Media type: E-Article Title: PROMIS: Global Analysis of PROtein‐Metabolite Interactions Contributor: Sokolowska, Ewelina M.; Schlossarek, Dennis; Luzarowski, Marcin; Skirycz, Aleksandra Published: Wiley, 2019 Published in: Current Protocols in Plant Biology, 4 (2019) 4 Language: English DOI: 10.1002/cppb.20101 ISSN: 2379-8068 Origination: Footnote: Description: AbstractSmall molecules are not only intermediates of metabolism, but also play important roles in signaling and in controlling cellular metabolism, growth, and development. Although a few systematic studies have been conducted, the true extent of protein–small molecule interactions in biological systems remains unknown. PROtein–metabolite interactions using size separation (PROMIS) is a method for studying protein–small molecule interactions in a non‐targeted, proteome‐ and metabolome‐wide manner. This approach uses size‐exclusion chromatography followed by proteomics and metabolomics liquid chromatography–mass spectrometry analysis of the collected fractions. Assuming that small molecules bound to proteins would co‐fractionate together, we found numerous small molecules co‐eluting with proteins, strongly suggesting the formation of stable complexes. Using PROMIS, we identified known small molecule–protein complexes, such as between enzymes and cofactors, and also found novel interactions. © 2019 The Authors.Basic Protocol 1: Preparation of native cell lysate from plant materialSupport Protocol: Bradford assay to determine protein concentrationBasic Protocol 2: Separation of molecular complexes using size‐exclusion chromatographyBasic Protocol 3: Simultaneous extraction of proteins and metabolites using single‐step extraction protocolBasic Protocol 4: Metabolomics analysisBasic Protocol 5: Proteomics analysis