Ligand-dependent cholesterol interactions with the human A(2A) adenosine receptor

Area of Science:

• Biochemistry
• Molecular Biology
• Computational Chemistry

Background:

• G-protein coupled receptors (GPCRs) are crucial drug targets.
• The human A2A adenosine receptor (A2AAR) is a key GPCR involved in various physiological processes.
• Cholesterol interactions with GPCRs, particularly at the cholesterol consensus motif (CCM), are implicated in receptor function and stability.

Purpose of the Study:

• To investigate ligand-specific cholesterol binding to the A2AAR CCM using all-atom simulations.
• To elucidate the role of Trp129 dynamics in mediating A2AAR-cholesterol interactions.
• To examine the effect of thermostabilizing mutations on A2AAR structure and cholesterol binding.

Main Methods:

• All-atom molecular dynamics simulations of the A2AAR bound to four different ligands.
• Analysis of molecular trajectories to assess cholesterol binding dynamics and protein conformational changes.
• Investigation of hydrogen bonding networks and rotameric states of key residues, such as Trp129.

Main Results:

• Evidence of ligand-specific cholesterol interactions with the A2AAR CCM, involving two cholesterol molecules.
• Binding of the synthetic agonist UK432097 disrupts Trp129-Ser47 hydrogen bonds, activating Trp129 rotameric dynamics and displacing one cholesterol molecule.
• Thermostabilizing mutations did not support the proposed mechanism of helix stabilization via cholesterol interaction; instead, a force-field dependent helix straightening was observed.

Conclusions:

• Ligand binding modulates cholesterol interactions at the A2AAR CCM through specific residue dynamics (Trp129).
• The observed effects challenge existing hypotheses regarding the role of cholesterol in thermostabilization of A2AAR via the CCM.
• Simulation data suggest a force-field dependency in helix conformational stability, impacting the interpretation of thermostabilizing mutations.