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High-resolution mapping of protein sequence-function relationships.

ABSTRACT: 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.

SUBMITTER: Fowler DM

PROVIDER: S-EPMC2938879 | biostudies-literature | 2010 Sep

REPOSITORIES: biostudies-literature

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High-resolution mapping of protein sequence-function relationships.

Fowler Douglas M DM Araya Carlos L CL Fleishman Sarel J SJ Kellogg Elizabeth H EH Stephany Jason J JJ Baker David D Fields Stanley S

Nature methods 20100815 9

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 proper ...[more]

PMID: 20711194

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