Metabolomics

, Volume 9, Issue 2, pp 403–417

Investigating alginate production and carbon utilization in Pseudomonas fluorescens SBW25 using mass spectrometry-based metabolic profiling

  • Stina K. Lien
  • Håvard Sletta
  • Trond E. Ellingsen
  • Svein Valla
  • Elon Correa
  • Royston Goodacre
  • Kai Vernstad
  • Sven Even Finborud Borgos
  • Per Bruheim
Original Article

Abstract

Metabolic profiling of Pseudomonas fluorescens SBW25 and various mutants derived thereof was performed to explore how the bacterium adapt to changes in carbon source and upon induction of alginate synthesis. The experiments were performed at steady-state conditions in nitrogen-limited chemostats using either fructose or glycerol as carbon source. Carbon source consumption was up-regulated in the alginate producing mutant with inactivated anti-sigma factor MucA. The mucA- mutants (also non-alginate producing mucA- control strains) had a higher dry weight yield on carbon source implying a change in carbon and energy metabolism due to the inactivation of the anti-sigma factor MucA. Both LC–MS/MS and GC–MS methods were used for quantitative metabolic profiling, and major reorganization of primary metabolite pools in both an alginate producing and a carbon source dependent manner was observed. Generally, larger changes were observed among the phosphorylated glycolytic metabolites, the pentose phosphate pathway metabolites and the nucleotide pool than among amino acids and citric acid cycle compounds. The most significant observation at the metabolite level was the significantly reduced energy charge of the mucA- mutants (both alginate producing and non-producing control strains) compared to the wild type strain. This reduction was caused more by a strong increase in the AMP pool than changes in the ATP and ADP pools. The alginate-producing mucA- mutant had a slightly increased GTP pool, while the GDP and GMP pools were strongly increased compared to non-producing mucA- strains and to the wild type. Thus, whilst changes in the adenosine phosphate nucleotide pool are attributed to the mucA inactivation, adjustments in the guanosine phosphate nucleotide pool are consequences of the GTP-dependent alginate production induced by the mucA inactivation. This metabolic profiling study provides new insight into carbon and energy metabolism of the alginate producer P. fluorescens.

Keywords

Pseudomonas fluorescens Metabolic profiling Mass spectrometry Alginate synthesis Anti-sigma factor MucA 

Supplementary material

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Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  • Stina K. Lien
    • 1
  • Håvard Sletta
    • 2
  • Trond E. Ellingsen
    • 2
  • Svein Valla
    • 1
  • Elon Correa
    • 3
  • Royston Goodacre
    • 3
    • 4
  • Kai Vernstad
    • 2
  • Sven Even Finborud Borgos
    • 1
    • 2
  • Per Bruheim
    • 1
  1. 1.Department of BiotechnologyNorwegian University of Science and TechnologyTrondheimNorway
  2. 2.Department of BiotechnologySINTEF Materials and ChemistryTrondheimNorway
  3. 3.School of Chemistry, Manchester Institute of BiotechnologyUniversity of ManchesterManchesterUK
  4. 4.Manchester Centre for Integrative Systems Biology, Manchester Institute of BiotechnologyUniversity of ManchesterManchesterUK

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