Structure of the LGI1–ADAM22ECD complex

a Domain organizations of LGI1 and ADAM22. LGI1 consists of the LRR (blue) and EPTP (purple) domains. The N-terminal secretion signal peptide (SP, enclosed by dotted lines) is removed in the secreted LGI1. The shaded blue boxes represent the N-and C-terminal caps, whereas the filled blue boxes represent the LRRs. The premature form of ADAM22 contains the N-terminal prosequence (enclosed by dotted lines). The mature ADAM22 consists of the metalloprotease-like (magenta), disintegrin (salmon pink), cysteine-rich (brown), EGF-like (orange), transmembrane (white), and cytoplasmic domains. The major ADAM22 isoform has a PDZ-binding motif in the C-terminal region of the cytoplasmic domain b Cryo-EM map and structure of the LGI1LRR–LGI1*EPTP–ADAM22ECD complex at 2.78 Å resolution. c Cryo-EM map and structure of the 3:3 LGI1–ADAM22ECD complex at 3.79 Å resolution. d Schematic diagram of the 3:3 LGI1–ADAM22ECD complex. Black circles indicate the LGI1LRR–LGI1*EPTP– ADAM22ECD complex.

Interactions between LGI1EPTP and ADAM22ECD

a Superposition of the cryo-EM structure of the present LGI1LRR–LGI1*EPTP–ADAM22ECD complex and the previously reported crystal structure of the LGI1EPTP–ADAM22ECD complex. b, c Overall view of the interface between LGI1EPTP and ADAM22ECD. The structure (b) and corresponding map (c) are shown. d, e Close-up view of the interaction between Arg330 of LGI1 and Asp405 of ADAM22. The structure (d) and corresponding map (e) are shown. f, g Close-up view of the interaction around Arg378 of LGI1. The structure (f) and corresponding map (g) are shown.

Structure and interaction of the higher-order LGI1–ADAM22ECD complex

a, b Overall view of the interface between LGI1LRR and LGI1*EPTP. The structure (a) and corresponding map (b) are shown. c Cryo-EM structure of the 3:3 LGI1–ADAM22ECD complex. c Superposition of the three LGI1-ADAM22ECD complexes in the cryo-EM structure of the 3:3 LGI1-ADAM22ECD complex. d Chain IDs of the individual LGI1 or ADAM22ECD molecules in the 3:3 LGI1-ADAM22ECD complex, assigned in this study.

HS-AFM observations of LGI1–ADAM22ECD complexes

a A representative HS-AFM image of the 3:3 LGI1–ADAM22ECD complex. The color bars on the right indicate height in nanometers. The frame rate was 1.0 frames/s. b, d Magnified HS-AFM images of the 3:3 (left in b) and 2:2 (left in d) LGI1–ADAM22ECD complexes. The simulated AFM images (middle) were derived from fitting to the experimental HS-AFM image (left). The well-fitting simulated AFM images and the coefficient of correlation (CC) are indicated. The tertiary structures of the 3:3 (b) and 2:2 (d) LGI1–ADAM22ECD complexes are shown in the same orientation as the simulated AFM images. c, e, f Sequential HS-AFM images of the 3:3 (c; see also Movie S1) and 2:2 (e, f; see also Movie S2) LGI1–ADAM22ECD complexes. A schematic illustration of the interpretation of HS-AFM images is shown at the bottom. Imaging parameters: scanning area = 120 × 96 nm2 (240 × 192 pixels); frame rate = 3.3 frames/s. HS-AFM experiments were repeated independently at least 3 times with consistent results.