Research ArticleBIOCHEMISTRY

Stereochemistry and amyloid inhibition: Asymmetric triplex metallohelices enantioselectively bind to Aβ peptide

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Science Advances  19 Jan 2018:
Vol. 4, no. 1, eaao6718
DOI: 10.1126/sciadv.aao6718
  • Fig. 1 The structures of metallohelices.

    (A) Symmetric metallohelices based on rigid ditopic bidentate ligands AB-BA yield D3-symmetric enantiomers [Fe2(AB-BA)3]. (B) The directional ligands AB-CD lead to C1-symmetric HHH and HHT “triplex” architectures. (C and D) Metallohelices self-assembled from various components of a range of functionalized helices with HHT structure. Me, methyl; napth, naphthyl; Bn, benzyl.

  • Fig. 2 The inhibition ability of metallohelices on Aβ.

    (A) The Aβ-ECFP fusion system used in screening Aβ aggregation inhibitors. (B and C) Screening results of metallohelices A and B from the Aβ-ECFP fusion screen system. (D) Aggregation kinetics of Aβ40 monitored by ThT assay with or without the incubation of A1 and B4. (E) The stopped-flow kinetic study of Aβ and A1. Values show means ± SD, and independent experiments were performed three times.

  • Fig. 3 AFM images of Aβ40/Aβ42 with or without the incubation of metallohelices (area corresponding to 3 μm × 3 μm).

    Control: 50 μM Aβ40/Aβ42 alone. ΛA1, ΔA1, ΛB4, and ΔB4: 50 μM Aβ40/Aβ42 with the incubation of 50 μM ΛA1, ΔA1, ΛB4, and ΔB4.

  • Fig. 4 The influence of metallohelices on the structure of Aβ40 and the binding affinity of metallohelices to Aβ40.

    (A and B) CD spectra of Aβ40 without or with the incubation of A1 (A) and B4 (B). (C and D) CD spectra of dialyzate were used to discriminate the enrichment of the enantiomer with lower binding affinity to Aβ40 in competition dialysis experiments. The concentrations of metallohelices were 50 μM.

  • Fig. 5 1H NMR spectra of Aβ40 with or without treatment of A1.

    (A) The signals caused by amide protons of K16, F19, F20, and E22 of Aβ40. (B) Locally amplified 1H NMR spectra of peak a. (C) Locally amplified 1H NMR spectra of peak b. (D) The peaks caused by the protons of F19 and F20 (black box) were obviously reduced in intensity with treatment of A1. The chemical shift values of Aβ40 were assigned on the basis of previous studies (4043).

  • Fig. 6 The interaction of Aβ40 and B4 by docking study.

    (A and B) Energy-minimized average models of ΛB4 (A) and ΔB4 (B) with Aβ40 interactions. (C and D) Hydrophobic interaction between Aβ40 and ΛB4 (C) and between Aβ40 and ΔB4 (D). (E) Locally amplified image of the black box in (C). (F) Locally amplified image of the black box in (D).

  • Fig. 7 The effect of A1 and B4 on the life span and Aβ plaques of the transgenic strain CL2006.

    (A and B) Kaplan-Meier survival curves of the transgenic strain CL2006 treated with or without metallohelices. Plots are representative of three independent experiments. (C to H) Representative images of Aβ plaques in the worm’s head region. Arrows indicate Aβ deposits. (C) Transgenic worm CL2006. Transgenic worm CL2006 with the incubation of ΛA1 (D) and ΛB4 (E). (F) Bristol N2 worm (wild type). Transgenic worm CL2006 with the incubation of ΔA1 (G) and ΔB4 (H).

Supplementary Materials

  • Supplementary material for this article is available at http://advances.sciencemag.org/cgi/content/full/4/1/eaao6718/DC1

    fig. S1. Influence of these metal complexes on the fluorescence of ECFP (a non-Aβ fusion system).

    fig. S2. The influence of these metallohelices on the fluorescence of ThT.

    fig. S3. Aggregation kinetics of Aβ42 monitored by ThT assay in the absence or presence of A1 and B4.

    fig. S4. Aggregation kinetics of Aβ40 monitored by ThT assay in the absence or presence of the ligands of A1 and B4.

    fig. S5. The inhibition effect of A1 and B4 on Aβ40/Aβ42 fibrillogenesis at different concentrations.

    fig. S6. The inhibition effect of the metallohelices on Aβ40 aggregation measured by SDS-PAGE.

    fig. S7. The influence of A1 and B4 on the second structures of Aβ42 monitored by CD.

    fig. S8. Fluorescence titration of Aβ40 (3 μM) with various concentrations of metallohelices in 20 mM tris buffer.

    fig. S9. ITC data for the Aβ40 titrations with metallohelices.

    fig. S10. SDS-PAGE analysis of the effect of metallohelices on tryptic digests of Aβ12–28.

    fig. S11. The aggregation kinetics of Aβ25–35 was monitored by the fluorescence of ThT in the absence or presence of A1 and B4.

    fig. S12. FTIR spectra of Aβ40 in different conditions.

    fig. S13. Structures of Aβ40 and metallohelices used for docking study.

    fig. S14. Energy-minimized average models of ΔA1 and ΔA1 with Aβ40 interactions.

    fig. S15. A1 and B4 scavenging ROS monitored by NBT and ABTS methods.

    fig. S16. Cyclic voltammograms corresponding to the O2/O2 redox couple.

    fig. S17. Effect of the metallohelices on ROS production in PC12 cells.

    fig. S18. Absorption spectra of 5 μM metallohelices in water and PBS.

    fig. S19. Effect of A1 and B4 on PC12 cell viability determined by MTT.

    fig. S20. Protection effects of metallohelices on Aβ40- and Aβ42-induced cytotoxicity of PC12 cells.

    table S1. IC50 values of metallohelices A1 and B4 for the inhibition of fibril formation and destabilization of the preformed fibrils.

    table S2. Analysis of fluorescence titration and ITC data.

    table S3. Enthalpy (ΔH), entropy (ΔS), and Gibbs free energy (ΔG) of the binding of Aβ with metallohelices at pH 7.3.

  • Supplementary Materials

    This PDF file includes:

    • fig. S1. Influence of these metal complexes on the fluorescence of ECFP (a non-Aβ fusion system).
    • fig. S2. The influence of these metallohelices on the fluorescence of ThT.
    • fig. S3. Aggregation kinetics of Aβ42 monitored by ThT assay in the absence or presence of A1 and B4.
    • fig. S4. Aggregation kinetics of Aβ40 monitored by ThT assay in the absence or presence of the ligands of A1 and B4.
    • fig. S5. The inhibition effect of A1 and B4 on Aβ40/Aβ42 fibrillogenesis at different concentrations.
    • fig. S6. The inhibition effect of the metallohelices on Aβ40 aggregation measured by SDS-PAGE.
    • fig. S7. The influence of A1 and B4 on the second structures of Aβ42 monitored by CD.
    • fig. S8. Fluorescence titration of Aβ40 (3 μM) with various concentrations of metallohelices in 20 mM tris buffer.
    • fig. S9. ITC data for the Aβ40 titrations with metallohelices.
    • fig. S10. SDS-PAGE analysis of the effect of metallohelices on tryptic digests of Aβ12–28.
    • fig. S11. The aggregation kinetics of Aβ25–35 was monitored by the fluorescence of ThT in the absence or presence of A1 and B4.
    • fig. S12. FTIR spectra of Aβ40 in different conditions.
    • fig. S13. Structures of Aβ40 and metallohelices used for docking study.
    • fig. S14. Energy-minimized average models of A1 with Aβ40 interactions.
    • fig. S15. A1 and B4 scavenging ROS monitored by NBT and ABTS methods.
    • fig. S16. Cyclic voltammograms corresponding to the O2/O2 redox couple.
    • fig. S17. Effect of the metallohelices on ROS production in PC12 cells.
    • fig. S18. Absorption spectra of 5 μM metallohelices in water and PBS.
    • fig. S19. Effect of A1 and B4 on PC12 cell viability determined by MTT.
    • fig. S20. Protection effects of metallohelices on Aβ40- and Aβ42-induced cytotoxicity of PC12 cells.
    • table S1. IC50 values of metallohelices A1 and B4 for the inhibition of fibril formation and destabilization of the preformed fibrils.
    • table S2. Analysis of fluorescence titration and ITC data.
    • table S3. Enthalpy (ΔH), entropy (ΔS), and Gibbs free energy (ΔG) of the binding of Aβ with metallohelices at pH 7.3.

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