Enantioselective protein affinity selection mass spectrometry (E-ASMS)
We report an enantioselective protein affinity selection mass spectrometry screening approach (E-ASMS) that enables the detection of weak binders, informs on selectivity, and generates orthogonal confirmation of binding. After method development with control proteins, we screen 31 human proteins against a designed library of 8,217 chiral compounds. We identify 16 binders to 12 targets, including many proteins predicted to be "challenging to ligand", and confirm their interactions through orthogonal biophysical assays. Seven binders to six targets display enantioselective binding, with K<sub>D</sub> values ranging from 3 to 20 µM. Binders for four targets (DDB1, WDR91, WDR55, and HAT1) are selected for in-depth characterization using X-ray crystallography. In all four cases, the mechanisms underlying enantioselectivity are readily explained. These results demonstrate that E-ASMS enables the identification and characterization of selective and weakly binding ligands for novel protein targets with unprecedented throughput and sensitivity.
Authors
- Wyatt W. Yue (ORCID: https://orcid.org/0000-0001-6959-6007)
- C.H. Arrowsmith (ORCID: https://orcid.org/0000-0002-4971-3250)
- Stefan Knapp (ORCID: https://orcid.org/0000-0001-5995-6494)
- A.M. Edwards (ORCID: https://orcid.org/0000-0002-4782-6016)
- Levon Halabelian (ORCID: https://orcid.org/0000-0003-4361-3619)
- Benjamin P. Brown (ORCID: https://orcid.org/0000-0002-3023-0427)
- Albina Bolotokova
- Tetiana Matviyuk
- Christopher Lenz (ORCID: https://orcid.org/0009-0000-2261-2823)
- Jianxian Sun (ORCID: https://orcid.org/0000-0002-0385-9959)
- Hongcheng Zeng (ORCID: https://orcid.org/0000-0002-4727-9980)
- Elisa Gibson (ORCID: https://orcid.org/0000-0002-7112-337X)
- Dafydd R. Owen (ORCID: https://orcid.org/0000-0003-3106-7809)
- Rachel Harding (ORCID: https://orcid.org/0000-0002-1134-391X)
- Ivan S. Kondratov (ORCID: https://orcid.org/0000-0003-0192-0725)
- Maria Kutera
- Conrad V. Simoben (ORCID: https://orcid.org/0000-0002-5958-6961)
- Susanne Müller (ORCID: https://orcid.org/0000-0003-2402-4157)
- Scott Houliston (ORCID: https://orcid.org/0000-0002-0113-0339)
- Vijayaratnam Santhakumar (ORCID: https://orcid.org/0000-0002-7001-557X)
- Stuart R. Green (ORCID: https://orcid.org/0000-0002-2960-9683)
- Fengling Li (ORCID: https://orcid.org/0000-0002-7962-9033)
- Matthieu Schapira (ORCID: https://orcid.org/0000-0002-1047-3309)
- Andreas C. Joerger (ORCID: https://orcid.org/0000-0002-1232-0138)
- Tyler S. Beyett (ORCID: https://orcid.org/0000-0001-5509-7004)
- Pegah Ghiabi
- Shabbir Ahmad (ORCID: https://orcid.org/0000-0003-2374-1197)
- Peter J. Brown (ORCID: https://orcid.org/0000-0002-8454-0367)
- Suzanne Ackloo (ORCID: https://orcid.org/0000-0002-9696-1839)
- Diwen Yang (ORCID: https://orcid.org/0000-0001-6951-9298)
- Madhushika Silva (ORCID: https://orcid.org/0009-0001-7060-0445)
- Lucy K Robbie
- R. L. Heath
- Aiping Dong
- Xiaoyun Wang (ORCID: https://orcid.org/0000-0002-7410-0667)
- Alexander Chuprina
- Danai Mavridi (ORCID: https://orcid.org/0009-0004-7379-9527)
- U Hang Chan
Institutions
- University of North Carolina at Chapel Hill (US)
- Goethe University Frankfurt (DE)
- University Health Network (CA)
- Harvard University (US)
- National Academy of Sciences of Ukraine (UA)
- Emory University (US)
- Pfizer (United States) (US)
- University of Toronto (CA)
- Scarborough Health Network (CA)
- Princess Margaret Cancer Centre (CA)
- Structural Genomics Consortium (CA)
- Institute of Bioorganic Chemistry and Petrochemistry V.P. Kukhar (UA)
- Lucht Probst Associates (Germany) (DE)
- Enamine (Ukraine) (UA)
- Newcastle University (GB)
Publication Details
- Journal
- Nature Communications
- Published
- 2025-12-17
- DOI
- https://doi.org/10.1038/s41467-025-67403-2
- Citations
- 1
- Primary Topic
- Mass Spectrometry Techniques and Applications
- Type
- article
- Field-Weighted Citation Impact
- 0.76
Funders
- Bristol-Myers Squibb
- Pfizer
- Bayer
- Emory University
- McGill University
- European Federation of Pharmaceutical Industries and Associations
- University of Toronto
- Alexander S. Onassis Public Benefit Foundation
- Deutsche Krebshilfe
- National Institutes of Health
- Genentech
- School of Medicine, Emory University
- Winship Cancer Institute
- Natural Sciences and Engineering Research Council of Canada