A stepwise mechanism for acetylcholine receptor channel gating

Muscle contraction is triggered by the opening of acetylcholine receptors at the vertebrate nerve-muscle synapse. The M2 helix of this allosteric membrane protein lines the channel, and contains a 'gate' that regulates the flow of ions through the pore. We used single-molecule kinetic analysis to probe the transition state of the gating conformational change and estimate the relative timing of M2 motions in the α-subunit of the murine acetylcholine receptor. This analysis produces a 'Φ-value' for a given residue that reflects its open-like versus closed-like character at the transition state. Here we show that most of the residues throughout the length of M2 have a Φ-value of ∼0.64 but that some near the middle have lower Φ-values of 0.52 or 0.31, suggesting that αM2 moves in three discrete steps. The core of the channel serves both as a gate that regulates ion flow and as a hub that directs the propagation of the gating isomerization through the membrane domain of the acetylcholine receptor.