Energetic Changes in the β Subunit during Gating of the Muscle Nicotinic Receptor

Abstract

The transmitter-binding sites in the adult muscle-type nicotinic receptor are located at the interfaces between the α subunit and the δ or ε subunits, while the β subunit does not contribute to binding. The β subunit is termed a "structural" subunit as a result. We made mutations to 27 residues in the mouse muscle β subunit (2 to 4 mutations at each location) to measure the consequences on the channel opening and closing rates. We analyzed the results in terms of the effect on the ratio of the opening to closing rates (θ) and, when sufficiently large changes occurred, on the slope of the logarithmic relation between the opening rate and θ (φ). φ can be interpreted in terms of the timing during channel activation when a given residue makes an energetic contribution to gating. We examined 12 positions in the extracellular domain (ECD) of the β subunit, chosen because mutations to homologous residues in other subunits had large effects on θ. In contrast, mutations at the locations in β produced small changes in gating. We examined 15 positions in the second transmembrane (TM2) and TM2-TM3 linker regions. The majority of locations showed larger changes in θ(typically > 1 kcal/mole). Estimates of φ were consistent with the idea that the energetic contributions from residues in the β subunit occurred later than for homologous residues in the α subunit but before residues in δ. Overall, the data suggest that the ECD and the interface between the ECD and TM domains of β make little contribution to the energetics of gating. However, channel-lining residues do contribute, although the data suggest that the timing is later than for the α subunit. (NS22356 & NS72770)