a, Schematic representation of the optimized GluD1 construct showing the C-terminal truncation at residue 851 and C-terminal thrombin cleavage site, along with GFP and octa-histidine tag (Supplementary Notes). b, Size-exclusion profile of the final purified protein showing receptor stability in optimized buffer conditions. c, Selected 2D class averages from reference-free 2D classification of GluD1 in complex with 1 mM 7-CKA and 1 mM Ca2+. The white arrows mark a few classes that show conformational heterogeneity of the extracellular receptor domains. d, 3D classification of GluD1 into seven classes reveals heterogeneity due to the movement of the two extracellular arms. More details are in Extended Data Fig. 2 and Supplementary Fig. 1.