Energy Maps of Acetylcholine Receptor Gating

Abstract

Occupancy of an AChR binding site by an agonist (A) shifts a pre-existing equilibrium from the Closed-channel towards the Open-channel conformation. We estimated energy changes in adult mouse A2C↔A2O gating (adult mouse) from single-channels. Mutations mainly cause local energy changes that can be mapped onto structures of related proteins. We made maps showing residue reaction progress (phi values) and approximate extents of energy changes in the channel-opening isomerization. Phi values range from 1 to 0, are approximately constant for each position and have a modal distribution (n=5 populations). Residues having similar phi values are clustered, with phi decreasing approximately longitudinally between the binding site and the gate. Two separated regions in the alpha subunits - the transmitter binding sites and M2-M3 linkers in the membrane domain - have the highest phi values (∼0.95), followed by the rest of the extracellular domain (∼0.8), most of the membrane domain (∼0.6) and the gate region of the pore (0.3). Residues at the extracellular-transmembrane domain interface undergo large energy changes in C→O. Largest energy changes occur at the binding sites, and in the alpha subunit at the M2-M3 linker, M1 and the gate. All of these energy changes, including those associated with the affinity change for the agonist, appear to arise from local ‘resettling’ events that do not transfer significant energy over distance. Expansion and flexure of the M2-M3 linkers in the alpha subunit appears to trigger the global allosteric transition, and the hydrophobic gate appears to unlock in three steps. The local character of side chain energy changes and the similar phi values of the binding sites and distant M2-M3 linkers suggest that the gating transition is not strictly a mechanical process.