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- Title
A Split-Luciferase Reporter Recognizing GFP and mCherry Tags to Facilitate Studies of Protein-Protein Interactions.
- Authors
Moustaqil, Mehdi; Bhumkar, Akshay; Gonzalez, Laura; Raoul, Lisa; Hunter, Dominic J. B.; Carrive, Pascal; Sierecki, Emma; Gambin, Yann
- Abstract
The use of fluorescently-tagged proteins in microscopy has become routine, and anti-GFP (Green fluorescent protein) affinity matrices are increasingly used in proteomics protocols. However, some protein-protein interactions assays, such as protein complementation assays (PCA), require recloning of each protein as a fusion with the different parts of the complementation system. Here we describe a generic system where the complementation is separated from the proteins and can be directly used with fluorescently-tagged proteins. By using nanobodies and performing tests in cell-free expression systems, we accelerated the development of multiple reporters, detecting heterodimers and homodimers or oligomers tagged with GFP or mCherry. We demonstrate that the system can detect interactions at a broad range of concentrations, from low nanomolar up to micromolar.
- Subjects
GREEN fluorescent protein; PROTEOMICS; PROTEIN-protein interactions; LEISHMANIA; LUCIFERASE genetics
- Publication
International Journal of Molecular Sciences, 2017, Vol 18, Issue 12, p2681
- ISSN
1661-6596
- Publication type
Article
- DOI
10.3390/ijms18122681