Structural Study Of Human Glutamate Receptors Glud Family

Human GluD1 and GluD2 proteins belong to the GluD subfamily of ionotropic glutamate receptors,which are widely distributed in the brain and play an important role in the development and maintenance of the nervous system.Previous studies reported that several human psychiatric diseases are related to mutations in the GRID1 and GRID2 genes.Since the identification of the two genes in 1997,there have been a lot of neurobiology researches focusing on them.However,due to the low protein expression and difficulties in crystallization,both the full-length protein structure and potential conformational changes of GluD1 and GluD2 are still unknown,which hinders our further understanding of its molecular mechanism and biological functions in synapses.In this study,in order to resolve the high-resolution structures of the full-length GluD family proteins and provide structural information for their potential conformational changes,we constructed the mammalian expression system and performed a series of optimization of the experimental procedure,and successfully obtained GluD1 in different small molecules bound states and GluD2 proteins in the apo state.Besides,we determined the affinity between GluD1/GluD2 and their potential ligands by Microscale thermophoresis(MST)experiment.After cryo-EM data collection and data processing,we finally resolved the structures of GluD2 protein in desensitized state as well as GluD1 protein in D-serine bound state,L-glutamate bound state and pentamidine bound state at overall resolutions of 5.2 ?,6 ?,8 ?,and 9 ?,respectively.Through structural analysis,we observed that the desensitized state GluD2 structure exhibits a F-shaped conformation whereas all the three ligand-bound state structures of GluD1 exhibit a Y-shaped conformation.By comparing with the previous reported crystal structures of GluD2 domains and performing homology analysis among different GluD homologs,we identified the conformational differences in the desensitized state of GluD2 and proposed a potential model of conformational changes in the GluD2 signaling.These findings not only advanced our understanding of the activation of GluD family proteins,but also shed light on the molecular mechanism of GluD2 signaling.Moreover,our findings about GluD family proteins will provide new insights into the treatment of various GluD-related human neurodevelopmental and psychiatric diseases and related drug discovery in the future.