ARTICLE pubs.acs.org/JPCB Quantifying Water-Mediated Protein in a Glutamate Receptor: A DFT Study Michelle A. Sahai and Philip C. Biggin* Ligand Interactions Structural Bioinformatics and Computational Biochemistry, University of Oxford, South Parks Road, Oxford OX1 3QU, United Kingdom bS Supporting Information ABSTRACT: It is becoming increasingly clear that careful treat- ment of water molecules in ligand protein interactions is required in many cases if the correct binding pose is to be identiﬁed in molecular docking. Water can form complex bridging networks and can play a critical role in dictating the binding mode of ligands. A particularly striking example of this can be found in the ionotropic glutamate receptors. Despite possessing similar chemical moieties, crystal structures of glutamate and R-amino-3-hydroxy-5-methyl-4- isoxazole-propionic acid (AMPA) in complex with the ligand- binding core of the GluA2 ionotropic glutamate receptor revealed, contrary to all expectation, two distinct modes of binding. The diﬀerence appears to be related to the position of water molecules within the binding pocket. However, it is unclear exactly what governs the preference for water molecules to occupy a particular site in any one binding mode. In this work we use density functional theory (DFT) calculations to investigate the interaction energies and polarization eﬀects of the various components of the binding pocket. Our results show (i) the energetics of a key water molecule are more favorable for the site found in the glutamate-bound mode compared to the alternative site observed in the AMPA-bound mode, (ii) polarization eﬀects are important for glutamate but less so for AMPA, (iii) ligand system interaction energies alone can predict the correct binding mode for glutamate, but for AMPA alternative modes of binding have similar interaction energies, and (iv) the internal energy is a signiﬁcant factor for AMPA but not for glutamate. We discuss the results within the broader context of rational drug-design. 1. INTRODUCTION Water constitutes the cellular environment where biomolecules interact. Speciﬁcally it plays a crucial role in mediating interactions between ligands and macromolecular receptors. Water is also important for protein folding and stability, and it is also known to actively participate in many catalytic processes in the cell. As such its importance has been increasingly recognized through protein protein and protein ligand interaction studies with respect to rational drug design.1 However, despite the wealth of structural information, the precise role of water molecules in mediating receptor ligand interactions remains unclear.2 In the context of drug design, one key problem is knowing whether a certain water molecule will be readily displaceable by functional groups designed to replace it. This issue has been the focus of recent computational studies.3 5 While pure molecular dynamics studies focused on identifying and characterizing the interactions between the protein, ligand, and conserved water molecules,6 free energy3,7 calculations were used to further elucidate the functions of these water molecules and probe their apparent stabilizing role in the protein binding site. A measure of the free energy is ultimately what is required in order to assess the stability of water molecules within binding sites. However, obtaining suﬃci- ent accuracy of these calculations, especially for charged systems (which encompasses most biological systems),8,9 is particularly challenging and an area of intense investigation.10 One factor in these calculations is the underlying simplicity of the force-ﬁeld which may or may not account for interactions at the necessary level required. Even with these simpliﬁed models, the calculations are still time-consuming enough to be prohibitive at the level of virtual screening. Nevertheless they do form the underlying physical basis of many scoring functions employed in virtual high-throughput screening methods, and consequently, it is of great interest to improve them as much as possible. Many binding pockets have a complex network of water molecules and in some instances this has already been exploited to give increased drug potency, for example, in the case of HIV-1 protease where cyclic urea derivatives were designed to displace a conserved water molecule.11 Related to the problem of displace- able water molecules is the concept that water molecules can be considered as part of the drug rather than part of the protein. This can lead to the situation where similar compounds that might be expected to adopt the same binding pose on the basis of their chemical structure, actually adopt diﬀerent poses within the binding pocket. A good example of this can be found in structures of the ionotropic glutamate receptors which are an important set of potential drug targets within the central nervous system. Received: Revised: Published: January 25, 2011 April 16, 2011 May 05, 2011 r 2011 American Chemical Society 7085 dx.doi.org/10.1021/jp200776t | J. Phys. Chem. B 2011, 115, 7085–7096