Figure 3.

Biophysical characterization of DDX3X protein and aggregation propensity induced by pathogenic mutations

(A) Circular dichroism (CD).

(B) Differential scanning fluorimetry (DSF) curves for D1D2 domain of WT-DDX3X and mutants during thermal denaturation. Tm values: WT - 51.6 ± 0.1°C, L556S monomer – 40.6 ± 0.1°C, L556S dimer – 44.5 ± 0.1°C, R376C – 40.1 ± 0.2°C.

(C) ANS fluorescence at 25°C.

(D) Rayleigh light scattering (RLS) at 350 nm of WT and mutants during incubation at 37°C.

(E) ThT fluorescence of protein samples before and after incubation at 37°C. ∗p < 0.05 when compared with WT 0 hr ∗∗p < 0.005 when compared with WT 24 hr.

(F) ANS fluorescence of proteins during incubation at 37°C.

(G) CD of L556S mutant at 10°C after incubation at 37°C during 24 hr.

(H) CD of R376C mutant before and after incubation at 37°C.

(I) Negative staining image of L556S-DDX3X after 3-hr-incubation at 37°C.

(J) Negative staining image of R376C-DDX3X after 3-hr-incubation at 37°C. Scale bar = 100 nm