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MicroScale Thermophoresis: A Rapid and Precise Method to Quantify Protein–Nucleic Acid Interactions in Solution

  • Adrian Michael Mueller
  • Dennis Breitsprecher
  • Stefan Duhr
  • Philipp Baaske
  • Thomas SchubertEmail author
  • Gernot LängstEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1654)

Abstract

Interactions between nucleic acids and proteins are driving gene expression programs and regulating the development of organisms. The binding affinities of transcription factors to their target sites are essential parameters to reveal their binding site occupancy and function in vivo. Microscale Thermophoresis (MST) is a rapid and precise method allowing for quantitative analysis of molecular interactions in solution on a microliter scale. The technique is based on the movement of molecules in temperature gradients, which is referred to as thermophoresis, and depends on molecule size, charge, and hydration shell. Since at least one of these parameters is typically affected upon binding of a ligand, the method can be used to analyze any kind of biomolecular interaction. This section provides a detailed protocol describing the analysis of DNA–protein interactions, using the transcription factor TTF-I as a model protein that recognizes a 10 bp long sequence motif.

Key words

Binding assay Dissociation constant DNA–protein interactions MicroScale thermophoresis Binding affinity 

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Copyright information

© Springer Science+Business Media LLC 2017

Authors and Affiliations

  • Adrian Michael Mueller
    • 1
  • Dennis Breitsprecher
    • 2
  • Stefan Duhr
    • 2
  • Philipp Baaske
    • 2
  • Thomas Schubert
    • 3
    Email author
  • Gernot Längst
    • 1
    Email author
  1. 1.Biochemistry IIIUniversity of RegensburgRegensburgGermany
  2. 2.NanoTemper Technologies GmbHMunichGermany
  3. 3.2bind GmbHRegensburgGermany

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