Exploration of LPS₂ agonist binding modes using the combination of a new hydrophobic scaffold and homology modeling

Lysophosphatidylserine (LysoPS) is an endogenous pan-agonist of three G-protein coupled receptors (GPCRs): LPS₁/GPR34, LPS₂/P2Y₁₀, and LPS₃/GPR174, and we previously reported a series of LysoPS-based agonists of these receptors. Interestingly, we found that LPS₁ agonist activity was very sensitive to structural change at the hydrophobic fatty acid moiety, whereas LPS₂ agonist activity was not. Here, to probe the molecular basis of LPS₂ agonist binding, we developed a new class of hydrophobic fatty acid surrogates having a biphenyl-ether scaffold. The LPS₂ agonist activity of these compounds proved sensitive to molecular modification of the hydrophobic skeleton. Thus, we next constructed an LPS₂ model by homology modeling and docking/molecular dynamics (MD) simulation, and validated it by means of SAR studies together with point mutations of selected receptor amino-acid residues. The putative ligand-binding site of LPS₂ is Γ-shaped, with a hydrophilic site horizontally embedded in the receptor transmembrane helix bundles and a perpendicular hydrophobic groove adjoining transmembrane domains 4 and 5 that is open to the membrane bilayer. The binding poses of LPS₂ agonists to this site are consistent with easy incorporation of various kinds of fatty acid surrogates. Structural development based on this model afforded a series of potent and selective LPS₂ full agonists, which showed enhanced in vitro actin stress fiber formation effect.