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Smart detection methods for protein interactions in living cells

 

Summary

In this webinar, we introduce the bioluminescent NanoBiT® and NanoBRETTM technology platforms and their use for studying protein interactions in live cells.  NanoLuc® Binary Technology (NanoBiT®) is a structural complementation reporter system based on NanoLuc® luciferase. It is composed of a large subunit called LgBiT (Large BiT; 18kDa) and a small subunit called SmBiT (Small BiT; 1.3 kDa) whose physical interaction reconstitutes the bright NanoBiT® luciferase. Their small size minimizes interference with normal protein function and the low intrinsic affinity mitigates unspecific signal while allowing to monitor interaction dynamics in real-time.

NanoBRET™ represents a powerful alternative to interrogate homo- and heteromeric protein interactions. In this proximity-based assay NanoLuc® is used as BRET energy donor and fluorescently labelled HaloTag® as the energy acceptor. When fused to proteins of interest, this technology enables sensitive, reproducible detection of protein interactions within their cellular context. Furthermore, we present how this live cell format can be adapted to quantitively assess target engagement.

We will highlight the advantages of NanoBiT® and NanoBRET™ and discuss the key steps of their workflows.

You will learn:

  • Principle of NanoBiT® and NanoBRET™ technologies
  • How to use NanoBiT® for studying protein:protein interaction
  • How to use NanoBRETTM for studying protein:protein interaction
  • How to use NanoBRET™ for studying target engagement


Speaker

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Erik Bonke, PhD

Application Specialist
Promega GmbH

Erik Bonke received a Diploma in biology from the Johannes-Gutenberg University in Mainz, Germany in 2012. In his diploma thesis at the University Hospital in Mainz he worked at the genetic manipulation of murine embryonic stem cells in order to generate different transgenic mouse strains. He did his doctoral thesis at the University Hospital in Frankfurt am Main, where he focused on the mechanistic fundamentals of mitochondrial ROS generation and their physiological implication as cellular second messenger molecules, a process termed redox signaling. After completion of the experimental part of his thesis, Mr. Bonke joined Promega Germany to work as an Application Specialist with a main focus on cellular reporter technologies. As part of this position, he is giving frequent practical as well as theoretical workshops/seminars on the application of Promega's current luminescent reporter portfolio. In 2018, he was awarded a doctoral degree by the Johann Wolfgang Goethe University Frankfurt.

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