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Science 31 January 1997:
Vol. 275. no. 5300, pp. 657 - 661
DOI: 10.1126/science.275.5300.657
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Reports

A General Strategy for Selecting High-Affinity Zinc Finger Proteins for Diverse DNA Target Sites

Harvey A. Greisman and Carl O. Pabo *

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.

Howard Hughes Medical Institute and Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
*   To whom correspondence should be addressed.

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Positive and negative regulation of endogenous genes by designed transcription factors.
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PNAS 97, 1495-1500
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A bacterial two-hybrid selection system for studying protein-DNA and protein-protein interactions.
J. K. Joung, E. I. Ramm, and C. O. Pabo (2000)
PNAS 97, 7382-7387
   Abstract »    Full Text »    PDF »


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