A single amino acid determines the specificity for the target sequence of two zinc-finger proteins in plants

The EPF family is a group of DNA-binding proteins with two canonical Cys2/His2 zinc-finger motifs in Petunia. These proteins are unique in terms of structure in that (i) the two zinc fingers are separated by spacers of various lengths and (ii) the sequence QALGGH is strongly conserved in the zinc-finger motifs of members of the family. In this study, domain-swapping and site-directed mutagenesis experiments with two members of the protein family, EPF2-5 and EPF2-7, which have different target sequences, revealed that only a single amino acid in the second zinc finger is responsible for the difference in target specificity. The position of this amino acid is different from those of determinants of target-sequence specificity in other zinc-finger proteins. Thus, the EPF family recognizes target sequences in a unique manner, together with the recognition of spacings in the target sequence that we demonstrated recently.