Figure 5

Nucleotide substitution analysis of Hlx1-B6 binding to PRDM9^Cst. (A) Competition assay for testing the binding of PRDM9^Cst to mutated Hlx1 binding sites. Substitutions of nucleotides 4 to 6; labeled oligos are indicated by asterisk. The compositions of reaction mixtures in the first three lanes are shown above each lane. Lanes 4 to 12 show the competition test with mutated oligos. The position and the nature of each mutation in the oligo are shown. The composition of the reaction mixtures in lanes 4 to 12 is mutated oligo + PRDM9^Cst + oligo* (see Materials and methods). The fraction of the shifted band is indicated below each lane. Red arrow, shifted band; black arrow, unbound fragment. (B) Graphic representation of the binding changes at each position produced by the nucleotide substitution analysis. (C) ZF domain of PRDM9^Cst. The letters in the boxes show the amino acids at positions -1, 3, and 6 relative to the α-helix of each finger.