Optimization of protein samples for NMR using thermal shift assays

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• 作者：Sandra Kozak ; Lukas Lercher ; Megha N. Karanth ; Rob Meijers…
• 刊名：Journal of Biomolecular NMR
• 出版年：2016
• 出版时间：April 2016
• 年：2016
• 卷：64
• 期：4
• 页码：281-289
• 全文大小：1,496 KB
• 刊物类别：Physics and Astronomy
• 刊物主题：Physics
  Biophysics and Biomedical Physics
  Polymer Sciences
  Biochemistry
  
• 出版者：Springer Netherlands
• ISSN：1573-5001
• 卷排序：64

文摘

Maintaining a stable fold for recombinant proteins is challenging, especially when working with highly purified and concentrated samples at temperatures >20 °C. Therefore, it is worthwhile to screen for different buffer components that can stabilize protein samples. Thermal shift assays or ThermoFluor® provide a high-throughput screening method to assess the thermal stability of a sample under several conditions simultaneously. Here, we describe a thermal shift assay that is designed to optimize conditions for nuclear magnetic resonance studies, which typically require stable samples at high concentration and ambient (or higher) temperature. We demonstrate that for two challenging proteins, the multicomponent screen helped to identify ingredients that increased protein stability, leading to clear improvements in the quality of the spectra. Thermal shift assays provide an economic and time-efficient method to find optimal conditions for NMR structural studies.KeywordsDifferential scanning fluorimetryNuclear magnetic resonanceProtein thermal stabilitySample optimizationThermal shift assayThermoFluor