To provide structural insights how 5b binds to the CTD of human Hsp90, we performed 40 independent molecular dynamics (MD) simulations of free diffusion of 5b in the presence of truncated monomeric Hsp90α (aa 294-699), using the Amber 18 suite of molecular simulation programs 46 and the ff14SB and a modified GAFF force field for protein and ligand. Initially, we generated 40 individual starting configurations by randomly placing 5b and the CTD structure, leaving at least 10 Å between atoms in 5b and the CTD structure. After minimization, thermalization, and density adaptation, we performed MD simulations of 500 ns length, in which the 5b molecule diffused freely. To counter the high flexibility of the C-terminal helix interface, we introduced positional restraints on the backbone atoms, adjusting the reference coordinates every 100 ns to allow for moderate protein movements.