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NanoBRET™ Assays for Monitoring Protein Interactions in Living Cells
Poster Title: NanoBRET™ Assays for Monitoring Protein Interactions in Living Cells
Submitted on 25 May 2017
Author(s): Jacqui L. Méndez, Kristin Riching, Sarah Mahan, Nancy Murphy, Marie Schwinn, Thomas Machleidt, Keith Wood, Marjeta Urh, and Danette L. Daniels
Affiliations: Promega Corporation, 2800 Woods Hollow Road, Madison, WI 53711, U.S.A
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Abstract: Identifying small molecule modulators, inhibitors or activators, of protein-protein interactions (PPI) remains challenging, largely due to the difficulty of developing robust, high-throughput screening tools that can used to interrogate these interactions within a biologically relevant context. Bioluminescent resonance energy transfer (BRET) has been used to monitor real-time protein:protein interactions in live cells, but current approaches suffer from limited sensitivity and narrow dynamic range. Here we present a new BRET method, termed NanoBRET, based on a small and extremely bright NanoLuc luciferase coupled to a HaloTag - long-wavelength fluorophore. This highly effective energy donor-acceptor combination boosts BRET performance, and the higher sensitivity facilitates application of the method in high density plates and high throughput screening. Here we present several examples of protein interaction biology that can be interrogated with NanoBRET including epigenetic interactions, adaptor recruitment to membrane receptors, and proteasomal recruitment of proteins targeted for degradation. Summary: NanoBRET PPI System is composed of two components

NanoBRET offers improved S:B over other BRET assays

NanoBRET provides sensitive live-cell method to study protein interactions
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