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Updated: Jul 2, 2025

Split-Ubiquitin Based Membrane Yeast Two-Hybrid MYTH System: A Powerful Tool For Identifying Protein-Protein Interactions
Published on: February 1, 2010
ER Membrane Protein Interactions Using the Split-Ubiquitin System (SUS)
Lisa Yasmin Asseck1, Niklas Wallmeroth1, Christopher Grefen2
1Centre for Plant Molecular Biology, Developmental Genetics, University of Tübingen, Tübingen, Germany.
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Protein-protein interactions (PPIs) play fundamental roles in all cellular processes. Especially membrane proteins facilitate a range of important biological functions in stimuli perception, signalling, and transport. Here we describe a detailed protocol for the yeast mating-based Split-Ubiquitin System (mbSUS) to study PPIs of ER membrane proteins in vivo. In contrast to the prominent yeast two hybrid, mbSUS enables analysis of full-length membrane proteins in their native cellular context. The system is based on the ubiquitin proteasome pathway leading to the release of an artificial transcription factor followed by activation of reporter genes to visualize PPIs. The mating-based approach is suitable for both small- and large-scale interaction studies. Additionally, we describe protocols to apply the recently established SUS Bridge assay (SUB), which is optimized for the detection of ternary protein interactions.
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