Dual-color click beetle luciferase heteroprotein fragment complementation assays

Victor Villalobos, Snehal Naik, Monique Bruinsma, Robin S. Dothager, Mei Hsiu Pan, Mustapha Samrakandi, Britney Moss, Adnan Elhammali, David Piwnica-Worms

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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.

Original languageEnglish
Pages (from-to)1018-1029
Number of pages12
JournalChemistry and Biology
Issue number9
StatePublished - Sep 24 2010

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    Villalobos, V., Naik, S., Bruinsma, M., Dothager, R. S., Pan, M. H., Samrakandi, M., Moss, B., Elhammali, A., & Piwnica-Worms, D. (2010). Dual-color click beetle luciferase heteroprotein fragment complementation assays. Chemistry and Biology, 17(9), 1018-1029. https://doi.org/10.1016/j.chembiol.2010.06.018