Acid-sensing ion channels (ASICs) are neuronal sodium-selective channels activated by reductions in extracellular pH. Structures of the three presumptive functional states, high-pH resting, low-pH desensitized, and toxin-stabilized open, have all been solved for chicken ASIC1. These structures, along with prior functional data, suggest that the isomerization or flipping of the β11–12 linker in the extracellular, ligand-binding domain is an integral component of the desensitization process. To test this, we combined fast perfusion electrophysiology, molecular dynamics simulations and state-dependent non-canonical amino acid cross-linking. We find that both desensitization and recovery can be accelerated by orders of magnitude by mutating resides in this linker or the surrounding region. Furthermore, desensitization can be suppressed by trapping the linker in the resting state, indicating that isomerization of the β11–12 linker is not merely a consequence of, but a necessity for the desensitization process in ASICs.

1 Acid-sensing ion channels (ASICs) are neuronal sodium-selective channels activated by reductions 2 in extracellular pH. Structures of the three presumptive functional states, high-pH resting, low-3 pH desensitized, and toxin-stabilized open, have all been solved for chicken ASIC1. These 4 structures, along with prior functional data, suggest that the isomerization or flipping of the β11-5 12 linker in the extracellular, ligand-binding domain is an integral component of the 6 desensitization process.To test this, we combined fast perfusion electrophysiology, molecular 7 dynamics simulations and state-dependent non-canonical amino acid cross-linking. We find that 8 both desensitization and recovery can be accelerated by orders of magnitude by mutating resides 9 in this linker or the surrounding region. Furthermore, desensitization can be suppressed by 10 trapping the linker in the resting state, indicating that isomerization of the β11-12 linker is not 11 merely a consequence of, but a necessity for the desensitization process in ASICs. 12

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