Successful structure-based drug discovery/design is dependent upon accurate protein:ligand structure determination and characterization. Gaining an understanding of the protein:ligand complex structure along with the proper protonation and explicit solvent effects is crucial for obtaining meaningful results from docking, thermodynamic calculations, active site exploration, and ultimately lead optimization. Our work has lead to a sea change in our ability to accurately refine protein:ligand structure through the tight integration of the DivCon, linear-scaling, semiempirical, quantum mechanics (SE-QM) engine with refinement methods. A significant and often neglected factor in SBDD is the protonation and tautomerism of the protein:ligand complex. Even with modern, atomistic structural biology methods, experimental protonation state determination remains elusive. It is generally extremely difficult to experimentally determine the tautomeric state of the ligand and of the surrounding active site - even at higher resolutions. Theoretically, protonation can be established using neutron diffraction; however, experimental requirements such as reliance on very large crystals and on deuterium exchange limit the method's suitability in SBDD. Recently, we found that SE-QM based macromolecular refinement is highly sensitive to the choice of a protonation state/tautomer form of ligands and residues. With this method, we are able determine the correct protonation states based on routine data even at lower resolutions commonly found in pharmaceutical research and development campaigns. The method and results as well as the effects of tautomerization and disorder in drug design will be discussed in detail. Finally, we will discuss the impact of improved protonation state determination on binding affinity prediction and lead optimization efforts.

The use of crystallographic and Cryo-EM data in accurate protein: Ligand protonation state determination and its impact on structure based drug discovery

ACS Fall 2021

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