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. 2017 Nov 20;5(1):16.
doi: 10.1007/s40203-017-0037-x. eCollection 2017.

Impact of protein-ligand solvation and desolvation on transition state thermodynamic properties of adenosine A2A ligand binding kinetics

Giuseppe Deganutti  1 , Andrei Zhukov  2 , Francesca Deflorian  2 , Stephanie Federico  3 , Giampiero Spalluto  3 , Robert M Cooke  2 , Stefano Moro  1 , Jonathan S Mason  2 , Andrea Bortolato  2
Affiliations

Impact of protein-ligand solvation and desolvation on transition state thermodynamic properties of adenosine A2A ligand binding kinetics

Giuseppe Deganutti et al. In Silico Pharmacol. .

Abstract

Ligand-protein binding kinetic rates are growing in importance as parameters to consider in drug discovery and lead optimization. In this study we analysed using surface plasmon resonance (SPR) the transition state (TS) properties of a set of six adenosine A2A receptor inhibitors, belonging to both the xanthine and the triazolo-triazine scaffolds. SPR highlighted interesting differences among the ligands in the enthalpic and entropic components of the TS energy barriers for the binding and unbinding events. To better understand at a molecular level these differences, we developed suMetaD, a novel molecular dynamics (MD)-based approach combining supervised MD and metadynamics. This method allows simulation of the ligand unbinding and binding events. It also provides the system conformation corresponding to the highest energy barrier the ligand is required to overcome to reach the final state. For the six ligands evaluated in this study their TS thermodynamic properties were linked in particular to the role of water molecules in solvating/desolvating the pocket and the small molecules. suMetaD identified kinetic bottleneck conformations near the bound state position or in the vestibule area. In the first case the barrier is mainly enthalpic, requiring the breaking of strong interactions with the protein. In the vestibule TS location the kinetic bottleneck is instead mainly of entropic nature, linked to the solvent behaviour.

Keywords: Biacore; Ligand binding kinetics; Metadynamics; Molecular dynamics; SPR; Supervised molecular dynamics.

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Figures

Fig. 1
Fig. 1
Chemical structures of the A2A AR ligands considered for the suMetaD test. ZM241385, Z80 and Z48 are [1,2,4]triazolo[1,5-a][1,3,5]triazine inhibitors; DPCPX, KW3902 and XAC are xanthine inhibitors
Fig. 2
Fig. 2
Eyring equation plots for the ligand binding association (left) and dissociation (right) from the SPR analysis for the 6 ligands included in this study. The values for ΔH‡ and ΔS‡ can be determined from kinetic data plotting ln(t/T) vs 1/T. In the resulting linear interpolation equation the slope corresponds to ΔH‡/R and the ΔS‡ can be calculated from the y-intercept
Fig. 3
Fig. 3
Schematic overview of the two alternative transition state locations detected by the suMetaD protocol for the ligand binding event. The ligand is represented by a blue circle, waters by smaller red circles, the pocket by a blue line divided by a black dotted line in the orthosteric site (bottom half) and the vestibule region (top half). Starting from the unbound state (left) with strong protein–water interactions and ordered waters the ligand reaches the bound state (right). Two alternative ligand locations corresponding to the transition state have been detected, the upper transition state showing the energy barrier is near the vestibule region and the lower one where it is deep in the orthosteric site. They are characterized by opposite enthalpic and entropic components related to the desolvation of the binding site
Fig. 4
Fig. 4
Schematic overview of the two alternative transition state locations detected by the suMetaD protocol for the ligand unbinding event. As in Fig. 3, the ligand is represented by a blue circle, waters by smaller red circles, the pocket by a blue line divided by a black dotted line in the orthosteric site (bottom half) and the vestibule region (top half). Starting from the bound state (left) with strong protein–ligand interactions and ordered waters the ligand reaches the unbound state (right). Two alternative ligand locations corresponding to the transition state have been detected: on the top, the energy barrier is near the vestibule region; on the bottom is deep in the orthosteric site. They are characterized by opposite enthalpic and entropic components related to the solvation of the binding site and of the small molecule
Fig. 5
Fig. 5
Protein–ligand locations/conformations corresponding to the binding kinetic bottlenecks detected by the suMetaD protocol for the 6 small molecules considered in this study. The ligand is shown in stick representation, the pocket as a mesh surface and waters as small spheres. Interactions among waters in the orthosteric site are shown as yellow dotted lines. The experimental energy of the enthalpic (ΔH‡) and entropic (−TΔS‡) components of the TS is reported
Fig. 6
Fig. 6
Protein–ligand positions/conformations corresponding to the unbinding kinetic bottlenecks detected by the suMetaD protocol for the 6 small molecules considered in this study. The ligand is shown in stick representation, the pocket as solid grey surface and waters as small spheres. Interactions among waters in the vestibule region near the ligand and among the extracellular loops are shown as yellow dotted lines. The experimental energy of the enthalpic (ΔH‡) and entropic (−TΔS‡) components of the TS is reported
Fig. 7
Fig. 7
Role of the solvent in the transition state entropic energy barrier. The experimental binding (blue) and unbinding (red) TS entropic barrier for the 6 ligands considered in this study is plotted on the X-axis. The Y-axis shows for the binding event (blue) the corresponding number of temporary trapped waters in the orthosteric site in the representative TS conformation. For the unbinding event (red), the Y-axis includes the number of waters in the extracellular side of the receptor at less than 4 Å from the ligand aliphatic carbon atoms in the representative TS conformation
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