Mapping protein-metabolite interactions in E. coli by integrating chromatographic techniques and co-fractionation mass spectrometry
Mateusz Wagner,
Jieun Kang,
Catherine Mercado,
Venkatesh P. Thirumlaikumar,
Michal Gorka,
Hanne Zillmer,
Jingzhe Guo,
Romina I. Minen,
Caroline F. Plecki,
Katayoon Dehesh,
Frank C. Schroeder,
Dirk Walther,
Aleksandra Skirycz
Affiliations
Mateusz Wagner
Boyce Thompson Institute, Ithaca, NY 14853, USA; Cornell University, Ithaca, NY 14853, USA
Jieun Kang
Boyce Thompson Institute, Ithaca, NY 14853, USA; Michigan State University, East Lansing, MI 48824, USA
Catherine Mercado
Boyce Thompson Institute, Ithaca, NY 14853, USA; Michigan State University, East Lansing, MI 48824, USA
Venkatesh P. Thirumlaikumar
Boyce Thompson Institute, Ithaca, NY 14853, USA; Purdue University, West Lafayette, IN 47907, USA
Michal Gorka
Celon Pharma, 05092 Lomianki, Poland
Hanne Zillmer
Max-Planck-Institute of Molecular Plant Physiology, 14476 Potsdam-Golm, Germany
Jingzhe Guo
University of California, Riverside, Riverside, CA 92521, USA
Romina I. Minen
Boyce Thompson Institute, Ithaca, NY 14853, USA; DKFZ German Cancer Research Center, 69120 Heidelberg, Germany
Caroline F. Plecki
Boyce Thompson Institute, Ithaca, NY 14853, USA; Syracuse University, Syracuse, NY 13244, USA
Katayoon Dehesh
University of California, Riverside, Riverside, CA 92521, USA
Frank C. Schroeder
Boyce Thompson Institute, Ithaca, NY 14853, USA; Cornell University, Ithaca, NY 14853, USA
Dirk Walther
Max-Planck-Institute of Molecular Plant Physiology, 14476 Potsdam-Golm, Germany
Aleksandra Skirycz
Boyce Thompson Institute, Ithaca, NY 14853, USA; Cornell University, Ithaca, NY 14853, USA; Michigan State University, East Lansing, MI 48824, USA; Corresponding author
Summary: Toward characterization of protein-metabolite interactomes, we recently introduced PROMIS, a co-fractionation-based mass spectrometry approach. However, the challenge lies in distinguishing true interactors from coincidental co-elution when a metabolite co-fractionates with numerous proteins. To address this, we integrated two chromatographic techniques—size exclusion and ion exchange—to enhance the mapping of protein-metabolite interactions (PMIs) in Escherichia coli. This integration aims to refine the PMI network by considering size and charge characteristics, resulting in 994 interactions involving 51 metabolites and 465 proteins. The PMI network is enriched for known and predicted interactions, providing validation. Furthermore, analyzing protein targets for different metabolites revealed functional insights, such as a connection between proteinogenic dipeptides and fatty acid biosynthesis. Notably, we uncovered an inhibitory interaction between the riboflavin degradation product lumichrome and orotate phosphoribosyltransferase, a key enzyme in de novo pyrimidine synthesis affecting biofilm formation. In summary, our integrated chromatographic approach significantly advances PMI mapping.
