Research Paper Volume 15, Issue 16 pp 8061—8089
Coumarin-chalcone hybrid LM-021 and indole derivative NC009-1 targeting inflammation and oxidative stress to protect BE(2)-M17 cells against α-synuclein toxicity
Figure 2. SNCA-His protein and aggregation. (A) SNCA-His protein. α-Synuclein (140 amino acids, marked in black) was in-frame fused to a 6-amino acid His tag (marked in red) via a 2 amino-acid linker (Leu and Glu, marked in blue). (B) SNCA-His aggregates formation. SNCA-His (20 μM in 300 μl buffer) was incubated at 37° C for 3 days and aggregates were examined by 10% SDS-PAGE gel and immunoblot probing with the α-synuclein antibody. (C) Cryo-TEM structure of aggregated SNCA-His. (D) Thioflavin T binding assay for SNCA-His aggregation. SNCA-His protein (20 μM) was incubated with trehalose, LM-021 or NC009-1 (1−10 μM) at 37° C for three days, and aggregation was analyzed by measuring thioflavin T fluorescence intensity (n = 3). P values: day 0 vs. day 3 (###: P < 0.001), or with vs. without compound treatment (***: P < 0.001).