another.
Table 1.
Details of the models of the VSD of the Kv1.2 channel.
Fig 2.
Starting configurations of the four models of the VSD with different molecular fragments.
(A) model-1. (B) model-2. (C) model-3. (D) model-4. The four TM helices, S1 (gray), S2 (yellow), S3 (red) and S4 (blue), are represented in transparent ribbons. Molecular fragments, I177C–Cd2+–R294C, I230C–Cd2+–R294C, I230D–Mg2+–F267D, F233W/E236–R294K, I230H–Zn2+–A291H, I268C–Cd2+–A287C, and T269C–Cd2+–A291C, are colored in red, yellow, green, magenta, cyan, orange, and blue, respectively, with amino acids being represented in sticks and ions in spheres.
Table 2.
Force field parameters for the metal ion bridges.
Fig 3.
Final configurations of the four models of the VSD with different molecular fragments.
The final configuration (left panel) and the correlation between the Cβ-Cβ distance and the Sγ-Cd2+, Oδ2-Mg2+, Nε-Zn2+, or Nζ-Oε2 distance in each molecular fragment (right panel) of (A) model-1, (B) model-2, (C) model-3, and (D) model-4. The last 10 ns trajectories of the restrained MD simulations were used to calculate the average values and standard deviations of the distances.
Fig 4.
The final coordinates of the molecular fragments in model-3 after the restrained MD simulation.
(A–F) The configurations of the four TM helices before and after the restrained MD simulation are shown in transparent and solid ribbons, respectively.
Fig 5.
Superimposition of the initial and final configurations of the four models of the VSD.
(A) model-1, (B) model-2, (C) model-3, and (D) model-4. The initial and final configurations of the backbone atoms are represented in transparent and solid sticks, respectively.
Table 3.
The backbone RMSD values of the TM helices S1–S4 between different models.
Fig 6.
Configurations of the spin-labels attached to T4 lysozyme.
(A) A snapshot of the T4 lysozyme system with 34 MTSSL spin-labels. The spin-label at each site has 25 copies and highlighted in the ball-and-stick representation. (B) Distance histograms of four pairs of spin-labels from ESR/DEER experiments (black solid line), and the molecular dynamics simulations without (blue dashed line) and with (red dashed line) an energy restraint. Please see the Supplementary Material, S3 and S4 Figs for the other 47 distance distributions.