Fig. 7: Stability Comparisons for Fc-Notch3 constructs with and without NTF-binding.
From: Oligomerization, trans-reduction, and instability of mutant NOTCH3 in inherited vascular dementia
a Demonstrates a collision-induced unfolding fingerprint of Fc-NOTCH3 WT protein bound to NTF. In all CIU fingerprints of Fc-constructs, four main features were observed. We selected the 22.5 ms feature for stability comparison (a, outlined in purple). NTF addition resulted in NTF-NOTCH3 binding identifiable by CIU (a). Dissociation of the NTF-NOTCH3 complex is tracked by the purple line in (a) and revealed increased amounts of unbound NOTCH3 protein at the feature traveling at a drift time of 22.5 ms (outlined in purple). Thus, we selected this final transition for comparison between WT and mutant proteins (a, pink box). CIU fingerprints for all proteins are displayed in Fig S7. Comparison of the Fc-Notch3 construct stability shifts (a, pink box) are displayed in (b). WT (n = 2) and R141C proteins (n = 2) required the highest onset voltages for the 22.5 ms feature (b). R90C (n = 4), C49Y (n = 4), and R75P mutants (n = 4) have a lower voltage requirement for the transition (b, orange asterisks). The addition of NTF resulted in a decrease in the onset voltage for the 22.5 ms feature in all constructs (b).
