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. 2023 May 19;299(6):104838. doi: 10.1016/j.jbc.2023.104838

Figure 3.

Figure 3

Effect of amino acid substitutions at residue 49 of NOTCH3.A, schematic showing residue 49 of NOTCH3 within the first EGF repeat of the Fc-NOTCH3(1–3) construct. The C49Y mutant has been linked to CADASIL (Fig. 1). The WT residue is a cysteine residue (black triangle), which was mutated to all 19 non-cysteine amino acids (red circles), resulting in 19 independent mutant clones, which were transfected into cultured cells, whose conditioned medium was analyzed for Fc protein mobility. B, Western blots of Protein A-agarose–enriched Fc-NOTCH3(1–3) proteins separated under nonreducing conditions. Wildtype protein was compared with 19 different mutants labeled with the amino acid shown. Y represents the C49Y mutant. C, quantification of the ratio of upper band signal to total signal is shown and is the result of at least three independent experiments (∗p < 0.05 compared with wildtype control).