Figure 2. Impact of ZBTB7A mutations on DNA binding.

(a) Model for the C-terminal zinc-finger domain of ZBTB7A comprising residues 382–488. The model is depicted as yellow ribbon with highlighted secondary structure. Zinc ions are shown as grey spheres. DNA is shown in brown with a grey molecular surface. R402 (purple) binds into the major groove and likely contributes to the affinity or sequence specificity of the DNA interaction of the zinc-finger domain. (b) Biotinylated oligonucleotides containing the ZBTB7A (alias: Pokemon) consensus binding motif (POK WT) or a mutant thereof (POK mut)¹⁴ used in DNA pull-down experiments. Spheres illustrate streptavidin-coated beads. (c) DNA pull-down using protein lysates from HEK293T cells expressing wild-type or mutant ZBTB7A. Western blot analysis shows that A175fs and R402H fail to bind oligonutides with a ZBTB7A-binding site (POK WT). Oligonucleotides with a mutated binding site (POK mut) were used as negative control. Input lanes were loaded with 10% of the protein lysate used for each binding reaction.