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. 2021 Apr;35(4):433-451.
doi: 10.1007/s10822-020-00354-6. Epub 2020 Oct 27.

The critical role of QM/MM X-ray refinement and accurate tautomer/protomer determination in structure-based drug design

Oleg Y Borbulevych  1 , Roger I Martin  1 , Lance M Westerhoff  2
Affiliations

The critical role of QM/MM X-ray refinement and accurate tautomer/protomer determination in structure-based drug design

Oleg Y Borbulevych et al. J Comput Aided Mol Des. 2021 Apr.

Abstract

Conventional protein:ligand crystallographic refinement uses stereochemistry restraints coupled with a rudimentary energy functional to ensure the correct geometry of the model of the macromolecule-along with any bound ligand(s)-within the context of the experimental, X-ray density. These methods generally lack explicit terms for electrostatics, polarization, dispersion, hydrogen bonds, and other key interactions, and instead they use pre-determined parameters (e.g. bond lengths, angles, and torsions) to drive structural refinement. In order to address this deficiency and obtain a more complete and ultimately more accurate structure, we have developed an automated approach for macromolecular refinement based on a two layer, QM/MM (ONIOM) scheme as implemented within our DivCon Discovery Suite and "plugged in" to two mainstream crystallographic packages: PHENIX and BUSTER. This implementation is able to use one or more region layer(s), which is(are) characterized using linear-scaling, semi-empirical quantum mechanics, followed by a system layer which includes the balance of the model and which is described using a molecular mechanics functional. In this work, we applied our Phenix/DivCon refinement method-coupled with our XModeScore method for experimental tautomer/protomer state determination-to the characterization of structure sets relevant to structure-based drug design (SBDD). We then use these newly refined structures to show the impact of QM/MM X-ray refined structure on our understanding of function by exploring the influence of these improved structures on protein:ligand binding affinity prediction (and we likewise show how we use post-refinement scoring outliers to inform subsequent X-ray crystallographic efforts). Through this endeavor, we demonstrate a computational chemistry ↔ structural biology (X-ray crystallography) "feedback loop" which has utility in industrial and academic pharmaceutical research as well as other allied fields.

Keywords: CSAR set; Ligand strain; Quantum mechanics X-ray refinement; Structure guided drug discovery; Structure-based drug discovery; X-ray crystallography; difference density Z-score; high throughput crystallography; protonation states; tautomers.

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Figures

Fig. 1
Fig. 1
Schematic view of the QM/MM two-layer (ONIOM) concept depicting two ligand QM regions with the balance of the receptor treated as a MM layer. This method can support any number of QM regions and may even treat the entire structure as a QM structure
Fig. 2
Fig. 2
Histogram of Ligand Strain Energy (LLSE) distributions for ligands from 55 CSAR structures refined using QM/MM method and conventional PHENIX. The lower the LLSE the less strain the ligand must accommodate to fit within its associated active site
Fig. 3
Fig. 3
Histogram of Ligand Z-score of the difference density (ZDD) distributions for ligands from 55 CSAR structures refined using QM/MM method and conventional PHENIX. The lower the ZDD the more accurate the model versus the experimental density
Fig. 4
Fig. 4
The σA-weighted mFo-DFc difference electron density map drawn at 3σ level around the ligand (ligand ID 60 K) in the PDB structure 4FKU refined with QM/MM (a) and conventional (b). The σA-weighted 2mFo-DFc electron density map is contoured at 1 σ. C is provided as an overlay of the two conformations
Fig. 5
Fig. 5
The regression lines of the correlation between experimental affinity (− logK) and computationally predicted GBVI/WSA scores for the 15 protein:ligand CDK2 complexes for PHENIX structures (Black), QM/MM structures (Red), hand-modified QM/MM structures (Green), and QM/MM refined structures with XModeScore chosen tautomers (Blue). Points involving structures discussed in the paper are labeled
Fig. 6
Fig. 6
The regression lines of correlation between experimental affinity (− logK) and computationally predicted GBVI/WSA scores for the 12 protein:ligand ERK2 complexes for PHENIX structures (Black), QM/MM structures (Red), and QM/MM refined structures with XModeScore chosen tautomers (Blue). Points involving structures discussed in the paper are labeled
Fig. 7
Fig. 7
The regression lines of correlation between experimental affinity (− logK) and computationally predicted GBVI/WSA scores for the 7 protein:ligand uPA complexes for PHENIX structures (Black), QM/MM structures (Red), hand-modified QM/MM structures (Green), and QM/MM refined structures with XModeScore chosen tautomers (Blue). Points involving structures discussed in the paper are labeled
Fig. 8
Fig. 8
The regression lines of correlation between experimental affinity (− logK) and computationally predicted GBVI/WSA scores for the protein target CHK1 for PHENIX structures (Black), QM/MM structures (Red), and QM/MM refined structures with XModeScore chosen tautomers (Blue). Points involving structures discussed in the paper are labeled
Fig. 9
Fig. 9
The σA-weighted mFo-DFc difference electron density map drawn at 3σ level around the ligand (ligand ID 46 K) in the PDB structure 4FKS refined with QM/MM (green) (a) and conventional (yellow) (b), as well as the superimposition of the two structures (c). The σA-weighted 2mFo-DFc electron density map is contoured at 1σ
Fig. 10
Fig. 10
Positive (green) and negative (red) peaks of the σA-weighted mFo-DFc difference electron density map around the ligand (ligand ID 675) and Wat526 in the binding pocket of the protein target uPA in the PDB structure 4FU9 refined with QM/MM before (a) and after (b) the manual fit. The σA-weighted 2mFo-DFc electron density map is contoured at 1 σ
Fig. 11
Fig. 11
Ligand Interaction diagram for the ligand ID 4CK in the PDB structure 4FKG after the QM/MM and conventional PHENIX refinements. Arrows added to underscore significant structural and interaction changes
Fig. 12
Fig. 12
The σA-weighted mFo-DFc difference electron density map peaks drawn at 3σ level around the ligand (ligand ID 239) in the PDB structure 4FUC refined with QM/MM for the default (a) and XModeScore best tautomers (b). The σA-weighted 2mFo-DFc electron density map is contoured at 1 σ
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