Parallelized Ligand Screening Using Dissolution Dynamic Nuclear Polarization

Analytical Chemistry
Yaewon KimChristian Hilty

Abstract

Protein-ligand interactions are frequently screened using nuclear magnetic resonance (NMR) spectroscopy. The dissociation constant (KD) of a ligand of interest can be determined via a spin-spin relaxation measurement of a reporter ligand in a single scan when using hyperpolarization by means of dissolution dynamic nuclear polarization (D-DNP). Despite nearly instantaneous signal acquisition, a limitation of D-DNP for the screening of protein-ligand interactions is the required polarization time on the order of tens of minutes. Here, we introduce a multiplexed NMR experiment, where a single hyperpolarized ligand sample is rapidly mixed with protein injected into two flow cells. NMR detection is achieved simultaneously on both channels, resulting in a chemical shift resolved spin relaxation measurement. Spectral resolution allows the use of reference compounds for accurate quantification of concentrations. Simultaneous use of two concentration ratios between protein and ligand broadens the range of KD that is accurately measurable in a single experiment to at least an order of magnitude. In a comparison of inhibitors for the protein trypsin, the average KD values of benzamidine and benzylamine were found to be 12.6 ± 1.4 μM and 2...Continue Reading

References

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Citations

Mar 31, 2018·Magnetic Resonance in Chemistry : MRC·Guannan Zhang, Christian Hilty
May 24, 2018·Chemistry, an Asian Journal·Kirill V KovtunovIgor V Koptyug
Jan 11, 2020·Chemphyschem : a European Journal of Chemical Physics and Physical Chemistry·Talia HarrisRachel Katz-Brull
Jun 30, 2019·Metabolites·Abdul-Hamid EmwasDavid S Wishart

Related Concepts

Ligands
Gene Products, Protein
Protein NMR Spectroscopy
Drug Prospecting
Benzamidines
Benzylamines
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