COMPREHENSIVE DNA RECOGNITION THROUGH CONCERTED INTERACTIONS FROM ADJACENT ZINC FINGERS

Zinc fingers are small DNA-binding modules noted for their occurrence in a large number of eukaryotic transcription factors, and their use i n protein engineering. Although it was expected that zinc fingers can bind to a wide diversity of DNA sequences, previous studies using mode l zinc finger domains from Zif268 (and Sp1) have revealed a potential Limitation to the DNA-binding specificity. For example, phage display selection of individual zinc fingers to recognize trinucleotide DNA su bsites returned fingers that bound specifically only to triplets of th e form GNN, i.e., triplets with guanine at the 5' end. Following our r ecently reported work [Isalan, M., Choo, Y., and Klug, A. (1997) Proc. Natl. Acad. Sci. U.S.A. 94, 5617-5621], we now show that this limitat ion can be overcome by the concerted randomization of certain amino ac id positions in adjacent zinc fingers that specify overlapping DNA sub sites. This illustrates an important mechanism underlying DNA recognit ion by arrays of zinc fingers, and points the way to improved strategi es for the design of highly specific zinc finger proteins that bind an y given nucleotide sequence.