Maintaining a stable fold for recombinant proteins is challenging, especially when working with highly purified and concentrated samples at temperatures >20Â*°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.
[NMR paper] Optimization of protein samples for NMR using thermal shift assays.
Optimization of protein samples for NMR using thermal shift assays.
Optimization of protein samples for NMR using thermal shift assays.
J Biomol NMR. 2016 Mar 17;
Authors: Kozak S, Lercher L, Karanth MN, Meijers R, Carlomagno T, Boivin S
Abstract
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...
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Increasing Throughput in Protein Thermal Shift Assays - Genetic Engineering & Biotechnology News
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Increasing Throughput in Protein Thermal Shift Assays
Genetic Engineering & Biotechnology News
Historically, protein interaction studies have been measured using an array of techniques, including nuclear magnetic resonance, differential scanning calorimetry, and surface plasma resonance imaging. Additionally, protein melt screening methods used ...
Increasing Throughput in Protein Thermal Shift Assays - Genetic Engineering & Biotechnology News
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[NMR paper] Practical use of chemical shift databases for protein solid-state NMR: 2D chemical shift maps and amino-acid assignment with secondary-structure information.
Practical use of chemical shift databases for protein solid-state NMR: 2D chemical shift maps and amino-acid assignment with secondary-structure information.
Practical use of chemical shift databases for protein solid-state NMR: 2D chemical shift maps and amino-acid assignment with secondary-structure information.
J Biomol NMR. 2013 Apr 28;
Authors: Fritzsching KJ, Yang Y, Schmidt-Rohr K, Hong M
Abstract
We introduce a Python-based program that utilizes the large database of (13)C and (15)N chemical shifts in the Biological Magnetic...
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04-30-2013 10:21 PM
[NMR paper] Protein structure prediction using global optimization by basin-hopping with NMR shift restraints.
Protein structure prediction using global optimization by basin-hopping with NMR shift restraints.
Related Articles Protein structure prediction using global optimization by basin-hopping with NMR shift restraints.
J Chem Phys. 2013 Jan 14;138(2):025102
Authors: Hoffmann F, Strodel B
Abstract
Computational methods that utilize chemical shifts to produce protein structures at atomic resolution have recently been introduced. In the current work, we exploit chemical shifts by combining the basin-hopping approach to global optimization with...
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02-03-2013 10:22 AM
[NMR paper] NMR assays for carbohydrate-based vaccines.
NMR assays for carbohydrate-based vaccines.
Related Articles NMR assays for carbohydrate-based vaccines.
J Pharm Biomed Anal. 2005 Aug 10;38(5):840-50
Authors: Jones C
Antibodies against the cell surface carbohydrates of many microbial pathogens protect against infection. This was initially exploited by the development of purified polysaccharide vaccines, but glycoconjugate vaccines, in which the cell surface carbohydrate of a microbial pathogen is covalently attached to an appropriate carrier protein, are proving the most effective means to...
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11-25-2010 08:21 PM
[NMR paper] Identification and optimization of protein domains for NMR studies.
Identification and optimization of protein domains for NMR studies.
Related Articles Identification and optimization of protein domains for NMR studies.
Methods Enzymol. 2005;394:3-16
Authors: Card PB, Gardner KH
The success of genomic sequencing projects in recent years has presented protein scientists with a formidable challenge in characterizing the vast number of gene products that have subsequently been identified. NMR has proven to be a valuable tool in the elucidation of various properties for many of these proteins, allowing versatile...
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[NMR paper] Thermal stability studies of a globular protein in aqueous poly(ethylene glycol) by (
Thermal stability studies of a globular protein in aqueous poly(ethylene glycol) by (1)H NMR.
http://www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--www3.interscience.wiley.com-aboutus-images-wiley_interscience_pubmed_logo_120x27.gif Related Articles Thermal stability studies of a globular protein in aqueous poly(ethylene glycol) by (1)H NMR.
Biotechnol Bioeng. 1996 Aug 20;51(4):410-21
Authors: Hancock TJ, Hsu JT
The reversible folding destabilization of hen lysozyme has been confirmed by a melting temperature (T(m)) decrease in aqueous...
Optimization of protein samples for NMR using thermal shift assays
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