Modelling and Molecular Dynamics Predict the Structure and Interactions of the Glycine Receptor Intracellular Domain

Glycine receptors (GlyRs) are glycine-gated inhibitory pentameric ligand-gated ion channels composed of alpha or alpha + beta subunits. A number of structures of these proteins have been reported, but to date, these have only revealed details of the extracellular and transmembrane domains, with the intracellular domain (ICD) remaining uncharacterised due to its high flexibility. The ICD is a region that can modulate function in addition to being critical for receptor localisation and clustering via proteins such as gephyrin. Here, we use modelling and molecular dynamics (MD) to reveal details of the ICDs of both homomeric and heteromeric GlyR. At their N and C ends, both the alpha and beta subunit ICDs have short helices, which are major sites of stabilising interactions; there is a large flexible loop between them capable of forming transient secondary structures. The alpha subunit can affect the beta subunit ICD structure, which is more flexible in a 4 alpha 2:1 beta than in a 4 alpha 1:1 beta GlyR. We also explore the effects of gephyrin binding by creating GlyR models bound to the gephyrin E domain; MD simulations suggest these are more stable than the unbound forms, and again there are alpha subunit-dependent differences, despite the fact the gephyrin binds to the beta subunit. The bound models also suggest that gephyrin causes compaction of the ICD. Overall, the data expand our knowledge of this important receptor protein and in particular clarify features of the underexplored ICD.