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Chem Biol. 2010 Sep 24;17(9):1018-29. doi: 10.1016/j.chembiol.2010.06.018.

Dual-color click beetle luciferase heteroprotein fragment complementation assays.

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1
Molecular Imaging Center, Mallinckrodt Institute of Radiology, Department of Developmental Biology, Washington University, St. Louis, MO 63110, USA.

Abstract

Understanding the functional complexity of protein interactions requires mapping biomolecular complexes within the cellular environment over biologically relevant time scales. Herein, we describe a set of reversible multicolored heteroprotein complementation fragments based on various firefly and click beetle luciferases that utilize the same substrate, D-luciferin. Luciferase heteroprotein fragment complementation systems enabled dual-color quantification of two discrete pairs of interacting proteins simultaneously or two distinct proteins interacting with a third shared protein in live cells. Using real-time analysis of click beetle green and click beetle red luciferase heteroprotein fragment complementation applied to β-TrCP, an E3-ligase common to the regulation of both β-catenin and IκBα, GSK3β was identified as a candidate kinase regulating IκBα processing. These dual-color protein interaction switches may enable directed dynamic analysis of a variety of protein interactions in living cells.

PMID:
20851351
PMCID:
PMC2943495
DOI:
10.1016/j.chembiol.2010.06.018
[Indexed for MEDLINE]
Free PMC Article
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