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Defining the structural relationship between kainate-receptor deactivation and desensitization

Abstract

Desensitization is an important mechanism curtailing the activity of ligand-gated ion channels (LGICs). Although the structural basis of desensitization is not fully resolved, it is thought to be governed by physicochemical properties of bound ligands. Here, we show the importance of an allosteric cation-binding pocket in controlling transitions between activated and desensitized states of rat kainate-type (KAR) ionotropic glutamate receptors (iGluRs). Tethering a positive charge to this pocket sustains KAR activation, preventing desensitization, whereas mutations that disrupt cation binding eliminate channel gating. These different outcomes explain the structural distinction between deactivation and desensitization. Deactivation occurs when the ligand unbinds before the cation, whereas desensitization proceeds if a ligand is bound without cation pocket occupancy. This sequence of events is absent from AMPA-type iGluRs; thus, cations are identified as gatekeepers of KAR gating, a role unique among even closely related LGICs.

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Figure 1: Kainate-receptor desensitization occurs with or without channel activation.
Figure 2: Mutation D776K eliminates GluK2 receptor desensitization.
Figure 3: Lys776 can act as a tethered ion at the GluK2 cation-binding pocket.
Figure 4: GluK2 D776K receptors gate in the absence of external ions.
Figure 5: Occupancy of the GluK2 cation-binding pocket is predicted to be disrupted by targeted mutation of the dimer interface.
Figure 6: Desensitization and deactivation are uncoupled in GluK2 KARs.
Figure 7: Desensitization does not substantially shift peak agonist potency of GluK2 KARs.

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Acknowledgements

This work was supported by operating grants from the Canadian Institutes of Health Research (CIHR) FRN-82804 (D.B.) and by the Leverhulme Trust RPG-059 (P.C.B.). G.B.D. and E.D.A. were supported by Natural Sciences and Engineering Research Council of Canada graduate fellowships, B.A.D. by a Chemical Biology CIHR postdoctoral award and M.R.P.A. by a CIHR Best & Banting doctoral award. D.B. is supported by a Canada Research Chair award. We thank the Oxford Supercomputing Centre for computer time. Finally, we wish to thank J. Johnson for comments on the manuscript and M. Fleck for discussions on AMPA receptors.

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G.B.D. designed and performed experiments, analyzed data and wrote the paper; M.M., B.A.D. and M.R.P.A. designed and performed experiments and analyzed data; E.D.A. analyzed data; P.C.B. designed experiments; and D.B. designed and performed experiments, analyzed data and wrote the paper.

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Correspondence to Philip C Biggin or Derek Bowie.

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Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–5 (PDF 953 kb)

Lys 776 replaces sodium at the cation binding pocket of GluK2 D776K.

MD simulation of the LBD dimer interface of the D776K receptor. (AVI 24165 kb)

Allosteric ions are unstably bound at the GluK2 Y521C L783C LBD dimer interface.

MD simulation of the LBD dimer interface of the Y521C L783C receptor. (AVI 19254 kb)

Mutation of the GluK2 cation binding pocket disrupts sodium binding.

MD simulation of the LBD dimer interface of the GluK2 E524G receptor. (AVI 14385 kb)

Allosteric ions form stable interactions at the LBD dimer interface of wildtype GluK2.

MD simulation of the LBD dimer interface of the wildtype GluK2 receptor. (AVI 18728 kb)

Mutation of the GluK2 LBD dimer interface disrupts sodium binding.

MD simulation of the LBD dimer interface of the GluK2 L783C receptor. (AVI 23966 kb)

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Dawe, G., Musgaard, M., Andrews, E. et al. Defining the structural relationship between kainate-receptor deactivation and desensitization. Nat Struct Mol Biol 20, 1054–1061 (2013). https://doi.org/10.1038/nsmb.2654

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