Data_Sheet_4_A Yeast-Based Functional Assay to Study Plant N-Degron – N-Recognin Interactions.pdf (84.38 kB)
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Data_Sheet_4_A Yeast-Based Functional Assay to Study Plant N-Degron – N-Recognin Interactions.pdf

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posted on 07.01.2022, 04:20 authored by Aida Kozlic, Nikola Winter, Theresia Telser, Jakob Reimann, Katrin Rose, Lilian Nehlin, Sophie Berckhan, Gunjan Sharma, Charlene Dambire, Tinne Boeckx, Michael J. Holdsworth, Andreas Bachmair

The N-degron pathway is a branch of the ubiquitin-proteasome system where amino-terminal residues serve as degradation signals. In a synthetic biology approach, we expressed ubiquitin ligase PRT6 and ubiquitin conjugating enzyme 2 (AtUBC2) from Arabidopsis thaliana in a Saccharomyces cerevisiae strain with mutation in its endogenous N-degron pathway. The two enzymes re-constitute part of the plant N-degron pathway and were probed by monitoring the stability of co-expressed GFP-linked plant proteins starting with Arginine N-degrons. The novel assay allows for straightforward analysis, whereas in vitro interaction assays often do not allow detection of the weak binding of N-degron recognizing ubiquitin ligases to their substrates, and in planta testing is usually complex and time-consuming.

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