Abstract
A method is described for selecting DNA-binding proteins that recognize desired sequences. The protocol involves gradually extending a new zinc finger protein across the desired 9- or 10-base pair target site, adding and optimizing one finger at a time. This procedure was tested with a TATA box, a p53 binding site, and a nuclear receptor element, and proteins were obtained that bind with nanomolar dissociation constants and discriminate effectively (greater than 20,000-fold) against nonspecific DNA. This strategy may provide important information about protein-DNA recognition as well as powerful tools for biomedical research.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Amino Acid Sequence
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Base Composition
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Base Sequence
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Binding Sites
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DNA / metabolism*
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DNA-Binding Proteins / chemistry
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DNA-Binding Proteins / metabolism*
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Genes, p53
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Hydrogen Bonding
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Models, Molecular
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Molecular Sequence Data
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Nucleic Acid Conformation
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Peptide Library
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Protein Conformation
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Protein Engineering*
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Protein Structure, Secondary
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Receptors, Cytoplasmic and Nuclear / genetics
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TATA Box
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Transcription Factors / chemistry
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Transcription Factors / metabolism
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Zinc Fingers*
Substances
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DNA-Binding Proteins
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Peptide Library
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Receptors, Cytoplasmic and Nuclear
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Transcription Factors
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DNA