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Mapping global effects of the anti-sigma factor MucA in Pseudomonas fluorescens SBW25 through genome-scale metabolic modeling

Borgos, Sven Even F.; Bordel, Sergio; Sletta, Håvard; Ertesvåg, Helga; Jakobsen, Øyvind Mejdell; Bruheim, Per; Ellingsen, Trond Erling; Nielsen, Jens; Valla, Svein
Journal article, Peer reviewed
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URI
http://hdl.handle.net/11250/2358401
Date
2013
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  • Institutt for bioteknologi og matvitenskap [910]
  • Publikasjoner fra CRIStin - NTNU [21869]
Original version
BMC Systems Biology 2013, 7(19):1-15   10.1186/1752-0509-7-19
Abstract
Background: Alginate is an industrially important polysaccharide, currently produced commercially by harvesting of

marine brown sea-weeds. The polymer is also synthesized as an exo-polysaccharide by bacteria belonging to the

genera Pseudomonas and Azotobacter, and these organisms may represent an alternative alginate source in the

future. The current work describes an attempt to rationally develop a biological system tuned for very high levels of

alginate production, based on a fundamental understanding of the system through metabolic modeling supported

by transcriptomics studies and carefully controlled fermentations.

Results: Alginate biosynthesis in Pseudomonas fluorescens was studied in a genomics perspective, using an alginate

over-producing strain carrying a mutation in the anti-sigma factor gene mucA. Cells were cultivated in chemostats

under nitrogen limitation on fructose or glycerol as carbon sources, and cell mass, growth rate, sugar uptake,

alginate and CO2 production were monitored. In addition a genome scale metabolic model was constructed and

samples were collected for transcriptome analyses. The analyses show that polymer production operates in a close

to optimal way with respect to stoichiometric utilization of the carbon source and that the cells increase the uptake

of carbon source to compensate for the additional needs following from alginate synthesis. The transcriptome

studies show that in the presence of the mucA mutation, the alg operon is upregulated together with genes

involved in energy generation, genes on both sides of the succinate node of the TCA cycle and genes encoding

ribosomal and other translation-related proteins. Strains expressing a functional MucA protein (no alginate

production) synthesize cellular biomass in an inefficient way, apparently due to a cycle that involves oxidation of

NADPH without ATP production. The results of this study indicate that the most efficient way of using a mucA

mutant as a cell factory for alginate production would be to use non-growing conditions and nitrogen deprivation.

Conclusions: The insights gained in this study should be very useful for a future efficient production of microbial

alginates.
Publisher
BioMed Central
Journal
BMC Systems Biology

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