Abstract
AbstractDifferential scanning fluorimetry (DSF) is an accessible, rapid, and economical biophysical technique that has seen many applications over the years, ranging from protein folding state detection to the identification of ligands that bind to the target protein. In this review, we discuss the theory, applications, and limitations of DSF, including the latest applications of DSF by ourselves and other researchers. We show that DSF is a powerful high-throughput tool in early drug discovery efforts. We place DSF in the context of other biophysical methods frequently used in drug discovery and highlight their benefits and downsides. We illustrate the uses of DSF in protein buffer optimization for stability, refolding, and crystallization purposes and provide several examples of each. We also show the use of DSF in a more downstream application, where it is used as an in vivo validation tool of ligand-target interaction in cell assays. Although DSF is a potent tool in buffer optimization and large chemical library screens when it comes to ligand-binding validation and optimization, orthogonal techniques are recommended as DSF is prone to false positives and negatives.
Publisher
Springer Science and Business Media LLC
Subject
Molecular Biology,Structural Biology,Biophysics
Reference114 articles.
1. Alexander CG, Wanner R, Johnson CM et al (2014) Novel microscale approaches for easy, rapid determination of protein stability in academic and commercial settings. Biochim Biophys Acta - Proteins Proteomics 1844:2241–2250. https://doi.org/10.1016/j.bbapap.2014.09.016
2. Alexandrov AI, Mileni M, Chien EYT et al (2008) Microscale fluorescent thermal stability assay for membrane proteins. Structure 16:351–359. https://doi.org/10.1016/j.str.2008.02.004
3. Andrews BT, Capraro DT, Sulkowska JI, Onuchic JN, Patricia AJ (2013) Hysteresis as a marker for complex, overlapping landscapes in proteins. J Phys Chem Lett 4:180–188. https://doi.org/10.1021/jz301893w
4. Arai K, Yasuda S, Kornberg A (1981) Mechanism of dnaB protein action. I. Crystallization and properties of dnaB protein, an essential replication protein in Escherichia coli. J Biol Chem 256(10):5247–5252
5. Arakawa T, Ejima D, Tsumoto K et al (2007) Suppression of protein interactions by arginine: a proposed mechanism of the arginine effects. Biophys Chem
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