Nucleic Acids Research Advance Access published online on December 4, 2008
Nucleic Acids Research, doi:10.1093/nar/gkn962
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MolecularBiology |
An affinity-based scoring scheme for predicting DNA-binding activities of modularly assembled zinc-finger proteins
1Department of Genetics, Development and Cell Biology, Bioinformatics and Computational Biology Program, Iowa State University, Ames, IA 50011, 2Molecular Pathology Unit, Center for Cancer Research, and Center for Computational and Integrative Biology, Massachusetts General Hospital, 149 13th Street, Charlestown, MA 02129, 3Department of Pathology, Harvard Medical School, Boston, MA 02115 and 4Department of Genetics, Cell Biology and Development and Center for Genome Engineering, University of Minnesota, MN 55455, USA
*To whom correspondence should be addressed. Tel: +1 515 294 4991; Fax: +1 515 294 6790; Email: jdsander{at}iastate.edu
To whom correspondence should be addressed. Tel: +1 515 294 4991; Fax: +1 515 294 6790; Email: ddobbs{at}iastate.edu
Received August 7, 2008. Revised November 10, 2008. Accepted November 12, 2008.
Zinc-finger proteins (ZFPs) have long been recognized for their potential to manipulate genetic information because they can be engineered to bind novel DNA targets. Individual zinc-finger domains (ZFDs) bind specific DNA triplet sequences; their apparent modularity has led some groups to propose methods that allow virtually any desired DNA motif to be targeted in vitro. In practice, however, ZFPs engineered using this ‘modular assembly’ approach do not always function well in vivo. Here we report a modular assembly scoring strategy that both identifies combinations of modules least likely to function efficiently in vivo and provides accurate estimates of their relative binding affinities in vitro. Predicted binding affinities for 53 ‘three-finger’ ZFPs, computed based on energy contributions of the constituent modules, were highly correlated (r = 0.80) with activity levels measured in bacterial two-hybrid assays. Moreover, Kd values for seven modularly assembled ZFPs and their intended targets, measured using fluorescence anisotropy, were also highly correlated with predictions (r = 0.91). We propose that success rates for ZFP modular assembly can be significantly improved by exploiting the score-based strategy described here.
The authors wish it to be known that, in their opinion, the first two authors should be regarded as joint First Authors.