Dataset Information
Functional analysis of isoprenoid precursors biosynthesis by quantitative metabolomics and isotopologue profiling
Ontology highlight
ABSTRACT: INTRODUCTION: Isoprenoids are amongst the most abundant and diverse biological molecules and are involved in a broad range of biological functions. Functional understanding of their biosynthesis is thus key in many fundamental and applicative fields, including systems biology, medicine and biotechnology. However, available methods do not yet allow accurate quantification and tracing of stable isotopes incorporation for all the isoprenoids precursors. OBJECTIVES: We developed and validated a complete methodology for quantitative metabolomics and isotopologue profiling of isoprenoid precursors in the yeast Saccharomyces cerevisiae. METHODS: This workflow covers all the experimental and computational steps from sample collection and preparation to data acquisition and processing. It also includes a novel quantification method based on liquid chromatography coupled to high-resolution mass spectrometry. Method validation followed the Metabolomics Standards Initiative guidelines. RESULTS: This workflow ensures accurate absolute quantification (RSD < 20%) of all mevalonate and prenyl pyrophosphates intermediates with a high sensitivity over a large linear range (from 0.1 to 50 pmol). In addition, we demonstrate that this workflow brings crucial information to design more efficient phytoene producers. Results indicate stable turnover rates of prenyl pyrophosphate intermediates in the constructed strains and provide quantitative information on the change of the biosynthetic flux of phytoene precursors. CONCLUSION: This methodology fills one of the last technical gaps for functional studies of isoprenoids biosynthesis and should be applicable to other eukaryotic and prokaryotic (micro)organisms after adaptation of some organism-dependent steps. This methodology also opens the way to 13C-metabolic flux analysis of isoprenoid biosynthesis.
INSTRUMENT(S): Liquid Chromatography MS - negative - reverse phase
SUBMITTER: floriant Bellvert
PROVIDER: MTBLS923 | MetaboLights | 2021-04-01
REPOSITORIES: MetaboLights
Publications
Functional analysis of isoprenoid precursors biosynthesis by quantitative metabolomics and isotopologue profiling.
Metabolomics : Official journal of the Metabolomic Society 20190821 9
<h4>Introduction</h4>Isoprenoids are amongst the most abundant and diverse biological molecules and are involved in a broad range of biological functions. Functional understanding of their biosynthesis is thus key in many fundamental and applicative fields, including systems biology, medicine and biotechnology. However, available methods do not yet allow accurate quantification and tracing of stable isotopes incorporation for all the isoprenoids precursors.<h4>Objectives</h4>We developed and val ...[more]
