Ligand interaction landscape of transcription factors and essential enzymes in E. coli

Hui Peng; Sergei Kotelnikov; Megan E Egbert; Shany Ofaim; Grant C Stevens; Sadhna Phanse; Tatiana Saccon; Mikhail Ignatov; Shubham Dutta; Zoe Istace; Mohamed Taha Moutaoufik; Hiroyuki Aoki; Neal Kewalramani; Jianxian Sun; Yufeng Gong; Dzmitry Padhorny; Gennady Poda; Andrey Alekseenko; Kathryn A Porter; George Jones; Irina Rodionova; Hongbo Guo; Oxana Pogoutse; Suprama Datta; Milton Saier; Mark Crovella; Sandor Vajda; Gabriel Moreno-Hagelsieb; John Parkinson; Daniel Segre; Mohan Babu; Dima Kozakov; Andrew Emili

doi:10.1016/j.cell.2025.01.003

Ligand interaction landscape of transcription factors and essential enzymes in E. coli

Cell. 2025 Mar 6;188(5):1441-1455.e15. doi: 10.1016/j.cell.2025.01.003. Epub 2025 Jan 24.

Authors

Hui Peng¹, Sergei Kotelnikov², Megan E Egbert³, Shany Ofaim⁴, Grant C Stevens⁵, Sadhna Phanse⁶, Tatiana Saccon⁷, Mikhail Ignatov², Shubham Dutta⁷, Zoe Istace⁷, Mohamed Taha Moutaoufik⁸, Hiroyuki Aoki⁷, Neal Kewalramani⁹, Jianxian Sun¹, Yufeng Gong¹, Dzmitry Padhorny², Gennady Poda¹⁰, Andrey Alekseenko², Kathryn A Porter³, George Jones², Irina Rodionova¹¹, Hongbo Guo⁵, Oxana Pogoutse⁵, Suprama Datta¹², Milton Saier¹¹, Mark Crovella¹³, Sandor Vajda¹⁴, Gabriel Moreno-Hagelsieb¹⁵, John Parkinson⁵, Daniel Segre¹⁶, Mohan Babu¹⁷, Dima Kozakov¹⁸, Andrew Emili¹⁹

Affiliations

¹ Department of Chemistry, University of Toronto, Toronto, ON M5S 3H6, Canada.
² Laufer Center for Physical and Quantitative Biology, Stony Brook University, Stony Brook, NY 11794, USA.
³ Department of Biomedical Engineering, Boston University, Boston, MA 02215, USA.
⁴ Program in Bioinformatics, Boston University, Boston, MA 02215, USA.
⁵ Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 1A8, Canada.
⁶ Center for Network Systems Biology, Boston University, Boston, MA 02218, USA; Department of Biochemistry, University of Regina, Regina, SK S4S 0A2, Canada.
⁷ Department of Biochemistry, University of Regina, Regina, SK S4S 0A2, Canada.
⁸ Department of Biochemistry, University of Regina, Regina, SK S4S 0A2, Canada; Faculty of Medical Sciences, University Mohammed VI Polytechnic, Benguerir, Morocco.
⁹ Program in Bioinformatics, Boston University, Boston, MA 02215, USA; Center for Network Systems Biology, Boston University, Boston, MA 02218, USA.
¹⁰ Drug Discovery Program, Ontario Institute for Cancer Research, Toronto, ON M5G 0A3, Canada; Leslie Dan Faculty of Pharmacy, University of Toronto, Toronto, ON M5S 3M2, Canada.
¹¹ Department of Molecular Biology, University of California, San Diego, La Jolla, San Diego, CA 920930, USA.
¹² Center for Network Systems Biology, Boston University, Boston, MA 02218, USA.
¹³ Program in Bioinformatics, Boston University, Boston, MA 02215, USA; Department of Computer Science, Boston University, Boston, MA 02215, USA.
¹⁴ Department of Biomedical Engineering, Boston University, Boston, MA 02215, USA; Department of Chemistry, Boston University, Boston, MA 02215, USA.
¹⁵ Department of Biology, Wilfrid Laurier University, Waterloo, ON N2L 3C5, Canada.
¹⁶ Department of Biomedical Engineering, Boston University, Boston, MA 02215, USA; Program in Bioinformatics, Boston University, Boston, MA 02215, USA; Department of Biology, Boston University, Boston, MA 02215, USA.
¹⁷ Department of Biochemistry, University of Regina, Regina, SK S4S 0A2, Canada. Electronic address: mohan.babu@uregina.ca.
¹⁸ Laufer Center for Physical and Quantitative Biology, Stony Brook University, Stony Brook, NY 11794, USA. Electronic address: midas@laufercenter.org.
¹⁹ Program in Bioinformatics, Boston University, Boston, MA 02215, USA; Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 1A8, Canada; Center for Network Systems Biology, Boston University, Boston, MA 02218, USA; Department of Chemistry, Boston University, Boston, MA 02215, USA; Department of Chemical Physiology and Biochemistry, Division of Oncological Sciences, Knight Cancer Institute, Oregon Health and Science University, Portland, OR, USA. Electronic address: emili@ohsu.edu.

PMID: 39862855
DOI: 10.1016/j.cell.2025.01.003

Abstract

Knowledge of protein-metabolite interactions can enhance mechanistic understanding and chemical probing of biochemical processes, but the discovery of endogenous ligands remains challenging. Here, we combined rapid affinity purification with precision mass spectrometry and high-resolution molecular docking to precisely map the physical associations of 296 chemically diverse small-molecule metabolite ligands with 69 distinct essential enzymes and 45 transcription factors in the gram-negative bacterium Escherichia coli. We then conducted systematic metabolic pathway integration, pan-microbial evolutionary projections, and independent in-depth biophysical characterization experiments to define the functional significance of ligand interfaces. This effort revealed principles governing functional crosstalk on a network level, divergent patterns of binding pocket conservation, and scaffolds for designing selective chemical probes. This structurally resolved ligand interactome mapping pipeline can be scaled to illuminate the native small-molecule networks of complete cells and potentially entire multi-cellular communities.

Keywords: LP/MS; chemical probe; dynamic network; enzymes; molecular docking; multi-modal interactome; precision structural modeling; protein-ligand binding pocket; protein-metabolite interactions; transcription factors.

MeSH terms

Binding Sites
Escherichia coli Proteins* / chemistry
Escherichia coli Proteins* / metabolism
Escherichia coli* / enzymology
Escherichia coli* / metabolism
Ligands
Metabolic Networks and Pathways
Molecular Docking Simulation
Protein Binding
Transcription Factors* / chemistry
Transcription Factors* / metabolism

Substances

Ligands
Transcription Factors
Escherichia coli Proteins