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Rational design of DNA sequence-specific zinc fingers

Kono, Hidetoshi; Imanishi, Miki*; Negi, Shigeru*; Tatsutani, Kazuya*; Sakaeda, Yui*; Hashimoto, Ayaka*; Nakayama, Chie*; Futaki, Shiro*; Sugiura, Yukio*

We developed a rational scheme for designing DNA binding proteins. The scheme was applied for a zinc finger protein and the designed sequences were experimentally characterized with high DNA sequence specificity. Starting with the backbone of a known finger structure, we initially calculated amino acid sequences compatible with that expected structure and the designed fingers were experimentally confirmed their expected secondary structures. The DNA-binding function was then added to the designed finger by reconsidering a section of the amino acid sequence and computationally selecting amino acids to have the lowest protein-DNA interaction energy for the target DNA sequences. Among the designed proteins, one had a gap between the lowest and second lowest protein-DNA interaction energies that was sufficient to give DNA sequence-specificity.

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Category:Biochemistry & Molecular Biology

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