Structure of the pore-farming transmembrane domain of a ligand-gated ion channel

The structure of the pore-forming transmembrane domain of the nicotinic acetylcholine receptor from Torpedo has been investigated by infrared spectroscopy. Treatment of affinity-purified receptor with either Pronase or proteinase K digests the extramembranous domains (roughly 75% of the protein mass), leaving hydrophobic membrane-imbedded peptides 3-6 kDa in size that are resistant to peptide H-1/H-2 exchange, Infrared spectra of the transmembrane domain preparations exhibit relatively sharp and symmetric amide I and amide II band contours centered near 1655 and 1545 cm(-1) respectively, in both (H2O)-H-1 and (H2O)-H-2. The amide I band is very similar to the amide I bands observed in the spectra of alpha -helical proteins, such as myoglobin and bacteriorhodopsin, that lack P structure and exhibit much less p-sheet character than is observed in proteins with as little as 20% P sheet. Curve-fitting estimates 75-80% cy-helical character, with the remaining peptides likely adopting extended and/or turn structures at the bilayer surface. Infrared dichroism spectra are consistent with transmembrane alpha -helices oriented perpendicular to the bilayer surface. The evidence strongly suggests that the transmembrane domain of the nicotinic receptor, the most intensively studied ligand-gated ion channel, is composed of five bundles of four transmembrane alpha-helices.