Figure 3. Effect of acetate ions & acridine derivatives on in vitro aggregation of TDP-43^2C.

(a) TDP-43^2C (400 μM) solubilized in aggregation buffer was added with increasing concentrations of sodium acetate (1:1 to 1:30, protein: compound, molar ratio) that was also pre-dissolved in the aggregation buffer but lacking the Tris-HCl & DMSO. Aggregation trend was monitored by ThT fluorescence at 37 °C for 15 hours with intermittent agitation (15 seconds agitation every 5 minutes). Increasing stoichiometry of sodium acetate caused concomitant marginal increase in the overall ThT fluorescence. (b) Aggregation trend of TDP-43^2C in presence of Acr and AIM was monitored by ThT fluorescence. TDP-43^2C (400 μM) was added with 125 μM Acr or 125 μM AIM (protein: compound ratio of 1: 0.3) in aggregation buffer lacking Tris-HCl and containing 8% DMSO final. The protein aggregation was examined at 37 °C for 15 hours with intermittent agitation by recording ThT fluorescence intensity. A sample lacking any compound was incubated similarly for comparison. (c) TDP-43^2C aggregation in presence of Acr-E was monitored by recording ThT fluorescence intensity. TDP-43^2C (400 μM) was added with 2000 μM Acr-E or in the control experiment with 2000 μM AIM4, in aggregation buffer containing 12 mM Tris-HCl and 8% DMSO final. Samples were incubated at 37 °C for 15 hours with intermittent agitation and ThT fluorescence was recorded. A sample lacking any compound was incubated & monitored similarly for comparison. Acr-E and AIM4 were found to respectively reduce the ThT fluorescence by 45% and 55%.