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High-throughput characterization of protein–protein interactions by reprogramming yeast mating

  1. Eric Klavinse,1
  1. aBioengineering Department, University of Washington, Seattle, WA 98105;
  2. bInstitute for Protein Design, University of Washington, Seattle, WA 98195;
  3. cBiochemistry Department, University of Washington, Seattle, WA 98195;
  4. dHoward Hughes Medical Institute, University of Washington, Seattle, WA 98195;
  5. eElectrical Engineering Department, University of Washington, Seattle, WA 98195
  1. Edited by Brenda Andrews, University of Toronto, Toronto, ON, Canada, and accepted by Editorial Board Member James J. Collins September 29, 2017 (received for review April 11, 2017)


De novo design of protein binders often requires experimental screening to select functional variants from a design library. We have achieved high-throughput, quantitative characterization of protein–protein binding interactions without requiring purified recombinant proteins, by linking interaction strength with yeast mating. Using a next-generation sequencing output, we have characterized protein networks consisting of thousands of pairwise interactions in a single tube and have demonstrated the effect of changing the binding environment. This approach addresses an existing bottleneck in protein binder design by enabling the high-throughput and quantitative characterization of binding strength between designed protein libraries and multiple target proteins in a fully defined environment.


High-throughput methods for screening protein–protein interactions enable the rapid characterization of engineered binding proteins and interaction networks. While existing approaches are powerful, none allow quantitative library-on-library characterization of protein interactions in a modifiable extracellular environment. Here, we show that sexual agglutination of Saccharomyces cerevisiae can be reprogrammed to link interaction strength with mating efficiency using synthetic agglutination (SynAg). Validation of SynAg with 89 previously characterized interactions shows a log-linear relationship between mating efficiency and protein binding strength for interactions with Kds ranging from below 500 pM to above 300 μM. Using induced chromosomal translocation to pair barcodes representing binding proteins, thousands of distinct interactions can be screened in a single pot. We demonstrate the ability to characterize protein interaction networks in a modifiable environment by introducing a soluble peptide that selectively disrupts a subset of interactions in a representative network by up to 800-fold. SynAg enables the high-throughput, quantitative characterization of protein–protein interaction networks in a fully defined extracellular environment at a library-on-library scale.


  • ?1To whom correspondence may be addressed. Email: dabaker{at}uw.edu or klavins{at}uw.edu.

Published under the PNAS license.

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