Supporting Information. Design of LVFFARK and LVFFARK-Functionalized Nanoparticles. for Inhibiting Amyloid β-protein Fibrillation and Cytotoxicity

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1 Supporting Information Design of LVFFARK and LVFFARK-Functionalized Nanoparticles for Inhibiting Amyloid β-protein Fibrillation and Cytotoxicity Neng Xiong, Xiao-Yan Dong, Jie Zheng, Fu-Feng Liu, * and Yan Sun, * Department of Biochemical Engineering and Key Laboratory of Systems Bioengineering of the Ministry of Education, School of Chemical Engineering and Technology, Tianjin University, Tianjin , China Department of Chemical and Biomolecular Engineering, The University of Akron, Akron, Ohio 44325, United States Supporting information in this document: Supporting figures S1-S12 1

2 Figure S1. The relative ThT fluorescence intensity of Aβ42 aggregates in different LVFFA concentrations after 72 h coincubation. Aβ42 concentration was 40 μm. The fluorescence of Aβ42 after incubation for 72 h was set to 100%. 2

3 Figure S2. A TEM image of the morphologies of Aβ42 aggregates coincubated with 200 μm LK7 after 24 h incubation. Aβ42 concentration was 40 μm. 3

4 Figure S3. The morphologies of LK7 self-aggregates observed by TEM. The concentrations of LK7 were (a) 40 μm and (b) 200 μm. 4

5 Figure S4. The relative ThT fluorescence intensity provided by LK7 aggregates after 72 h incubation. The fluorescence of Aβ42 after incubation for 72 h was set to 100%. 5

6 Figure S5 Inhibitory effect of LK7 on Aβ42-induced cytotoxicity towards PC12 cell line using (a) MTT assay and (b) LDH leakage assay. Aβ concentration was 40 μm. In the MTT assay, the cell viability treated with PBS buffer only was set to 100%. In the LDH leakage assay, the cells were incubated with 1% (v/v) Triton X-100 in FBS-free medium at 37 C for 1 h to obtain a representative maximal LDH release as the positive control with 100% cytotoxicity. The control (cell viability treated with PBS buffer only) was set to 100%. p<0.001, compared to the control group. # p<0.05, ## p<0.01, ### p<0.001, compared to the Aβ42-treated group (LK7:Aβ=0). 6

7 Figure S6. Cytotoxicity of PC12 induced by the self-assembly of LK7 using (a) MTT assay and (b) LDH leakage assay. In the MTT assay, the cell viability treated with PBS buffer only was set to 100%. In the LDH leakage assay, the cells were incubated with 1% (v/v) Triton X-100 in FBS-free medium at 37 C for 1 h to obtain a representative maximal LDH release as the positive control with 100% cytotoxicity. The control (cell viability treated with PBS buffer only) was set to 100%. p<0.05, p<0.001, compared to the control group. 7

8 Figure S7. Synthesis process for LK7 was anchored to PLGA-NPs via the conventional EDC-NHS reaction between carboxyl groups on the surface of PLGA-NPs and ε- amino group of Lysine residues in LK7, which is indicated in red. 8

9 Figure S8. An SEM image of PLGA-NPs. 9

10 Figure S9. Size distributions of Aβ42 aggregates in the presence of (a) PLGA-NPs and (b) determined at 0, 6, and 24 h by DLS. The concentrations of both PLGA-NPs and LK7@PLGA-NPs were all 100 µg/ml. Aβ42 concentration was 40 μm. 10

11 Figure S10. Far-UV circular dichroism spectra of 100 μg/ml PLGA-NPs and incubated for 0 and 24 h. 11

12 Figure S11. Cytotoxicity of PC12 induced by (a and b) PLGA-NPs and (c and d) NPs. The cell viability were estimated by both (a and c) MTT and (b and d) LDH leakage assays. In the MTT assay, the cell viability treated with PBS buffer alone was set to 100%. In the LDH leakage assay, the cells were incubated with 1% (v/v) Triton X-100 in FBS-free medium at 37 C for 1 h to obtain a representative maximal LDH release as the positive control with 100% cytotoxicity. The control (cell viability treated with PBS buffer alone) was set to 100%. p<0.05, p<0.001, compared to the control group. 12

13 Figure S12. Inhibitory effect of (a and b) PLGA-NPs and (c and d) on Aβ42- induced cytotoxicity toward PC12 cells. Aβ42 concentration was 40 μm. The cell viability was estimated by both (a and c) MTT and (b and d) LDH assay. In the MTT assay, the cell viability treated with PBS buffer only was set to 100%. In the LDH leakage assay, the cells were incubated with 1% (v/v) Triton X-100 in FBS-free medium at 37 C for 1 h to obtain a representative maximal LDH release as the positive control with 100% cytotoxicity. The control (cell viability treated with PBS buffer alone) was set to 100%. p<0.001, compared to control groups. # p<0.05, ## p<0.01, ### p<0.01, compared to Aβ42-treated group. 13