In the bound systems, the number of direct inter-subunit hydrogen bonds obtained by summing the average number for the five subunits (see Figure in the Supporting Information) slightly increases as the number of bound ligand decreases from five (42.6±1.8) to three (43.7±2.0) to two (44.7±2.1). However, the more ligands are bound, the more additional interactions mediated by the ligands are present, as previously discussed, strongly enhancing the adhesion of adjacent subunits. The Apo structure shows a substantially reduced number of inter-subunit hydrogen bonds (32.4±2.4) with respect to the ligated models, consistently with a less compact structure, with subunits in the ECD undergoing an outward tilt from the channel axis when transitioning from the open state to the Apo state. Some interactions involving specific residues observed in the ligated models are still detectable in the zero-ligand model, although with lower occurrences. In the zero-ligand model, some interface residues often formed interactions with multiple residues on the adjacent subunit, instead of a single residue with high occurrence. Selected hydrogen bonds between conserved residue pairs across bound pockets in the bound models are shown in Figure .