Chapter title |
TIPI: TEV Protease-Mediated Induction of Protein Instability.
|
---|---|
Chapter number | 43 |
Book title |
Ubiquitin Family Modifiers and the Proteasome
|
Published in |
Methods in molecular biology, January 2012
|
DOI | 10.1007/978-1-61779-474-2_43 |
Pubmed ID | |
Book ISBNs |
978-1-61779-473-5, 978-1-61779-474-2
|
Authors |
Christof Taxis, Michael Knop |
Editors |
R. Jürgen Dohmen, Martin Scheffner |
Abstract |
Reverse genetics approaches require methods to inactivate a specific protein. One possibility is to modify the target protein with a degradation signal (degron). Degrons are short, transferable sequences that confer protein instability. They target proteins for degradation either constitutively or after activation, e.g., by phosphorylation, presence of a binding partner, or conformational rearrangements in the substrate. In this chapter, we describe a synthetic way to activate a degron. It employs the generation of an N-degron by cleavage of a substrate with the site-specific tobacco etch virus (TEV) protease. Subsequently, the substrate is targeted for degradation by the ubiquitin-proteasome system. This TEV protease-induced protein instability system provides a powerful approach to generate conditional mutants for synthetic biology or for the investigation of protein functions in a specific cellular context. |
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