Cholesterol impacts chemokine CCR5 receptor ligand‐binding activity

The chemokine CCR5 receptor is target of maraviroc, a negative allosteric modulator of CCR5 that blocks the HIV protein gp120 from associating with the receptor, thereby inhibiting virus cellular entry. As noted with other G‐protein‐coupled receptor family members, the role of the lipid environment in CCR5 signaling remains obscure and very modestly investigated. Controversial literature on the impact of cholesterol (Chol) depletion in HIV infection and CCR5 signaling, including the hypothesis that Chol depletion could inhibit HIV infection, lead us to focus on the understanding of Chol impact in the first stages of receptor activation. To address this aim, the approach chosen was to employ reconstituted model lipid systems of controlled lipid composition containing CCR5 from two distinct expression systems: Pichia pastoris and cell‐free expression. The characterization of receptor/ligand interaction in terms of total binding or competition binding assays was independently performed by plasmon waveguide resonance and fluorescence anisotropy, respectively. Maraviroc, a potent receptor antagonist, was the ligand investigated. Additionally, coarse‐grained molecular dynamics simulation was employed to investigate Chol impact in the receptor‐conformational flexibility and dynamics. Results obtained with receptor produced by different expression systems and using different biophysical approaches clearly demonstrate a considerable impact of Chol in the binding affinity of maraviroc to the receptor and receptor‐conformational dynamics. Chol considerably decreases maraviroc binding affinity to the CCR5 receptor. The mechanisms by which this effect occurs seem to involve the adoption of distinct receptor‐conformational states with restrained structural dynamics and helical motions in the presence of Chol. The chemokine CCR5 receptor is a target of maraviroc, a negative allosteric modulator of CCR5 that blocks the HIV protein gp120 from associating with the receptor, thereby inhibiting virus cellular entry. Cholesterol (Chol) is suggested to impact CCR5‐mediated responses. Here, we investigate the impact of Chol in ligand recognition by maraviroc in cell membrane fragments and in reconstituted lipid model systems using biophysical approaches. The presence of Chol in the membrane decreases ligand affinity for CCR5 and restrains its dynamics.