Shen_2012_J.Clin.Invest_122_2613

Congenital myasthenic syndromes (CMSs) are neuromuscular disorders that can be caused by defects in ace-tylcholine receptor (AChR) function. Disease-associated point mutants can reveal the unsuspected functional significance of mutated residues. We identified two pathogenic mutations in the extracellular domain of the AChR alpha subunit (AChRalpha) in a patient with myasthenic symptoms since birth: a V188M mutation in the C-loop and a heteroallelic G74C mutation in the main immunogenic region. The G74C mutation markedly reduced surface AChR expression in cultured cells, whereas the V188M mutant was expressed robustly but had severely impaired kinetics. Single-channel patch-clamp analysis indicated that V188M markedly decreased the apparent AChR channel opening rate and gating efficiency. Mutant cycle analysis of energetic coupling among conserved residues within or dispersed around the AChRalpha C-loop revealed that V188 is functionally linked to Y190 in the C-loop and to D200 in beta-strand 10, which connects to the M1 transmembrane domain. Furthermore, V188M weakens inter-residue coupling of K145 in beta-strand 7 with Y190 and with D200. Cumulatively, these results indicate that V188 of AChRalpha is part of an interdependent tetrad that contributes to rearrangement of the C-loop during the initial coupling of agonist binding to channel gating.