Journal article
Glycine Receptor Complex Analysis Using Immunoprecipitation‐Blue Native Gel Electrophoresis‐Mass Spectrometry
Proteomics, Vol.20(3-4), 1900403
2020
Abstract
The pentameric glycine receptor (GlyR), comprising the α1 and β subunits, is a major inhibitory ionotropic receptor in brainstem and spinal cord. GlyRs interact with gephyrin (GPHN), a scaffold protein that anchors the GlyR in the plasma membrane and enables it to form clusters in glycinergic postsynapses. Using an interaction proteomics approach, evidence of the ArfGEFs IQ motif and Sec7 domain 3 (IQSEC3) and IQ motif and Sec7 domain 2 (IQSEC2) as two novel synaptic proteins interacting with GlyR complexes is provided. When the affinity-isolated GlyR complexes are fractionated by blue native gel electrophoresis and characterized by mass spectrometry, GlyR α1β-GPHN appears as the most abundant complex with a molecular weight of ≈1 MDa, and GlyR α1β-GPHN-IQSEC3 as a minor protein complex of ≈1.2 MDa. A third GlyR α1β-GPHN-IQSEC2 complex exists at the lowest amount with a mass similar to the IQSEC3 containing complex. Using yeast two-hybrid it is demonstrated that IQSEC3 interacts with the GlyR complex by binding to the GPHN G domain at the N-terminal of the IQSEC3 IQ-like domain. The data provide direct evidence of the interaction of IQSEC3 with GlyR-GPHN complexes, underscoring a potential role of these ArfGEFs in the function of glycinergic synapses.
Details
- Title
- Glycine Receptor Complex Analysis Using Immunoprecipitation‐Blue Native Gel Electrophoresis‐Mass Spectrometry
- Authors
- Sophie J F van der Spek (Author) - Vrije Universiteit AmsterdamFrank Koopmans (Author) - Vrije Universiteit AmsterdamIryna Paliukhovich (Author) - Vrije Universiteit AmsterdamSarah L Ramsden (Author) - University College LondonKirsten Harvey (Author) - University College LondonRobert J Harvey (Author) - University of the Sunshine Coast - School of Health and Sport SciencesAugust B Smit (Author) - Vrije Universiteit AmsterdamKa Wan Li (Corresponding Author) - Vrije Universiteit Amsterdam
- Publication details
- Proteomics, Vol.20(3-4), 1900403
- Publisher
- Wiley - V C H Verlag GmbH & Co. KGaA
- Date published
- 2020
- DOI
- 10.1002/pmic.201900403
- ISSN
- 1615-9853
- Copyright note
- Copyright © 2020 The Authors. Proteomics published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim. This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.
- Organisation Unit
- School of Health; University of the Sunshine Coast, Queensland; School of Health and Sport Sciences - Legacy; Centre for Bioinnovation; School of Health and Behavioural Sciences - Legacy
- Language
- English
- Record Identifier
- 99450840302621
- Output Type
- Journal article
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