Fig. 3
Free-energy landscape underlying GlpG’s unfolding and extraction from a bilayer under high applied force (Top) Two-dimensional free energy landscape at high applied force as a function of Z, the average of the z-coordinates of the Cα atoms, and D, the distance between the N-terminus and C-terminus of GlpG. Relative free energies are indicated with colors in units of kT, where blue indicates a low-free energy and red indicates a high-free energy. An unfolding path is shown as a purple line drawn from the folded state, N, (Z ≈ −3 Å, D ≈ 37 Å) to the unfolded state, U (Z ≈ −17 Å, D ≈ 270 Å). (Bottom) One-dimensional free energy profile along the path shown in the top panel as a function of the end-to-end distance, D. Representative structures from TS and I1 are shown in Fig. 2. Representative structures from I2, U1, and U2 are shown in Fig. 4. Under these high force conditions, the folded state is metastable and is separated by a large barrier from the first intermediate state, I1. The barriers separating the intermediate states I1, I2, and U are significantly smaller than the barrier between N and I1. At this particular value of the applied force, the completely extended state U2 is slightly higher in free energy than U1, which has TM1–2 inserted in the membrane but unfolded (see Fig. 4). At larger values of the applied force, U2 becomes the global free energy minimum
