Binding Properties of the Artificial Zinc Fingers Coding Gene Sint1

Abstract

On the basis of the recognition “code” that suggests specific rules between zinc finger's primary structure and the finger's potential DNA binding sites, we have constructed a new three-zinc finger coding gene to target the nine base pair DNA sequence: 5′-TGG-ATG-GAC-3′. This artificial gene named “Sint1” belongs to the Cys2-His2zinc finger type. The amino acid positions, crucial for DNA binding, have been specifically chosen on the basis of the amino acid/base contacts more frequently represented in the available list of the proposed recognition “code”. Here we demonstrate that Sint1 protein binds specifically the double strand “code” DNA target, with a dissociation constant (Kd) comparable to the Kd of the well known Zif268 protein. Sint1 “code” deduced and the “experimental” selected DNA binding sites share five nucleotide positions. Interestingly, Sint1 shows both high affinity and specificity toward the single strand “code” DNA binding site, with a Kd comparable to the corresponding double strand DNA target. Moreover, we prove that Sint1 is able to bind RNA similarly to several natural zinc finger proteins.