| Title: | High-throughput characterization of protein-protein interactions by reprogramming yeast mating |
| Author(s): | Younger D; Berger S; Baker D; Klavins E; |
| Address: | "Bioengineering Department, University of Washington, Seattle, WA 98105. Institute for Protein Design, University of Washington, Seattle, WA 98195. Institute for Protein Design, University of Washington, Seattle, WA 98195; dabaker@uw.edu klavins@uw.edu. Biochemistry Department, University of Washington, Seattle, WA 98195. Howard Hughes Medical Institute, University of Washington, Seattle, WA 98195. Electrical Engineering Department, University of Washington, Seattle, WA 98195 dabaker@uw.edu klavins@uw.edu" |
| ISSN/ISBN: | 1091-6490 (Electronic) 0027-8424 (Print) 0027-8424 (Linking) |
| Abstract: | "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 K(d)s ranging from below 500 pM to above 300 muM. 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" |
| Keywords: | "Agglutination/genetics *Gene Expression Regulation, Fungal Gene Library Mating Factor/*genetics/metabolism Protein Binding Protein Interaction Mapping/*methods Saccharomyces cerevisiae/*genetics/metabolism *Translocation, Genetic Two-Hybrid System Techniq;" |
| Notes: | "MedlineYounger, David Berger, Stephanie Baker, David Klavins, Eric eng HHMI/Howard Hughes Medical Institute/ Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S. 2017/11/01 Proc Natl Acad Sci U S A. 2017 Nov 14; 114(46):12166-12171. doi: 10.1073/pnas.1705867114. Epub 2017 Oct 31" |
