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Water Networks Repopulate Protein-Ligand Interfaces with Temperature.

ABSTRACT: High-resolution crystal structures highlight the importance of water networks in protein-ligand interactions. However, as these are typically determined at cryogenic temperature, resulting insights may be structurally precise but not biologically accurate. By collecting 10 matched room-temperature and cryogenic datasets of the biomedical target Hsp90α, we identified changes in water networks that impact protein conformations at the ligand binding interface. Water repositioning with temperature repopulates protein ensembles and ligand interactions. We introduce Flipper conformational barcodes to identify temperature-sensitive regions in electron density maps. This revealed that temperature-responsive states coincide with ligand-responsive regions and capture unique binding signatures that disappear upon cryo-cooling. Our results have implications for discovering Hsp90 selective ligands, and, more generally, for the utility of hidden protein and water conformations in drug discovery.

SUBMITTER: Stachowski TR

PROVIDER: S-EPMC9329195 | biostudies-literature | 2022 Aug

REPOSITORIES: biostudies-literature

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Water Networks Repopulate Protein-Ligand Interfaces with Temperature.

Stachowski Timothy R TR Vanarotti Murugendra M Seetharaman Jayaraman J Lopez Karlo K Fischer Marcus M

Angewandte Chemie (International ed. in English) 20220621 31

High-resolution crystal structures highlight the importance of water networks in protein-ligand interactions. However, as these are typically determined at cryogenic temperature, resulting insights may be structurally precise but not biologically accurate. By collecting 10 matched room-temperature and cryogenic datasets of the biomedical target Hsp90α, we identified changes in water networks that impact protein conformations at the ligand binding interface. Water repositioning with temperature r ...[more]

PMID: 35648650

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Project description:Rational drug design focuses on the explanation and prediction of complex formation between therapeutic targets and small-molecule ligands. As a third and often overlooked interacting partner, water molecules play a critical role in the thermodynamics of protein-ligand binding, impacting both the entropy and enthalpy components of the binding free energy and by extension, on-target affinity and bioactivity. The community has realized the importance of binding site waters, as evidenced by the number of computational tools to predict the structure and thermodynamics of their networks. However, quantitative experimental characterization of relevant protein-ligand-water systems, and consequently the validation of these modeling methods, remains challenging. Here, we investigated the impact of solvent exchange from light (H2O) to heavy water (D2O) to provide complete thermodynamic profiling of these ternary systems. Utilizing the solvent isotope effects, we gain a deeper understanding of the energetic contributions of various components. Specifically, we conducted isothermal titration calorimetry experiments on trypsin with a series of p-substituted benzamidines, as well as carbonic anhydrase II (CAII) with a series of aromatic sulfonamides. Significant differences in binding enthalpies found between light vs heavy water indicate a substantial role of the binding site water network in protein-ligand binding. Next, we challenged two conceptually distinct modeling methods, the grid-based WaterFLAP and the molecular dynamics-based MobyWat, by predicting and scoring relevant water networks. The predicted water positions accurately reproduce those in available high-resolution X-ray and neutron diffraction structures of the relevant protein-ligand complexes. Estimated energetic contributions of the identified water networks were corroborated by the experimental thermodynamics data. Besides providing a direct validation for the predictive power of these methods, our findings confirmed the importance of considering binding site water networks in computational ligand design.

Dataset Information

Water Networks Repopulate Protein-Ligand Interfaces with Temperature.

Publications

Water Networks Repopulate Protein-Ligand Interfaces with Temperature.

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