Improving analytical methods for protein-protein interaction through implementation of chemically inducible dimerization
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Improving analytical methods for protein-protein interaction through implementation of chemically inducible dimerization. / Andersen, Tonni Grube; Nintemann, Sebastian; Marek, Magdalena; Halkier, Barbara Ann; Schulz, Alexander; Burow, Meike.
I: Scientific Reports, Bind 6, 2016.Publikation: Bidrag til tidsskrift › Tidsskriftartikel › Forskning › fagfællebedømt
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TY - JOUR
T1 - Improving analytical methods for protein-protein interaction through implementation of chemically inducible dimerization
AU - Andersen, Tonni Grube
AU - Nintemann, Sebastian
AU - Marek, Magdalena
AU - Halkier, Barbara Ann
AU - Schulz, Alexander
AU - Burow, Meike
N1 - Do3qv Times Cited:0 Cited References Count:50
PY - 2016
Y1 - 2016
N2 - When investigating interactions between two proteins with complementary reporter tags in yeast two-hybrid or split GFP assays, it remains troublesome to discriminate true-from false-negative results and challenging to compare the level of interaction across experiments. This leads to decreased sensitivity and renders analysis of weak or transient interactions difficult to perform. In this work, we describe the development of reporters that can be chemically induced to dimerize independently of the investigated interactions and thus alleviate these issues. We incorporated our reporters into the widely used split ubiquitin-, bimolecular fluorescence complementation (BiFC)- and Forster resonance energy transfer (FRET)-based methods and investigated different protein-protein interactions in yeast and plants. We demonstrate the functionality of this concept by the analysis of weakly interacting proteins from specialized metabolism in the model plant Arabidopsis thaliana. Our results illustrate that chemically induced dimerization can function as a built-in control for split-based systems that is easily implemented and allows for direct evaluation of functionality.
AB - When investigating interactions between two proteins with complementary reporter tags in yeast two-hybrid or split GFP assays, it remains troublesome to discriminate true-from false-negative results and challenging to compare the level of interaction across experiments. This leads to decreased sensitivity and renders analysis of weak or transient interactions difficult to perform. In this work, we describe the development of reporters that can be chemically induced to dimerize independently of the investigated interactions and thus alleviate these issues. We incorporated our reporters into the widely used split ubiquitin-, bimolecular fluorescence complementation (BiFC)- and Forster resonance energy transfer (FRET)-based methods and investigated different protein-protein interactions in yeast and plants. We demonstrate the functionality of this concept by the analysis of weakly interacting proteins from specialized metabolism in the model plant Arabidopsis thaliana. Our results illustrate that chemically induced dimerization can function as a built-in control for split-based systems that is easily implemented and allows for direct evaluation of functionality.
KW - fkbp12-rapamycin-associated protein glucosinolate biosynthesis arabidopsis-thaliana fluorescent protein split-ubiquitin genetic system living cells in-vivo rapamycin target
U2 - 10.1038/srep27766
DO - 10.1038/srep27766
M3 - Journal article
C2 - 27282591
VL - 6
JO - Scientific Reports
JF - Scientific Reports
SN - 2045-2322
ER -
ID: 167093023