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Gene Regulatory Networks

Volume 786 of the series Methods in Molecular Biology pp 115-130

Date:

Detecting Protein–Protein Interactions with the Split-Ubiquitin Sensor

  • Alexander DünklerAffiliated withDepartment of Biology, Institute of Molecular Genetics and Cell Biology, Ulm University
  • , Judith MüllerAffiliated withDepartment of Biology, Institute of Molecular Genetics and Cell Biology, Ulm University
  • , Nils JohnssonAffiliated withDepartment of Biology, Institute of Molecular Genetics and Cell Biology, Ulm University Email author 

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Abstract

A detailed understanding of a cellular process requires the knowledge about the interactions between its protein constituents. The Split-Ubiquitin technique allows to monitor and detect interactions of very diverse proteins, including transcription factors and membrane-associated proteins. The technique is based on unique features of ubiquitin, the enzymes of the ubiquitin pathway, and the reconstitution of a native-like ubiquitin from its N- and C-terminal fragments. Using Ura3p as a reporter for the reconstitution of the ubiquitin fragments, methods are presented that enable to screen in yeast for interaction partners of a given protein with either a randomly generated expression library or a defined but more limited array of protein fusions.

Key words

Protein–protein interaction Split-protein sensor Split-ubiquitin Protein interaction screens Systems biology Protein complementation assay Yeast Binary interactions