Journal
NATURE METHODS
Volume 7, Issue 9, Pages 741-U108Publisher
NATURE PUBLISHING GROUP
DOI: 10.1038/nmeth.1492
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Funding
- NATIONAL CENTER FOR RESEARCH RESOURCES [P41RR011823] Funding Source: NIH RePORTER
- NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES [F32GM084699] Funding Source: NIH RePORTER
- Howard Hughes Medical Institute Funding Source: Medline
- NCRR NIH HHS [P41 RR011823, P41 RR11823, P41 RR011823-05] Funding Source: Medline
- NIGMS NIH HHS [P41 GM103533, F32GM084699, F32 GM084699, F32 GM084699-03] Funding Source: Medline
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We present a large-scale approach to investigate the functional consequences of sequence variation in a protein. The approach entails the display of hundreds of thousands of protein variants, moderate selection for activity and high-throughput DNA sequencing to quantify the performance of each variant. Using this strategy, we tracked the performance of > 600,000 variants of a human WW domain after three and six rounds of selection by phage display for binding to its peptide ligand. Binding properties of these variants defined a high-resolution map of mutational preference across the WW domain; each position had unique features that could not be captured by a few representative mutations. Our approach could be applied to many in vitro or in vivo protein assays, providing a general means for understanding how protein function relates to sequence.
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