SorLA is a neuronal sorting receptor considered to be a major risk factor for Alzheimer's disease. We have recently reported that it directs lysosomal targeting of nascent neurotoxic amyloid-β (Aβ) peptides by directly binding Aβ. Here, we determined the crystal structure of the human sorLA domain responsible for Aβ capture, Vps10p, in an unbound state and in complex with two ligands. Vps10p assumes a ten-bladed β-propeller fold with a large tunnel at the center. An internal ligand derived from the sorLA propeptide bound inside the tunnel to extend the β-sheet of one of the propeller blades. The structure of the sorLA Vps10p–Aβ complex revealed that the same site is used. Peptides are recognized by sorLA Vps10p in redundant modes without strict dependence on a particular amino acid sequence, thus suggesting a broad specificity toward peptides with a propensity for β-sheet formation.
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We would like to thank the staff of the beamlines at Photon Factory and SPring-8 for their help with X-ray data collection, K. Yamashita for help in the setup of MD simulations and for discussions on data analysis, S. Thompson for critical reading and editing of the manuscript, K. Tamura-Kawakami for excellent technical assistance and M. Sakai for preparation of the manuscript. This work was supported by the Grant-in-Aid for Scientific Research on Innovative Areas 'Analysis and Synthesis of Multidimensional Immune Organ Network' (no. 24111006 to J.T.) and 'Dynamic Ordering of Biomolecular Systems for Creation of Integrated Functions' (no. 25102008 to K.K.) from the Ministry of Education, Culture, Sports, Science and Technology of Japan (MEXT), by the 'Platform for Drug Discovery, Informatics, and Structural Life Science' grant from the MEXT (no. 12736015 to J.T.), by the 'X-ray Free Electron Laser Priority Strategy Program' grant from the MEXT (no. 12004060 to J.T.), by the Nanotechnology Platform Project (no. S-14-MS-1039 to J.T.) from the MEXT, by the Okazaki ORION project (to K.K.) and by the Research Funding for Longevity Sciences from the National Center for Geriatrics and Gerontology of Japan (no. 25-19 to K.K.).
The authors declare no competing financial interests.
Integrated supplementary information
Supplementary Figure 1 Quantitation of the binding of pro53 peptide to sorLA Vps10p measured by the AP reporter assay.
The graph shows tracings of the typical chromogenic AP reaction observed with AP-pro53 (orange) or control AP-MycHis (blue) eluted from the sorLA Vps10p-beads. Note that gradual increase in the absorbance with the AP-MycHis is indistinguishable from that with mock sample (where no AP activity is present in the reaction mixture), indicating that the background nonspecific binding of AP protein to the beads is negligible.
Amino acid sequences are from human sorLA (SORLA_hu, NP_003096.1), human sortilin (sort_hu, CCA66904), mouse SorCS1 (sorCS1_mo, Q9JLC4), and yeast Vps10p (Vps10p_sp, O42930). Secondary structure elements are denoted by straight (strands) or wavy (helices) lines below each sequence. Strand designation is shown above the alignment with the same color code as in Fig. 3. The L1 and L2 segments are highlighted in salmon and cyan, respectively. Cysteines are shown with a grey background with lines connecting the disulfide-bonded pair. The Gly511 mutated in a familial AD patient is marked by a red box. Propeptide cleavage sites are indicated by “//”.
The Vps10p propeller domain (gray) is shown in surface (left) or ribbon (right) presentations, with L1, L2, and the AD-causing mutation residue Gly511 colored in salmon, cyan, and red, respectively. Propeptide ligand bound inside the tunnel is shown in CPK model in both panels.
(a) Structural changes that accompany propeptide binding. The residue number is plotted against distance between mainchain Cα atoms in the ligand-free (at pH 4.5) and the propeptide-bound (at pH 6.5) forms of the sorLA Vps10p domain after structural superposition. Regions disordered in either structure are indicated by horizontal light blue bars. Note that the largest structural differences are found in L1 and the 10CC-b segments. (b) RMSF values for each residue during the MD simulation of the propeptide-bound form of sorLA Vps10p domain (see the legend to Figure 5d) are plotted similarly to the (a).
Supplementary Figure 5 Prediction of β-aggregation tendency of various sorLA Vps10p ligand peptides with the PASTA server.
The amino acid sequences of (a) the sorLA propeptide (53 residues), (b) Aβ40 (40 residues), (c) NT (13 residues), (d) HA (11 residues), and the sortilin propeptide (44 residues) were fed to the PASTA server (http://biocomp.bio.unipd.it/pasta/) and the resultant per-residue aggregation probability scores, h(k), are plotted.
Supplementary Figures 1–5 and Supplementary Tables 1–4 (PDF 2120 kb)
Uncropped gel images (PDF 259 kb)
The movie depicts the trajectory from a representative 10-ns MD simulation of sorLA Vps10p-propeptide complex. Structure is color-coded as in Fig. 4a, where L1, L2, and bound peptide is shown in magenta, cyan, and blue, respectively (MOV 1107 kb)
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Kitago, Y., Nagae, M., Nakata, Z. et al. Structural basis for amyloidogenic peptide recognition by sorLA. Nat Struct Mol Biol 22, 199–206 (2015). https://doi.org/10.1038/nsmb.2954
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