Abstract
Ionotropic glutamate receptors (iGluRs) are responsible for fast synaptic transmission throughout the nervous system. Conformational changes of the transmembrane domain (TMD) underlying ion channel activation and desensitization remain poorly understood. Here, we explored the dynamics of the TMD of AMPA-type iGluRs using genetically-encoded unnatural amino acid (UAA) photo-crosslinkers, p-benzoyl-L-phenylalanine (BzF) and p-azido-L-phenylalanine (AzF). We introduced UAAs at sites throughout the TMD of the GluA2 receptor and characterized these mutants in patch-clamp recordings, exposing them to glutamate and UV light. This approach revealed a range of optical effects on the activity of mutant receptors. We found evidence that an interaction between the Pre-M1 and the M4 TMD helix was essential for normal activation and desensitization. Photoactivation at F579AzF, a residue behind the selectivity filter, had extraordinarily broad effects on gating and desensitization. This observation suggests coupling to other parts of the receptor and like in other tetrameric channels, selectivity filter gating.