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. Author manuscript; available in PMC: 2010 Mar 4.
Published in final edited form as: Gastroenterology. 2008 Apr 4;135(1):152–162. doi: 10.1053/j.gastro.2008.03.074

Figure 5. Mutation of DNA-binding motifs within APC impede DNA binding and inhibition of replication.

Figure 5

A. Schematic diagram of S(T)PXX binding motifs within APC 4-2 and the sites altered by mutations. Mutation sites are indicated with stars.

B. Coomassie blue staining and western blotting of purified mutant APC segments. Purified proteins were separated by 10% SDS-PAGE, stained with Coomassie bright blue (upper panel) and probed with a monoclonal anti-His antibody (lower panel).

C. Gel mobility shifts of radioactively labeled oligonucleotides in the presence of wild-type or mutant APC4-2. A radiolabelled 68-base single-stranded oligonucleotide (2 nM) was incubated with the corresponding proteins, separated by 8% PAGE and analyzed by phosphorimager. The protein concentrations from low to high are: 0, 18.75, 37.5, 75, 150, 200, 250, and 300 nM, respectively.

D. Agarose gel electrophoresis of PCR products prepared in the presence of wild-type or mutant APC4-2 at different quantitative ratio of [protein]/[DNA (bp)] as Figure 3A. PCR products were separated using 1% agarose gel electrophoresis and visualized by ethidium bromide staining.