Fig. 1: Structures of human, mouse, and rat P2X7Rs in the apo closed state conformation reveal distinct allosteric ligand-binding sites.

A Ribbon representation of the apo closed state structures of human (shades of blue and gray), mouse (shades of purple and gray), and rat (shades of green and gray, PDB code: 8TR5) P2X7Rs, aligned with ChimeraX³⁵. The classical allosteric ligand-binding site is boxed in red. Two CHS molecules per protomer (inner and outer; both tan), located at the interface between TM1 and TM2 on the extracellular side of the membrane, are found only in the hP2X7R. GDP (tan) and Zn²⁺ ions (slate gray) are labeled and shown within the cytoplasmic ballast (Supplementary Fig. 2 A). B Magnified and 120° rotated view of A, highlighting cryo-EM density for two molecules of CHS bound to one protomer of the hP2X7R. C Same view as B, highlighting the hydrophobic and hydrogen bonding interactions between the inner CHS molecule and the hP2X7R. D 50° rotated view from C highlighting the hydrophobic and hydrogen bonding interactions between the outer CHS molecule and the hP2X7R. E-G Magnified view of the classical allosteric ligand-binding site from A, highlighting the differences between human (E, light blue and gray), mouse (F, light purple and gray), and rat (G, light green and gray) P2X7R orthologs³⁵. Residues 95, 108, and 312 (red labels) are the key residue differences in the classical allosteric pockets between the three orthologs. The larger V312 in hP2X7Rs (A312 in mP2X7Rs and rP2X7Rs) forces the neighboring residue Y295 (orange labels) to adopt an alternative rotameric conformation that condenses the classical allosteric pocket in the human ortholog only (Supplementary Fig. 7).