Figure 5.

Energetic instability and structural unfolding of the R114G T1 tetramer. A. Tetrameric T1 structures derived from MD simulations for WT and R114G at 1 ns and 3.0 μs viewed from the intracellular side. In each subunit (A, B, C, D), helices are colored red, β strands yellow, loops green. Structures are rendered with PyMol. Symbols (Δ,Δ) represent one of three independent MD simulations of WT and R114G, respectively (See Supplementary Figure S5A for all three simulation-based structures for WT and for R114G). B. Root-mean-square deviations (RMSDs) of non-hydrogen atoms from three independent WT (blue) and three independent R114G (red) simulations. Symbols to the right of the traces indicate different simulation runs. C. Mean nonbonded potential energies (E_nb) of the WT (blue) and R114G (red) T1 tetramers from three independent simulations (n = 3 for each curve) was calculated as the sum of the van der Waals and electrostatic energies. Lightly shaded areas represent standard deviation. D. Dihedral angles and distance moved for WT and R114G variants. Left: Changes in φ and ψ angles for select residues in a subunit from one illustrative WT system (blue) and a subunit from one illustrative R114G system (red) during the 3-μs simulation are shown superimposed. For each residue indicated along the ordinate, time progresses from 0 to 3 μs downward. Secondary conformations are assigned according to PyMol as in Figure 1B. Right: Distance moved for the α-carbon of each selected residue from its initial location at 1 ns to its location at 3 μs in the four subunits (A, B, C, D) in the systems shown in (A). Vertical arrow indicates that each plot progresses from N-terminal to C-terminal residues. The measured intersubunit distance for crosslinkable cysteines at R170 and D178 was 7–10 Å for WT vs. 25–27 Å for variant R114G.