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. Author manuscript; available in PMC: 2020 Nov 17.
Published in final edited form as: Biochemistry. 2020 Jul 21;59(30):2796–2812. doi: 10.1021/acs.biochem.0c00514

Figure 5.

Figure 5.

Split-ubiquitin protein interaction assays with human transcription factors ATF3, CREB1, and REST vs human UBR1 and UBR2 E3 ubiquitin ligases. (A) Design of split-Ub assays44,98,99,112 (see Materials and Methods). (B) Row 1, hsATF3 vs full-length hsUBR1. Row 2, hsATF3 vs the Nt-fragment of hsUBR1 (hsUBR11–1059). Row 3, hsATF3 vs the Ct-fragment of hsUBR1 (hsUBR11060–1749). Row 4, hsATF3 vs the vector alone. (C) Row 1, hsATF3 vs full-length hsUBR2. Row 2, hsATF3 vs the Nt-fragment of hsUBR2 (hsUBR21–1071). Row 3, hsATF3 vs the Ct-fragment of hsUBR2 (hsUBR11072–1755). Row 4, hsATF3 vs the vector alone. (D) Row 1, hsCREB1 vs full-length hsUBR1. Row 2, hsCREB1 vs the Nt-fragment of hsUBR1 (hsUBR11–1059). Row 3, hsCREB1 vs the Ct-fragment of hsUBR1 (hsUBR11060–1749). Row 4, hsCREB1 vs the vector alone. Row 5, hsATF3 vs full-length hsUBR1 (the same as in row 1 of panel A but an independent split-Ub assay). Row 6, hsATF3 vs the Nt-fragment of hsUBR1 (hsUBR11–1059) (the same as in row 2 of panel B but an independent split-Ub assay). (E) Row 1, hsREST vs full-length hsUBR1. Row 2, hsREST vs the Nt-fragment of hsUBR1 (hsUBR11–1059). Row 3, hsREST vs the Ct-fragment of hsUBR1 (hsUBR11060–1749). Row 4, hsREST vs the vector alone. Row 5, vectors alone. Row 6, hsATF3 vs the Nt-fragment of hsUBR1 (hsUBR11–1059) (the same as in row 2 of panel B but an independent split-Ub assay). Row 7, hsATF3 vs full-length hsUBR1 (the same as in row 1 of panel A but an independent split-Ub assay).