The Proton Responsiveness in the Extracellular Domain of GLIC Differs in the Presence of the ELIC Transmembrane Domain.

Prokaryotic homologues of Cys-loop receptors have proven to be useful in understanding their eukaryotic counterparts, but even the best studied of these, Gloeobacter ligand-gated ion channel (GLIC), is still not yet fully understood. GLIC is activated by protons with a pH50 between 5 and 6, implicating a histidine residue in its activation, but although a histidine residue (His11') in the pore-forming α-helix (M2) is known to be involved in gating, the His in the extracellular domain (ECD), His127, is not. Nevertheless, there is evidence from a GLIC-glycine chimera for a proton sensitive residue or region in the GLIC extracellular domain. Here we create a novel chimeric receptor with the ECD of GLIC and the transmembrane domain of ELIC (GELIC). Expression of this receptor in oocytes reveals proton activation, although the pH50 (6.7) differs from that of GLIC (5.4). Exploration of protonatable residues in the ECD reveals that the pKa s of five Asp residues (31, 49, 91, 136, and 178) differ between the open and closed states of GLIC. Substitution of these residues with Ala or Asn shows somewhat similar effects for GLIC and GELIC in Asp91 mutants, but different effects for the others. Overall, the data suggest that protonation of residues in the ECD is a requirement for channel opening in GELIC but plays only a minor role in GLIC, where gating may be largely driven via protonation of the His residue in its pore.