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. 2017 Jun 12;36(14):2047–2060. doi: 10.15252/embj.201796664

Figure EV1. Purification and validation of nuclease activity of human recombinant XPF‐ERCC1 (XE).

Figure EV1

  1. Purified XPF‐ERCC1 (XE) analysed on an SDS–PAGE gel (4–12%) stained with InstantBlue (left‐hand panel) and Western blot analysis (right‐hand panel). WT = wild type XE; D676A = XE mutated to substitute aspartic acid residue 676 of XPF with alanine.
  2. Nuclease activity of WT and D676A forms of XE on a “simple fork” substrate. The substitution of metal‐binding residue in XPF (D676A) renders the XE complex devoid of any nuclease activity. Red circles denote 3′[32P]‐radiolabelled nucleotides.
  3. (Top panel) Nuclease activity of XE on a “simple fork” substrate over a time course. (Bottom panel) Quantification of intact substrate and incision products expressed as a percentage of initial substrate as in top panel, n = 2.