Complementation of a CHO
43-3B extract with ERCC1 protein. (A) Extract (60 µg
protein) derived from the CHO 43-3B ERCC1 mutant cell line was incubated
with plasmid DNA molecules containing a single cisplatin lesion
(lane 1) and various amounts of recombinant ERCC1 and/or
recombinant XPF (lanes 2–6). Reactions in lanes 2–5
received 300, 262.5, 150 and 37.5 ng of ERCC1, respectively. Reactions
in lanes 3–6 received 37.5, 150, 262.5 and 300 ng of XPF, respectively.
Lane 7 is a control reaction with 18.5 ng of the ERCC1–XPF
complex produced in E.coli with a dicistronic vector
encoding both subunits. Excised oligonucleotides are indicated by
the bracket on the left. (B) The activity of Δ88ERCC1 was compared to the activity
of full-length ERCC1. Reactions in lanes 2–4 received 38,
150 and 300 ng of rERCC1, respectively, and in lanes 5–7
they received 38, 150 and 300 ng of Δ88ERCC1,
respectively. (C) Quantification of the relative
complementation levels reached with ERCC1 (dotted line) and Δ88ERCC1 (solid line).