Estimation of Protein–Ligand Unbinding Kinetics Using Non-Equilibrium Targeted Molecular Dynamics Simulations

Abstract

We here report on non-equilibrium targeted Molecular Dynamics simulations as tool for the estimation of protein-ligand unbinding kinetics. With this method, we furthermore investigate the molecular basis determining unbinding rates, correlating simulations with experimental data from SPR kinetics measurements and X-ray crystallography on two small molecule compound libraries bound to the N-terminal domain of the chaperone Hsp90. Within the investigated libraries, we find ligand conformational changes and protein-ligand nonbonded interactions as discriminators for unbinding rates. Ligands with flexible chemical scaffold may remain longer at the protein target if they need to pass through extended conformations upon unbinding, or if they exhibit strong electrostatic and/or van der Waals interactions with the target. Ligands with rigid chemical scaffold can exhibit longer residence times if they need to perform any kind of conformational change for unbinding, while electrostatic interactions with the protein can facilitate unbinding. Our resultsshow that understanding the unbinding pathway and the protein-ligand interactions along this path is crucial for the prediction of small molecule ligands with defined unbinding kinetics. Supporting InformationFour supporting tables, nine supporting figures and additional references (PDF) SMILES annotations (CSV) Accession CodesThe crystallographic coordinates of novel compounds are deposited in the Protein Data Bank under the accession codes 5LRL (2d) and 5LO1 (2j). Authors will release the atomic coordinates and experimental data upon article publication.

Multisecond ligand dissociation dynamics from atomistic simulations

“…The second investigated protein complex is the N-terminal domain of heat shock protein 90 (Hsp90) bound to a resorcinol scaffold-based inhibitor (1j in ref. 47 ). This protein has recently been established as a test system for investigating the molecular effects influencing binding kinetics [47][48][49][50] , and the selected inhibitor unbinds on a scale of half a minute.…”

“…47 ). This protein has recently been established as a test system for investigating the molecular effects influencing binding kinetics [47][48][49][50] , and the selected inhibitor unbinds on a scale of half a minute. From the overall appearance of free energy and friction profiles (Fig.…”

Hierarchical coupling between ATP hydrolysis and Hsp90’s client binding site

“…To accelerate structural changes leading to closed state B in simulations and to overcome the timescale limitations of unbiased simulations, we employed nonequilibrium targeted MD simulations 56,57 with implicit solvent for one of the ADP simulations (see Methods for details). The simulations indeed show that the distance distributions evolve towards the experimentally measured distances (see Fig.…”

“…Visualization of molecular data was performed with vmd. 78 For nonequilibrium targeted molecular dynamics simulations, 56,57 we employed the PULL code from Gromacs. Prior to these simulations, we removed bulk solvent and ions from the final structure of one of the ADP+P i simulations after 1 µs simulation time, mimicking the influence of water by setting the relative permittivity r = 78 in the simulation, while lowering the overall friction of the system.…”

Exhaustive search of the configurational space of heat‐shock protein 90 with its inhibitor by multicanonical molecular dynamics based dynamic docking

“…[ 23 ] Therefore, Hsp90 has attracted much attention in the field of biochemistry and drug design. [ 24–28 ] Structurally, Hsp90 consists of three domains: the N‐terminal domain, the middle domain, and the C‐terminal domain. The N‐terminal domain contains a nucleotide binding site that binds adenosine triphosphate (ATP), which is hydrolyzed to adenosine diphosphate (ADP) after an immature client protein has bound to Hsp90, upon which ADP dissociates from the site.…”