Fig. 7. Fucosylation of EGF repeats from wild-type and Fringe. (A) Proteins containing EGF repeats. Protein EGF13-15 includes EGF repeats 13-15 from wild type N, flanked by V5 and 6His tags. There is a potential O-fucosylation site in EGF repeat 13 (T540; blue T). Protein EGF13-15 I578T carries the Ile578 to Thr578 substitution (red T) from the spl allele. There are potential O-fucosylation sites in EGF repeat 13 and 14. Protein EGF13-15T540I resembles protein EGF13-15 with Thr540 substituted by Ile. This protein has no potential O-fucosylation site. Protein EGF13-15T540I,I578T is same as protein EGF13-15I578T, except that Thr540 is substituted into Ile. Ile578 is the only potential fucosylation site. (B) Autoradiographs of radiolabeled proteins after membrane transfer. EGF13-15T540I protein is not labeled (lane 3), unlike the other proteins. (C) Immunoblotting with anti-His₆ control for equal protein loading. Red asterisk indicates mutation.