Time dependent fibrillation of WT and 112-syn.

<p>Time dependent fibrillation of WT-syn (<b>A</b>) and 112-syn (<b>B</b>) was monitored by the enhancement of ThT fluorescence. WT and 112-syn (0.5 mg/mL) at 37°C were incubated in 20 mM Tris buffer, pH 7.5 for indicated time intervals as described in the “<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0098657#s2" target="_blank">Methods</a>” section and the ThT fluorescence emission spectrum was recorded. (<b>C</b>) Fold change in the ThT fluorescence of WT and 112-syn incubated for different time intervals. Data presented are the mean ± SD of three separate experiments. (<b>D</b>) Aliquots from reaction mixtures incubated at 37°C for 24 h were adsorbed onto carbon coated grids and then stained with 1% uranyl acetate for 1–2 min. Negatively stained TEM images of WT and 112-syn are shown. <i>Scale bars</i>, 500 nm and data is a representative of three different fields of view. *p<0.01 as compared to 0 h readings for 112-syn.</p

Effect of WT-syn on time dependent fibrillation of 112-syn.

<p>(<b>A</b>) 112-syn (0.5 mg/mL) was incubated in the presence or absence of different ratios of WT-syn at 37°C in 20 mM Tris buffer, pH 7.5 for 12 h. The increase in ThT fluorescence emission spectrum (470–600 nm) was recorded following excitation at 450 nm. (<b>B</b>) Fold change in the ThT fluorescence of 112-syn, incubated for 12 h in the presence or absence of different ratios of WT-syn. Data presented are the mean ± SD of three separate experiments. *p<0.01 as compared to 0 h reading for 112-syn; #p<0.01 as compared to112-syn alone.</p

Effect of WT-syn on heat induced aggregation of 112-syn.

<p>(<b>A</b>) The chaperone-like activity of WT-syn was monitored by incubating 112-syn (0.4 mg/mL) in the presence or absence of different ratios of WT-syn as described in the “<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0098657#s2" target="_blank">Methods</a>” section. The time dependent increase in absorbance at 360 nm was recorded over a period of 10 min. (<b>B</b>) Fold change in the net absorbance values from (A) before and after 10 min of incubation at 65°C recorded at 360 nm.*p<0.01 as compared to 112-syn before heating.</p

Effect of WT and 112-syn on heat induced aggregation of aldolase (ALD).

<p>(<b>A</b>) <i>In vitro</i> protein aggregation assay employing aldolase (0.1 mg/mL in PBS) was performed at 65°C in the absence or presence of WT and 112-syn at different ratios (1∶2 and 1∶4) and the light scattering was monitored at 360 nm for a period of 10 min. (<b>B</b>) The net change in the absorbance of reaction mixtures of (A) before and after 10 min of incubation at 65°C. Data are the mean ± SD of three separate experiments. * indicates p<0.01 as compared to aldolase (decrease); #p<0.01 as compared to aldolase (increase) and $ p<0.01 as compared to the respective controls (before heating).</p

Temperature dependent aggregation of 112-syn.

<p>(<b>A</b>) 112-syn at different concentrations (0.2, 0.4, 0.6 mg/mL in PBS) was heated at 45°C, 55°C, and 65°C and the light scattering was monitored at 360 nm for a period of 10 min as described under “<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0098657#s2" target="_blank">Methods</a>” section. (<b>B</b>) Fold change in the net absorbance values from (A) at 360 nm before and after heating the reaction mixtures. (<b>C</b>) ThT fluorescence of reaction mixtures as indicated in (A) before (0 min) and after exposing the samples at indicated temperatures for 10 min.*p<0.01 as compared to 0 min readings for both (B) and (C). (<b>D</b>) Negatively stained TEM images of WT and 112-syn before and after exposure to 65°C for 10 min.</p