13C-Fingerprinting and Metabolic Flux Analysis of Bacterial Metabolisms

Methods Mol Biol. 2019:1927:215-230. doi: 10.1007/978-1-4939-9142-6_15.

Abstract

13C-assisted metabolism analysis provides rigorous calculations of the intracellular reaction rates (i.e., fluxes) within the central metabolism of microbial hosts. This mapping of the intracellular network within microbes has proven to be essential for understanding the cell physiology. The approach is also a key to identifying central metabolic nodes, probing the rigidity of a metabolic network, revealing cofactor balances, and delineating hidden pathways. Here we present the methodology of using stable isotopic carbon substrates for both qualitative (13C-fingerprinting of functional pathways) and quantitative (Metabolic Flux Analysis) metabolism studies on bacterial species. In this methodology, we include step-by-step instructions to use the open source WUflux software for the steady-state flux calculations based on labeling information of amino acids or free metabolites.

Keywords: 13C-MFA; Flux ratio; Metabolism; Metabolites; Proteinogenic amino acids; WUflux.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Amino Acids / metabolism
  • Bacteria / metabolism*
  • Carbon Isotopes* / metabolism
  • Computational Biology / methods
  • Energy Metabolism*
  • Gas Chromatography-Mass Spectrometry
  • Glucose / metabolism
  • Metabolic Flux Analysis* / methods
  • Metabolic Networks and Pathways
  • Metabolomics* / methods
  • Software
  • User-Computer Interface
  • Workflow

Substances

  • Amino Acids
  • Carbon Isotopes
  • Glucose