Metabolomics approach for enzyme discovery

Natsumi Saito, Martin Robert, Sayaka Kitamura, Richard Baran, Tomoyoshi Soga, Hirotada Mori, Takaaki Nishioka, Masaru Tomita

Research output: Contribution to journalArticle

61 Citations (Scopus)

Abstract

The search for novel enzymes is an important but difficult task in functional genomics. Here, we present a systematic method based on in vitro assays in combination with metabolite profiling to discover novel enzymatic activities. A complex mixture of metabolites is incubated with purified candidate proteins and the reaction mixture is subsequently profiled by capillary electrophoresis electrospray ionization mass spectrometry (CE-MS). Specific changes in the metabolite composition can directly suggest the presence of an enzymatic activity while subsequent identification of the compounds whose level changed specifically can pinpoint the actual substrate(s) and product(s) of the reaction. We first evaluated the method using several Escherichia coli metabolic enzymes and then applied it to the functional screening of uncharacterized proteins. In this manner, YbhA and YbiV proteins were found to display both phosphotransferase and phosphatase activity toward different sugars/sugar phosphates. Our approach should be broadly applicable and useful for enzyme discovery in any system.

Original languageEnglish
Pages (from-to)1979-1987
Number of pages9
JournalJournal of Proteome Research
Volume5
Issue number8
DOIs
Publication statusPublished - 2006 Aug 1

Keywords

  • Capillary electrophoresis
  • Enzyme
  • Functional genomics
  • Mass spectrometry
  • Metabolomics
  • Screening

ASJC Scopus subject areas

  • Biochemistry
  • Chemistry(all)

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    Saito, N., Robert, M., Kitamura, S., Baran, R., Soga, T., Mori, H., Nishioka, T., & Tomita, M. (2006). Metabolomics approach for enzyme discovery. Journal of Proteome Research, 5(8), 1979-1987. https://doi.org/10.1021/pr0600576