A Split-Luciferase Reporter Recognizing GFP and mCherry Tags to Facilitate Studies of Protein-Protein Interactions.

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.

SUBMITTER: Moustaqil M 

PROVIDER: S-EPMC5751283 | biostudies-literature | 2017 Dec

REPOSITORIES: biostudies-literature