Ewelina M. Sokolowska, Dennis Schlossarek, Marcin Luzarowski, Aleksandra Skirycz
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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.</p><p><b>Basic Protocol 1</b>: Preparation of native cell lysate from plant material</p><p><b>Support Protocol</b>: Bradford assay to determine protein concentration</p><p><b>Basic Protocol 2</b>: Separation of molecular complexes using size-exclusion chromatography</p><p><b>Basic Protocol 3</b>: Simultaneous extraction of proteins and metabolites using single-step extraction protocol</p><p><b>Basic Protocol 4</b>: Metabolomics analysis</p><p><b>Basic Protocol 5</b>: Proteomics analysis</p>","PeriodicalId":10932,"journal":{"name":"Current protocols in plant biology","volume":"4 4","pages":""},"PeriodicalIF":0.0000,"publicationDate":"2019-11-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/cppb.20101","citationCount":"13","resultStr":"{\"title\":\"PROMIS: Global Analysis of PROtein-Metabolite Interactions\",\"authors\":\"Ewelina M. 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引用次数: 13
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PROMIS: Global Analysis of PROtein-Metabolite Interactions
Small 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 material
Support Protocol : Bradford assay to determine protein concentration
Basic Protocol 2 : Separation of molecular complexes using size-exclusion chromatography
Basic Protocol 3 : Simultaneous extraction of proteins and metabolites using single-step extraction protocol
Basic Protocol 4 : Metabolomics analysis
Basic Protocol 5 : Proteomics analysis