Asymmetry and Ion Selectivity Properties of Bacterial Channel NaK Mutants Derived from Ionotropic Glutamate Receptors

Ionotropic glutamate receptors are ligand-gated cation channels that play essential roles in the excitatory synaptic transmission throughout the central nervous system. A number of open-pore structures of a-a mino-3-hydroxy-5-methyl-4-isoxazolepropionic-acid (AMPA)-type glutamate receptors became available recently by cryo-electron microscopy (cryo-EM). These structures provide valuable insights into the con-formation of the selectivity filter (SF), the part of the ion channel that determines the ion selectivity. Nonetheless, due to the moderate resolution of the cryo-EM structures, detailed information such as ion occupancy of monovalent and divalent cations as well as positioning of the side-chains in the SF is still missing. Here, in an attempt to obtain high-resolution information about glutamate receptor SFs, we incorporated partial SF sequences of the AMPA and kainate receptors into the bacterial tetrameric cation channel NaK, which served as a structural scaffold. We determined a series of X-ray structures of NaK-CDI, NaK-SDI and NaK-SELM mutants at 1.42-2.10 angstrom resolution, showing distinct ion occupation of different monovalent cations. Molecular dynamics (MD) simulations of NaK-CDI indicated the channel to be conductive to monovalent cations, which agrees well with our electrophysiology recordings. More-over, previously unobserved structural asymmetry of the SF was revealed by the X-ray structures and MD simulations, implying its importance in ion non-selectivity of tetrameric cation channels. (c) 2023 Elsevier Ltd. All rights reserved.

Automated summary

Ionotropic glutamate receptors are ligand-gated cation channels crucial for excitatory synaptic transmission in the central nervous system. Recent cryo-EM studies have revealed open-pore structures of AMPA-type receptors, providing insights into ion selectivity. However, detailed information regarding ion occupancy and SF side-chain positioning is still lacking. Here, we incorporated partial SF sequences into the bacterial cation channel NaK and obtained high-resolution X-ray structures, revealing distinct ion occupation and structural asymmetry.