Insights from Fragment Hit Binding Assays by Molecular Simulations

Abstract

Novel bioactive molecules can be rationally designed by growing and linking small fragments. Because fragments are fast and promiscuous, it is common to have contradictory hit results between different experimental screening techniques. Here, we simultaneously determine fragment binding poses, affinities, and kinetics on a focused library of 42 fragments against the serine protease factor Xa using multimillisecond molecular dynamics simulations. We predict experimental poses of 12 over 15 S1 crystal structures, and affinities are recovered for 4 out of 6. A kinetic map of protein cavities is computed in terms of on- and off-rates as well as insights into secondary ligand poses. The results suggest that the approach can be useful to recapitulate discordant fragment screening data.

Binding Kinetics in Drug Discovery

“…Aside from the limitations we have mentioned when using biasing methods, the use of unbiased simulations, although theoretically more correct, still present some obstacles. We came across some of these challenges during the study of the large set of fragments focused for the factor Xa protease . For instance, one of the fragments, the largest in the library, lacked enough sampling to produce good statistics for a converged model.…”

“…However, although visualization of these ground‐level species is experimentally plausible, determination of short‐lived species, as meta‐stable states, or the TS, is experimentally very complicated. To date, MD has already proven to be a powerful tool for the quantitative estimation of kinetics of fast associating fragments,,, but it has also been very interesting for the understanding of binding pathways, intermediates and other kinetic determinants, difficult to characterize by experiments. Shaw et al .…”

“…More interestingly,t his study permitted to characterize the role of al oop in the vicinities of the pocket during the binding pathway:t he loop was able to explore both opena nd closed conformations in absence of the ligand, however, the ligand stabilizedt he open conformer while entering the pocket, and the closed one once insidei t. [65] More recently we focusedo na nl argerl ibrary of 42 fragments against factor Xa, as erine protease with 2k nown targetable pockets. [66] Using more than2ms of total simulation time it was possible to obtain thermodynamic, poses and kinetic estimates in gooda greement with experimental results. Interestingly,S 1f ragments with amine or amidine groups showed slower on and off rates compared to S1 neutral binders (Fig.…”

“…In some cases, very complex rearrangement of the protein should be captured in order to obtain this. However,e ven [66].I nteraction rate plot as obtained in for as et of small fragments against the serine protease factor Xa. Most potent and resident molecules contained amidine and basic moieties, whereas the rapidly dissociating fragments, encircled in orange, were neutral.…”

Recent advances in dynamic docking for drug discovery

“…However, with the future advent of even more powerful hardware and software for MD simulations, it is only a matter of time before we see unbiased MD being routinely used to screen mid‐sized compound libraries in the hit identification phase of drug discovery . Noticeably, multimillisecond MD simulations have been recently used to screen a small library of fragments, returning binding poses, affinities, and kinetics for a set of ligand poses, which might be helpful to recapitulate discordant fragment screening data . Although we may expect extensive MD simulations to be performed routinely in the near future, kinetics could remain an issue.…”