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Anal Chem. 2019 Jul 2;91(13):8162-8169. doi: 10.1021/acs.analchem.9b00477. Epub 2019 Jun 12.

Accelerating the Throughput of Affinity Mass Spectrometry-Based Ligand Screening toward a G Protein-Coupled Receptor.

Lu Y1,2,3, Qin S1, Zhang B1, Dai A4, Cai X4, Ma M1,2,3, Gao ZG5, Yang D4, Stevens RC1,2, Jacobson KA5, Wang MW3,4,6, Shui W1,2.

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iHuman Institute , ShanghaiTech University , 201210 Shanghai , China.
School of Life Science and Technology , ShanghaiTech University , 201210 Shanghai , China.
University of Chinese Academy of Sciences , 100049 Beijing , China.
The National Center for Drug Screening and the CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica , Chinese Academy of Sciences , 201203 Shanghai , China.
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK) , National Institutes of Health , Bethesda , Maryland 20892 United States.
School of Pharmacy , Fudan University , 201203 Shanghai , China.


Affinity mass spectrometry (MS) enables rapid screening of compound mixtures for ligands bound to a specific protein target, yet its current throughput is limited to individually assay pools of 400-2000 compounds. Typical affinity MS screens implemented in pharmaceutical industry laboratories identify putative ligands based on qualitative analysis of compound binding to the target whereas no quantitative information is acquired to discriminate high- and low-affinity ligands in the screening phase. Furthermore, these screens require purification of a stabilized form of the protein target, which poses a great challenge for membrane receptor targets. Here, we describe a new, potentially general affinity MS strategy that allows screening of 20,000 compounds in one pool for highly efficient ligand discovery toward a G protein-coupled receptor (GPCR) target. Quantitative measurement of compound binding to the receptor enables high-affinity ligand selection using both the purified receptor and receptor-embedded cell membranes. This high-throughput, label-free and quantitative affinity MS screen resulted in discovery of three new antagonists of the A2A adenosine receptor.

[Available on 2020-07-02]

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