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. 2018 Sep 13;293(43):16697–16708. doi: 10.1074/jbc.RA118.003781

Figure 4.

Figure 4.

Ndfip proteins can activate Su(dx). A, domain organization of Drosophila Notch. B, sequence alignment of Drosophila Ndfip (dNdfip) and human Ndfip1 (hNdfip1). Three PY motifs are indicated with a red rectangular box. C, hNdfip1 can efficiently activate Su(dx). D, effects of dNdfip WT or PY mutants on Su(dx) autoubiquitination. E, GST-dNdfip can robustly pull down Su(dx) WW12L34, WW12L, and LWW34, whereas GST-Notch PY can only pull down WW12L34 and LWW34 but not WW12L. F, ITC-based measurement of the binding affinities between dNdfip and Su(dx) WW34 or WW12L34. G and H, MBP pulldown assay of MBP-tagged 12L34HECT WT and various mutants with an increasing amount of Trx-dNdfip. In this assay, the molar ratios of dNdfip and Su(dx) were 1:1, 2:1, and 5:1, and the quantification data of dNdfip pulled down by Su(dx) are shown at the bottom.