Probing a model of a GPCR/ligand complex in an explicit membrane environment. The human cholecystokinin-1 receptor

¹ Université Henri Poincaré
² INSERM

^* To whom correspondence should be addressed. E-mail: christophe.chipot{at}edam.uhp-nancy.fr.

Submitted on July 12, 2005
Revised on August 17, 2005
Accepted on 24 October 2005

	Abstract

A three-dimensional model structure of a complex formed by a G protein-coupled receptor (GPCR) and an agonist ligand is probed and refined using molecular dynamics (MD) simulations and free energy calculations in a realistic environment. The model of the human receptor of cholecystokinin (CCK) associated to agonist ligand CCK9 was obtained from a synergistic procedure combining site-directed mutagenesis experiments and in silico modeling [Archer--Lahlou et al., J. Med. Chem. 2005, 48, 180-191]. The 31 ns MD simulation in an explicit membrane environment indicates that both the structure of the receptor and its interactions with the ligand are robust. Whereas the secondary structure of the -helix bundle is well preserved, the region of the intracellular loops exhibits a significant flexibility likely to be ascribed to the absence of G protein subunits in the model. New insight into the structural features of the binding pocket is gained, in particular, the interplay of the ligand with both the receptor and internal water molecules. Water-mediated interactions are shown to participate in the binding, hence, suggesting additional site-directed mutagenesis experiments. Accurate free energy calculations on mutated ligands provide differences in the receptor-ligand binding affinity, thus offering a direct, quantitative comparison to experiment. We propose that this detailed consistency-checking procedure be used as a routine refinement step of in vacuo GPCR models, prior to further investigation and application to structure-based drug design.

Key Words: Free energy calculations, G protein-coupled receptor, Membrane, Molecular dynamics

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