Proteomics

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Experimental strategies to improve drug-target identification in mass spectrometry-based thermal stability assays


ABSTRACT: Mass spectrometry (MS)-based thermal stability assays have recently emerged as one of the most promising solutions for the identification of protein-ligand interactions. Here, we have investigated eight combinations of several recently introduced MS-based advancements, including the Phase-Constrained Spectral Deconvolution Method, Field Asymmetric Ion Mobility Spectrometry, and the implementation of a carrier sample as improved MS-based acquisition approaches for thermal stability assays (iMAATSA). We used intact Jurkat cells treated with a commercially available MEK inhibitor, followed by heat treatment, to prepare a set of unfractionated isobarically-labeled proof-of-concept samples to compare the performance of eight different iMAATSAs. Finally, the best-performing iMAATSA was compared to a conventional approach and evaluated in a fractionation experiment. Improvements of up to 82% and 86% were demonstrated in protein identifications and high-quality melting curves, respectively, over the conventional approach in the proof-of-concept study, while an approximately 12% improvement in melting curve comparisons was achieved in the fractionation experiment.

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): T Cell, Cell Culture

SUBMITTER: Clifford Phaneuf  

LAB HEAD: Dr. Alexander R. Ivanov

PROVIDER: PXD040284 | Pride | 2023-05-10

REPOSITORIES: pride

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Experimental strategies to improve drug-target identification in mass spectrometry-based thermal stability assays.

Phaneuf Clifford G CG   Aizikov Konstantin K   Grinfeld Dmitry D   Kreutzmann Arne A   Mourad Daniel D   Lange Oliver O   Dai Daniel D   Zhang Bailin B   Belenky Alexei A   Makarov Alexander A AA   Ivanov Alexander R AR  

Communications chemistry 20230406 1


Mass spectrometry (MS)-based thermal stability assays have recently emerged as one of the most promising solutions for the identification of protein-ligand interactions. Here, we have investigated eight combinations of several recently introduced MS-based advancements, including the Phase-Constrained Spectral Deconvolution Method, Field Asymmetric Ion Mobility Spectrometry, and the implementation of a carrier sample as improved MS-based acquisition approaches for thermal stability assays (iMAATS  ...[more]

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