(a) Conformational ensemble view of αS and αS-Fasudil ensemble, (b) A schematic of β-Variational Autoencoder (β-VAE), (c) Training and validation loss for β-VAE and (d) RMSE as a function of the β parameter. The red-annotated β parameter was used for the investigation

The free energy landscape of (a) αS and (b) αS-Fasudil system as determined from the latent dimensions of β-VAE

(a) A flowchart of the process of building a Markov State Model (MSM). (b, c) Implied timescales plot of the αS and αS-Fasudil systems. (c) Macrostate populations of the 3-state and 6-state MSM of the αS and αS-Fasudil systems. Bootstrapping was used to estimate the mean and standard deviations of the macrostate populations.

Intrapeptide residue-wise contact probability maps of (a) six macrostates (FS1 to FS6) in the presence of fasudil. A contact is considered if the Cα atoms of two residues are within a distance of 8 Å of eachother. Axes denote the residue numbers. The color scale for the contact probability is shown at the extreme right of each panel of maps. The color bar along the axes of the plots represents the segments in the αS monomer. Specific contact regions are marked by boxes and numbered. These contacts are illustrated by representative snapshots and the corresponding contacts are similarly marked and numbered.

Intrapeptide residue-wise contact probability maps of the three macrostates (MS1 to MS3) in neat water. A contact is considered if the Cα atoms of two residues are within a distance of 8 Å of eachother. Axes denote the residue numbers. The color scale for the contact probability is shown at the extreme right. The color bar along the axes of the plots represents the segments in the αS monomer. Specific contact regions are marked by boxes and numbered. These contacts are illustrated by representative snapshots and the corresponding contacts are similarly marked and numbered.

(a) Schematic of denoising convolutional variational autoencoder (b) Latent space of the contact map of αS and αS-Fasudil metastable states. (c) SSIM and PSNR values for αS and αS-Fasudil metastable states.

(a) Contact probability map of residue-wise interaction of fasudil with αS. (b) Overlay of representative conformations from the six macrostates (FS1 to FS6) along with the bound fasudil molecules. The conformations are colored segment-wise as shown in the legend. The fasudil molecules, in licorice representation, are colored yellow.

(a) Total Protein entropy of the metastable states of αS and αS-Fasudil calculated from the torsion angles, relative to a fully flexible chain (b) Total water entropy in αS and αS-Fasudil system and (c) Residue-wise backbone entropy within the αS and αS-Fasudil states.

The rate network obtained from MFPT analysis for the (a) 3-state MSM of the αS ensemble in neat water and (b) 6-state MSM of the αS-Fasudil ensemble. The sizes of the discs is proportional to the stationary population of the respective state. The thickness of the arrows connecting the states is proportional to the transition rate (1/MFPT) between the two states and the MFPT values are shown on the arrows. (c, d) Projection of conformation sub-ensemble PCCA+ for αS and αS-Fasudil system in latent space, with respective protein entropy values (in unit of J mol−1 K −1) annotated on top of it. (e) Correlation between protein entropy and transition time to the major state for αS and αS-Fasudil system. The inset plot corresponds to the correlation between entropy and transition to the macrostate for αS and αS-Fasudl system.